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Sample records for early larval zebrafish

  1. The behavior of larval zebrafish reveals stressor-mediated anorexia during early vertebrate development

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    De Marco, Rodrigo J.; Groneberg, Antonia H.; Yeh, Chen-Min; Treviño, Mario; Ryu, Soojin

    2014-01-01

    The relationship between stress and food consumption has been well documented in adults but less so in developing vertebrates. Here we demonstrate that an encounter with a stressor can suppress food consumption in larval zebrafish. Furthermore, we provide indication that food intake suppression cannot be accounted for by changes in locomotion, oxygen consumption and visual responses, as they remain unaffected after exposure to a potent stressor. We also show that feeding reoccurs when basal levels of cortisol (stress hormone in humans and teleosts) are re-established. The results present evidence that the onset of stress can switch off the drive for feeding very early in vertebrate development, and add a novel endpoint for analyses of metabolic and behavioral disorders in an organism suitable for high-throughput genetics and non-invasive brain imaging. PMID:25368561

  2. A zebrafish larval model reveals early tissue-specific innate immune responses to Mucor circinelloides.

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    Voelz, Kerstin; Gratacap, Remi L; Wheeler, Robert T

    2015-11-01

    Mucormycosis is an emerging fungal infection that is clinically difficult to manage, with increasing incidence and extremely high mortality rates. Individuals with diabetes, suppressed immunity or traumatic injury are at increased risk of developing disease. These individuals often present with defects in phagocytic effector cell function. Research using mammalian models and phagocytic effector cell lines has attempted to decipher the importance of the innate immune system in host defence against mucormycosis. However, these model systems have not been satisfactory for direct analysis of the interaction between innate immune effector cells and infectious sporangiospores in vivo. Here, we report the first real-time in vivo analysis of the early innate immune response to mucormycete infection using a whole-animal zebrafish larval model system. We identified differential host susceptibility, dependent on the site of infection (hindbrain ventricle and swim bladder), as well as differential functions of the two major phagocyte effector cell types in response to viable and non-viable spores. Larval susceptibility to mucormycete spore infection was increased upon immunosuppressant treatment. We showed for the first time that macrophages and neutrophils were readily recruited in vivo to the site of infection in an intact host and that spore phagocytosis can be observed in real-time in vivo. While exploring innate immune effector recruitment dynamics, we discovered the formation of phagocyte clusters in response to fungal spores that potentially play a role in fungal spore dissemination. Spores failed to activate pro-inflammatory gene expression by 6 h post-infection in both infection models. After 24 h, induction of a pro-inflammatory response was observed only in hindbrain ventricle infections. Only a weak pro-inflammatory response was initiated after spore injection into the swim bladder during the same time frame. In the future, the zebrafish larva as a live whole

  3. Cardiac and Metabolic Physiology of Early Larval Zebrafish (Danio rerio) Reflects Parental Swimming Stamina.

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    Gore, Matthew; Burggren, Warren W

    2012-01-01

    Swimming stamina in adult fish is heritable, it is unknown if inherited traits that support enhanced swimming stamina in offspring appear only in juveniles and/or adults, or if these traits actually appear earlier in the morphologically quite different larvae. To answer this question, mature adult zebrafish (Danio rerio) were subjected to a swimming performance test that allowed separation into low swimming stamina or high swimming stamina groups. Adults were then bred within their own performance groups. Larval offspring from each of the two groups, designated high (L(HSD)) and low stamina-derived larvae (L(LSD)), were then reared at 27°C in aerated water (21% O(2)). Routine (f(H),r) and active (f(H),a) heart rate, and routine [Formula: see text] and active [Formula: see text] mass-specific oxygen consumption were recorded from 5 days post fertilization (dpf) through 21 dpf, and gross cost of transport and factorial aerobic metabolic scope were derived from [Formula: see text] measurements. Heart rate generally ranged between 150 and 225 bpm in both L(HSD) and L(LSD) populations. However, significant (P stamina in adult parents also appear in their larval offspring well before attainment of juvenile or adult features.

  4. Cardiac and metabolic physiology of early larval zebrafish (Danio rerio reflects parental swimming stamina

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    Warren W Burggren

    2012-02-01

    Full Text Available Swimming stamina in adult fish is heritable, it is unknown if inherited traits that support enhanced swimming stamina in offspring appear only in juveniles and/or adults, or if these traits actually appear earlier in the morphologically quite different larvae. To answer this question, mature adult zebrafish (Danio rerio were subjected to a swimming performance test that allowed separation into low swimming stamina or high swimming stamina groups. Adults were then bred within their own performance groups. Larval offspring from each of the two groups, designated high (LHSD and low stamina-derived larvae (LLSD, were then reared at 27°C in aerated water (21% O2. Routine (fH,r and active (fH,a heart rate, and routine (M.O2,r and active (M.O2,a mass-specific oxygen consumption were recorded from 5 days post fertilization (dpf through 21 dpf, and gross cost of transport and factorial aerobic metabolic scope were derived from M.O2 measurements. Heart rate generally ranged between 150 and 225 b•min-1 in both LHSD and LLSD populations. However, significant (P<0.05 differences existed between the LLSD and LHSD populations at 5 and 14 dpf in fH,r and at days 10 and 15 dpf in fH,a. M.O2,r was 0.04 to 0.32 μmol•mg-1•hr-1, while M.O2,a was 0.2 to 1.2 μmol•mg-1•hr-1. Significant (P<0.05 differences between the LLSD and LHSD populations in M.O2,r occurred at 7, 10 and 21 dpf and in M.O2,a at 7 dpf. Gross cost of transport was ~6-10 µmol O2 . µg-1 . m-1 at 5 dpf, peaking at 14-19 µmol O2 . µg-1 . m-1 at 7-10 dpf, before falling again to 5-6 µmol O2 . µg-1 . m-1 at 21 dpf, with gross cost of transport significantly higher in the LLSD population at 7 dpf. Collectively, these data indicate that inherited physiological differences contributing to enhanced stamina in adult parents appear in their larval offspring well before attainment of juvenile or adult features.

  5. Adaptive locomotor behavior in larval zebrafish.

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    Portugues, Ruben; Engert, Florian

    2011-01-01

    In this study we report that larval zebrafish display adaptive locomotor output that can be driven by unexpected visual feedback. We develop a new assay that addresses visuomotor integration in restrained larval zebrafish. The assay involves a closed-loop environment in which the visual feedback a larva receives depends on its own motor output in a way that resembles freely swimming conditions. The experimenter can control the gain of this closed feedback loop, so that following a given motor output the larva experiences more or less visual feedback depending on whether the gain is high or low. We show that increases and decreases in this gain setting result in adaptive changes in behavior that lead to a generalized decrease or increase of motor output, respectively. Our behavioral analysis shows that both the duration and tail beat frequency of individual swim bouts can be modified, as well as the frequency with which bouts are elicited. These changes can be implemented rapidly, following an exposure to a new gain of just 175 ms. In addition, modifications in some behavioral parameters accumulate over tens of seconds and effects last for at least 30 s from trial to trial. These results suggest that larvae establish an internal representation of the visual feedback expected from a given motor output and that the behavioral modifications are driven by an error signal that arises from the discrepancy between this expectation and the actual visual feedback. The assay we develop presents a unique possibility for studying visuomotor integration using imaging techniques available in the larval zebrafish.

  6. GROWTH AND BEHAVIOR OF LARVAL ZEBRAFISH Danio ...

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    Because Zebrafish (Danio rerio) have become a popular and important model for scientific research, the capability to rear larval zebrafish to adulthood is of great importance. Recently research examining the effects of diet (live versus processed) have been published. In the current study we examined whether the larvae can be reared on a processed diet alone, live food alone, or the combination while maintaining normal locomotor behavior, and acceptable survival, length and weight at 14 dpf in a static system. A 14 day feeding trial was conducted in glass crystallizing dishes containing 500 ml of 4 ppt Instant Ocean. On day 0 pdf 450 embryos were selected as potential study subjects and placed in a 26○C incubator on a 14:10 (light:dark) light cycle. At 4 dpf 120 normally developing embryos were selected per treatment and divided into 3 bowls of 40 embryos (for an n=3 per treatment; 9 bowls total). Treatment groups were: G (Gemma Micro 75 only), R (L-type marine rotifers (Brachionus plicatilis) only) or B (Gemma and rotifers). Growth (length), survival, water quality and rotifer density were monitored on days 5-14. On day 14, weight of larva in each bowl was measured and 8 larva per bowl were selected for use in locomotor testing. This behavior paradigm tests individual larval zebrafish under both light and dark conditions in a 24-well plate.After 14 dpf, survival among the groups was not different (92-98%). By days 7 -14 R and B larvae were ~2X longer

  7. The neural basis of visual behaviors in the larval zebrafish.

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    Portugues, Ruben; Engert, Florian

    2009-12-01

    We review visually guided behaviors in larval zebrafish and summarise what is known about the neural processing that results in these behaviors, paying particular attention to the progress made in the last 2 years. Using the examples of the optokinetic reflex, the optomotor response, prey tracking and the visual startle response, we illustrate how the larval zebrafish presents us with a very promising model vertebrate system that allows neurocientists to integrate functional and behavioral studies and from which we can expect illuminating insights in the near future. Copyright 2009 Elsevier Ltd. All rights reserved.

  8. Hypothalamic Projections to the Optic Tectum in Larval Zebrafish

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    Heap, Lucy A.; Vanwalleghem, Gilles C.; Thompson, Andrew W.; Favre-Bulle, Itia; Rubinsztein-Dunlop, Halina; Scott, Ethan K.

    2018-01-01

    The optic tectum of larval zebrafish is an important model for understanding visual processing in vertebrates. The tectum has been traditionally viewed as dominantly visual, with a majority of studies focusing on the processes by which tectal circuits receive and process retinally-derived visual information. Recently, a handful of studies have shown a much more complex role for the optic tectum in larval zebrafish, and anatomical and functional data from these studies suggest that this role extends beyond the visual system, and beyond the processing of exclusively retinal inputs. Consistent with this evolving view of the tectum, we have used a Gal4 enhancer trap line to identify direct projections from rostral hypothalamus (RH) to the tectal neuropil of larval zebrafish. These projections ramify within the deepest laminae of the tectal neuropil, the stratum album centrale (SAC)/stratum griseum periventriculare (SPV), and also innervate strata distinct from those innervated by retinal projections. Using optogenetic stimulation of the hypothalamic projection neurons paired with calcium imaging in the tectum, we find rebound firing in tectal neurons consistent with hypothalamic inhibitory input. Our results suggest that tectal processing in larval zebrafish is modulated by hypothalamic inhibitory inputs to the deep tectal neuropil. PMID:29403362

  9. Hypothalamic Projections to the Optic Tectum in Larval Zebrafish

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    Lucy A. Heap

    2018-01-01

    Full Text Available The optic tectum of larval zebrafish is an important model for understanding visual processing in vertebrates. The tectum has been traditionally viewed as dominantly visual, with a majority of studies focusing on the processes by which tectal circuits receive and process retinally-derived visual information. Recently, a handful of studies have shown a much more complex role for the optic tectum in larval zebrafish, and anatomical and functional data from these studies suggest that this role extends beyond the visual system, and beyond the processing of exclusively retinal inputs. Consistent with this evolving view of the tectum, we have used a Gal4 enhancer trap line to identify direct projections from rostral hypothalamus (RH to the tectal neuropil of larval zebrafish. These projections ramify within the deepest laminae of the tectal neuropil, the stratum album centrale (SAC/stratum griseum periventriculare (SPV, and also innervate strata distinct from those innervated by retinal projections. Using optogenetic stimulation of the hypothalamic projection neurons paired with calcium imaging in the tectum, we find rebound firing in tectal neurons consistent with hypothalamic inhibitory input. Our results suggest that tectal processing in larval zebrafish is modulated by hypothalamic inhibitory inputs to the deep tectal neuropil.

  10. Lipid Uptake, Metabolism, and Transport in the Larval Zebrafish

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    Vanessa H. Quinlivan

    2017-11-01

    Full Text Available The developing zebrafish is a well-established model system for studies of energy metabolism, and is amenable to genetic, physiological, and biochemical approaches. For the first 5 days of life, nutrients are absorbed from its endogenous maternally deposited yolk. At 5 days post-fertilization, the yolk is exhausted and the larva has a functional digestive system including intestine, liver, gallbladder, pancreas, and intestinal microbiota. The transparency of the larval zebrafish, and the genetic and physiological similarity of its digestive system to that of mammals make it a promising system in which to address questions of energy homeostasis relevant to human health. For example, apolipoprotein expression and function is similar in zebrafish and mammals, and transgenic animals may be used to examine both the transport of lipid from yolk to body in the embryo, and the trafficking of dietary lipids in the larva. Additionally, despite the identification of many fatty acid and lipid transport proteins expressed by vertebrates, the cell biological processes that mediate the transport of dietary lipids from the intestinal lumen to the interior of enterocytes remain to be elucidated. Genetic tractability and amenability to live imaging and a range of biochemical methods make the larval zebrafish an ideal model in which to address open questions in the field of lipid transport, energy homeostasis, and nutrient metabolism.

  11. Investigating the embryo/larval toxic and genotoxic effects of {gamma} irradiation on zebrafish eggs

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    Simon, O., E-mail: olivier.simon@irsn.fr [Laboratoire de Radioecologie et d' Ecotoxicologie, Institut de Radioprotection et de Surete Nucleaire, Cadarache, Bat 186, BP3, 13115 Saint-Paul-lez-Durance Cedex (France); Massarin, S. [Laboratoire de Modelisation Environnementale, Institut de Radioprotection et de Surete Nucleaire, Cadarache, Bat 159, BP3, 13115 Saint-Paul-lez-Durance Cedex (France); Coppin, F. [Laboratoire de Radioecologie et d' Ecotoxicologie, Institut de Radioprotection et de Surete Nucleaire, Cadarache, Bat 186, BP3, 13115 Saint-Paul-lez-Durance Cedex (France); Hinton, T.G. [Service d' Etude du Comportement des Radionucleides dans les Ecosystemes, Institut de Radioprotection et de Surete Nucleaire, Cadarache, Bat 159, BP3, 13115 Saint-Paul-lez-Durance Cedex (France); Gilbin, R. [Laboratoire de Radioecologie et d' Ecotoxicologie, Institut de Radioprotection et de Surete Nucleaire, Cadarache, Bat 186, BP3, 13115 Saint-Paul-lez-Durance Cedex (France)

    2011-11-15

    Eggs/larval of freshwater fish (Danio rerio) were exposed to low dose rates of external gamma radiation (from 1 to 1000 mGy d{sup -1}) over a 20-day period, with the objective of testing the appropriateness of the 10 mGy d{sup -1} guideline suggested by the IAEA. The present study examines different endpoints, mortality and hatching time and success of embryos as well as the genotoxicity of {gamma}-irradiations (after 48 h). The 20-day embryo-larval bioassay showed an enhanced larval resistance to starvation after chronic exposure to {gamma} irradiation (from low 1 mGy d{sup -1} to high dose rate 1000 mGy d{sup -1}) and an acceleration in hatching time. Gamma irradiation led to increased genotoxic damage Ito zebrafish egg (40-50% DNA in tail in Comet assay) from the lowest dose rate (1 mGy d{sup -1}). Possible mechanisms of {gamma} radiotoxicity and implications for radioprotection are discussed. - Highlights: > Relevant information on the {gamma} radiation impact on early life stage biota is scarce. > The eggs of zebrafish Danio rerio were selected as biological model. > We test the appropriateness of the 10 mGy d{sup -1} guideline (IAEA). > We observed effects measured at individual levels (starvation, hatching time). > Chronic gamma irradiation led to increased genotoxic damage to zebrafish egg. > {gamma} radiotoxicity mechanisms and implications for radioprotection are discussed.

  12. Properties of the Visible Light Phototaxis and UV Avoidance Behaviors in the Larval Zebrafish.

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    Guggiana-Nilo, Drago A; Engert, Florian

    2016-01-01

    For many organisms, color is an essential source of information from visual scenes. The larval zebrafish has the potential to be a model for the study of this topic, given its tetrachromatic retina and high dependence on vision. In this study we took a step toward understanding how the larval zebrafish might use color sensing. To this end, we used a projector-based paradigm to force a choice of a color stimulus at every turn of the larva. The stimuli used spanned most of the larval spectral range, including activation of its Ultraviolet (UV) cone, which has not been described behaviorally before. We found that zebrafish larvae swim toward visible wavelengths (>400 nm) when choosing between them and darkness, as has been reported with white light. However, when presented with UV light and darkness zebrafish show an intensity dependent avoidance behavior. This UV avoidance does not interact cooperatively with phototaxis toward longer wavelengths, but can compete against it in an intensity dependent manner. Finally, we show that the avoidance behavior depends on the presence of eyes with functional UV cones. These findings open future avenues for studying the neural circuits that underlie color sensing in the larval zebrafish.

  13. Short-term developmental effects and potential mechanisms of azoxystrobin in larval and adult zebrafish (Danio rerio).

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    Cao, Fangjie; Wu, Peizhuo; Huang, Lan; Li, Hui; Qian, Le; Pang, Sen; Qiu, Lihong

    2018-05-01

    Previous study indicated that azoxystrobin had high acute toxicity to zebrafish, and larval zebrafish were more sensitive to azoxystrobin than adult zebrafish. The objective of the present study was to investigate short-term developmental effects and potential mechanisms of azoxystrobin in larval and adult zebrafish. After zebrafish embryos and adults were exposed to 0.01, 0.05 and 0.20 mg/L azoxystrobin (equal to 25, 124 and 496 nM azoxystrobin, respectively) for 8 days, the lethal effect, physiological responses, liver histology, mitochondrial ultrastructure, and expression alteration of genes related to mitochondrial respiration, oxidative stress, cell apoptosis and innate immune response were determined. The results showed that there was no significant effect on larval and adult zebrafish after exposure to 0.01 mg/L azoxystrobin. However, increased ROS, MDA concentration and il1b in larval zebrafish, as well as increased il1b, il8 and cxcl-c1c in adult zebrafish were induced after exposure to 0.05 mg/L azoxystrobin. Reduced mitochondrial complex III activity and ATP concentration, increased SOD activity, ROS and MDA concentration, decreased cytb, as well as increased sod1, sod2, cat, il1b, il8 and cxcl-c1c were observed both in larval and adult zebrafish after exposure to 0.20 mg/L azoxystrobin; meanwhile, increased p53, bax, apaf1 and casp9, alteration of liver histology and mitochondrial ultrastructure in larval zebrafish, and alteration of mitochondrial ultrastructure in adult zebrafish were also induced. The results demonstrated that azoxytrobin induced short-term developmental effects on larval zebrafish and adult zebrafish, including mitochondrial dysfunction, oxidative stress, cell apoptosis and innate immune response. Statistical analysis indicated that azoxystrobin induced more negative effects on larval zebrafish, which might be the reason for the differences of developmental toxicity between larval and adult zebrafish caused by

  14. Correlating Whole Brain Neural Activity with Behavior in Head-Fixed Larval Zebrafish.

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    Orger, Michael B; Portugues, Ruben

    2016-01-01

    We present a protocol to combine behavioral recording and imaging using 2-photon laser-scanning microscopy in head-fixed larval zebrafish that express a genetically encoded calcium indicator. The steps involve restraining the larva in agarose, setting up optics that allow projection of a visual stimulus and infrared illumination to monitor behavior, and analysis of the neuronal and behavioral data.

  15. Optogenetics in a transparent animal: circuit function in the larval zebrafish.

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    Portugues, Ruben; Severi, Kristen E; Wyart, Claire; Ahrens, Misha B

    2013-02-01

    Optogenetic tools can be used to manipulate neuronal activity in a reversible and specific manner. In recent years, such methods have been applied to uncover causal relationships between activity in specified neuronal circuits and behavior in the larval zebrafish. In this small, transparent, genetic model organism, noninvasive manipulation and monitoring of neuronal activity with light is possible throughout the nervous system. Here we review recent work in which these new tools have been applied to zebrafish, and discuss some of the existing challenges of these approaches. Copyright © 2012. Published by Elsevier Ltd.

  16. GROWTH AND BEHAVIOR OF LARVAL ZEBRAFISH Danio rerio FED A PROCESSED DIET, LIVE FOOD, OR THE COMBINATION

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    Because Zebrafish (Danio rerio) have become a popular and important model for scientific research, the capability to rear larval zebrafish to adulthood is of great importance. Recently research examining the effects of diet (live versus processed) have been published. In the cu...

  17. 3D Finite Element Electrical Model of Larval Zebrafish ECG Signals

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    Crowcombe, James; Dhillon, Sundeep Singh; Hurst, Rhiannon Mary; Egginton, Stuart; Müller, Ferenc; Sík, Attila; Tarte, Edward

    2016-01-01

    Assessment of heart function in zebrafish larvae using electrocardiography (ECG) is a potentially useful tool in developing cardiac treatments and the assessment of drug therapies. In order to better understand how a measured ECG waveform is related to the structure of the heart, its position within the larva and the position of the electrodes, a 3D model of a 3 days post fertilisation (dpf) larval zebrafish was developed to simulate cardiac electrical activity and investigate the voltage distribution throughout the body. The geometry consisted of two main components; the zebrafish body was modelled as a homogeneous volume, while the heart was split into five distinct regions (sinoatrial region, atrial wall, atrioventricular band, ventricular wall and heart chambers). Similarly, the electrical model consisted of two parts with the body described by Laplace’s equation and the heart using a bidomain ionic model based upon the Fitzhugh-Nagumo equations. Each region of the heart was differentiated by action potential (AP) parameters and activation wave conduction velocities, which were fitted and scaled based on previously published experimental results. ECG measurements in vivo at different electrode recording positions were then compared to the model results. The model was able to simulate action potentials, wave propagation and all the major features (P wave, R wave, T wave) of the ECG, as well as polarity of the peaks observed at each position. This model was based upon our current understanding of the structure of the normal zebrafish larval heart. Further development would enable us to incorporate features associated with the diseased heart and hence assist in the interpretation of larval zebrafish ECGs in these conditions. PMID:27824910

  18. Exploratory behaviour in the open field test adapted for larval zebrafish: impact of environmental complexity.

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    Ahmad, Farooq; Richardson, Michael K

    2013-01-01

    This study aimed to develop and characterize a novel (standard) open field test adapted for larval zebrafish. We also developed and characterized a variant of the same assay consisting of a colour-enriched open field; this was used to assess the impact of environmental complexity on patterns of exploratory behaviours as well to determine natural colour preference/avoidance. We report the following main findings: (1) zebrafish larvae display characteristic patterns of exploratory behaviours in the standard open field, such as thigmotaxis/centre avoidance; (2) environmental complexity (i.e. presence of colours) differentially affects patterns of exploratory behaviours and greatly attenuates natural zone preference; (3) larvae displayed the ability to discriminate colours. As reported previously in adult zebrafish, larvae showed avoidance towards blue and black; however, in contrast to the reported adult behaviour, larvae displayed avoidance towards red. Avoidance towards yellow and preference for green and orange are shown for the first time, (4) compared to standard open field tests, exposure to the colour-enriched open field resulted in an enhanced expression of anxiety-like behaviours. To conclude, we not only developed and adapted a traditional rodent behavioural assay that serves as a gold standard in preclinical drug screening, but we also provide a version of the same test that affords the possibility to investigate the impact of environmental stress on behaviour in larval zebrafish while representing the first test for assessment of natural colour preference/avoidance in larval zebrafish. In the future, these assays will improve preclinical drug screening methodologies towards the goal to uncover novel drugs. This article is part of a Special Issue entitled: insert SI title. Copyright © 2012 Elsevier B.V. All rights reserved.

  19. Screening in larval zebrafish reveals tissue-specific distribution of fifteen fluorescent compounds

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    Yuxiao Yao

    2017-09-01

    Full Text Available The zebrafish is a prominent vertebrate model for low-cost in vivo whole organism screening. In our recent screening of the distribution patterns of fluorescent compounds in live zebrafish larvae, fifteen compounds with tissue-specific distributions were identified. Several compounds were observed to accumulate in tissues where they were reported to induce side-effects, and compounds with similar structures tended to be enriched in the same tissues, with minor differences. In particular, we found three novel red fluorescent bone-staining dyes: purpurin, lucidin and 3-hydroxy-morindone; purpurin can effectively label bones in both larval and adult zebrafish, as well as in postnatal mice, without significantly affecting bone mass and density. Moreover, two structurally similar chemotherapeutic compounds, doxorubicin and epirubicin, were observed to have distinct distribution preferences in zebrafish. Epirubicin maintained a relatively higher concentration in the liver, and performed better in inhibiting hepatic hyperplasia caused by the over-expression of krasG12V. In total, our study suggests that the transparent zebrafish larvae serve as valuable tools for identifying tissue-specific distributions of fluorescent compounds.

  20. Effects of larval-juvenile treatment with perchlorate and co-treatment with thyroxine on zebrafish sex ratios

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    Mukhi, S.; Torres, L.; Patino, R.

    2007-01-01

    The objective of this study was to determine the effect of larval-juvenile exposure to perchlorate, a thyroid hormone synthesis inhibitor, on the establishment of gonadal sex ratios in zebrafish. Zebrafish were exposed to untreated water or water containing perchlorate at 100 or 250 ppm for a period of 30 days starting at 3 days postfertilization (dpf). Recovery treatments consisted of a combination of perchlorate and exogenous thyroxine (T4; 10 nM). Thyroid histology was assessed at the end of the treatment period (33 dpf), and gonadal histology and sex ratios were determined in fish that were allowed an additional 10-day period of growth in untreated water. As expected, exposure to perchlorate caused changes in thyroid histology consistent with hypothyroidism and these effects were reversed by co-treatment with exogenous T4. Perchlorate did not affect fish survival but co-treatment with T4 induced higher mortality. However, relative to the corresponding perchlorate concentration, co-treatment with T4 caused increased mortality only at a perchlorate concentration of 100 ppm. Perchlorate alone or in the presence of T4 suppressed body length at 43 dpf relative to control values. Perchlorate exposure skewed the sex ratio toward female in a concentration-dependent manner, and co-treatment with T4 not only blocked the feminizing effect of perchlorate but also overcompensated by skewing the sex ratio towards male. Moreover, co-treatment with T4 advanced the onset of spermatogenesis in males. There was no clear association between sex ratios and larval survival or growth. We conclude that endogenous thyroid hormone plays a role in the establishment of gonadal sex phenotype during early development in zebrafish. ?? 2006 Elsevier Inc. All rights reserved.

  1. Spinal interneurons differentiate sequentially from those driving the fastest swimming movements in larval zebrafish to those driving the slowest ones.

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    McLean, David L; Fetcho, Joseph R

    2009-10-28

    Studies of neuronal networks have revealed few general principles that link patterns of development with later functional roles. While investigating the neural control of movements, we recently discovered a topographic map in the spinal cord of larval zebrafish that relates the position of motoneurons and interneurons to their order of recruitment during swimming. Here, we show that the map reflects an orderly pattern of differentiation of neurons driving different movements. First, we use high-speed filming to show that large-amplitude swimming movements with bending along much of the body appear first, with smaller, regional swimming movements emerging later. Next, using whole-cell patch recordings, we demonstrate that the excitatory circuits that drive large-amplitude, fast swimming movements at larval stages are present and functional early on in embryos. Finally, we systematically assess the orderly emergence of spinal circuits according to swimming speed using transgenic fish expressing the photoconvertible protein Kaede to track neuronal differentiation in vivo. We conclude that a simple principle governs the development of spinal networks in which the neurons driving the fastest, most powerful swimming in larvae develop first with ones that drive increasingly weaker and slower larval movements layered on over time. Because the neurons are arranged by time of differentiation in the spinal cord, the result is a topographic map that represents the speed/strength of movements at which neurons are recruited and the temporal emergence of networks. This pattern may represent a general feature of neuronal network development throughout the brain and spinal cord.

  2. Masking of a circadian behavior in larval zebrafish involves the thalamo-habenula pathway.

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    Lin, Qian; Jesuthasan, Suresh

    2017-06-22

    Changes in illumination can rapidly influence behavior that is normally controlled by the circadian clock. This effect is termed masking. In mice, masking requires melanopsin-expressing retinal ganglion cells that detect blue light and project to the thalamus. It is not known whether masking is wavelength-dependent in other vertebrates, nor is it known whether the thalamus is also involved or how it influences masking. Here, we address these questions in zebrafish. We find that diel vertical migration, a circadian behavior in larval zebrafish, is effectively triggered by blue, but not by red light. Two-photon calcium imaging reveals that a thalamic nucleus and a downstream structure, the habenula, have a sustained response to blue but not to red light. Lesioning the habenula reduces light-evoked climbing. These data suggest that the thalamo-habenula pathway is involved in the ability of blue light to influence a circadian behavior.

  3. Methods for culturing saltwater rotifers (Brachionus plicatilis) for rearing larval zebrafish.

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    Lawrence, Christian; Sanders, Erik; Henry, Eric

    2012-09-01

    The saltwater rotifer, Brachionus plicatilis, is widely used in the aquaculture industry as a prey item for first-feeding fishes due to its ease of culture, small size, rapid reproductive rate, and amenability to enrichment with nutrients. Despite the distinct advantages of this approach, rotifers have only been sporadically utilized for rearing larval zebrafish, primarily because of the common misconception that maintaining cultures of rotifers is difficult and excessively time-consuming. Here we present simple methods for maintaining continuous cultures of rotifers capable of supporting even the very largest zebrafish aquaculture facility, with minimal investments in materials, time, labor, and space. Examples of the methods' application in one large, existing facility is provided, and troubleshooting of common problems is discussed.

  4. Quantification of larval zebrafish motor function in multiwell plates using open-source MATLAB applications.

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    Zhou, Yangzhong; Cattley, Richard T; Cario, Clinton L; Bai, Qing; Burton, Edward A

    2014-07-01

    This article describes a method to quantify the movements of larval zebrafish in multiwell plates, using the open-source MATLAB applications LSRtrack and LSRanalyze. The protocol comprises four stages: generation of high-quality, flatly illuminated video recordings with exposure settings that facilitate object recognition; analysis of the resulting recordings using tools provided in LSRtrack to optimize tracking accuracy and motion detection; analysis of tracking data using LSRanalyze or custom MATLAB scripts; and implementation of validation controls. The method is reliable, automated and flexible, requires plate format suitable for high-throughput applications.

  5. Restoration of anatomical continuity after spinal cord transection depends on Wnt/β-catenin signaling in larval zebrafish

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    Daniel Wehner

    2018-02-01

    Full Text Available This data article contains descriptive and experimental data on spinal cord regeneration in larval zebrafish and its dependence on Wnt/β-catenin signaling. Analyzing spread of intraspinally injected fluorescent dextran showed that anatomical continuity is rapidly restored after complete spinal cord transection. Pharmacological interference with Wnt/β-catenin signaling (IWR-1 impaired restoration of spinal continuity. For further details and experimental findings please refer to the research article by Wehner et al. Wnt signaling controls pro-regenerative Collagen XII in functional spinal cord regeneration in zebrafish (Wehner et al., 2017 [1]. Keywords: Wnt, Beta-catenin, Regeneration, Spinal cord, Zebrafish

  6. Whole-field visual motion drives swimming in larval zebrafish via a stochastic process.

    Science.gov (United States)

    Portugues, Ruben; Haesemeyer, Martin; Blum, Mirella L; Engert, Florian

    2015-05-01

    Caudo-rostral whole-field visual motion elicits forward locomotion in many organisms, including larval zebrafish. Here, we investigate the dependence on the latency to initiate this forward swimming as a function of the speed of the visual motion. We show that latency is highly dependent on speed for slow speeds (1.5 s, which is much longer than neuronal transduction processes. What mechanisms underlie these long latencies? We propose two alternative, biologically inspired models that could account for this latency to initiate swimming: an integrate and fire model, which is history dependent, and a stochastic Poisson model, which has no history dependence. We use these models to predict the behavior of larvae when presented with whole-field motion of varying speed and find that the stochastic process shows better agreement with the experimental data. Finally, we discuss possible neuronal implementations of these models. © 2015. Published by The Company of Biologists Ltd.

  7. Prey capture behaviour evoked by simple visual stimuli in larval zebrafish

    Directory of Open Access Journals (Sweden)

    Isaac Henry Bianco

    2011-12-01

    Full Text Available Understanding how the nervous system recognises salient stimuli in the environ- ment and selects and executes the appropriate behavioural responses is a fundamen- tal question in systems neuroscience. To facilitate the neuroethological study of visually-guided behaviour in larval zebrafish, we developed virtual reality assays in which precisely controlled visual cues can be presented to larvae whilst their behaviour is automatically monitored using machine-vision algorithms. Freely swimming larvae responded to moving stimuli in a size-dependent manner: they directed multiple low amplitude orienting turns (∼ 20◦ towards small moving spots (1◦ but reacted to larger spots (10◦ with high-amplitude aversive turns (∼ 60◦. The tracking of small spots led us to examine how larvae respond to prey during hunting routines. By analysing movie sequences of larvae hunting parame- cia, we discovered that all prey capture routines commence with eye convergence and larvae maintain their eyes in a highly converged position for the duration of the prey-tracking and capture swim phases. We adapted our virtual reality assay to deliver artificial visual cues to partially restrained larvae and found that small moving spots evoked convergent eye movements and J-turns of the tail, which are defining features of natural hunting. We propose that eye convergence represents the engagement of a predatory mode of behaviour in larval fish and serves to increase the region of binocular visual space to enable stereoscopic targeting of prey.

  8. Quercetin, a natural product supplement, impairs mitochondrial bioenergetics and locomotor behavior in larval zebrafish (Danio rerio).

    Science.gov (United States)

    Zhang, Ji-Liang; Laurence Souders, Christopher; Denslow, Nancy D; Martyniuk, Christopher J

    2017-07-15

    Quercetin is a natural product that is sold as a supplement in health food stores. While there are reported benefits for this flavonoid as a dietary supplement due to antioxidant properties, the full scope of its biological interactions has not been fully addressed. To learn more about the mechanisms of action related to quercetin, we exposed zebrafish (Danio rerio) embryos to 1 and 10μg/L quercetin for 96h starting at 3h post fertilization. Quercetin up to 10μg/L did not induce significant mortality in developing fish, but did increase prevalence of an upward-curved dorsal plane in hatched larvae. To determine whether this developmental defect was potentially related to mitochondrial bioenergetics during development, we measured oxygen consumption rate in whole embryos following a 24-hour exposure to quercetin. Basal mitochondrial and ATP-linked respiration were decreased at 1 and 10μg/L quercetin, and maximal respiration was decreased at 10μg/L quercetin, suggesting that quercetin impairs mitochondrial bioenergetics. This is proposed to be related to the deformities observed during development. Due to the fact that ATP production was affected by quercetin, larval behaviors related to locomotion were investigated, as well as transcriptional responses of six myogenesis transcripts. Quercetin at 10μg/L significantly reduced the swimming velocity of zebrafish larvae. The expression levels of both myostatin A (mstna) and myogenic differentiation (myoD) were also altered by quercetin. Mstna, an inhibitory factor for myogenesis, was significantly increased at 1μg/L quercetin exposure, while myoD, a stimulatory factor for myogenesis, was significantly increased at 10μg/L quercetin exposure. There were no changes in transcripts related to apoptosis (bcl2, bax, casp3, casp7), but we did observe a decrease in mRNA levels for catalase (cat) in fish exposed to each dose, supporting an oxidative stress response. Our data support the hypothesis that quercetin may affect

  9. Direction selectivity in the larval zebrafish tectum is mediated by asymmetric inhibition

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    Abhinav eGrama

    2012-09-01

    Full Text Available The extraction of the direction of motion is an important computation performed by many sensory systems and in particular, the mechanism by which direction selective ganglion cells (DS-RGCs in the retina acquire their selective properties, has been studied extensively. However, whether DS-RGCs simply relay this information to downstream areas or whether additional and potentially de-novo processing occurs in these recipient structures is a matter of great interest. Neurons in the larval zebrafish tectum, the largest retino-recipent area in this animal, show direction selective responses to moving visual stimuli but how these properties are acquired is still unknown. In order to study this, we first used two-photon calcium imaging to classify the population responses of tectal cells to bars moving at different speeds and in different directions. Subsequently, we performed in-vivo whole cell electrophysiology on these direction selective tectal neurons and we found that their inhibitory inputs were strongly biased towards the null direction of motion, whereas the excitatory inputs showed little selectivity. In addition, we found that excitatory currents evoked by a stimulus moving in the preferred direction occurred before the inhibitory currents whereas a stimulus moving in the null direction evoked currents in the reverse temporal order. The membrane potential modulations resulting from these currents were enhanced by the spike generation mechanism to generate amplified direction selectivity in the spike output. Thus our results implicate a local inhibitory circuit in generating direction selectivity in tectal neurons.

  10. Whole-brain serial-section electron microscopy in larval zebrafish.

    Science.gov (United States)

    Hildebrand, David Grant Colburn; Cicconet, Marcelo; Torres, Russel Miguel; Choi, Woohyuk; Quan, Tran Minh; Moon, Jungmin; Wetzel, Arthur Willis; Scott Champion, Andrew; Graham, Brett Jesse; Randlett, Owen; Plummer, George Scott; Portugues, Ruben; Bianco, Isaac Henry; Saalfeld, Stephan; Baden, Alexander David; Lillaney, Kunal; Burns, Randal; Vogelstein, Joshua Tzvi; Schier, Alexander Franz; Lee, Wei-Chung Allen; Jeong, Won-Ki; Lichtman, Jeff William; Engert, Florian

    2017-05-18

    High-resolution serial-section electron microscopy (ssEM) makes it possible to investigate the dense meshwork of axons, dendrites, and synapses that form neuronal circuits. However, the imaging scale required to comprehensively reconstruct these structures is more than ten orders of magnitude smaller than the spatial extents occupied by networks of interconnected neurons, some of which span nearly the entire brain. Difficulties in generating and handling data for large volumes at nanoscale resolution have thus restricted vertebrate studies to fragments of circuits. These efforts were recently transformed by advances in computing, sample handling, and imaging techniques, but high-resolution examination of entire brains remains a challenge. Here, we present ssEM data for the complete brain of a larval zebrafish (Danio rerio) at 5.5 days post-fertilization. Our approach utilizes multiple rounds of targeted imaging at different scales to reduce acquisition time and data management requirements. The resulting dataset can be analysed to reconstruct neuronal processes, permitting us to survey all myelinated axons (the projectome). These reconstructions enable precise investigations of neuronal morphology, which reveal remarkable bilateral symmetry in myelinated reticulospinal and lateral line afferent axons. We further set the stage for whole-brain structure-function comparisons by co-registering functional reference atlases and in vivo two-photon fluorescence microscopy data from the same specimen. All obtained images and reconstructions are provided as an open-access resource.

  11. A novel mechanism for mechanosensory-based rheotaxis in larval zebrafish.

    Science.gov (United States)

    Oteiza, Pablo; Odstrcil, Iris; Lauder, George; Portugues, Ruben; Engert, Florian

    2017-07-27

    When flying or swimming, animals must adjust their own movement to compensate for displacements induced by the flow of the surrounding air or water. These flow-induced displacements can most easily be detected as visual whole-field motion with respect to the animal's frame of reference. Despite this, many aquatic animals consistently orient and swim against oncoming flows (a behaviour known as rheotaxis) even in the absence of visual cues. How animals achieve this task, and its underlying sensory basis, is still unknown. Here we show that, in the absence of visual information, larval zebrafish (Danio rerio) perform rheotaxis by using flow velocity gradients as navigational cues. We present behavioural data that support a novel algorithm based on such local velocity gradients that fish use to avoid getting dragged by flowing water. Specifically, we show that fish use their mechanosensory lateral line to first sense the curl (or vorticity) of the local velocity vector field to detect the presence of flow and, second, to measure its temporal change after swim bouts to deduce flow direction. These results reveal an elegant navigational strategy based on the sensing of flow velocity gradients and provide a comprehensive behavioural algorithm, also applicable for robotic design, that generalizes to a wide range of animal behaviours in moving fluids.

  12. Coordination of fictive motor activity in the larval zebrafish is generated by non-segmental mechanisms.

    Directory of Open Access Journals (Sweden)

    Timothy D Wiggin

    Full Text Available The cellular and network basis for most vertebrate locomotor central pattern generators (CPGs is incompletely characterized, but organizational models based on known CPG architectures have been proposed. Segmental models propose that each spinal segment contains a circuit that controls local coordination and sends longer projections to coordinate activity between segments. Unsegmented/continuous models propose that patterned motor output is driven by gradients of neurons and synapses that do not have segmental boundaries. We tested these ideas in the larval zebrafish, an animal that swims in discrete episodes, each of which is composed of coordinated motor bursts that progress rostrocaudally and alternate from side to side. We perturbed the spinal cord using spinal transections or strychnine application and measured the effect on fictive motor output. Spinal transections eliminated episode structure, and reduced both rostrocaudal and side-to-side coordination. Preparations with fewer intact segments were more severely affected, and preparations consisting of midbody and caudal segments were more severely affected than those consisting of rostral segments. In reduced preparations with the same number of intact spinal segments, side-to-side coordination was more severely disrupted than rostrocaudal coordination. Reducing glycine receptor signaling with strychnine reversibly disrupted both rostrocaudal and side-to-side coordination in spinalized larvae without disrupting episodic structure. Both spinal transection and strychnine decreased the stability of the motor rhythm, but this effect was not causal in reducing coordination. These results are inconsistent with a segmented model of the spinal cord and are better explained by a continuous model in which motor neuron coordination is controlled by segment-spanning microcircuits.

  13. Fibroblast growth factor signaling is required for early somatic gonad development in zebrafish.

    Science.gov (United States)

    Leerberg, Dena M; Sano, Kaori; Draper, Bruce W

    2017-09-01

    The vertebrate ovary and testis develop from a sexually indifferent gonad. During early development of the organism, primordial germ cells (the gamete lineage) and somatic gonad cells coalesce and begin to undergo growth and morphogenesis to form this bipotential gonad. Although this aspect of development is requisite for a fertile adult, little is known about the genetic regulation of early gonadogenesis in any vertebrate. Here, we provide evidence that fibroblast growth factor (Fgf) signaling is required for the early growth phase of a vertebrate bipotential gonad. Based on mutational analysis in zebrafish, we show that the Fgf ligand 24 (Fgf24) is required for proliferation, differentiation, and morphogenesis of the early somatic gonad, and as a result, most fgf24 mutants are sterile as adults. Additionally, we describe the ultrastructural elements of the early zebrafish gonad and show that distinct somatic cell populations can be identified soon after the gonad forms. Specifically, we show that fgf24 is expressed in an epithelial population of early somatic gonad cells that surrounds an inner population of mesenchymal somatic gonad cells that are in direct contact with the germ cells, and that fgf24 is required for stratification of the somatic tissue. Furthermore, based on gene expression analysis, we find that differentiation of the inner mesenchymal somatic gonad cells into functional cell types in the larval and early juvenile-stage gonad is dependent on Fgf24 signaling. Finally, we argue that the role of Fgf24 in zebrafish is functionally analogous to the role of tetrapod FGF9 in early gonad development.

  14. UPLC/MS MS data of testosterone metabolites in human and zebrafish liver microsomes and whole zebrafish larval microsomes

    Directory of Open Access Journals (Sweden)

    Moayad Saad

    2018-02-01

    Full Text Available This article represents data regarding a study published in Toxicology in vitro entitled “ in vitro CYP-mediated drug metabolism in the zebrafish (embryo using human reference compounds” (Saad et al., 2017 [1]. Data were acquired with ultra-performance liquid chromatography – accurate mass mass spectrometry (UPLC-amMS. A full spectrum scan was conducted for the testosterone (TST metabolites from the microsomal stability assay in zebrafish and humans. The microsomal proteins were extracted from adult zebrafish male (MLM and female (FLM livers, whole body homogenates of 96 h post fertilization larvae (EM and a pool of human liver microsomes from 50 donors (HLM. Data are expressed as the abundance from the extracted ion chromatogram of the metabolites.

  15. Homeostatic response to sleep/rest deprivation by constant water flow in larval zebrafish in both dark and light conditions.

    Science.gov (United States)

    Aho, Vilma; Vainikka, Maija; Puttonen, Henri A J; Ikonen, Heidi M K; Salminen, Tiia; Panula, Pertti; Porkka-Heiskanen, Tarja; Wigren, Henna-Kaisa

    2017-06-01

    Sleep-or sleep-like states-have been reported in adult and larval zebrafish using behavioural criteria. These reversible quiescent periods, displaying circadian rhythmicity, have been used in pharmacological, genetic and neuroanatomical studies of sleep-wake regulation. However, one of the important criteria for sleep, namely sleep homeostasis, has not been demonstrated unequivocally. To study rest homeostasis in zebrafish larvae, we rest-deprived 1-week-old larvae with a novel, ecologically relevant method: flow of water. Stereotyped startle responses to sensory stimuli were recorded after the rest deprivation to study arousal threshold using a high-speed camera, providing an appropriate time resolution to detect species-specific behavioural responses occurring in a millisecond time-scale. Rest-deprived larvae exhibited fewer startle responses than control larvae during the remaining dark phase and the beginning of the light phase, which can be interpreted as a sign of rest homeostasis-often used as equivalent of sleep homeostasis. To address sleep homeostasis further, we probed the adenosinergic system, which in mammals regulates sleep homeostasis. The adenosine A1 receptor agonist, cyclohexyladenosine, administered during the light period, decreased startle responses and increased immobility bouts, while the adenosine antagonist, caffeine, administered during the dark period, decreased immobility bouts. These results suggest that the regulation of sleep homeostasis in zebrafish larvae consists of the same elements as that of other species. © 2017 European Sleep Research Society.

  16. Toxicity of hexahydro-1,3,5-trinitro-1,3,5-triazine to larval zebrafish (Danio rerio)

    Science.gov (United States)

    Mukhi, S.; Pan, X.; Cobb, G.P.; Patino, R.

    2005-01-01

    Hexahydro-1,3,5-trinitro-1,3,5-triazine, a cyclonitramine commonly known as RDX, is used in the production of military munitions. Contamination of soil, sediment, and ground and surface waters with RDX has been reported in different places around the world. Acute and subacute toxicities of RDX have been relatively well documented in terrestrial vertebrates, but among aquatic vertebrates the information available is limited. The objective of this study was to characterize the acute toxicity of RDX to larval zebrafish. Mortality (LC50) and incidence of vertebral column deformities (EC50) were two of the end points measured in this study. The 96-h LC50 was estimated at 22.98 and 25.64 mg l-1 in two different tests. The estimated no-observed-effective- concentration (NOEC) values of RDX on lethality were 13.27 ?? 0.05 and 15.32 ?? 0.30 mg l-1; and the lowest-observed-effective- concentration (LOEC) values were 16.52 ?? 0.05 and 19.09 ?? 0.23 mg l-1 in these two tests, respectively. The 96-h EC50 for vertebral deformities on survivors from one of the acute lethality tests was estimated at 20.84 mg l-1, with NOEC and LOEC of 9.75 ?? 0.34 and 12.84 ?? 0.34 mg l-1, respectively. Behavioral aberrations were also noted in this acute toxicity study, including the occurrence of whirling movement and lethargic behavior. The acute effects of RDX on survival, incidence of deformities, and behavior of larval zebrafish occurred at the high end of the most frequently reported concentrations of RDX in aquatic environments. The chronic effects of RDX in aquatic vertebrates need to be determined for an adequate assessment of the ecological risk of environmental RDX. ?? 2005 Elsevier Ltd. All rights reserved.

  17. The power of projectomes: genetic mosaic labeling in the larval zebrafish brain reveals organizing principles of sensory circuits.

    Science.gov (United States)

    Robles, Estuardo

    2017-09-01

    In no vertebrate species do we possess an accurate, comprehensive tally of neuron types in the brain. This is in no small part due to the vast diversity of neuronal types that comprise complex vertebrate nervous systems. A fundamental goal of neuroscience is to construct comprehensive catalogs of cell types defined by structure, connectivity, and physiological response properties. This type of information will be invaluable for generating models of how assemblies of neurons encode and distribute sensory information and correspondingly alter behavior. This review summarizes recent efforts in the larval zebrafish to construct sensory projectomes, comprehensive analyses of axonal morphologies in sensory axon tracts. Focusing on the olfactory and optic tract, these studies revealed principles of sensory information processing in the olfactory and visual systems that could not have been directly quantified by other methods. In essence, these studies reconstructed the optic and olfactory tract in a virtual manner, providing insights into patterns of neuronal growth that underlie the formation of sensory axon tracts. Quantitative analysis of neuronal diversity revealed organizing principles that determine information flow through sensory systems in the zebrafish that are likely to be conserved across vertebrate species. The generation of comprehensive cell type classifications based on structural, physiological, and molecular features will lead to testable hypotheses on the functional role of individual sensory neuron subtypes in controlling specific sensory-evoked behaviors.

  18. A novel method for rearing first-feeding larval zebrafish: polyculture with Type L saltwater rotifers (Brachionus plicatilis).

    Science.gov (United States)

    Best, Jason; Adatto, Isaac; Cockington, Jason; James, Althea; Lawrence, Christian

    2010-09-01

    Promoting high rates of growth and survival can be a major challenge in zebrafish culture, especially during the first-feeding stage. Here we describe a new rearing technique in which zebrafish larvae are polycultured in static tanks with Type "L" saltwater rotifers (Brachionus plicatilis) for the first 5 days of feeding (days 5-9 postfertilization). To demonstrate the effectiveness of this technique, we conducted rearing trials using fish from two different strains: AB and nacre. Growth, survival, water quality, and rotifer density were assayed daily through the polyculture phase (days 5-9), and during the transition to standard rearing conditions (days 10-12). After that point, once the fish were fully integrated onto recirculating systems, parameters were measured once per week out to day 30. In all trials, the fish displayed high rates of growth and survival throughout the three phases (polyculture, transition, and recirculating flow), indicating that this method may be employed during the critical first-feeding stage to help improve rearing performance in zebrafish facilities. Additionally, water quality parameters observed during the polyculture phase of the trials reveal that early zebrafish larvae are much more tolerant of elevated levels of ammonia and salinity than previously believed.

  19. Assessing Locomotor Activity in Larval Zebrafish: Influence of Extrinsic and Intrinsic Variables

    Science.gov (United States)

    The U.S. Environmental Protection Agency is evaluating methods to screen and prioritize large numbers of chemicals for developmental toxicity. We are exploring methods to detect developmentally neurotoxic chemicals using zebrafish behavior at 6 days of age. The behavioral paradig...

  20. High-Throughput Light Sheet Microscopy for the Automated Live Imaging of Larval Zebrafish

    Science.gov (United States)

    Baker, Ryan; Logan, Savannah; Dudley, Christopher; Parthasarathy, Raghuveer

    The zebrafish is a model organism with a variety of useful properties; it is small and optically transparent, it reproduces quickly, it is a vertebrate, and there are a large variety of transgenic animals available. Because of these properties, the zebrafish is well suited to study using a variety of optical technologies including light sheet fluorescence microscopy (LSFM), which provides high-resolution three-dimensional imaging over large fields of view. Research progress, however, is often not limited by optical techniques but instead by the number of samples one can examine over the course of an experiment, which in the case of light sheet imaging has so far been severely limited. Here we present an integrated fluidic circuit and microscope which provides rapid, automated imaging of zebrafish using several imaging modes, including LSFM, Hyperspectral Imaging, and Differential Interference Contrast Microscopy. Using this system, we show that we can increase our imaging throughput by a factor of 10 compared to previous techniques. We also show preliminary results visualizing zebrafish immune response, which is sensitive to gut microbiota composition, and which shows a strong variability between individuals that highlights the utility of high throughput imaging. National Science Foundation, Award No. DBI-1427957.

  1. Drugs that Target Dopamine Receptors: Changes in Locomotor Activity in Larval Zebrafish

    Science.gov (United States)

    As part of an effort at the US Environmental Protection Agency to develop a rapid in vivo screen for prioritization of toxic chemicals, we have begun to characterize the locomotor activity of zebrafish (Danio rerio) larvae. This includes assessing the acute effects of drugs known...

  2. Targeting the-Dopaminergic Nervous System: Altering Behavior in Larval Zebrafish

    Science.gov (United States)

    Zebrafish (Dania rerio) are becoming an important model system in studying the effects of environmental chemicals on behavior. In order to develop a rapid in vivo screen to prioritize toxic chemicals, we have begun assessing the acute locomotor effects of drugs that act on the do...

  3. Kita driven expression of oncogenic HRAS leads to early onset and highly penetrant melanoma in zebrafish.

    Directory of Open Access Journals (Sweden)

    Cristina Santoriello

    2010-12-01

    Full Text Available Melanoma is the most aggressive and lethal form of skin cancer. Because of the increasing incidence and high lethality of melanoma, animal models for continuously observing melanoma formation and progression as well as for testing pharmacological agents are needed.Using the combinatorial Gal4-UAS system, we have developed a zebrafish transgenic line that expresses oncogenic HRAS under the kita promoter. Already at 3 days transgenic kita-GFP-RAS larvae show a hyper-pigmentation phenotype as earliest evidence of abnormal melanocyte growth. By 2-4 weeks, masses of transformed melanocytes form in the tail stalk of the majority of kita-GFP-RAS transgenic fish. The adult tumors evident between 1-3 months of age faithfully reproduce the immunological, histological and molecular phenotypes of human melanoma, but on a condensed time-line. Furthermore, they show transplantability, dependence on mitfa expression and do not require additional mutations in tumor suppressors. In contrast to kita expressing melanocyte progenitors that efficiently develop melanoma, mitfa expressing progenitors in a second Gal4-driver line were 4 times less efficient in developing melanoma during the three months observation period.This indicates that zebrafish kita promoter is a powerful tool for driving oncogene expression in the right cells and at the right level to induce early onset melanoma in the presence of tumor suppressors. Thus our zebrafish model provides a link between kita expressing melanocyte progenitors and melanoma and offers the advantage of a larval phenotype suitable for large scale drug and genetic modifier screens.

  4. Habituation of the C-start response in larval zebrafish exhibits several distinct phases and sensitivity to NMDA receptor blockade.

    Directory of Open Access Journals (Sweden)

    Adam C Roberts

    Full Text Available The zebrafish larva has been a valuable model system for genetic and molecular studies of development. More recently, biologists have begun to exploit the surprisingly rich behavioral repertoire of zebrafish larvae to investigate behavior. One prominent behavior exhibited by zebrafish early in development is a rapid escape reflex (the C-start. This reflex is mediated by a relatively simple neural circuit, and is therefore an attractive model behavior for neurobiological investigations of simple forms of learning and memory. Here, we describe two forms of short-lived habituation of the C-start in response to brief pulses of auditory stimuli. A rapid form, persisting for ≥1 min but <15 min, was induced by 120 pulses delivered at 0.5-2.0 Hz. A more extended form (termed "short-term habituation" here, which persisted for ≥25 min but <1 h, was induced by spaced training. The spaced training consisted of 10 blocks of auditory pulses delivered at 1 Hz (5 min interblock interval, 900 pulses per block. We found that these two temporally distinguishable forms of habituation are mediated by different cellular mechanisms. The short-term form depends on activation of N-methyl-d-aspartate receptors (NMDARs, whereas the rapid form does not.

  5. Impact of co-exposure with butachlor and triadimefon on thyroid endocrine system in larval zebrafish.

    Science.gov (United States)

    Cao, Chuyan; Wang, Qiangwei; Jiao, Fang; Zhu, Guonian

    2016-09-01

    Butachlor (BTL) and triadimefon (TDF), the widely used herbicide and fungicide, are unavoidable enter into the aquatic environment. However, there were limited study regarding to the joint toxicity of these two pesticides on fish at present. To evaluate the potential thyroid-disrupting toxicity and exposed to different concentrations of BTL mixed with TDF. Zebrafish embryo (n=3) were exposed to 0.01 and 0.05 fold of LC50 from the acute joint toxicity test, of which 0.32mg/L (BTL) and 9.41mg/L (TDF) for single or mixture agents (BTL: 0.0064mg/L, 0.032mg/L; TDF: 0.1882mg/L, 0.9410mg/L; co-exposure: 0.0032mg/L BTL+0.0941mg/L TDF, 0.016mg/l BTL+0.4705mg/L TDF) after 10-day post-fertilization. Hatching, malformation, survival rates and thyroid hormones (THs), genes expression involved in HPT-axis of embryos were measured and detected in control and separately/co-exposure treatments. THs contents were evaluated by ELISA kit and the expression levels of genes were determined by RT-PCR. Hatching, malformation and survival rates of embryos exposed to single BTL exhibited no statistically significant difference from the control besides decreased of high concentration in survival rates. Exposure to TDF reduced hatching, survival rate and increased malformation. The combined exposure to BTL and TDF resulted in greater adverse effects on embryonic development. BTL exposure significantly increased free T3 and T4 contents. Elevated free T3 content was also observed in the larvae exposed with single BTL. Co-exposure of the two pesticides caused greater enhanced of T3 and T4 levels. Furthermore, gene data showed BTL up-regulated the mRNA expression of tpo, tshβ, tg, ttr, dio2, TDF up-regulated the mRNA expression of tpo, trα, ttr, dio2 and down-regulated trβ gene. The mixture of the two pesticides caused up-regulation mRNA expression of trα, trβ, tg, ttr, dio2. BTL and TDF resulted in adverse effects on zebrafish embryonic development and caused thyroid endocrine disruption

  6. Integrated toxic evaluation of sulfamethazine on zebrafish: Including two lifespan stages (embryo-larval and adult) and three exposure periods (exposure, post-exposure and re-exposure).

    Science.gov (United States)

    Yan, Zhengyu; Yang, Qiulian; Jiang, Weili; Lu, Jilai; Xiang, Zhongrun; Guo, Ruixin; Chen, Jianqiu

    2018-03-01

    Persistence of antibiotics in aquatic environment may pose a risk to the non-target aquatic organisms. This study provided an integrated evaluation to analyze the toxic stress of sulfamethazine (SMZ) on zebrafish in two lifespan stages (embryo-larval and adult) and three exposure periods (exposure, post-exposure and re-exposure). Zebrafish embryos and adult zebrafish were exposed to SMZ at 0.2, 20 and 2000 μg/L, respectively. The results showed that SMZ at any given concentration inhibited the hatching of embryos at 58-96 hpf (hours post-fertilization). Our result also indicated that two major kinds of the malformation, which was induced by the antibiotic, were edema and spinal curvature. Additionally, the antibiotic stimulated the heartbeat while reduced the body length of the embryo at 72 hpf. Superoxide dismutase (SOD) activities and malondialdehyde (MDA) contents significantly increased at 120 hpf when the embryos were exposed to the lowest concentration (0.2 μg/L) of the antibiotic. On the other hand, the antibiotic induced SOD activities and MDA contents in adult zebrafish in the exposure and re-exposure periods. The MDA contents could recover while SOD activities still increased in 2 d after the exposure. Both SOD activities and MDA contents could recover in 7 d after the exposure. Levels of SOD and MDA in the re-exposure were higher than those in the first exposure. Our results suggested that SMZ had toxic effects on both embryos and adult zebrafish, and provided an integrated evaluation of the toxic effects of SMZ on zebrafish at a new perspective. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Neural control and modulation of swimming speed in the larval zebrafish.

    Science.gov (United States)

    Severi, Kristen E; Portugues, Ruben; Marques, João C; O'Malley, Donald M; Orger, Michael B; Engert, Florian

    2014-08-06

    Vertebrate locomotion at different speeds is driven by descending excitatory connections to central pattern generators in the spinal cord. To investigate how these inputs determine locomotor kinematics, we used whole-field visual motion to drive zebrafish to swim at different speeds. Larvae match the stimulus speed by utilizing more locomotor events, or modifying kinematic parameters such as the duration and speed of swimming bouts, the tail-beat frequency, and the choice of gait. We used laser ablations, electrical stimulation, and activity recordings in descending neurons of the nucleus of the medial longitudinal fasciculus (nMLF) to dissect their contribution to controlling forward movement. We found that the activity of single identified neurons within the nMLF is correlated with locomotor kinematics, and modulates both the duration and oscillation frequency of tail movements. By identifying the contribution of individual supraspinal circuit elements to locomotion kinematics, we build a better understanding of how the brain controls movement. Copyright © 2014 Elsevier Inc. All rights reserved.

  8. Global gene expression in larval zebrafish (Danio rerio) exposed to selective serotonin reuptake inhibitors (fluoxetine and sertraline) reveals unique expression profiles and potential biomarkers of exposure

    International Nuclear Information System (INIS)

    Park, June-Woo; Heah, Tze Ping; Gouffon, Julia S.; Henry, Theodore B.; Sayler, Gary S.

    2012-01-01

    Larval zebrafish (Danio rerio) were exposed (96 h) to selective serotonin reuptake inhibitors (SSRIs) fluoxetine and sertraline and changes in transcriptomes analyzed by Affymetrix GeneChip ® Zebrafish Array were evaluated to enhance understanding of biochemical pathways and differences between these SSRIs. The number of genes differentially expressed after fluoxetine exposure was 288 at 25 μg/L and 131 at 250 μg/L; and after sertraline exposure was 33 at 25 μg/L and 52 at 250 μg/L. Same five genes were differentially regulated in both SSRIs indicating shared molecular pathways. Among these, the gene coding for FK506 binding protein 5, annotated to stress response regulation, was highly down-regulated in all treatments (results confirmed by qRT-PCR). Gene ontology analysis indicated at the gene expression level that regulation of stress response and cholinesterase activities were influenced by these SSRIs, and suggested that changes in transcription of these genes could be used as biomarkers of SSRI exposure. - Highlights: ► Exposure of zebrafish to selective serotonin reuptake inhibitors (SSRIs). ► Fluoxetine and sertraline generate different global gene expression profiles. ► Genes linked to stress response and acetylcholine esterase affected by both SSRIs. - Global gene expression profiles in zebrafish exposed to selective serotonin reuptake inhibitors.

  9. Alternative methods for toxicity assessments in fish: comparison of the fish embryo toxicity and the larval growth and survival tests in zebrafish and fathead minnows.

    Science.gov (United States)

    Jeffries, Marlo K Sellin; Stultz, Amy E; Smith, Austin W; Rawlings, Jane M; Belanger, Scott E; Oris, James T

    2014-11-01

    An increased demand for chemical toxicity evaluations has resulted in the need for alternative testing strategies that address animal welfare concerns. The fish embryo toxicity (FET) test developed for zebrafish (Danio rerio) is one such alternative, and the application of the FET test to other species such as the fathead minnow (Pimephales promelas) has been proposed. In the present study, the performances of the FET test and the larval growth and survival (LGS; a standard toxicity testing method) test in zebrafish and fathead minnows were evaluated. This required that testing methods for the fathead minnow FET and zebrafish LGS tests be harmonized with existing test methods and that the performance of these testing strategies be evaluated by comparing the median lethal concentrations of 2 reference toxicants, 3,4-dicholoraniline and ammonia, obtained via each of the test types. The results showed that procedures for the zebrafish FET test can be adapted and applied to the fathead minnow. Differences in test sensitivity were observed for 3,4-dicholoraniline but not ammonia; therefore, conclusions regarding which test types offer the least or most sensitivity could not be made. Overall, these results show that the fathead minnow FET test has potential as an alternative toxicity testing strategy and that further analysis with other toxicants is warranted in an effort to better characterize the sensitivity and feasibility of this testing strategy. © 2014 SETAC.

  10. The toxicity of chlorpyrifos on the early life stage of zebrafish: a survey on the endpoints at development, locomotor behavior, oxidative stress and immunotoxicity.

    Science.gov (United States)

    Jin, Yuanxiang; Liu, Zhenzhen; Peng, Tao; Fu, Zhengwei

    2015-04-01

    Chlorpyrifos (CPF) is one of the most toxic pesticides in aquatic ecosystem, but its toxicity mechanisms to fish are still not fully understood. This study examined the toxicity targets of CPF in early life stage of zebrafish on the endpoints at developmental toxicity, neurotoxicity, oxidative stress and immunotoxicity. Firstly, CPF exposure decreased the body length, inhibited the hatchability and heart rate, and resulted in a number of morphological abnormalities, primarily spinal deformities (SD) and pericardial edema (PE), in larval zebrafish. Secondly, the free swimming activities and the swimming behaviors of the larvae in response to the stimulation of light-to-dark photoperiod transition were significantly influenced by the exposure to 100 and 300 μg/L CPF. In addition, the activity of acetylcholinesterase (AChE) and the transcription of some genes related to neurotoxicity were also influenced by CPF exposure. Thirdly, CPF exposure induced oxidative stress in the larval zebrafish. The malondialdehyde (MDA) levels increased and the glutathione (GSH) contents decreased significantly in a concentration-dependent manner after the exposure to CPF for 96 hours post fertilization (hpf). CPF affected not only the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) and glutathione S-transferase (GST), but also the transcriptional levels of their respective genes. Finally, the mRNA levels of the main cytokines including tumor necrosis factor α (Tnfα), interferon (Ifn), interleukin-1 beta (Il-1β), interleukin 6 (Il6), complement factor 4 (C4) in the larvae increased significantly after the exposure to 100 or 300 μg/L CPF for 96 hpf, suggesting that the innate immune system disturbed by CPF in larvae. Taken together, our results suggested that CPF had the potential to cause developmental toxicity, behavior alterations, oxidative stress and immunotoxicity in the larval zebrafish. Copyright © 2015 Elsevier Ltd. All rights

  11. Sensorimotor Representations in Cerebellar Granule Cells in Larval Zebrafish Are Dense, Spatially Organized, and Non-temporally Patterned.

    Science.gov (United States)

    Knogler, Laura D; Markov, Daniil A; Dragomir, Elena I; Štih, Vilim; Portugues, Ruben

    2017-05-08

    A fundamental question in neurobiology is how animals integrate external sensory information from their environment with self-generated motor and sensory signals in order to guide motor behavior and adaptation. The cerebellum is a vertebrate hindbrain region where all of these signals converge and that has been implicated in the acquisition, coordination, and calibration of motor activity. Theories of cerebellar function postulate that granule cells encode a variety of sensorimotor signals in the cerebellar input layer. These models suggest that representations should be high-dimensional, sparse, and temporally patterned. However, in vivo physiological recordings addressing these points have been limited and in particular have been unable to measure the spatiotemporal dynamics of population-wide activity. In this study, we use both calcium imaging and electrophysiology in the awake larval zebrafish to investigate how cerebellar granule cells encode three types of sensory stimuli as well as stimulus-evoked motor behaviors. We find that a large fraction of all granule cells are active in response to these stimuli, such that representations are not sparse at the population level. We find instead that most responses belong to only one of a small number of distinct activity profiles, which are temporally homogeneous and anatomically clustered. We furthermore identify granule cells that are active during swimming behaviors and others that are multimodal for sensory and motor variables. When we pharmacologically change the threshold of a stimulus-evoked behavior, we observe correlated changes in these representations. Finally, electrophysiological data show no evidence for temporal patterning in the coding of different stimulus durations. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  12. Ovarian development and early larval survival of Stenopus zanzibaricus (Bruce, 1976

    Directory of Open Access Journals (Sweden)

    Daniel Marques

    2014-06-01

    Full Text Available Despite economically valuable, ornamental shrimps are poorly studied and there is a lack of protocols for their captive breeding. Stenopus is one of the most important genera of ornamental shrimps, being Stenopus zanzibaricus one of the species with less information about captive breeding and larviculture. For a better knowledge of its reproductive cycle, we evaluated morphological and color changes during ovarian development of adult females through daily photographs taken during all the cycle. The effect of three diets (Brachionus plicatilis + Tetraselmis chuii; newly artemia nauplii + Tetraselmis chuii; newly artemia nauplii and two different temperatures (25ºC and 27ºC on early larval development were also evaluated. With this study, it was expected to obtain some insight about Stenopus zanzibaricus reproductive cycle and early larval development, in order to develop captive breeding and larval rearing protocols for this economic valuable species.

  13. Optimisation of Embryonic and Larval ECG Measurement in Zebrafish for Quantifying the Effect of QT Prolonging Drugs

    Science.gov (United States)

    Dhillon, Sundeep Singh; Dóró, Éva; Magyary, István; Egginton, Stuart; Sík, Attila; Müller, Ferenc

    2013-01-01

    Effective chemical compound toxicity screening is of paramount importance for safe cardiac drug development. Using mammals in preliminary screening for detection of cardiac dysfunction by electrocardiography (ECG) is costly and requires a large number of animals. Alternatively, zebrafish embryos can be used as the ECG waveform is similar to mammals, a minimal amount of chemical is necessary for drug testing, while embryos are abundant, inexpensive and represent replacement in animal research with reduced bioethical concerns. We demonstrate here the utility of pre-feeding stage zebrafish larvae in detection of cardiac dysfunction by electrocardiography. We have optimised an ECG recording system by addressing key parameters such as the form of immobilization, recording temperature, electrode positioning and developmental age. Furthermore, analysis of 3 days post fertilization (dpf) zebrafish embryos treated with known QT prolonging drugs such as terfenadine, verapamil and haloperidol led to reproducible detection of QT prolongation as previously shown for adult zebrafish. In addition, calculation of Z-factor scores revealed that the assay was sensitive and specific enough to detect large drug-induced changes in QTc intervals. Thus, the ECG recording system is a useful drug-screening tool to detect alteration to cardiac cycle components and secondary effects such as heart block and arrhythmias in zebrafish larvae before free feeding stage, and thus provides a suitable replacement for mammalian experimentation. PMID:23579446

  14. Cloning of zebrafish Mustn1 orthologs and their expression during early development.

    Science.gov (United States)

    Camarata, Troy; Vasilyev, Aleksandr; Hadjiargyrou, Michael

    2016-11-15

    Mustn1 is a small nuclear protein that is involved in the development and regeneration of the musculoskeletal system. Previous work established a role for Mustn1 in myogenic and chondrogenic differentiation. In addition, recent evidence suggests a potential role for Mustn1 in cilia function in zebrafish. A detailed study of Mustn1 expression has yet to be conducted in zebrafish. As such, we report herein the cloning of the zebrafish Mustn1 orthologs, mustn1a and mustn1b, and their expression during zebrafish embryonic and larval development. Results indicate a 44% nucleotide identity between the two paralogs. Phylogenetic analysis further confirmed that the Mustn1a and 1b predicted proteins were highly related to other vertebrate members of the Mustn1 protein family. Whole mount in situ hybridization revealed expression of both mustn1a and 1b at the 7-somite stage through 72hpf in structures such as Kupffer's vesicle, segmental mesoderm, head structures, and otic vesicle. Additionally, in 5day old larva, mustn1a and 1b expression is detected in the neurocranium, otic capsule, and the gut. Although both were expressed in the neurocranium, mustn1a was localized in the hypophyseal fenestra whereas mustn1b was found near the posterior basicapsular commissure. mustn1b also displayed expression in the ceratohyal and ceratobranchial elements of the pharyngeal skeleton. These expression patterns were verified temporally by q-PCR analysis. Taken together, we conclude that Mustn1 expression is conserved in vertebrates and that the variations in expression of the two zebrafish paralogs suggest different modes of molecular regulation. Copyright © 2016 Elsevier B.V. All rights reserved.

  15. Early gonad development in zebrafish ( Danio rerio ) | Okuthe ...

    African Journals Online (AJOL)

    Gonadogenesis in zebrafish goes through an initial ovarian phase then subsequently into either ovarian or testicular phases. How germ cells choose to commit to an oogenic fate and enter meiosis or alternatively not enter meiosis and commit to a spermatogenetic fate remains a key question. This study investigated events ...

  16. The effects of waterborne uranium on the hatching success, development, and survival of early life stages of zebrafish (Danio rerio)

    International Nuclear Information System (INIS)

    Bourrachot, Stephanie; Simon, Olivier; Gilbin, Rodolphe

    2008-01-01

    In this study, we investigated the effects of the radioactive metal uranium (U) on the embryonic development, hatching success, growth rate, and survival of juvenile zebrafish (Danio rerio). We studied the effects of depleted uranium (20-500 μg L -1 of DU), inducing mainly chemical toxicity due to its low specific activity, and the combined effects of chemical and radiological toxicity by using a higher specific activity uranium isotope (20 and 100 μg L -1 of 233 U). Results showed that early life stages are significantly affected by uranium exposure through both chemical and combined (chemical and radiological) toxicity. Experiments showed significant effects of U on hatching success starting at the concentration of 250 μg L -1 of DU, causing a 42% delay in median hatching times relative to control. Furthermore, a reduction of growth (decrease in body length and weight) was observed followed by a high mortality of pro-larvae stage (up to 100% at DU concentrations of 250 μg L -1 upon a 15 day exposure). Bioaccumulation measurements highlighted that U was mainly localised in the chorion but penetrated in the embryo inside eggs at a higher concentration. The effects differed depending on the isotopic composition of the uranium: sublethal defects in the tail detachment process were more pronounced for 233 U than DU exposure, while the presence of 233 U specifically affected embryo development and led to higher mortality rates of the prolarvae. The results from this study showed that the early life stages of zebrafish seems to be more sensitive to uranium contamination than more mature stages, and underline the importance of including pro-larval stages into toxicity tests in order to improve the relevancy for environmental risk assessments

  17. External gamma irradiation-induced effects in early-life stages of zebrafish, Danio rerio

    International Nuclear Information System (INIS)

    Gagnaire, B.; Cavalié, I.; Pereira, S.; Floriani, M.; Dubourg, N.; Camilleri, V.; Adam-Guillermin, C.

    2015-01-01

    Highlights: • The present study aimed to evaluate the effects of gamma rays on zebrafish larvae. • Different techniques were used: gene expression, biochemistry, microscopy and macroscopical observations. • The results showed that gamma irradiation can alter embryo-larval development at several levels of organization. - Abstract: In the general context of validation of tools useful for the characterization of ecological risk linked to ionizing radiation, the effects of an external gamma irradiation were studied in zebrafish larvae irradiated for 96 h with two dose rates: 0.8 mGy/d, which is close to the level recommended to protect ecosystems from adverse effects of ionizing radiation (0.24 mGy/d) and a higher dose rate of 570 mGy/d. Several endpoints were investigated, such as mortality, hatching, and some parameters of embryo-larval development, immunotoxicity, apoptosis, genotoxicity, neurotoxicity and histological alterations. Results showed that an exposure to gamma rays induced an acceleration of hatching for both doses and a decrease of yolk bag diameter for the highest dose, which could indicate an increase of global metabolism. AChE activity decreased with the low dose rate of gamma irradiation and alterations were also shown in muscles of irradiated larvae. These results suggest that gamma irradiation can induce damages on larval neurotransmission, which could have repercussions on locomotion. DNA damages, basal ROS production and apoptosis were also induced by irradiation, while ROS stimulation index and EROD biotransformation activity were decreased and gene expression of acetylcholinesterase, choline acetyltransferase, cytochrome p450 and myeloperoxidase increased. These results showed that ionizing radiation induced an oxidative stress conducting to DNA damages. This study characterized further the modes of action of ionizing radiation in fish.

  18. External gamma irradiation-induced effects in early-life stages of zebrafish, Danio rerio

    Energy Technology Data Exchange (ETDEWEB)

    Gagnaire, B., E-mail: beatrice.gagnaire@irsn.fr [Institut de Radioprotection et de Sureté Nucléaire (IRSN), PRP-ENV/SERIS/LECO, Cadarache, Saint-Paul-lez-Durance 13115 (France); Cavalié, I. [Institut de Radioprotection et de Sureté Nucléaire (IRSN), PRP-ENV/SERIS/LECO, Cadarache, Saint-Paul-lez-Durance 13115 (France); Pereira, S. [Neolys Diagnostics, Lyon 69373 (France); Floriani, M.; Dubourg, N.; Camilleri, V.; Adam-Guillermin, C. [Institut de Radioprotection et de Sureté Nucléaire (IRSN), PRP-ENV/SERIS/LECO, Cadarache, Saint-Paul-lez-Durance 13115 (France)

    2015-12-15

    Highlights: • The present study aimed to evaluate the effects of gamma rays on zebrafish larvae. • Different techniques were used: gene expression, biochemistry, microscopy and macroscopical observations. • The results showed that gamma irradiation can alter embryo-larval development at several levels of organization. - Abstract: In the general context of validation of tools useful for the characterization of ecological risk linked to ionizing radiation, the effects of an external gamma irradiation were studied in zebrafish larvae irradiated for 96 h with two dose rates: 0.8 mGy/d, which is close to the level recommended to protect ecosystems from adverse effects of ionizing radiation (0.24 mGy/d) and a higher dose rate of 570 mGy/d. Several endpoints were investigated, such as mortality, hatching, and some parameters of embryo-larval development, immunotoxicity, apoptosis, genotoxicity, neurotoxicity and histological alterations. Results showed that an exposure to gamma rays induced an acceleration of hatching for both doses and a decrease of yolk bag diameter for the highest dose, which could indicate an increase of global metabolism. AChE activity decreased with the low dose rate of gamma irradiation and alterations were also shown in muscles of irradiated larvae. These results suggest that gamma irradiation can induce damages on larval neurotransmission, which could have repercussions on locomotion. DNA damages, basal ROS production and apoptosis were also induced by irradiation, while ROS stimulation index and EROD biotransformation activity were decreased and gene expression of acetylcholinesterase, choline acetyltransferase, cytochrome p450 and myeloperoxidase increased. These results showed that ionizing radiation induced an oxidative stress conducting to DNA damages. This study characterized further the modes of action of ionizing radiation in fish.

  19. Early development and larval behaviour of a minnow, Barbus ...

    African Journals Online (AJOL)

    The chubbyhead barb, Barbus anoplus, underwent a population explosion in the early phases of filling of Lake be Roux on the Orange River. This successful colonization was possibly related to the survival strategy of the young stages of this minnow. It is suggested that some of the development traits of B. anoplus enabled ...

  20. Changes in Neurotransmitter Profiles during Early Zebrafish (Danio rerio) Development and after Pesticide Exposure.

    Science.gov (United States)

    Tufi, Sara; Leonards, Pim; Lamoree, Marja; de Boer, Jacob; Legler, Juliette; Legradi, Jessica

    2016-03-15

    During early development, neurotransmitters are important stimulants for the development of the central nervous system. Although the development of different neuronal cell types during early zebrafish (Danio rerio) development is well-studied, little is known of the levels of neurotransmitters, their precursors and metabolites during development, and how these levels are affected by exposure to environmental contaminants. A method based on hydrophilic interaction liquid chromatography coupled to tandem mass spectrometry has been applied for the first time to zebrafish embryos and larvae to study five neurotransmitter systems in parallel, including the dopaminergic-andrenergic, glutaminergic-GABAnergic, serotoninergic, histaminergic, and cholinergic systems. Our method enables the quantification of neurotransmitters and their precursors and metabolites in whole zebrafish from the period of zygote to free-swimming larvae 6 days postfertilization (dpf). We observed a developmental stage-dependent pattern with clear differences between the first 2 days of development and the following days. Whereas the neurotransmitter levels steadily increased, the precursors showed a peak at 3 dpf. After exposure to several pesticides, significant differences in concentrations of neurotransmitters and precursors were observed. Our study revealed new insights about neurotransmitter systems during early zebrafish development and showed the usefulness of our approach for environmental neurotoxicity studies.

  1. Diatom diet selectivity by early post-larval abalone Haliotis diversicolor supertexta under hatchery conditions

    Science.gov (United States)

    Zhang, Yuyu; Gao, Yahui; Liang, Junrong; Chen, Changping; Zhao, Donghai; Li, Xuesong; Li, Yang; Wu, Wenzhong

    2010-11-01

    Benthic diatoms constitute the primary diet of abalone during their early stages of development. To evaluate the dietary preferences of early post-larval abalone, Haliotis diversicolor supertexta, we analyzed the gut contents of post-larvae that settled on diatom films. We compared the abundance and species diversity of diatom assemblages in the gut to those of the epiphytic diatom assemblages on the attachment films, and identified 40 benthic diatom species in the gut contents of post-larvae 12 to 24 d after settlement. The most abundant taxa in the gut contents were Navicula spp., Amphora copulate, and Amphora coffeaeformis. Navicula spp. accounted for 64.0% of the cell density. In the attachment films, we identified 110 diatom species belonging to 38 genera. Pennate diatoms were the dominant members including the species Amphiprora alata, Cocconeis placentula var. euglypta, Cylindrotheca closterium, Navicula sp. 2, and A. coffeaeformis. Nano-diatoms (abalone seed. The difference of the composition and abundance of diatoms between in the guts and on the biofilms suggests that early post-larval grazing was selective. An early post-larval abalone preferred nano-diatoms and the genera Navicula and Amphora during the month after settlement.

  2. Early detection monitoring for larval dreissenid mussels: how much plankton sampling is enough?

    Science.gov (United States)

    Counihan, Timothy D; Bollens, Stephen M

    2017-03-01

    The development of quagga and zebra mussel (dreissenids) monitoring programs in the Pacific Northwest provides a unique opportunity to evaluate a regional invasive species detection effort early in its development. Recent studies suggest that the ecological and economic costs of a dreissenid infestation in the Pacific Northwest of the USA would be significant. Consequently, efforts are underway to monitor for the presence of dreissenids. However, assessments of whether these efforts provide for early detection are lacking. We use information collected from 2012 to 2014 to characterize the development of larval dreissenid monitoring programs in the states of Idaho, Montana, Oregon, and Washington in the context of introduction and establishment risk. We also estimate the effort needed for high-probability detection of rare planktonic taxa in four Columbia and Snake River reservoirs and assess whether the current level of effort provides for early detection. We found that the effort expended to monitor for dreissenid mussels increased substantially from 2012 to 2014, that efforts were distributed across risk categories ranging from high to very low, and that substantial gaps in our knowledge of both introduction and establishment risk exist. The estimated volume of filtered water required to fully census planktonic taxa or to provide high-probability detection of rare taxa was high for the four reservoirs examined. We conclude that the current level of effort expended does not provide for high-probability detection of larval dreissenids or other planktonic taxa when they are rare in these reservoirs. We discuss options to improve early detection capabilities.

  3. Comparison of Dissolved Nickel and Nickel Nanoparticles Toxicity in Larval Zebrafish in Terms of Gene Expression and DNA Damage.

    Science.gov (United States)

    Boran, Halis; Şaffak, Savaş

    2018-01-01

    With the use of nanoparticles (NPs) in many industrial activities and consumer products, it is important to evaluate the effects of their release into the environment. Metal NPs (e.g., Ni-NPs or Cu-NPs) can release metal ions that are toxic to aquatic organisms; however, whether the toxicity is from metal ions rather than unique "nano-scale" effects of the NPs is unresolved. This research investigated Ni-NP toxicity in zebrafish Danio rerio larvae to clarify whether toxic effects are attributable to release of Ni ions. First, the acute (96-h lethal) toxicity of Ni-NPs was determined in comparison to aqueous Ni in fish exposed to Ni(II) by water-soluble NiCl 2 . Subsequently, sublethal experiments with Ni-NPs and Ni(II) were conducted to assess changes in expression of stress-related genes (mt2, rad51, and p53) by quantitative PCR. Acute toxicity of Ni in fish exposed to Ni(II) was higher (96-h LC 50  = 32.6 mg/L) than for fish exposed to Ni-NPs (96-h LC 50  = 122.2 mg/L). Also, DNA strand breaks were higher in Ni(II)- than Ni-NPs-exposed larvae. Induction of stress-related genes in larvae was complex and was not directly related to Ni-NPs and Ni(II) concentration, although there was a significant induction in the mt2 and p53 gene of the larvae exposed to Ni-NPs and Ni(II) relative to controls. Results indicated that while Ni-NPs induced gene expression (presumably by the release of Ni ions), the differences in concentration relationships of gene expression between Ni-NPs and Ni(II) suggest that factors in addition to the release of Ni ions from Ni-NPs influence acute toxicity of Ni-NPs.

  4. Expression of voltage-activated calcium channels in the early zebrafish embryo.

    Science.gov (United States)

    Sanhueza, Dayán; Montoya, Andro; Sierralta, Jimena; Kukuljan, Manuel

    2009-05-01

    Increases in cytosolic calcium concentrations regulate many cellular processes, including aspects of early development. Calcium release from intracellular stores and calcium entry through non-voltage-gated channels account for signalling in non-excitable cells, whereas voltage-gated calcium channels (CaV) are important in excitable cells. We report the expression of multiple transcripts of CaV, identified by its homology to other species, in the early embryo of the zebrafish, Danio rerio, at stages prior to the differentiation of excitable cells. CaV mRNAs and proteins were detected as early as the 2-cell stages, which indicate that they arise from both maternal and zygotic transcription. Exposure of embryos to pharmacological blockers of CaV does not perturb early development significantly, although late effects are appreciable. These results suggest that CaV may have a role in calcium homeostasis and control of cellular process during early embryonic development.

  5. Metabolomics and transcriptomics reveal the toxicity of difenoconazole to the early life stages of zebrafish (Danio rerio).

    Science.gov (United States)

    Teng, Miaomiao; Zhu, Wentao; Wang, Dezhen; Qi, Suzhen; Wang, Yao; Yan, Jin; Dong, Kai; Zheng, Mingqi; Wang, Chengju

    2018-01-01

    Difenoconazole is widely used to inhibit the growth of fungi, but its residue in the water environment may threaten ecosystem and human health. Here, 1 H nuclear magnetic resonance (NMR) and LC-MS/MS based metabolomics and transcriptomics approaches were used to assess the response of zebrafish to difenoconazole exposure. Early life stages of zebrafish were exposed to difenoconazole at environmentally relevant concentrations for 168h. Their comparison with the control group suggested an adverse development and disturbance of steroid hormones and VTG. KEGG pathway analysis identified five biological processes on the basis of differentially expressed genes (DEGs), as well as altered metabolites and amino acids in zebrafish following difenoconazole exposure. These affected processes included energy metabolism, amino acids metabolism, lipid metabolism, nucleotide metabolism, and an immune-related pathway. Collectively, these results bring us closer to an incremental understanding of the toxic effects of difenoconazole on zebrafish in its early development, and lend support to the continued use of the early life stages of zebrafish as a classical model to evaluate underlying environmental risks of xenobiotics in aquatic organisms. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Comparison of Baseline Behavior and the Developmental Effects of Diazepam on Locomotor Activity in 3 Strains of Larval Zebrafish (Danio rerio)

    Science.gov (United States)

    Choice of strain is an important consideration in zebrafish husbandry and research. In the scientific literature there is concern that zebrafish strains may behave and respond differently to toxicants. A few studies have compared the baseline behavior of various strains of larv...

  7. Zebrafish foxo3b negatively regulates canonical Wnt signaling to affect early embryogenesis.

    Directory of Open Access Journals (Sweden)

    Xun-wei Xie

    Full Text Available FOXO genes are involved in many aspects of development and vascular homeostasis by regulating cell apoptosis, proliferation, and the control of oxidative stress. In addition, FOXO genes have been showed to inhibit Wnt/β-catenin signaling by competing with T cell factor to bind to β-catenin. However, how important of this inhibition in vivo, particularly in embryogenesis is still unknown. To demonstrate the roles of FOXO genes in embryogenesis will help us to further understand their relevant physiological functions. Zebrafish foxo3b gene, an orthologue of mammalian FOXO3, was expressed maternally and distributed ubiquitously during early embryogenesis and later restricted to brain. After morpholino-mediated knockdown of foxo3b, the zebrafish embryos exhibited defects in axis and neuroectoderm formation, suggesting its critical role in early embryogenesis. The embryo-developmental marker gene staining at different stages, phenotype analysis and rescue assays revealed that foxo3b acted its role through negatively regulating both maternal and zygotic Wnt/β-catenin signaling. Moreover, we found that foxo3b could interact with zebrafish β-catenin1 and β-catenin2 to suppress their transactivation in vitro and in vivo, further confirming its role relevant to the inhibition of Wnt/β-catenin signaling. Taken together, we revealed that foxo3b played a very important role in embryogenesis and negatively regulated maternal and zygotic Wnt/β-catenin signaling by directly interacting with both β-catenin1 and β-catenin2. Our studies provide an in vivo model for illustrating function of FOXO transcription factors in embryogenesis.

  8. Egg and early larval development of laboratory reared dusky grouper, Epinephelus marginatus (Lowe, 1834 (Picies, Serranidae

    Directory of Open Access Journals (Sweden)

    Branko Glamuzina

    1998-12-01

    Full Text Available The embryonic and early larval development of the laboratory-reared dusky grouper, Epinephelus marginatus (Lowe, 1834 are described and illustrated. The eggs, with a mean diameter of 846.68 ± 41 µm and a range from 736-940 µm, were spherical and transparent with transparent chorion. Embryonic development lasted 30 hours at 23°C. Newly-hatched larvae were 1.52 ± 0.066 mm in length. Absorption of the yolk sac was complete after the fourth day, when larvae reached 2.63 ± 0.123 mm in total length. The mouth opened 72 hours after hatching, and was in function after 96 hours, with an opening diameter ranging from 250-300 µm. Larvae had two fields of intensive pigmentation, one above the intestine, and the other between the anus and the end of the notochord.

  9. Toxicity and cardiac effects of carbaryl in early developing zebrafish (Danio rerio) embryos

    International Nuclear Information System (INIS)

    Lin, C.C.; Hui, Michelle N.Y.; Cheng, S.H.

    2007-01-01

    Carbaryl, an acetylcholinesterase inhibitor, is known to be moderately toxic to adult zebrafish and has been reported to cause heart malformations and irregular heartbeat in medaka. We performed experiments to study the toxicity of carbaryl, specifically its effects on the heart, in early developing zebrafish embryos. LC50 and EC50 values for carbaryl at 28 h post-fertilization were 44.66 μg/ml and 7.52 μg/ml, respectively, and 10 μg/ml carbaryl was used in subsequent experiments. After confirming acetylcholinesterase inhibition by carbaryl using an enzymatic method, we observed red blood cell accumulation, delayed hatching and pericardial edema, but not heart malformation as described in some previous reports. Our chronic exposure data also demonstrated carbaryl-induced bradycardia, which is a common effect of acetylcholinesterase inhibitors due to the accumulation of acetylcholine, in embryos from 1 day post-fertilization (dpf) to 5 dpf. The distance between the sinus venosus, the point where blood enters the atrium, and the bulbus arteriosus, the point where blood leaves the ventricle, indicated normal looping of the heart tube. Immunostaining of myosin heavy chains with the ventricle-specific antibody MF20 and the atrium-specific antibody S46 showed normal development of heart chambers. At the same time, acute exposure resulted in carbaryl-induced bradycardia. Heart rate dropped significantly after a 10-min exposure to 100 μg/ml carbaryl but recovered when carbaryl was removed. The novel observation of carbaryl-induced bradycardia in 1- and 2-dpf embryos suggested that carbaryl affected cardiac function possibly through an alternative mechanism other than acetylcholinesterase inhibition such as inhibition of calcium ion channels, since acetylcholine receptors in zebrafish are not functional until 3 dpf. However, the exact nature of this mechanism is currently unknown, and thus further studies are required

  10. Alternative haplotypes of antigen processing genes in zebrafish diverged early in vertebrate evolution

    Science.gov (United States)

    McConnell, Sean C.; Hernandez, Kyle M.; Wcisel, Dustin J.; Kettleborough, Ross N.; Stemple, Derek L.; Andrade, Jorge; de Jong, Jill L. O.

    2016-01-01

    Antigen processing and presentation genes found within the MHC are among the most highly polymorphic genes of vertebrate genomes, providing populations with diverse immune responses to a wide array of pathogens. Here, we describe transcriptome, exome, and whole-genome sequencing of clonal zebrafish, uncovering the most extensive diversity within the antigen processing and presentation genes of any species yet examined. Our CG2 clonal zebrafish assembly provides genomic context within a remarkably divergent haplotype of the core MHC region on chromosome 19 for six expressed genes not found in the zebrafish reference genome: mhc1uga, proteasome-β 9b (psmb9b), psmb8f, and previously unknown genes psmb13b, tap2d, and tap2e. We identify ancient lineages for Psmb13 within a proteasome branch previously thought to be monomorphic and provide evidence of substantial lineage diversity within each of three major trifurcations of catalytic-type proteasome subunits in vertebrates: Psmb5/Psmb8/Psmb11, Psmb6/Psmb9/Psmb12, and Psmb7/Psmb10/Psmb13. Strikingly, nearby tap2 and MHC class I genes also retain ancient sequence lineages, indicating that alternative lineages may have been preserved throughout the entire MHC pathway since early diversification of the adaptive immune system ∼500 Mya. Furthermore, polymorphisms within the three MHC pathway steps (antigen cleavage, transport, and presentation) are each predicted to alter peptide specificity. Lastly, comparative analysis shows that antigen processing gene diversity is far more extensive than previously realized (with ancient coelacanth psmb8 lineages, shark psmb13, and tap2t and psmb10 outside the teleost MHC), implying distinct immune functions and conserved roles in shaping MHC pathway evolution throughout vertebrates. PMID:27493218

  11. Embryonic, Larval, and Early Juvenile Development of the Tropical Sea Urchin, Salmacis sphaeroides (Echinodermata: Echinoidea

    Directory of Open Access Journals (Sweden)

    M. Aminur Rahman

    2012-01-01

    Full Text Available Salmacis sphaeroides (Linnaeus, 1758 is one of the regular echinoids, occuring in the warm Indo-West Pacific, including Johor Straits, between Malaysia and Singapore. In order to investigate the developmental basis of morphological changes in embryos and larvae, we documented the ontogeny of S. sphaeroides in laboratory condition. Gametes were obtained from adult individuals by 0.5 M KCl injection into the coelomic cavity. Fertilization rate at limited sperm concentration (10−5 dilution was 96.6±1.4% and the resulting embryos were reared at 24°C. First cleavage (2-cell, 4-cell, 8-cell, 16-cell, 32-cell, and multicell (Morulla stages were achieved 01.12, 02.03, 02.28, 02.51, 03.12, and 03.32 h postfertilization. Ciliated blastulae with a mean length of 174.72±4.43 μm hatched 08.45 h after sperm entry. The gastrulae formed 16.15 h postfertilization and the archenteron elongated constantly while ectodermal red-pigmented cells migrated synchronously to the apical plate. Pluteus larva started to feed unicellular algae in 2 d, grew continuously, and finally attained metamorphic competence in 35 d after fertilization. Metamorphosis took approximately 1 h 30 min from attachment to the complete resorption of larval tissues and the development of complete juvenile structure with adult spines, extended tubefeet and well-developed pedicellaria, the whole event of which usually took place within 1 d postsettlement. This study represents the first successful investigation on embryonic, larval, and early juvenile development of S. sphaeroides. The findings would greatly be helpful towards the understanding of ontogeny and life-history strategies, which will facilitate us to develop the breeding, seed production, and culture techniques of sea urchins in captive condition.

  12. Redescription of the early larval stages of the pandalid shrimp Chlorotocus crassicornis (Decapoda: Caridea: Pandalidae).

    Science.gov (United States)

    Landeira, Jose M; Jiang, Guo-Chen; Chan, Tin-Yam; Shih, Tung-Wei; Gozález-Gordillo, J Ignacio

    2015-09-07

    The first four larval stages of the pandalid shrimp Chlorotocus crassicornis (A. Costa, 1871) are described and illustrated from laboratory-reared material obtained from ovigerous females collected in the southwestern Spain and south Taiwan. The second to fourth larval stages of this species are reported for the first time to science. Detailed examination of the first larval stages reveals that previous description misidentified some key larval characters which have prevented its identification in plankton samples. It is found that the zoeal morphology of Chlorotocus is not very different from other pandalid larvae, and in fact closely resembles Plesionika and Heterocarpus.

  13. Light impacts embryonic and early larval development of the European eel, Anguilla anguilla

    DEFF Research Database (Denmark)

    Politis, Sebastian Nikitas; Butts, Ian; Tomkiewicz, Jonna

    2014-01-01

    Little is known about the natural ecology of European eel during early life history. Weextend our understandings on the ecology of this species by studying howearly life stages perform under various light regimes.We assessed the effects of intensity, photoperiod (12:12 and 24:0 h light/dark) and ......Little is known about the natural ecology of European eel during early life history. Weextend our understandings on the ecology of this species by studying howearly life stages perform under various light regimes.We assessed the effects of intensity, photoperiod (12:12 and 24:0 h light...... stages. In particular, for the 12:12 h photoperiod, embryonic survival, until 26 h post-fertilization was significantly higher when reared under low (62 ± 13%) than those reared under high intensity light (42 ± 13%). Furthermore, embryos reared in low light had a higher hatch success (16 ± 7%) than those...... in high intensity light (12 ± 7%). Larval yolk-sac area was significantly affected by photoperiod and body area was significantly affected by the interaction between intensity × photoperiod. The highest incidence of deformities (75%) occurred when embryos were reared in high intensity white light under...

  14. Impacts of triclosan exposure on zebrafish early-life stage: Toxicity and acclimation mechanisms.

    Science.gov (United States)

    Falisse, Elodie; Voisin, Anne-Sophie; Silvestre, Frédéric

    2017-08-01

    Triclosan (TCS) is a broad spectrum antibacterial agent widely used in personal care products and present in most aquatic ecosystems. This study investigated the occurrence of triclosan acclimation and the biological mechanisms underlying the stress response triggered in early-life stage of zebrafish. Zebrafish eggs were first exposed to four different sublethal concentrations of TCS (2, 20, 50 and 100μg/L) for 7days following fertilization and subsequently exposed to a lethal concentration of TCS (1000μg/L). During the time-to-death exposure (TTD), mortality was continuously recorded to evaluate if increased resistance occurred. Overall, larvae exposed to 50μg/L of TCS demonstrated higher sensitivity, with delayed hatching and increased mortality during the sub-lethal exposure and significant lower mean time-to-death (TTD) value compared to the other groups. Interestingly, fish exposed to the highest concentration of TCS (100μg/L) presented a similar mean TTD value as controls and a significantly better survival in comparison with embryos exposed to 50μg/L, suggesting that acclimation process has been triggered at this concentration. Proteomic and enzymatic analyses were conducted on 7days post fertilization (dpf) larvae exposed to 50μg/L and 100μg/L of TCS giving insights into the functional changes triggered at those specific concentrations. TCS seemed to affect proteins involved in cytoskeleton, stress response, eyes and neuronal development. This was endorsed by the enzymatic results, which suggest impairment in glutathione metabolism and acute neurotoxicity. A significant 2.5-fold and 3-fold increase of AChE activity was observed following TCS exposure. Moreover, GPx activity was significantly increased whereas a significant inhibition of GR activity was observed, suggesting that de novo synthesis of reduced GSH might occur in order to maintain the ratio between reduced and oxidized GSH. Proteomic results revealed possible candidate protein involved in

  15. Intrinsic Xenobiotic Metabolizing Enzyme Activities in Early Life Stages of Zebrafish (Danio rerio).

    Science.gov (United States)

    Otte, Jens C; Schultz, Bernadette; Fruth, Daniela; Fabian, Eric; van Ravenzwaay, Bennard; Hidding, Björn; Salinas, Edward R

    2017-09-01

    Early life stages of zebrafish (Danio rerio, zf) are gaining attention as an alternative invivo test system for drug discovery, early developmental toxicity screenings and chemical testing in ecotoxicological and toxicological testing strategies. Previous studies have demonstrated transcriptional evidence for xenobiotic metabolizing enzymes (XME) during early zf development. However, elaborate experiments on XME activities during development are incomplete. In this work, the intrinsic activities of representative phase I and II XME were monitored by transformation of putative zf model substrates analyzed using photometry and high pressure liquid chromatography techniques. Six different defined stages of zf development (between 2.5 h postfertilization (hpf) to 120 hpf) were investigated by preparing a subcellular fraction from whole organism homogenates. We demonstrated that zf embryos as early as 2.5 hpf possess intrinsic metabolic activities for esterase, Aldh, Gst, and Cyp1a above the methodological detection limit. The activities of the enzymes Cyp3a and Nat were measurable during later stages in development. Activities represent dynamic patterns during development. The role of XME activities revealed in this work is relevant for the assessing toxicity in this test system and therefore contributes to a valuable characterization of zf embryos as an alternative testing organism in toxicology. © The Author 2017. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  16. Apoptosis-related genes induced in response to ketamine during early life stages of zebrafish.

    Science.gov (United States)

    Félix, Luís M; Serafim, Cindy; Valentim, Ana M; Antunes, Luís M; Matos, Manuela; Coimbra, Ana M

    2017-09-05

    Increasing evidence supports that ketamine, a widely used anaesthetic, potentiates apoptosis during development through the mitochondrial pathway of apoptosis. Defects in the apoptotic machinery can cause or contribute to the developmental abnormalities previously described in ketamine-exposed zebrafish. The involvement of the apoptotic machinery in ketamine-induced teratogenicity was addressed by assessing the apoptotic signals at 8 and 24 hpf following 20min exposure to ketamine at three stages of early zebrafish embryo development (256 cell, 50% epiboly and 1-4 somites stages). Exposure at the 256-cell stage to ketamine induced an up-regulation of casp8 and pcna at 8 hpf while changes in pcna at the mRNA level were observed at 24 hpf. After the 50% epiboly stage exposure, the mRNA levels of casp9 were increased at 8 and 24 hpf while aifm1 was affected at 24 hpf. Both tp53 and pcna expressions were increased at 8 hpf. After exposure during the 1-4 somites stage, no meaningful changes on transcript levels were observed. The distribution of apoptotic cells and the caspase-like enzymatic activities of caspase-3 and -9 were not affected by ketamine exposure. It is proposed that ketamine exposure at the 256-cell stage induced a cooperative mechanism between proliferation and cellular death while following exposure at the 50% epiboly, a p53-dependent and -independent caspase activation may occur. Finally, at the 1-4 somites stage, the defence mechanisms are already fully in place to protect against ketamine-insult. Thus, ketamine teratogenicity seems to be dependent on the functional mechanisms present in each developmental stage. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Lnx2 ubiquitin ligase is essential for exocrine cell differentiation in the early zebrafish pancreas.

    Science.gov (United States)

    Won, Minho; Ro, Hyunju; Dawid, Igor B

    2015-10-06

    The gene encoding the E3 ubiquitin ligase Ligand of Numb protein-X (Lnx)2a is expressed in the ventral-anterior pancreatic bud of zebrafish embryos in addition to its expression in the brain. Knockdown of Lnx2a by using an exon 2/intron 2 splice morpholino resulted in specific inhibition of the differentiation of ventral bud derived exocrine cell types, with little effect on endocrine cell types. A frame shifting null mutation in lnx2a did not mimic this phenotype, but a mutation that removed the exon 2 splice donor site did. We found that Lnx2b functions in a redundant manner with its paralog Lnx2a. Inhibition of lnx2a exon 2/3 splicing causes exon 2 skipping and leads to the production of an N-truncated protein that acts as an interfering molecule. Thus, the phenotype characterized by inhibition of exocrine cell differentiation requires inactivation of both Lnx2a and Lnx2b. Human LNX1 is known to destabilize Numb, and we show that inhibition of Numb expression rescues the Lnx2a/b-deficient phenotype. Further, Lnx2a/b inhibition leads to a reduction in the number of Notch active cells in the pancreas. We suggest that Lnx2a/b function to fine tune the regulation of Notch through Numb in the differentiation of cell types in the early zebrafish pancreas. Further, the complex relationships among genotype, phenotype, and morpholino effect in this case may be instructive in the ongoing consideration of morpholino use.

  18. Using the Larval Zebrafish Locomotor Assay in Functional Neurotoxicity Screening: Light Intensity and the Order of Stimulus Presentation Affect the Outcome

    Science.gov (United States)

    The U.S. Environmental Protection Agency is evaluating methods to screen and prioritize large numbers of chemicals using 6 day old zebrafish (Danio rerio) as an alternative test model for detecting neurotoxic chemicals. We use a behavioral testing paradigm that simultaneously tes...

  19. Inhibition of signaling between human CXCR4 and zebrafish ligands by the small molecule IT1t impairs the formation of triple-negative breast cancer early metastases in a zebrafish xenograft model

    Directory of Open Access Journals (Sweden)

    Claudia Tulotta

    2016-02-01

    Full Text Available Triple-negative breast cancer (TNBC is a highly aggressive and recurrent type of breast carcinoma that is associated with poor patient prognosis. Because of the limited efficacy of current treatments, new therapeutic strategies need to be developed. The CXCR4-CXCL12 chemokine signaling axis guides cell migration in physiological and pathological processes, including breast cancer metastasis. Although targeted therapies to inhibit the CXCR4-CXCL12 axis are under clinical experimentation, still no effective therapeutic approaches have been established to block CXCR4 in TNBC. To unravel the role of the CXCR4-CXCL12 axis in the formation of TNBC early metastases, we used the zebrafish xenograft model. Importantly, we demonstrate that cross-communication between the zebrafish and human ligands and receptors takes place and human tumor cells expressing CXCR4 initiate early metastatic events by sensing zebrafish cognate ligands at the metastatic site. Taking advantage of the conserved intercommunication between human tumor cells and the zebrafish host, we blocked TNBC early metastatic events by chemical and genetic inhibition of CXCR4 signaling. We used IT1t, a potent CXCR4 antagonist, and show for the first time its promising anti-tumor effects. In conclusion, we confirm the validity of the zebrafish as a xenotransplantation model and propose a pharmacological approach to target CXCR4 in TNBC.

  20. Effects of Simulated Microgravity on Otolith Growth of Larval Zebrafish using a Rotating-Wall Vessel: Appropriate Rotation Speed and Fish Developmental Stage

    Science.gov (United States)

    Li, Xiaoyan; Anken, Ralf; Liu, Liyue; Wang, Gaohong; Liu, Yongding

    2017-02-01

    Stimulus dependence is a general feature of developing animal sensory systems. In this respect, it has extensively been shown earlier that fish inner ear otoliths can act as test masses as their growth is strongly affected by altered gravity such as hypergravity obtained using centrifuges, by (real) microgravity achieved during spaceflight or by simulated microgravity using a ground-based facility. Since flight opportunities are scarce, ground-based simulators of microgravity, using a wide variety of physical principles, have been developed to overcome this shortcoming. Not all of them, however, are equally well suited to provide functional weightlessness from the perspective of the biosystem under evaluation. Therefore, the range of applicability of a particular simulator has to be extensively tested. Earlier, we have shown that a Rotating-Wall Vessel (RWV) can be used to provide simulated microgravity for developing Zebrafish regarding the effect of rotation on otolith development. In the present study, we wanted to find the most effective speed of rotation and identify the appropriate developmental stage of Zebrafish, where effects are the largest, in order to provide a methodological basis for future in-depth analyses dedicated to the physiological processes underlying otolith growth at altered gravity. Last not least, we compared data on the effect of simulated microgravity on the size versus the weight of otoliths, since the size usually is measured in related studies due to convenience, but the weight more accurately approximates the physical capacity of an otolith. Maintaining embryos at 10 hours post fertilization for three days in the RWV, we found that 15 revolutions per minute (rpm) yielded the strongest effects on otolith growth. Maintenance of Zebrafish staged at 10 hpf, 1 day post fertilization (dpf), 4 dpf, 7 dpf and 14 dpf for three days at 15 rpm resulted in the most prominent effects in 7 dpf larvae. Weighing versus measuring the size of otoliths

  1. Distribution of the early larval stages of cod, plaice and lesser sandeel across haline fronts in the North Sea

    DEFF Research Database (Denmark)

    Munk, Peter; Wright, P.J.; Pihl, Niels Jørgen

    2002-01-01

    A number of commercially important fish species spawn in the coastal areas of the North Sea in the late winter, including cod (Gadus morhua), plaice (Pleuronectes platessa) and lesser sandeel (Ammodytes marinus). The distribution of the early stages of these species overlap to some extent...... Influence (ROFI), predominantly in the Dogger Bank and German Bight areas. There was a high degree of overlap between the distributions of cod and plaice, while the maximal abundance of lesser sandeel was found inshore of the other species. Larval distributions were to a large extent confined by the frontal...

  2. Gene Expression Patterns during the Early Stages of Chemically Induced Larval Metamorphosis and Settlement of the Coral Acropora millepora

    Science.gov (United States)

    Siboni, Nachshon; Abrego, David; Motti, Cherie A.; Tebben, Jan; Harder, Tilmann

    2014-01-01

    The morphogenetic transition of motile coral larvae into sessile primary polyps is triggered and genetically programmed upon exposure to environmental biomaterials, such as crustose coralline algae (CCA) and bacterial biofilms. Although the specific chemical cues that trigger coral larval morphogenesis are poorly understood there is much more information available on the genes that play a role in this early life phase. Putative chemical cues from natural biomaterials yielded defined chemical samples that triggered different morphogenetic outcomes: an extract derived from a CCA-associated Pseudoalteromonas bacterium that induced metamorphosis, characterized by non-attached metamorphosed juveniles; and two fractions of the CCA Hydrolithon onkodes (Heydrich) that induced settlement, characterized by attached metamorphosed juveniles. In an effort to distinguish the genes involved in these two morphogenetic transitions, competent larvae of the coral Acropora millepora were exposed to these predictable cues and the expression profiles of 47 coral genes of interest (GOI) were investigated after only 1 hour of exposure using multiplex RT–qPCR. Thirty-two GOI were differentially expressed, indicating a putative role during the early regulation of morphogenesis. The most striking differences were observed for immunity-related genes, hypothesized to be involved in cell recognition and adhesion, and for fluorescent protein genes. Principal component analysis of gene expression profiles resulted in separation between the different morphogenetic cues and exposure times, and not only identified those genes involved in the early response but also those which influenced downstream biological changes leading to larval metamorphosis or settlement. PMID:24632854

  3. Epilepsy, Behavioral Abnormalities, and Physiological Comorbidities in Syntaxin-Binding Protein 1 (STXBP1 Mutant Zebrafish.

    Directory of Open Access Journals (Sweden)

    Brian P Grone

    Full Text Available Mutations in the synaptic machinery gene syntaxin-binding protein 1, STXBP1 (also known as MUNC18-1, are linked to childhood epilepsies and other neurodevelopmental disorders. Zebrafish STXBP1 homologs (stxbp1a and stxbp1b have highly conserved sequence and are prominently expressed in the larval zebrafish brain. To understand the functions of stxbp1a and stxbp1b, we generated loss-of-function mutations using CRISPR/Cas9 gene editing and studied brain electrical activity, behavior, development, heart physiology, metabolism, and survival in larval zebrafish. Homozygous stxbp1a mutants exhibited a profound lack of movement, low electrical brain activity, low heart rate, decreased glucose and mitochondrial metabolism, and early fatality compared to controls. On the other hand, homozygous stxbp1b mutants had spontaneous electrographic seizures, and reduced locomotor activity response to a movement-inducing "dark-flash" visual stimulus, despite showing normal metabolism, heart rate, survival, and baseline locomotor activity. Our findings in these newly generated mutant lines of zebrafish suggest that zebrafish recapitulate clinical phenotypes associated with human syntaxin-binding protein 1 mutations.

  4. Toxicity of effluents from gasoline stations oil-water separators to early life stages of zebrafish Danio rerio.

    Science.gov (United States)

    Alves, Romulo Nepomuceno; Mariz, Célio Freire; Paulo, Driele Ventura de; Carvalho, Paulo S M

    2017-07-01

    Used petroleum hydrocarbons and gasoline stations runoff are significant sources of polycyclic aromatic hydrocarbons (PAHs) to aquatic ecosystems. Samples of the final effluent of oil-water-separators were collected at gasoline stations in the metropolitan region of Recife, Brazil, before release to sewage or rainwater systems. Effluent soluble fractions (ESF) were prepared and bioassays were performed according to the Fish Embryo Toxicity Test. The test involved exposing zebrafish Danio rerio embryos to dilutions of the ESFs for 96 h, with daily examination of lethality and sublethal morphological effects integrated through the General Morphology Score (GMS), based on the achievement of developmental hallmarks. Frequencies of abnormalities were recorded after exposures. ESF LC50-96h (lethal concentration to 50% of exposed embryos) in the most toxic effluent achieved 8.9% (v/v), equivalent to 11 μg phenanthrene equivalents L -1 . GMS scores indicated significantly delayed embryo-larval development at ESF dilutions of 10% and 20% from effluents of all gas stations. Major abnormalities detected after the 96 h exposure included the presence of a yolk sac not fully absorbed coupled with the lack of an inflated swim bladder, lack of both pectoral fins, and the failure to develop a protruding mouth. Effective equivalent PAH concentrations that induce a 50% frequency of larvae without an inflated swim bladder (EC50) were 4.9 μg phenanthrene L -1 , 21.8 μg naphthalene L -1 , and 34.1 μg chrysene L -1 . This study shows that PAHs in ESFs from gas stations oil water separators are toxic to zebrafish, contributing to the toxicity of urban storm waters. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Kaempferol suppresses lipid accumulation by inhibiting early adipogenesis in 3T3-L1 cells and zebrafish.

    Science.gov (United States)

    Lee, Yeon-Joo; Choi, Hyeon-Son; Seo, Min-Jung; Jeon, Hui-Jeon; Kim, Kui-Jin; Lee, Boo-Yong

    2015-08-01

    Kaempferol is a flavonoid present in Kaempferia galanga and Opuntia ficus indica var. saboten. Recent studies have suggested that it has anti-oxidant, anti-inflammatory, anti-cancer, and anti-obesity effects. In this study, we focused on the anti-adipogenic effects of kaempferol during adipocyte differentiation. The results showed that kaempferol inhibits lipid accumulation in adipocytes and zebrafish. Oil Red O and Nile Red staining showed that the number of intracellular lipid droplets decreased in adipocytes and zebrafish treated with kaempferol. LPAATθ (lysophosphatidic acid acyltransferase), lipin1, and DGAT1 (triglyceride synthetic enzymes) and FASN and SREBP-1C (fatty acid synthetic proteins) showed decreased expression levels in the presence of kaempferol. In addition, treatment of kaempferol showed an inhibitory activity on cell cycle progression. Kaempferol delayed cell cycle progression from the S to G2/M phase through the regulation of cyclins in a dose-dependent manner. Kaempferol blocked the phosphorylation of AKT (protein kinase B) and mammalian target of rapamycin (mTOR) signaling pathway during the early stages of adipogenesis. In addition, kaempferol down-regulated pro-early adipogenic factors such as CCAAT-enhancer binding proteins β (C/EBPβ), and Krüppel-like factors (KLFs) 4 and 5, while anti-early adipogenic factors, such as KLF2 and pref-1(preadipocyte factor-1), were upregulated. These kaempferol-mediated regulations of early adipogenic factors resulted in the attenuation of late adipogenic factors such as C/EBPα and peroxisome proliferator-activated receptor γ (PPARγ). These results were supported in zebrafish based on the decrease in lipid accumulation and expression of adipogenic factors. Our results indicated that kaempferol might have an anti-obesity effect by regulating lipid metabolism.

  6. Learning and memory in zebrafish larvae

    Science.gov (United States)

    Roberts, Adam C.; Bill, Brent R.; Glanzman, David L.

    2013-01-01

    Larval zebrafish possess several experimental advantages for investigating the molecular and neural bases of learning and memory. Despite this, neuroscientists have only recently begun to use these animals to study memory. However, in a relatively short period of time a number of forms of learning have been described in zebrafish larvae, and significant progress has been made toward their understanding. Here we provide a comprehensive review of this progress; we also describe several promising new experimental technologies currently being used in larval zebrafish that are likely to contribute major insights into the processes that underlie learning and memory. PMID:23935566

  7. Deiodinase knockdown during early zebrafish development affects growth, development, energy metabolism, motility and phototransduction.

    Directory of Open Access Journals (Sweden)

    Enise Bagci

    Full Text Available Thyroid hormone (TH balance is essential for vertebrate development. Deiodinase type 1 (D1 and type 2 (D2 increase and deiodinase type 3 (D3 decreases local intracellular levels of T3, the most important active TH. The role of deiodinase-mediated TH effects in early vertebrate development is only partially understood. Therefore, we investigated the role of deiodinases during early development of zebrafish until 96 hours post fertilization at the level of the transcriptome (microarray, biochemistry, morphology and physiology using morpholino (MO knockdown. Knockdown of D1+D2 (D1D2MO and knockdown of D3 (D3MO both resulted in transcriptional regulation of energy metabolism and (muscle development in abdomen and tail, together with reduced growth, impaired swim bladder inflation, reduced protein content and reduced motility. The reduced growth and impaired swim bladder inflation in D1D2MO could be due to lower levels of T3 which is known to drive growth and development. The pronounced upregulation of a large number of transcripts coding for key proteins in ATP-producing pathways in D1D2MO could reflect a compensatory response to a decreased metabolic rate, also typically linked to hypothyroidism. Compared to D1D2MO, the effects were more pronounced or more frequent in D3MO, in which hyperthyroidism is expected. More specifically, increased heart rate, delayed hatching and increased carbohydrate content were observed only in D3MO. An increase of the metabolic rate, a decrease of the metabolic efficiency and a stimulation of gluconeogenesis using amino acids as substrates may have been involved in the observed reduced protein content, growth and motility in D3MO larvae. Furthermore, expression of transcripts involved in purine metabolism coupled to vision was decreased in both knockdown conditions, suggesting that both may impair vision. This study provides new insights, not only into the role of deiodinases, but also into the importance of a correct

  8. Influence of the temperature on the early larval development of the Pacific red snapper, Lutjanus peru (Nichols & Murphy, 1922

    Directory of Open Access Journals (Sweden)

    José Antonio Estrada-Godínez

    2015-03-01

    Full Text Available The Pacific red snapper, Lutjanus peru, is a commercially important species throughout its distribution range, making it a good alternative for aquaculture; however, there is few information regarding environmental conditions and their influence on early development of this species. Temperature is one of the main factors affecting embryo and larval development in marine fishes. In this paper, the effects of different temperatures upon hatching rate, growth, consumption of yolk sac and oil droplet and the formation of the digestive system and eye pigmentation were evaluated in larvae of this species under experimental conditions. Eggs incubated between 20 and 32°C showed hatching rates higher than 90%. However, larvae maintained at 26°C showed significantly larger notochord length and were the first to complete the pigmentation of the eyes and the formation of the digestive system when still possessing enough reserves in the yolk sac. Therefore, according to the results obtained, it is recommended that the incubation of eggs and larval rearing in Pacific red snapper takes place between 25 and 26°C.

  9. Caffeine induces high expression of cyp-35A family genes and inhibits the early larval development in Caenorhabditis elegans.

    Science.gov (United States)

    Min, Hyemin; Kawasaki, Ichiro; Gong, Joomi; Shim, Yhong-Hee

    2015-03-01

    Intake of caffeine during pregnancy can cause retardation of fetal development. Although the significant influence of caffeine on animal development is widely recognized, much remains unknown about its mode of action because of its pleiotropic effects on living organisms. In the present study, by using Caenorhabditis elegans as a model organism, the effects of caffeine on development were examined. Brood size, embryonic lethality, and percent larval development were investigated, and caffeine was found to inhibit the development of C. elegans at most of the stages in a dosage-dependent fashion. Upon treatment with 30 mM caffeine, the majority (86.1 ± 3.4%) of the L1 larvae were irreversibly arrested without further development. In contrast, many of the late-stage larvae survived and grew to adults when exposed to the same 30 mM caffeine. These results suggest that early-stage larvae are more susceptible to caffeine than later-stage larvae. To understand the metabolic responses to caffeine treatment, the levels of expression of cytochrome P450 (cyp) genes were examined with or without caffeine treatment using comparative micro-array, and it was found that the expression of 24 cyp genes was increased by more than 2-fold (p family was the most prominent. Interestingly, depletion of the cyp-35A family genes one-by-one or in combination through RNA interference resulted in partial rescue from early larval developmental arrest caused by caffeine treatment, suggesting that the high-level induction of cyp-35A family genes can be fatal to the development of early-stage larvae.

  10. Biomarkers as a tool to assess effects of chromium (VI): comparison of responses in zebrafish early life stages and adults.

    Science.gov (United States)

    Domingues, Inês; Oliveira, Rhaul; Lourenço, Joana; Grisolia, Cesar Koppe; Mendo, Sónia; Soares, A M V M

    2010-09-01

    The present work aims to compare the sensitivity of embryos and adult zebrafish to chromium (VI) (as potassium dichromate) focusing on biomarkers (cholinesterase, glutathione S-transferase and lactate dehydrogenase) as endpoints. Zebrafish eggs showed less sensitivity to Cr (VI) (96 h-LC50=145.7 mg/L) than adults (96 h-LC50=39.4 mg/L) probably due to the protective action of the chorion. However, biomarkers were much more responsive in larvae than in adults and gave clear indications about Cr (VI) mode of action: it seems to be neurotoxic (inhibited cholinesterase), to inhibit glutathione S-transferase activity and to interfere with cellular metabolic activity (changes in lactate dehydrogenase activity) in larvae. In adults, only glutathione S-transferase was responsive, showing a clear inhibition. The responsiveness of the analyzed biomarkers in larvae reinforces the idea of the usefulness of early life stage assays in the assessment of chemicals effects. Moreover, early life stage assays also contributed with relevant information regarding anomalies in larvae development and behavior. Further research should focus on the use of biomarkers to assess long term effects which are ecologically more relevant. Copyright (c) 2010 Elsevier Inc. All rights reserved.

  11. Pharmacologic modeling of primary mitochondrial respiratory chain dysfunction in zebrafish.

    Science.gov (United States)

    Byrnes, James; Ganetzky, Rebecca; Lightfoot, Richard; Tzeng, Michael; Nakamaru-Ogiso, Eiko; Seiler, Christoph; Falk, Marni J

    2017-07-18

    Mitochondrial respiratory chain (RC) disease is a heterogeneous and highly morbid group of energy deficiency disorders for which no proven effective therapies exist. Robust vertebrate animal models of primary RC dysfunction are needed to explore the effects of variation in RC disease subtypes, tissue-specific manifestations, and major pathogenic factors contributing to each disorder, as well as their pre-clinical response to therapeutic candidates. We have developed a series of zebrafish (Danio rerio) models that inhibit, to variable degrees, distinct aspects of RC function, and enable quantification of animal development, survival, behaviors, and organ-level treatment effects as well as effects on mitochondrial biochemistry and physiology. Here, we characterize four pharmacologic inhibitor models of mitochondrial RC dysfunction in early larval zebrafish, including rotenone (complex I inhibitor), azide (complex IV inhibitor), oligomycin (complex V inhibitor), and chloramphenicol (mitochondrial translation inhibitor that leads to multiple RC complex dysfunction). A range of concentrations and exposure times of each RC inhibitor were systematically evaluated on early larval development, animal survival, integrated behaviors (touch and startle responses), organ physiology (brain death, neurologic tone, heart rate), and fluorescence-based analyses of mitochondrial physiology in zebrafish skeletal muscle. Pharmacologic RC inhibitor effects were validated by spectrophotometric analysis of Complex I, II and IV enzyme activities, or relative quantitation of ATP levels in larvae. Outcomes were prioritized that utilize in vivo animal imaging and quantitative behavioral assessments, as may optimally inform the translational potential of pre-clinical drug screens for future clinical study in human mitochondrial disease subjects. The RC complex inhibitors each delayed early embryo development, with short-term exposures of these three agents or chloramphenicol from 5 to 7 days

  12. Exposure to low dose benzo[a]pyrene during early life stages causes symptoms similar to cardiac hypertrophy in adult zebrafish.

    Science.gov (United States)

    Huang, Lixing; Gao, Dongxu; Zhang, Youyu; Wang, Chonggang; Zuo, Zhenghong

    2014-07-15

    Growing evidence indicates that polycyclic aromatic hydrocarbons (PAHs) can lead to cardiac hypertrophy and recent research indicates that exposure to low dose crude oil during early embryonic development may lead to impacts on heart health at later life stages. The aim of this study was to evaluate whether exposure during early life stages to low dose benzo[a]pyrene (BaP), as a high-ring PAH, would lead to cardiac hypertrophy at later life stages. Zebrafish were exposed to low dose BaP until 96 hpf, then transferred to clean water and maintained for a year before histological and molecular biological analysis. Our results showed that exposure to low level BaP during early life stages increased heart weight to body weight ratios and deposited collagen in the heart of adult zebrafish. ANP, BNP and c-Myc were also induced in the heart of adult zebrafish by BaP. These results proved that low level BaP exposure during early life stages caused symptoms similar to cardiac hypertrophy in adult zebrafish. Our results displayed an elevated expression of CdC42, RhoA, p-ERK1, 2 and Rac1. Therefore, the mechanism of the cardiac hypertrophy caused by BaP exposure during early life stages may be through inducing the expression of CdC42, RhoA and Rac1, together with activating ERK1, 2. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Human amyloidogenic light chain proteins result in cardiac dysfunction, cell death, and early mortality in zebrafish.

    Science.gov (United States)

    Mishra, Shikha; Guan, Jian; Plovie, Eva; Seldin, David C; Connors, Lawreen H; Merlini, Giampaolo; Falk, Rodney H; MacRae, Calum A; Liao, Ronglih

    2013-07-01

    Systemic amyloid light-chain (AL) amyloidosis is associated with rapidly progressive and fatal cardiomyopathy resulting from the direct cardiotoxic effects of circulating AL light chain (AL-LC) proteins and the indirect effects of AL fibril tissue infiltration. Cardiac amyloidosis is resistant to standard heart failure therapies, and, to date, there are limited treatment options for these patients. The mechanisms underlying the development of cardiac amyloidosis and AL-LC cardiotoxicity are largely unknown, and their study has been limited by the lack of a suitable in vivo model system. Here, we establish an in vivo zebrafish model of human AL-LC-induced cardiotoxicity. AL-LC isolated from AL cardiomyopathy patients or control nonamyloidogenic LC protein isolated from multiple myeloma patients (Con-LC) was directly injected into the circulation of zebrafish at 48 h postfertilization. AL-LC injection resulted in impaired cardiac function, pericardial edema, and increased cell death relative to Con-LC, culminating in compromised survival with 100% mortality within 2 wk, independent of AL fibril deposition. Prior work has implicated noncanonical p38 MAPK activation in the pathogenesis of AL-LC-induced cardiotoxicity, and p38 MAPK inhibition via SB-203580 rescued AL-LC-induced cardiac dysfunction and cell death and attenuated mortality in zebrafish. This in vivo zebrafish model of AL-LC cardiotoxicity demonstrates that antagonism of p38 MAPK within the AL-LC cardiotoxic signaling response may serve to improve cardiac function and mortality in AL cardiomyopathy. Furthermore, this in vivo model system will allow for further study of the molecular underpinnings of AL cardiotoxicity and identification of novel therapeutic strategies.

  14. EVALUATING THE EFFECTS OF FLY ASH EXPOSURE ON FISH EARLY LIFE STAGES: FATHEAD MINNOW EMBRYO-LARVAL TESTS

    Energy Technology Data Exchange (ETDEWEB)

    Greeley Jr, Mark Stephen [ORNL; Elmore, Logan R [ORNL; McCracken, Kitty [ORNL

    2012-05-01

    On December 22, 2008, a dike containing fly ash and bottom ash in an 84-acre complex of the Tennessee Valley Authority's (TVA) Kingston Steam Plant in East Tennessee failed and released a large quantity of ash into the adjacent Emory River. Ash deposits extended as far as 4 miles upstream (Emory River mile 6) of the Plant, and some ash was carried as far downstream as Tennessee River mile 564 ({approx}4 miles downstream of the Tennessee River confluence with the Clinch River). A byproduct of coal burning power plants, fly ash contains a variety of metals and other elements which, at sufficient concentrations and in specific forms, can be toxic to biological systems. The effects of fly ash contamination on exposed fish populations depend on the magnitude and duration of exposure, with the most significant risk considered to be the effects of specific ash constituents, especially selenium, on fish early life stages. Uptake by adult female fish of fly ash constituents through the food chain and subsequent maternal transfer of contaminants to the developing eggs is thought to be the primary route of selenium exposure to larval fish (Woock and others 1987, Coyle and others 1993, Lemly 1999, Moscatello and others 2006), but direct contact of the fertilized eggs and developing embryos to ash constituents in river water and sediments is also a potential risk factor (Woock and others 1987, Coyle and others 1993, Jezierska and others 2009). To address the risk of fly ash from the Kingston spill to the reproductive health of downstream fish populations, ORNL has undertaken a series of studies in collaboration with TVA including: (1) a field study of the bioaccumulation of fly ash constituents in fish ovaries and the reproductive condition of sentinel fish species in reaches of the Emory and Clinch Rivers affected by the fly ash spill; (2) laboratory tests of the potential toxicity of fly ash from the spill area on fish embryonic and larval development (reported in the

  15. Seeing the whole picture: A comprehensive imaging approach to functional mapping of circuits in behaving zebrafish.

    Science.gov (United States)

    Feierstein, C E; Portugues, R; Orger, M B

    2015-06-18

    In recent years, the zebrafish has emerged as an appealing model system to tackle questions relating to the neural circuit basis of behavior. This can be attributed not just to the growing use of genetically tractable model organisms, but also in large part to the rapid advances in optical techniques for neuroscience, which are ideally suited for application to the small, transparent brain of the larval fish. Many characteristic features of vertebrate brains, from gross anatomy down to particular circuit motifs and cell-types, as well as conserved behaviors, can be found in zebrafish even just a few days post fertilization, and, at this early stage, the physical size of the brain makes it possible to analyze neural activity in a comprehensive fashion. In a recent study, we used a systematic and unbiased imaging method to record the pattern of activity dynamics throughout the whole brain of larval zebrafish during a simple visual behavior, the optokinetic response (OKR). This approach revealed the broadly distributed network of neurons that were active during the behavior and provided insights into the fine-scale functional architecture in the brain, inter-individual variability, and the spatial distribution of behaviorally relevant signals. Combined with mapping anatomical and functional connectivity, targeted electrophysiological recordings, and genetic labeling of specific populations, this comprehensive approach in zebrafish provides an unparalleled opportunity to study complete circuits in a behaving vertebrate animal. Copyright © 2014. Published by Elsevier Ltd.

  16. Environmental conditions, early life stages distributions and larval feeding of patagonian sprat Sprattusfuegensis and common sardine Strangomerabentincki in fjords and channels of the northern Chilean patagonia

    Science.gov (United States)

    Contreras, Tabit; Castro, Leonardo R.; Montecinos, Sandra; Gonzalez, Humberto E.; Soto, Samuel; Muñoz, Maria I.; Palma, Sergio

    2014-12-01

    We assessed ontogenetic changes in distribution and feeding of the Patagonian sprat Sprattus fuegensis and common sardine Strangomera bentincki, and their association with environmental characteristics (hydrography, larval food, gelatinous zooplankton predators), and actual feeding from inshore to offshore areas of the Chilean Patagonia. During the springs of 2007 and 2008, S. bentincki egg and larvae were present north of the Taitao Peninsula (47°S) and S. fuegensis was found to the south of the peninsula. Along the inshore-offshore axis, distributions also differed: while eggs and early larval stages of S. bentincki occurred inshore and seawards, larger larvae occurred mostly seawards. The opposite was observed in S. fuegensis. However, distributions of both species followed the same rule, determined by the size of their prey: eggs and early larval stages occurred in areas of higher abundance of small prey sizes, and larger larvae coincided with the highest abundances of larger prey sizes. No relationship was detected between potential gelatinous predators and the egg and larval distributions of both fish species. Mean ingested prey sizes in both species increased as larvae grew, while maintaining the capacity to feed on small sized items. This ontogenetic feeding pattern and the distributions linked to prey seem to be beneficial in order to take advantage of short term food pulses and to overcome the strong changes in environmental conditions east to west from fjords to open waters.

  17. The effect of excess expression of GFP in a novel heart-specific green fluorescence zebrafish regulated by nppa enhancer at early embryonic development.

    Science.gov (United States)

    Huang, Wen; Deng, Yun; Dong, Wei; Yuan, Wuzhou; Wan, Yongqi; Mo, Xiaoyan; Li, Yongqing; Wang, Zequn; Wang, Yuequn; Ocorr, Karen; Zhang, Bo; Lin, Shuo; Wu, Xiushan

    2011-02-01

    In order to study the impalpable effect of GFP in homozygous heart-specific GFP-positive zebrafish during the early stage, the researchers analyzed the heart function of morphology and physiology at the first 3 days after fertilization. This zebrafish line was produced by a large-scale Tol2 transposon mediated enhancer trap screen that generated a transgenic zebrafish with a heart-specific expression of green fluorescent protein (GFP)-tagged under control of the nppa enhancer. In situ hybridization experiments showed that the nppa:GFP line faithfully recapitulated both the spatial and temporal expressions of the endogenous nppa. Green fluorescence was intensively and specifically expressed in the myocardial cells located both in the heart chambers and in the atrioventricular canal. The embryonic heart of nppa:GFP line developed normally compared with those in the wild type. There was no difference between the nappa:GFP and wild type lines with respect to heart rate, overall size, ejection volume, and fractional shortening. Thus the excess expression of GFP in this transgenic line seemed to exert no detrimental effects on zebrafish hearts during the early stages.

  18. Temperature, paternity and asynchronous hatching influence early developmental characteristics of larval Atlantic cod, Gadus morhua

    DEFF Research Database (Denmark)

    Politis, Sebastian Nikitas; Dahlke, Flemming T.; Butts, Ian A.E.

    2014-01-01

    Offspring, especially during early development, are influenced by both intrinsic properties endowed to them by their parents, extrinsic environmental factors as well as the interplay between genes and the environment. We investigated the effects of paternity (P), temperature (T), and asynchronous...

  19. Maternal topoisomerase II alpha, not topoisomerase II beta, enables embryonic development of zebrafish top2a-/- mutants

    LENUS (Irish Health Repository)

    Sapetto-Rebow, Beata

    2011-11-23

    Abstract Background Genetic alterations in human topoisomerase II alpha (TOP2A) are linked to cancer susceptibility. TOP2A decatenates chromosomes and thus is necessary for multiple aspects of cell division including DNA replication, chromosome condensation and segregation. Topoisomerase II alpha is also required for embryonic development in mammals, as mouse Top2a knockouts result in embryonic lethality as early as the 4-8 cell stage. The purpose of this study was to determine whether the extended developmental capability of zebrafish top2a mutants arises from maternal expression of top2a or compensation from its top2b paralogue. Results Here, we describe bloody minded (blm), a novel mutant of zebrafish top2a. In contrast to mouse Top2a nulls, zebrafish top2a mutants survive to larval stages (4-5 day post fertilization). Developmental analyses demonstrate abundant expression of maternal top2a but not top2b. Inhibition or poisoning of maternal topoisomerase II delays embryonic development by extending the cell cycle M-phase. Zygotic top2a and top2b are co-expressed in the zebrafish CNS, but endogenous or ectopic top2b RNA appear unable to prevent the blm phenotype. Conclusions We conclude that maternal top2a enables zebrafish development before the mid-zygotic transition (MZT) and that zebrafish top2a and top2b are not functionally redundant during development after activation of the zygotic genome.

  20. Maternal topoisomerase II alpha, not topoisomerase II beta, enables embryonic development of zebrafish top2a-/- mutants

    Directory of Open Access Journals (Sweden)

    Sapetto-Rebow Beata

    2011-11-01

    Full Text Available Abstract Background Genetic alterations in human topoisomerase II alpha (TOP2A are linked to cancer susceptibility. TOP2A decatenates chromosomes and thus is necessary for multiple aspects of cell division including DNA replication, chromosome condensation and segregation. Topoisomerase II alpha is also required for embryonic development in mammals, as mouse Top2a knockouts result in embryonic lethality as early as the 4-8 cell stage. The purpose of this study was to determine whether the extended developmental capability of zebrafish top2a mutants arises from maternal expression of top2a or compensation from its top2b paralogue. Results Here, we describe bloody minded (blm, a novel mutant of zebrafish top2a. In contrast to mouse Top2a nulls, zebrafish top2a mutants survive to larval stages (4-5 day post fertilization. Developmental analyses demonstrate abundant expression of maternal top2a but not top2b. Inhibition or poisoning of maternal topoisomerase II delays embryonic development by extending the cell cycle M-phase. Zygotic top2a and top2b are co-expressed in the zebrafish CNS, but endogenous or ectopic top2b RNA appear unable to prevent the blm phenotype. Conclusions We conclude that maternal top2a enables zebrafish development before the mid-zygotic transition (MZT and that zebrafish top2a and top2b are not functionally redundant during development after activation of the zygotic genome.

  1. BISPHENOL A EXPOSURE DURING EARLY DEVELOPMENT INDUCES SEX-SPECIFIC CHANGES IN ADULT ZEBRAFISH SOCIAL INTERACTIONS

    Science.gov (United States)

    Weber, Daniel N.; Hoffmann, Raymond G.; Hoke, Elizabeth S.; Tanguay, Robert L.

    2014-01-01

    Developmental bisphenol A (BPA) exposure is associated with adverse behavioral effects, although underlying modes of action remain unclear. Because BPA is a suspected xenoestrogen, the objective was to identify sex-based changes in adult zebrafish social behavior developmentally exposed to BPA (0.0, 0.1 or 1 μM) or one of two control compounds (0.1μM 17β-estradiol [E2], and 0.1 μM GSK4716, a synthetic estrogen-related receptor γ ligand). A test chamber was divided lengthwise so each arena held one fish unable to detect the presence of the other fish. A mirror was inserted at one end of each arena; baseline activity levels were determined without mirror. Arenas were divided into 3, computer-generated zones to represent different distances from mirror image. Circadian rhythm patterns were evaluated at 1–3 (= AM) and 5–8 (= PM) hr postprandial. Adult zebrafish were placed into arenas and monitored by digital camera for 5 min. Total distance traveled, % time spent at mirror image, and number of attacks on mirror image were quantified. E2, GSK4716, and all BPA treatments dampened male activity and altered male circadian activity patterns; there was no marked effect on female activity. BPA induced non-monotonic effects (response curve changes direction within range of concentrations examined) on male % time at mirror only in AM. All treatments produced increased % time at the mirror during PM. Male attacks on the mirror were reduced by BPA exposure only during AM. There were sex-specific effects of developmental BPA on social interactions and time-of-day of observation affected results. PMID:25424546

  2. FishNet: an online database of zebrafish anatomy

    Directory of Open Access Journals (Sweden)

    Gibson Abigail J

    2007-08-01

    Full Text Available Abstract Background Over the last two decades, zebrafish have been established as a genetically versatile model system for investigating many different aspects of vertebrate developmental biology. With the credentials of zebrafish as a developmental model now well recognized, the emerging new opportunity is the wider application of zebrafish biology to aspects of human disease modelling. This rapidly increasing use of zebrafish as a model for human disease has necessarily generated interest in the anatomy of later developmental phases such as the larval, juvenile, and adult stages, during which many of the key aspects of organ morphogenesis and maturation take place. Anatomical resources and references that encompass these stages are non-existent in zebrafish and there is therefore an urgent need to understand how different organ systems and anatomical structures develop throughout the life of the fish. Results To overcome this deficit we have utilized the technique of optical projection tomography to produce three-dimensional (3D models of larval fish. In order to view and display these models we have created FishNet http://www.fishnet.org.au, an interactive reference of zebrafish anatomy spanning the range of zebrafish development from 24 h until adulthood. Conclusion FishNet contains more than 36 000 images of larval zebrafish, with more than 1 500 of these being annotated. The 3D models can be manipulated on screen or virtually sectioned. This resource represents the first complete embryo to adult atlas for any species in 3D.

  3. Quantification of vestibular-induced eye movements in zebrafish larvae

    Directory of Open Access Journals (Sweden)

    Mo Weike

    2010-09-01

    Full Text Available Abstract Background Vestibular reflexes coordinate movements or sensory input with changes in body or head position. Vestibular-evoked responses that involve the extraocular muscles include the vestibulo-ocular reflex (VOR, a compensatory eye movement to stabilize retinal images. Although an angular VOR attributable to semicircular canal stimulation was reported to be absent in free-swimming zebrafish larvae, recent studies reveal that vestibular-induced eye movements can be evoked in zebrafish larvae by both static tilts and dynamic rotations that tilt the head with respect to gravity. Results We have determined herein the basis of sensitivity of the larval eye movements with respect to vestibular stimulus, developmental stage, and sensory receptors of the inner ear. For our experiments, video recordings of larvae rotated sinusoidally at 0.25 Hz were analyzed to quantitate eye movements under infrared illumination. We observed a robust response that appeared as early as 72 hours post fertilization (hpf, which increased in amplitude over time. Unlike rotation about an earth horizontal axis, rotation about an earth vertical axis at 0.25 Hz did not evoke eye movements. Moreover, vestibular-induced responses were absent in mutant cdh23 larvae and larvae lacking anterior otoliths. Conclusions Our results provide evidence for a functional vestibulo-oculomotor circuit in 72 hpf zebrafish larvae that relies upon sensory input from anterior/utricular otolith organs.

  4. Maternal Cortisol Mediates Hypothalamus-Pituitary-Interrenal Axis Development in Zebrafish

    Science.gov (United States)

    Nesan, Dinushan; Vijayan, Mathilakath M.

    2016-01-01

    In zebrafish (Danio rerio), de novo synthesis of cortisol in response to stressor exposure commences only after hatch. Maternally deposited cortisol is present during embryogenesis, but a role for this steroid in early development is unclear. We tested the hypothesis that maternal cortisol is essential for the proper development of hypothalamus-pituitary-interrenal (HPI) axis activity and the onset of the stressor-induced cortisol response in larval zebrafish. In this study, zygotic cortisol content was manipulated by microinjecting antibody to sequester this steroid, thereby making it unavailable during embryogenesis. This was compared with embryos containing excess cortisol by microinjection of exogenous steroid. The resulting larval phenotypes revealed distinct treatment effects, including deformed mesoderm structures when maternal cortisol was unavailable and cardiac edema after excess cortisol. Maternal cortisol unavailability heightened the cortisol stress response in post-hatch larvae, whereas excess cortisol abolished the stressor-mediated cortisol elevation. This contrasting hormonal response corresponded with altered expression of key HPI axis genes, including crf, 11B hydroxylase, pomca, and star, which were upregulated in response to reduced cortisol availability and downregulated when embryos had excess cortisol. These findings for the first time underscore a critical role for maternally deposited cortisol in programming HPI axis development and function in zebrafish. PMID:26940285

  5. In Vitro Biotransformation of Two Human CYP3A Probe Substrates and Their Inhibition during Early Zebrafish Development.

    Science.gov (United States)

    Verbueken, Evy; Alsop, Derek; Saad, Moayad A; Pype, Casper; Van Peer, Els M; Casteleyn, Christophe R; Van Ginneken, Chris J; Wilson, Joanna; Van Cruchten, Steven J

    2017-01-22

    At present, the zebrafish embryo is increasingly used as an alternative animal model to screen for developmental toxicity after exposure to xenobiotics. Since zebrafish embryos depend on their own drug-metabolizing capacity, knowledge of their intrinsic biotransformation is pivotal in order to correctly interpret the outcome of teratogenicity assays. Therefore, the aim of this in vitro study was to assess the activity of cytochrome P450 (CYP)-a group of drug-metabolizing enzymes-in microsomes from whole zebrafish embryos (ZEM) of 5, 24, 48, 72, 96 and 120 h post-fertilization (hpf) by means of a mammalian CYP substrate, i.e., benzyloxy-methyl-resorufin (BOMR). The same CYP activity assays were performed in adult zebrafish liver microsomes (ZLM) to serve as a reference for the embryos. In addition, activity assays with the human CYP3A4-specific Luciferin isopropyl acetal (Luciferin-IPA) as well as inhibition studies with ketoconazole and CYP3cide were carried out to identify CYP activity in ZLM. In the present study, biotransformation of BOMR was detected at 72 and 96 hpf; however, metabolite formation was low compared with ZLM. Furthermore, Luciferin-IPA was not metabolized by the zebrafish. In conclusion, the capacity of intrinsic biotransformation in zebrafish embryos appears to be lacking during a major part of organogenesis.

  6. In Vitro Biotransformation of Two Human CYP3A Probe Substrates and Their Inhibition during Early Zebrafish Development

    Directory of Open Access Journals (Sweden)

    Evy Verbueken

    2017-01-01

    Full Text Available At present, the zebrafish embryo is increasingly used as an alternative animal model to screen for developmental toxicity after exposure to xenobiotics. Since zebrafish embryos depend on their own drug-metabolizing capacity, knowledge of their intrinsic biotransformation is pivotal in order to correctly interpret the outcome of teratogenicity assays. Therefore, the aim of this in vitro study was to assess the activity of cytochrome P450 (CYP—a group of drug-metabolizing enzymes—in microsomes from whole zebrafish embryos (ZEM of 5, 24, 48, 72, 96 and 120 h post-fertilization (hpf by means of a mammalian CYP substrate, i.e., benzyloxy-methyl-resorufin (BOMR. The same CYP activity assays were performed in adult zebrafish liver microsomes (ZLM to serve as a reference for the embryos. In addition, activity assays with the human CYP3A4-specific Luciferin isopropyl acetal (Luciferin-IPA as well as inhibition studies with ketoconazole and CYP3cide were carried out to identify CYP activity in ZLM. In the present study, biotransformation of BOMR was detected at 72 and 96 hpf; however, metabolite formation was low compared with ZLM. Furthermore, Luciferin-IPA was not metabolized by the zebrafish. In conclusion, the capacity of intrinsic biotransformation in zebrafish embryos appears to be lacking during a major part of organogenesis.

  7. Tyrosine pathway regulation is host-mediated in the pea aphid symbiosis during late embryonic and early larval development.

    Science.gov (United States)

    Rabatel, Andréane; Febvay, Gérard; Gaget, Karen; Duport, Gabrielle; Baa-Puyoulet, Patrice; Sapountzis, Panagiotis; Bendridi, Nadia; Rey, Marjolaine; Rahbé, Yvan; Charles, Hubert; Calevro, Federica; Colella, Stefano

    2013-04-10

    Nutritional symbioses play a central role in insects' adaptation to specialized diets and in their evolutionary success. The obligatory symbiosis between the pea aphid, Acyrthosiphon pisum, and the bacterium, Buchnera aphidicola, is no exception as it enables this important agricultural pest insect to develop on a diet exclusively based on plant phloem sap. The symbiotic bacteria provide the host with essential amino acids lacking in its diet but necessary for the rapid embryonic growth seen in the parthenogenetic viviparous reproduction of aphids. The aphid furnishes, in exchange, non-essential amino acids and other important metabolites. Understanding the regulations acting on this integrated metabolic system during the development of this insect is essential in elucidating aphid biology. We used a microarray-based approach to analyse gene expression in the late embryonic and the early larval stages of the pea aphid, characterizing, for the first time, the transcriptional profiles in these developmental phases. Our analyses allowed us to identify key genes in the phenylalanine, tyrosine and dopamine pathways and we identified ACYPI004243, one of the four genes encoding for the aspartate transaminase (E.C. 2.6.1.1), as specifically regulated during development. Indeed, the tyrosine biosynthetic pathway is crucial for the symbiotic metabolism as it is shared between the two partners, all the precursors being produced by B. aphidicola. Our microarray data are supported by HPLC amino acid analyses demonstrating an accumulation of tyrosine at the same developmental stages, with an up-regulation of the tyrosine biosynthetic genes. Tyrosine is also essential for the synthesis of cuticular proteins and it is an important precursor for cuticle maturation: together with the up-regulation of tyrosine biosynthesis, we observed an up-regulation of cuticular genes expression. We were also able to identify some amino acid transporter genes which are essential for the switch

  8. Comparative Developmental Toxicity of Flavonoids Using an Integrative Zebrafish System.

    Science.gov (United States)

    Bugel, Sean M; Bonventre, Josephine A; Tanguay, Robert L

    2016-11-01

    Flavonoids are a large, structurally diverse class of bioactive naturally occurring chemicals commonly detected in breast milk, soy based infant formulas, amniotic fluid, and fetal cord blood. The potential for pervasive early life stage exposures raises concerns for perturbation of embryogenesis, though developmental toxicity and bioactivity information is limited for many flavonoids. Therefore, we evaluated a suite of 24 flavonoid and flavonoid-like chemicals using a zebrafish embryo-larval toxicity bioassay-an alternative model for investigating developmental toxicity of environmentally relevant chemicals. Embryos were exposed to 1-50 µM of each chemical from 6 to 120 h postfertilization (hpf), and assessed for 26 adverse developmental endpoints at 24, 72, and 120 hpf. Behavioral changes were evaluated in morphologically normal animals at 24 and 72 hpf, at 120 hpf using a larval photomotor response (LPR) assay. Gene expression was comparatively evaluated for all compounds for effects on biomarker transcripts indicative of AHR (cyp1a) and ER (cyp19a1b, esr1, lhb, vtg) pathway bioactivity. Overall, 15 of 24 flavonoids elicited adverse effects on one or more of the developmental or behavioral endpoints. Hierarchical clustering and principle component analyses compared toxicity profiles and identified 3 distinct groups of bioactive flavonoids. Despite robust induction of multiple estrogen-responsive biomarkers, co-exposure with ER and GPER antagonists did not ameliorate toxicity, suggesting ER-independence and alternative modes of action. Taken together, these studies demonstrate that development is sensitive to perturbation by bioactive flavonoids in zebrafish that are not related to traditional estrogen receptor mode of action pathways. This integrative zebrafish platform provides a useful framework for evaluating flavonoid developmental toxicity and hazard prioritization. © The Author 2016. Published by Oxford University Press on behalf of the Society of

  9. Functional analysis of a tyrosinase gene involved in early larval shell biogenesis in Crassostrea angulata and its response to ocean acidification.

    Science.gov (United States)

    Yang, Bingye; Pu, Fei; Li, Lingling; You, Weiwei; Ke, Caihuan; Feng, Danqing

    2017-04-01

    The formation of the primary shell is a vital process in marine bivalves. Ocean acidification largely influences shell formation. It has been reported that enzymes involved in phenol oxidation, such as tyrosinase and phenoloxidases, participate in the formation of the periostracum. In the present study, we cloned a tyrosinase gene from Crassostrea angulata named Ca-tyrA1, and its potential function in early larval shell biogenesis was investigated. The Ca-tyrA1 gene has a full-length cDNA of 2430bp in size, with an open reading frame of 1896bp in size, which encodes a 631-amino acid protein that includes a 24-amino acid putative signal peptide. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis revealed that Ca-tyrA1 transcription mainly occurs at the trochophore stage, and the Ca-tyrA1 mRNA levels in the 3000ppm treatment group were significantly upregulated in the early D-veliger larvae. WMISH and electron scanning microscopy analyses showed that the expression of Ca-tyrA1 occurs at the gastrula stage, thereby sustaining the early D-veliger larvae, and the shape of its signal is saddle-like, similar to that observed under an electron scanning microscope. Furthermore, the RNA interference has shown that the treatment group has a higher deformity rate than that of the control, thereby indicating that Ca-tyrA1 participates in the biogenesis of the primary shell. In conclusion, and our results indicate that Ca-tyrA1 plays a vital role in the formation of the larval shell and participates in the response to larval shell damages in Crassostrea angulata that were induced by ocean acidification. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Growth and survival of larval and early juvenile lesser sandeel in patchy prey field in the North Sea: An examination using individual-based modelling

    DEFF Research Database (Denmark)

    Gürkan, Zeren; Christensen, Asbjørn; Deurs, Mikael van

    2012-01-01

    -stages in the North Sea. Simulations of patchiness related starvation mortality are able to explain observed patterns of variation in sandeel growth. Reduced prey densities within patches decrease growth and survival rate of larvae and match–mismatch affect growth and survival of larvae with different hatch time due...... by modeling copepod size spectra dynamics and patchiness based on particle count transects and Continuous Plankton Recorder time series data. The study analyzes the effects of larval hatching time, presence of zooplankton patchiness and within patch abundance on growth and survival of sandeel early life...

  11. Effects of prolonged exposure to perchlorate on thyroid and reproductive function in zebrafish

    Science.gov (United States)

    Mukhi, S.; Patino, R.

    2007-01-01

    The objectives of this study were to determine the effects of prolonged exposure to perchlorate on (1) thyroid status and reproductive performance of adult zebrafish (Danio rerio) and (2) F1 embryo survival and early larval development. Using a static-renewal procedure, mixed sex populations of adult zebrafish were exposed to 0, 10, and 100 mg/l nominal concentrations of waterborne perchlorate for 10 weeks. Thyroid histology was qualitatively assessed, and females and males were separated and further exposed to their respective treatments for six additional weeks. Eight females in each tank replicate (n = 3) were paired weekly with four males from the same respective treatment, and packed-egg (spawn) volume (PEV) was measured each of the last five weeks. At least once during weeks 14-16 of exposure, other end points measured included fertilization rate, fertilized egg diameter, hatching rate, standard length, and craniofacial development of 4-day-postfertilization larvae and thyroid hormone content of 3.5-h embryos and of exposed mothers. At 10 weeks of exposure, perchlorate at both concentrations caused thyroidal hypertrophy and colloid depletion. A marked reduction in PEV was observed toward the end of the 6-week spawning period, but fertilization and embryo hatching rates were unaffected. Fertilized egg diameter and larval length were increased by parental exposure to perchlorate. Larval head depth was unaffected but the forward protrusion of the lower jaw-associated cartilage complexes, Meckel's and ceratohyal, was decreased. Exposure to both concentrations of perchlorate inhibited whole-body thyroxine content in mothers and embryos, but triiodothyronine content was unchanged. In conclusion, prolonged exposure of adult zebrafish to perchlorate not only disrupts their thyroid endocrine system but also impairs reproduction and influences early F1 development. ?? 2007 Oxford University Press.

  12. The Role of Maternal Nutrition on Oocyte Size and Quality, with Respect to Early Larval Development in The Coral-Eating Starfish, Acanthaster planci.

    Directory of Open Access Journals (Sweden)

    Ciemon Frank Caballes

    Full Text Available Variation in local environmental conditions can have pronounced effects on the population structure and dynamics of marine organisms. Previous studies on crown-of-thorns starfish, Acanthaster planci, have primarily focused on effects of water quality and nutrient availability on larval growth and survival, while the role of maternal nutrition on reproduction and larval development has been overlooked. To examine the effects of maternal nutrition on oocyte size and early larval development in A. planci, we pre-conditioned females for 60 days on alternative diets of preferred coral prey (Acropora abrotanoides versus non-preferred coral prey (Porites rus and compared resulting gametes and progeny to those produced by females that were starved over the same period. Females fed ad libitum with Acropora increased in weight, produced heavier gonads and produced larger oocytes compared to Porites-fed and starved females. Fed starfish (regardless of whether it was Acropora or Porites produced bigger larvae with larger stomachs and had a higher frequency of normal larvae that reached the late bipinnaria / early brachiolaria stage compared to starved starfish. Females on Acropora diet also produced a higher proportion of larvae that progressed to more advanced stages faster compared to Porites-fed starfish, which progressed faster than starved starfish. These results suggest that maternal provisioning can have important consequences for the quality and quantity of progeny. Because food quality (coral community structure and quantity (coral abundance varies widely among reef locations and habitats, local variation in maternal nutrition of A. planci is likely to moderate reproductive success and may explain temporal and spatial fluctuations in abundance of this species.

  13. The Role of Maternal Nutrition on Oocyte Size and Quality, with Respect to Early Larval Development in The Coral-Eating Starfish, Acanthaster planci.

    Science.gov (United States)

    Caballes, Ciemon Frank; Pratchett, Morgan S; Kerr, Alexander M; Rivera-Posada, Jairo A

    2016-01-01

    Variation in local environmental conditions can have pronounced effects on the population structure and dynamics of marine organisms. Previous studies on crown-of-thorns starfish, Acanthaster planci, have primarily focused on effects of water quality and nutrient availability on larval growth and survival, while the role of maternal nutrition on reproduction and larval development has been overlooked. To examine the effects of maternal nutrition on oocyte size and early larval development in A. planci, we pre-conditioned females for 60 days on alternative diets of preferred coral prey (Acropora abrotanoides) versus non-preferred coral prey (Porites rus) and compared resulting gametes and progeny to those produced by females that were starved over the same period. Females fed ad libitum with Acropora increased in weight, produced heavier gonads and produced larger oocytes compared to Porites-fed and starved females. Fed starfish (regardless of whether it was Acropora or Porites) produced bigger larvae with larger stomachs and had a higher frequency of normal larvae that reached the late bipinnaria / early brachiolaria stage compared to starved starfish. Females on Acropora diet also produced a higher proportion of larvae that progressed to more advanced stages faster compared to Porites-fed starfish, which progressed faster than starved starfish. These results suggest that maternal provisioning can have important consequences for the quality and quantity of progeny. Because food quality (coral community structure) and quantity (coral abundance) varies widely among reef locations and habitats, local variation in maternal nutrition of A. planci is likely to moderate reproductive success and may explain temporal and spatial fluctuations in abundance of this species.

  14. An Fgf-Shh signaling hierarchy regulates early specification of the zebrafish skull.

    Science.gov (United States)

    McCarthy, Neil; Sidik, Alfire; Bertrand, Julien Y; Eberhart, Johann K

    2016-07-15

    The neurocranium generates most of the craniofacial skeleton and consists of prechordal and postchordal regions. Although development of the prechordal is well studied, little is known of the postchordal region. Here we characterize a signaling hierarchy necessary for postchordal neurocranial development involving Fibroblast growth factor (Fgf) signaling for early specification of mesodermally-derived progenitor cells. The expression of hyaluron synthetase 2 (has2) in the cephalic mesoderm requires Fgf signaling and Has2 function, in turn, is required for postchordal neurocranial development. While Hedgehog (Hh)-deficient embryos also lack a postchordal neurocranium, this appears primarily due to a later defect in chondrocyte differentiation. Inhibitor studies demonstrate that postchordal neurocranial development requires early Fgf and later Hh signaling. Collectively, our results provide a mechanistic understanding of early postchordal neurocranial development and demonstrate a hierarchy of signaling between Fgf and Hh in the development of this structure. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  15. A zebrafish model of congenital disorders of glycosylation with phosphomannose isomerase deficiency reveals an early opportunity for corrective mannose supplementation

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    Jaime Chu

    2013-01-01

    Individuals with congenital disorders of glycosylation (CDG have recessive mutations in genes required for protein N-glycosylation, resulting in multi-systemic disease. Despite the well-characterized biochemical consequences in these individuals, the underlying cellular defects that contribute to CDG are not well understood. Synthesis of the lipid-linked oligosaccharide (LLO, which serves as the sugar donor for the N-glycosylation of secretory proteins, requires conversion of fructose-6-phosphate to mannose-6-phosphate via the phosphomannose isomerase (MPI enzyme. Individuals who are deficient in MPI present with bleeding, diarrhea, edema, gastrointestinal bleeding and liver fibrosis. MPI-CDG patients can be treated with oral mannose supplements, which is converted to mannose-6-phosphate through a minor complementary metabolic pathway, restoring protein glycosylation and ameliorating most symptoms, although liver disease continues to progress. Because Mpi deletion in mice causes early embryonic lethality and thus is difficult to study, we used zebrafish to establish a model of MPI-CDG. We used a morpholino to block mpi mRNA translation and established a concentration that consistently yielded 13% residual Mpi enzyme activity at 4 days post-fertilization (dpf, which is within the range of MPI activity detected in fibroblasts from MPI-CDG patients. Fluorophore-assisted carbohydrate electrophoresis detected decreased LLO and N-glycans in mpi morphants. These deficiencies resulted in 50% embryonic lethality by 4 dpf. Multi-systemic abnormalities, including small eyes, dysmorphic jaws, pericardial edema, a small liver and curled tails, occurred in 82% of the surviving larvae. Importantly, these phenotypes could be rescued with mannose supplementation. Thus, parallel processes in fish and humans contribute to the phenotypes caused by Mpi depletion. Interestingly, mannose was only effective if provided prior to 24 hpf. These data provide insight into treatment efficacy

  16. Generation and characterization of Kctd15 mutations in zebrafish.

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    Alison Heffer

    Full Text Available Potassium channel tetramerization domain containing 15 (Kctd15 was previously found to have a role in early neural crest (NC patterning, specifically delimiting the region where NC markers are expressed via repression of transcription factor AP-2a and inhibition of Wnt signaling. We used transcription activator-like effector nucleases (TALENs to generate null mutations in zebrafish kctd15a and kctd15b paralogs to study the in vivo role of Kctd15. We found that while deletions producing frame-shift mutations in each paralog showed no apparent phenotype, kctd15a/b double mutant zebrafish are smaller in size and show several phenotypes including some affecting the NC, such as expansion of the early NC domain, increased pigmentation, and craniofacial defects. Both melanophore and xanthophore pigment cell numbers and early markers are up-regulated in the double mutants. While we find no embryonic craniofacial defects, adult mutants have a deformed maxillary segment and missing barbels. By confocal imaging of mutant larval brains we found that the torus lateralis (TLa, a region implicated in gustatory networks in other fish, is absent. Ablation of this brain tissue in wild type larvae mimics some aspects of the mutant growth phenotype. Thus kctd15 mutants show deficits in the development of both neural crest derivatives, and specific regions within the central nervous system, leading to a strong reduction in normal growth rates.

  17. Low Doses of Gamma-Irradiation Induce an Early Bystander Effect in Zebrafish Cells Which Is Sufficient to Radioprotect Cells

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    Pereira, Sandrine; Malard, Véronique; Ravanat, Jean-Luc; Davin, Anne-Hélène; Armengaud, Jean; Foray, Nicolas; Adam-Guillermin, Christelle

    2014-01-01

    The term “bystander effect” is used to describe an effect in which cells that have not been exposed to radiation are affected by irradiated cells though various intracellular signaling mechanisms. In this study we analyzed the kinetics and mechanisms of bystander effect and radioadaptation in embryonic zebrafish cells (ZF4) exposed to chronic low dose of gamma rays. ZF4 cells were irradiated for 4 hours with total doses of gamma irradiation ranging from 0.01–0.1 Gy. In two experimental conditions, the transfer of irradiated cells or culture medium from irradiated cells results in the occurrence of DNA double strand breaks in non-irradiated cells (assessed by the number of γ-H2AX foci) that are repaired at 24 hours post-irradiation whatever the dose. At low total irradiation doses the bystander effect observed does not affect DNA repair mechanisms in targeted and bystander cells. An increase in global methylation of ZF4 cells was observed in irradiated cells and bystander cells compared to control cells. We observed that pre-irradiated cells which are then irradiated for a second time with the same doses contained significantly less γ-H2AX foci than in 24 h gamma-irradiated control cells. We also showed that bystander cells that have been in contact with the pre-irradiated cells and then irradiated alone present less γ-H2AX foci compared to the control cells. This radioadaptation effect is significantly more pronounced at the highest doses. To determine the factors involved in the early events of the bystander effect, we performed an extensive comparative proteomic study of the ZF4 secretomes upon irradiation. In the experimental conditions assayed here, we showed that the early events of bystander effect are probably not due to the secretion of specific proteins neither the oxidation of these secreted proteins. These results suggest that early bystander effect may be due probably to a combination of multiple factors. PMID:24667817

  18. Swim-training changes the spatio-temporal dynamics of skeletogenesis in zebrafish larvae (Danio rerio.

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    Ansa W Fiaz

    Full Text Available Fish larvae experience many environmental challenges during development such as variation in water velocity, food availability and predation. The rapid development of structures involved in feeding, respiration and swimming increases the chance of survival. It has been hypothesized that mechanical loading induced by muscle forces plays a role in prioritizing the development of these structures. Mechanical loading by muscle forces has been shown to affect larval and embryonic bone development in vertebrates, but these investigations were limited to the appendicular skeleton. To explore the role of mechanical load during chondrogenesis and osteogenesis of the cranial, axial and appendicular skeleton, we subjected zebrafish larvae to swim-training, which increases physical exercise levels and presumably also mechanical loads, from 5 until 14 days post fertilization. Here we show that an increased swimming activity accelerated growth, chondrogenesis and osteogenesis during larval development in zebrafish. Interestingly, swim-training accelerated both perichondral and intramembranous ossification. Furthermore, swim-training prioritized the formation of cartilage and bone structures in the head and tail region as well as the formation of elements in the anal and dorsal fins. This suggests that an increased swimming activity prioritized the development of structures which play an important role in swimming and thereby increasing the chance of survival in an environment where water velocity increases. Our study is the first to show that already during early zebrafish larval development, skeletal tissue in the cranial, axial and appendicular skeleton is competent to respond to swim-training due to increased water velocities. It demonstrates that changes in water flow conditions can result into significant spatio-temporal changes in skeletogenesis.

  19. Expression pattern of cdkl5 during zebrafish early development: implications for use as model for atypical Rett syndrome.

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    Vitorino, Marta; Cunha, Nídia; Conceição, Natércia; Cancela, M Leonor

    2018-05-11

    Atypical Rett syndrome is a child neurodevelopmental disorder induced by mutations in CDKL5 gene and characterized by a progressive regression in development with loss of purposeful use of the hands, slowed brain and head growth, problems with walking, seizures, and intellectual disability. At the moment, there is no cure for this pathology and little information is available concerning animal models capable of mimicking its phenotypes, thus the development of additional animal models should be of interest to gain more knowledge about the disease. Zebrafish has been used successfully as model organism for many human genetic diseases; however, no information is available concerning the spatial and temporal expression of cdkl5 orthologous in this organism. In the present study, we identified the developmental expression patterns of cdkl5 in zebrafish by quantitative PCR and whole-mount in situ hybridization. cdkl5 is expressed maternally at low levels during the first 24 h of development. After that the expression of the gene increases significantly and it starts to be expressed mainly in the nervous system and in several brain structures, such as telencephalon, mesencephalon and diencephalon. The expression patterns of cdkl5 in zebrafish is in accordance with the tissues known to be affected in humans and associated to symptoms and deficits observed in Rett syndrome patients thus providing the first evidence that zebrafish could be an alternative model to study the molecular pathways of this disease as well as to test possible therapeutic approaches capable of rescuing the phenotype.

  20. Excess dietary cholesterol may have an adverse effect on growth performance of early post-larval Litopenaeus vannamei

    Science.gov (United States)

    2012-01-01

    One experiment was conducted to determine the nutritive value of cholesterol for post-larval shrimp, Litopenaeus vannamei. Four isoenergetic and isonitrogenous diets supplemented with four levels of cholesterol (D1, D2, D3 and D4 with 0, 0.5%, 1% and 2% cholesterol, respectively) were fed to triplicate groups of L. vannamei shrimp (mean initial wet weight 0.8 mg) for 27 days. After the trial, shrimp fed the D1 diet had the best growth performance (final body weights: FBW; weight gain: WG; specific growth rate: SGR), while there was no significant difference between diet treatments with respect to survival. The whole body crude protein level in the shrimp decreased with the increase in dietary cholesterol levels, while the whole body crude lipid level in shrimps in the D4 diet treatment was significantly higher (P vannamei; excess dietary cholesterol may thus lead to adverse effects on the growth performance of post-larval shrimp. PMID:22958647

  1. Excess dietary cholesterol may have an adverse effect on growth performance of early post-larval Litopenaeus vannamei

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    Niu Jin

    2012-06-01

    Full Text Available Abstract One experiment was conducted to determine the nutritive value of cholesterol for post-larval shrimp, Litopenaeus vannamei. Four isoenergetic and isonitrogenous diets supplemented with four levels of cholesterol (D1, D2, D3 and D4 with 0, 0.5%, 1% and 2% cholesterol, respectively were fed to triplicate groups of L. vannamei shrimp (mean initial wet weight 0.8 mg for 27 days. After the trial, shrimp fed the D1 diet had the best growth performance (final body weights: FBW; weight gain: WG; specific growth rate: SGR, while there was no significant difference between diet treatments with respect to survival. The whole body crude protein level in the shrimp decreased with the increase in dietary cholesterol levels, while the whole body crude lipid level in shrimps in the D4 diet treatment was significantly higher (P L. vannamei; excess dietary cholesterol may thus lead to adverse effects on the growth performance of post-larval shrimp.

  2. Simultaneous quantification of T4, T3, rT3, 3,5-T2 and 3,3'-T2 in larval zebrafish (Danio rerio) as a model to study exposure to polychlorinated biphenyls.

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    Chen, Xiaopeng; Walter, Kyla M; Miller, Galen W; Lein, Pamela J; Puschner, Birgit

    2018-06-01

    Environmental toxicants that interfere with thyroid hormone (TH) signaling can impact growth and development in animals and humans. Zebrafish represent a model to study chemically induced TH disruption, prompting the need for sensitive detection of THs. Simultaneous quantification of 3,3',5-triiodo-l-thyronine (T3), thyroxine (T4), 3,3',5'-triiodo-l-thyronine (rT3), 3,5-diiodo-l-thyronine (3,5-T2) and 3,3'-diiodo-l-thyronine (3,3'-T2) in zebrafish larvae was achieved by ultra-performance liquid chromatography-tandem mass spectrometry in positive ion mode. Solid-phase extraction with SampliQ cartridges and derivatization with 3 m hydrochloric acid in n-butanol reduced matrix effects. Derivatized compounds were separated on an Acquity UPLC BEH C 18 column with mobile phases consisting of 0.1% acetic acid in deionized water and 0.1% acetic acid in methanol. The limits of detection ranged from 0.5 to 0.6 pg injected on column. The method was validated by evaluating recovery (77.1-117.2%), accuracy (87.3-123.9%) and precision (0.5-12.4%) using diluted homogenized zebrafish embryos spiked with all target compounds. This method was then applied to zebrafish larvae collected after 114 h of exposure to polychlorinated biphenyls (PCBs), including PCB 28, PCB 66 and PCB 95, or the technical mixture Aroclor 1254. Exposure to PCB 28 and PCB 95 increased the T4:T3 ratio and decreased the T3:rT3 ratio, demonstrating that this method can effectively detect PCB-induced alterations in THs. Copyright © 2018 John Wiley & Sons, Ltd.

  3. Myosin-1 inhibition by PClP affects membrane shape, cortical actin distribution and lipid droplet dynamics in early Zebrafish embryos.

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    Prabuddha Gupta

    Full Text Available Myosin-1 (Myo1 represents a mechanical link between the membrane and actin-cytoskeleton in animal cells. We have studied the effect of Myo1 inhibitor PClP in 1-8 cell Zebrafish embryos. Our results indicate a unique involvement of Myo1 in early development of Zebrafish embryos. Inhibition of Myo1 (by PClP and Myo2 (by Blebbistatin lead to arrest in cell division. While Myo1 isoforms appears to be important for both the formation and the maintenance of cleavage furrows, Myo2 is required only for the formation of furrows. We found that the blastodisc of the embryo, which contains a thick actin cortex (~13 μm, is loaded with cortical Myo1. Myo1 appears to be crucial for maintaining the blastodisc morphology and the actin cortex thickness. In addition to cell division and furrow formation, inhibition of Myo1 has a drastic effect on the dynamics and distribution of lipid droplets (LDs in the blastodisc near the cleavage furrow. All these results above are effects of Myo1 inhibition exclusively; Myo2 inhibition by blebbistatin does not show such phenotypes. Therefore, our results demonstrate a potential role for Myo1 in the maintenance and formation of furrow, blastodisc morphology, cell-division and LD organization within the blastodisc during early embryogenesis.

  4. Critical early roles for col27a1a and col27a1b in zebrafish notochord morphogenesis, vertebral mineralization and post-embryonic axial growth.

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    Christiansen, Helena E; Lang, Michael R; Pace, James M; Parichy, David M

    2009-12-29

    Fibrillar collagens are well known for their links to human diseases, with which all have been associated except for the two most recently identified fibrillar collagens, type XXIV collagen and type XXVII collagen. To assess functions and potential disease phenotypes of type XXVII collagen, we examined its roles in zebrafish embryonic and post-embryonic development. We identified two type XXVII collagen genes in zebrafish, col27a1a and col27a1b. Both col27a1a and col27a1b were expressed in notochord and cartilage in the embryo and early larva. To determine sites of type XXVII collagen function, col27a1a and col27a1b were knocked down using morpholino antisense oligonucleotides. Knockdown of col27a1a singly or in conjunction with col27a1b resulted in curvature of the notochord at early stages and formation of scoliotic curves as well as dysmorphic vertebrae at later stages. These defects were accompanied by abnormal distributions of cells and protein localization in the notochord, as visualized by transmission electron microscopy, as well as delayed vertebral mineralization as detected histologically. Together, our findings indicate a key role for type XXVII collagen in notochord morphogenesis and axial skeletogenesis and suggest a possible human disease phenotype.

  5. Tritiated water exposure disrupts myofibril structure and induces mis-regulation of eye opacity and DNA repair genes in zebrafish early life stages.

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    Arcanjo, Caroline; Armant, Olivier; Floriani, Magali; Cavalie, Isabelle; Camilleri, Virginie; Simon, Olivier; Orjollet, Daniel; Adam-Guillermin, Christelle; Gagnaire, Béatrice

    2018-04-27

    Tritium ( 3 H) is a radioactive isotope of hydrogen. In the environment, the most common form of tritium is tritiated water (HTO). The present study aimed to identify early biomarkers of HTO contamination through the use of an aquatic model, the zebrafish (Danio rerio). We used the zebrafish embryo-larvae model to investigate the modes of action of HTO exposure at dose rates of 0.4 and 4 mGy/h, dose rates expected to induce deleterious effects on fish. Zebrafish were exposed to HTO from 3 hpf (hours post fertilization) to 96 hpf. The transcriptomic effects were investigated 24 h and 96 h after the beginning of the contamination, using mRNAseq. Results suggested an impact of HTO contamination, regardless of the dose rate, on genes involved in muscle contraction (tnnt2d, tnni2a.4, slc6a1a or atp2a1l) and eye opacity (crygm2d9, crygmxl1, mipb or lim2.3) after 24 h of contamination. Interestingly, an opposite differential expression was highlighted in genes playing a role in muscle contraction and eye opacity in 24 hpf embryos when comparing dose rates, suggesting an onset of DNA protective mechanisms. The expression of h2afx and ddb2 involved in DNA repair was enhanced in response to HTO exposure. The entrainment of circadian clock and the response to H 2 O 2 signalling pathways were enriched at 96 hpf at 0.4 mGy/h and in both stages after 4 mGy/h. Genes involved in ROS scavenging were differentially expressed only after 24 h of exposure for the lowest dose rate, suggesting the onset of early protective mechanisms against oxidative stress. Effects highlighted on muscle at the molecular scale were confirmed at a higher biological scale, as electron microscopy observations revealed sarcomere impairments in 96 hpf larvae for both dose rates. Together with other studies, the present work provides useful data to better understand modes of action of tritium on zebrafish embryos-larvae. Copyright © 2018 Elsevier B.V. All rights reserved.

  6. Transcriptome analysis of severe hypoxic stress during development in zebrafish

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    I.G. Woods

    2015-12-01

    Full Text Available Hypoxia causes critical cellular injury both in early human development and in adulthood, leading to cerebral palsy, stroke, and myocardial infarction. Interestingly, a remarkable phenomenon known as hypoxic preconditioning arises when a brief hypoxia exposure protects target organs against subsequent, severe hypoxia. Although hypoxic preconditioning has been demonstrated in several model organisms and tissues including the heart and brain, its molecular mechanisms remain poorly understood. Accordingly, we used embryonic and larval zebrafish to develop a novel vertebrate model for hypoxic preconditioning, and used this model to identify conserved hypoxia-regulated transcripts for further functional study as published in Manchenkov et al. (2015 in G3: Genes|Genomes|Genetics. In this Brief article, we provide extensive annotation for the most strongly hypoxia-regulated genes in zebrafish, including their human orthologs, and describe in detail the methods used to identify, filter, and annotate hypoxia-regulated transcripts for downstream functional and bioinformatic assays using the source data provided in Gene Expression Omnibus Accession GSE68473.

  7. The utility of zebrafish to study the mechanisms by which ethanol affects social behavior and anxiety during early brain development.

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    Parker, Matthew O; Annan, Leonette V; Kanellopoulos, Alexandros H; Brock, Alistair J; Combe, Fraser J; Baiamonte, Matteo; Teh, Muy-Teck; Brennan, Caroline H

    2014-12-03

    Exposure to moderate levels of ethanol during brain development has a number of effects on social behavior but the molecular mechanisms that mediate this are not well understood. Gaining a better understanding of these factors may help to develop therapeutic interventions in the future. Zebrafish offer a potentially useful model in this regard. Here, we introduce a zebrafish model of moderate prenatal ethanol exposure. Embryos were exposed to 20mM ethanol for seven days (48hpf-9dpf) and tested as adults for individual social behavior and shoaling. We also tested their basal anxiety with the novel tank diving test. We found that the ethanol-exposed fish displayed reductions in social approach and shoaling, and an increase in anxiety in the novel tank test. These behavioral differences corresponded to differences in hrt1aa, slc6a4 and oxtr expression. Namely, acute ethanol caused a spike in oxtr and ht1aa mRNA expression, which was followed by down-regulation at 7dpf, and an up-regulation in slc6a4 at 72hpf. This study confirms the utility of zebrafish as a model system for studying the molecular basis of developmental ethanol exposure. Furthermore, it proposes a putative developmental mechanism characterized by ethanol-induced OT inhibition leading to suppression of 5-HT and up-regulation of 5-HT1A, which leads, in turn, to possible homeostatic up-regulation of 5-HTT at 72hpf and subsequent imbalance of the 5-HT system. Copyright © 2014 Elsevier Inc. All rights reserved.

  8. Immunolocalization and expression of Na(+)/K(+) -ATPase in embryos, early larval stages and adults of the freshwater shrimp Palaemonetes argentinus (Decapoda, Caridea, Palaemonidae).

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    Ituarte, Romina Belén; Lignot, Jehan-Hervé; Charmantier, Guy; Spivak, Eduardo; Lorin-Nebel, Catherine

    2016-06-01

    The euryhaline shrimp Palaemonetes argentinus exemplifies an evolutionary transition from brackish to freshwater habitats that requires adequate osmoregulatory capacities. Hyperosmoregulation is functional at hatching and it likely begins during the embryonic phase allowing this species to develop entirely in fresh water. Here, we investigated the Na(+)/K(+)-ATPase α-subunit gene (nka-α) expression using quantitative real-time PCR and localized Na(+)/K(+)-ATPase (NKA) in ion-transporting epithelia through immunofluorescence microscopy. We reared shrimps from spawning to juvenile stages at two salinities (1, 15 ‰) and maintained adults for 3 weeks at three salinity treatments (1, 15, 25 ‰). nka-α gene expression was measured in: (1) embryos at an early (SI), intermediate (SII) and late (SIII) stage of embryonic development; (2) newly hatched larvae (Zoea I, ZI); and (3) isolated gill tissue of adults. The nka-α expression was low in SI and SII embryos and reached maximum levels prior to hatching (SIII), which were similar to expression levels detected in the ZI. The nka-α expression in SIII and ZI was highest at 15 ‰, whereas salinity did not affect expression in earlier embryos. In SIII, in ZI and in a later zoeal stage ZIV, NKA was localized in epithelial cells of pleurae, in the inner-side epithelium of branchiostegite and in the antennal glands. Gills appeared in the ZIV but NKA immunolabeling of the cells of the gill shaft occurred in a subsequent developmental larval stage, the decapodid. Extrabranchial organs constitute the main site of osmoregulation in early ontogenetic stages of this freshwater shrimp.

  9. Lactobacillus rhamnosus accelerates zebrafish backbone calcification and gonadal differentiation through effects on the GnRH and IGF systems.

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    Matteo A Avella

    Full Text Available Endogenous microbiota play essential roles in the host's immune system, physiology, reproduction and nutrient metabolism. We hypothesized that a continuous administration of an exogenous probiotic might also influence the host's development. Thus, we treated zebrafish from birth to sexual maturation (2-months treatment with Lactobacillus rhamnosus, a probiotic species intended for human use. We monitored for the presence of L. rhamnosus during the entire treatment. Zebrafish at 6 days post fertilization (dpf exhibited elevated gene expression levels for Insulin-like growth factors -I and -II, Peroxisome proliferator activated receptors -α and -β, VDR-α and RAR-γ when compared to untreated-10 days old zebrafish. Using a gonadotropin-releasing hormone 3 GFP transgenic zebrafish (GnRH3-GFP, higher GnRH3 expression was found at 6, 8 and 10 dpf upon L. rhamnosus treatment. The same larvae exhibited earlier backbone calcification and gonad maturation. Noteworthy in the gonad development was the presence of first testes differentiation at 3 weeks post fertilization in the treated zebrafish population -which normally occurs at 8 weeks- and a dramatic sex ratio modulation (93% females, 7% males in control vs. 55% females, 45% males in the treated group. We infer that administration of L. rhamnosus stimulated the IGF system, leading to a faster backbone calcification. Moreover we hypothesize a role for administration of L. rhamnosus on GnRH3 modulation during early larval development, which in turn affects gonadal development and sex differentiation. These findings suggest a significant role of the microbiota composition on the host organism development profile and open new perspectives in the study of probiotics usage and application.

  10. An α-smooth muscle actin (acta2/αsma zebrafish transgenic line marking vascular mural cells and visceral smooth muscle cells.

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    Thomas R Whitesell

    Full Text Available Mural cells of the vascular system include vascular smooth muscle cells (SMCs and pericytes whose role is to stabilize and/or provide contractility to blood vessels. One of the earliest markers of mural cell development in vertebrates is α smooth muscle actin (acta2; αsma, which is expressed by pericytes and SMCs. In vivo models of vascular mural cell development in zebrafish are currently lacking, therefore we developed two transgenic zebrafish lines driving expression of GFP or mCherry in acta2-expressing cells. These transgenic fish were used to trace the live development of mural cells in embryonic and larval transgenic zebrafish. acta2:EGFP transgenic animals show expression that largely mirrors native acta2 expression, with early pan-muscle expression starting at 24 hpf in the heart muscle, followed by skeletal and visceral muscle. At 3.5 dpf, expression in the bulbus arteriosus and ventral aorta marks the first expression in vascular smooth muscle. Over the next 10 days of development, the number of acta2:EGFP positive cells and the number of types of blood vessels associated with mural cells increases. Interestingly, the mural cells are not motile and remain in the same position once they express the acta2:EGFP transgene. Taken together, our data suggests that zebrafish mural cells develop relatively late, and have little mobility once they associate with vessels.

  11. Distinct patterns of notochord mineralization in zebrafish coincide with the localization of Osteocalcin isoform 1 during early vertebral centra formation

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    Bensimon-Brito Anabela

    2012-10-01

    Full Text Available Abstract Background In chondrichthyans, basal osteichthyans and tetrapods, vertebral bodies have cartilaginous anlagen that subsequently mineralize (chondrichthyans or ossify (osteichthyans. Chondrocytes that form the vertebral centra derive from somites. In teleost fish, vertebral centrum formation starts in the absence of cartilage, through direct mineralization of the notochord sheath. In a second step, the notochord is surrounded by somite-derived intramembranous bone. In several small teleost species, including zebrafish (Danio rerio, even haemal and neural arches form directly as intramembranous bone and only modified caudalmost arches remain cartilaginous. This study compares initial patterns of mineralization in different regions of the vertebral column in zebrafish. We ask if the absence or presence of cartilaginous arches influences the pattern of notochord sheath mineralization. Results To reveal which cells are involved in mineralization of the notochord sheath we identify proliferating cells, we trace mineralization on the histological level and we analyze cell ultrastructure by TEM. Moreover, we localize proteins and genes that are typically expressed by skeletogenic cells such as Collagen type II, Alkaline phosphatase (ALP and Osteocalcin (Oc. Mineralization of abdominal and caudal vertebrae starts with a complete ring within the notochord sheath and prior to the formation of the bony arches. In contrast, notochord mineralization of caudal fin centra starts with a broad ventral mineral deposition, associated with the bases of the modified cartilaginous arches. Similar, arch-related, patterns of mineralization occur in teleosts that maintain cartilaginous arches throughout the spine. Throughout the entire vertebral column, we were able to co-localize ALP-positive signal with chordacentrum mineralization sites, as well as Collagen II and Oc protein accumulation in the mineralizing notochord sheath. In the caudal fin region, ALP and

  12. Distinct patterns of notochord mineralization in zebrafish coincide with the localization of Osteocalcin isoform 1 during early vertebral centra formation.

    Science.gov (United States)

    Bensimon-Brito, Anabela; Cardeira, João; Cancela, Maria Leonor; Huysseune, Ann; Witten, Paul Eckhard

    2012-10-09

    In chondrichthyans, basal osteichthyans and tetrapods, vertebral bodies have cartilaginous anlagen that subsequently mineralize (chondrichthyans) or ossify (osteichthyans). Chondrocytes that form the vertebral centra derive from somites. In teleost fish, vertebral centrum formation starts in the absence of cartilage, through direct mineralization of the notochord sheath. In a second step, the notochord is surrounded by somite-derived intramembranous bone. In several small teleost species, including zebrafish (Danio rerio), even haemal and neural arches form directly as intramembranous bone and only modified caudalmost arches remain cartilaginous. This study compares initial patterns of mineralization in different regions of the vertebral column in zebrafish. We ask if the absence or presence of cartilaginous arches influences the pattern of notochord sheath mineralization. To reveal which cells are involved in mineralization of the notochord sheath we identify proliferating cells, we trace mineralization on the histological level and we analyze cell ultrastructure by TEM. Moreover, we localize proteins and genes that are typically expressed by skeletogenic cells such as Collagen type II, Alkaline phosphatase (ALP) and Osteocalcin (Oc). Mineralization of abdominal and caudal vertebrae starts with a complete ring within the notochord sheath and prior to the formation of the bony arches. In contrast, notochord mineralization of caudal fin centra starts with a broad ventral mineral deposition, associated with the bases of the modified cartilaginous arches. Similar, arch-related, patterns of mineralization occur in teleosts that maintain cartilaginous arches throughout the spine.Throughout the entire vertebral column, we were able to co-localize ALP-positive signal with chordacentrum mineralization sites, as well as Collagen II and Oc protein accumulation in the mineralizing notochord sheath. In the caudal fin region, ALP and Oc signals were clearly produced both by the

  13. Effects of different rearing temperatures on muscle development and stress response in the early larval stages of Acipenser baerii

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    Lucia Aidos

    2017-11-01

    Full Text Available The present study aims at investigating muscle development and stress response in early stages of Siberian sturgeon when subjected to different rearing temperatures, by analysing growth and development of the muscle and by assessing the stress response of yolk-sac larvae. Siberian sturgeon larvae were reared at 16°C, 19°C and 22°C until the yolk-sac was completely absorbed. Sampling timepoints were: hatching, schooling and complete yolk-sac absorption stage. Histometrical, histochemical and immunohistochemical analyses were performed in order to characterize muscle growth (total muscle area, TMA; slow muscle area, SMA; fast muscle area, FMA, development (anti-proliferating cell nuclear antigen -PCNA or anticaspase as well as stress conditions by specific stress biomarkers (heat shock protein 70 or 90, HSP70 or HSP90. Larvae subjected to the highest water temperature showed a faster yolk-sac absorption. Histometry revealed that both TMA and FMA were larger in the schooling stage at 19°C while no differences were observed in the SMA at any of the tested rearing temperatures. PCNA quantification revealed a significantly higher number of proliferating cells in the yolk-sac absorption phase at 22°C than at 16°C. HSP90 immunopositivity seems to be particularly evident at 19°C. HPS70 immunopositivity was never observed in the developing lateral muscle.

  14. A Zebrafish Heart Failure Model for Assessing Therapeutic Agents.

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    Zhu, Xiao-Yu; Wu, Si-Qi; Guo, Sheng-Ya; Yang, Hua; Xia, Bo; Li, Ping; Li, Chun-Qi

    2018-03-20

    Heart failure is a leading cause of death and the development of effective and safe therapeutic agents for heart failure has been proven challenging. In this study, taking advantage of larval zebrafish, we developed a zebrafish heart failure model for drug screening and efficacy assessment. Zebrafish at 2 dpf (days postfertilization) were treated with verapamil at a concentration of 200 μM for 30 min, which were determined as optimum conditions for model development. Tested drugs were administered into zebrafish either by direct soaking or circulation microinjection. After treatment, zebrafish were randomly selected and subjected to either visual observation and image acquisition or record videos under a Zebralab Blood Flow System. The therapeutic effects of drugs on zebrafish heart failure were quantified by calculating the efficiency of heart dilatation, venous congestion, cardiac output, and blood flow dynamics. All 8 human heart failure therapeutic drugs (LCZ696, digoxin, irbesartan, metoprolol, qiliqiangxin capsule, enalapril, shenmai injection, and hydrochlorothiazide) showed significant preventive and therapeutic effects on zebrafish heart failure (p failure model developed and validated in this study could be used for in vivo heart failure studies and for rapid screening and efficacy assessment of preventive and therapeutic drugs.

  15. Shaped 3D Singular Spectrum Analysis for Quantifying Gene Expression, with Application to the Early Zebrafish Embryo

    Directory of Open Access Journals (Sweden)

    Alex Shlemov

    2015-01-01

    Full Text Available Recent progress in microscopy technologies, biological markers, and automated processing methods is making possible the development of gene expression atlases at cellular-level resolution over whole embryos. Raw data on gene expression is usually very noisy. This noise comes from both experimental (technical/methodological and true biological sources (from stochastic biochemical processes. In addition, the cells or nuclei being imaged are irregularly arranged in 3D space. This makes the processing, extraction, and study of expression signals and intrinsic biological noise a serious challenge for 3D data, requiring new computational approaches. Here, we present a new approach for studying gene expression in nuclei located in a thick layer around a spherical surface. The method includes depth equalization on the sphere, flattening, interpolation to a regular grid, pattern extraction by Shaped 3D singular spectrum analysis (SSA, and interpolation back to original nuclear positions. The approach is demonstrated on several examples of gene expression in the zebrafish egg (a model system in vertebrate development. The method is tested on several different data geometries (e.g., nuclear positions and different forms of gene expression patterns. Fully 3D datasets for developmental gene expression are becoming increasingly available; we discuss the prospects of applying 3D-SSA to data processing and analysis in this growing field.

  16. Towards a comprehensive catalog of zebrafish behavior 1.0 and beyond.

    Science.gov (United States)

    Kalueff, Allan V; Gebhardt, Michael; Stewart, Adam Michael; Cachat, Jonathan M; Brimmer, Mallorie; Chawla, Jonathan S; Craddock, Cassandra; Kyzar, Evan J; Roth, Andrew; Landsman, Samuel; Gaikwad, Siddharth; Robinson, Kyle; Baatrup, Erik; Tierney, Keith; Shamchuk, Angela; Norton, William; Miller, Noam; Nicolson, Teresa; Braubach, Oliver; Gilman, Charles P; Pittman, Julian; Rosemberg, Denis B; Gerlai, Robert; Echevarria, David; Lamb, Elisabeth; Neuhauss, Stephan C F; Weng, Wei; Bally-Cuif, Laure; Schneider, Henning

    2013-03-01

    Zebrafish (Danio rerio) are rapidly gaining popularity in translational neuroscience and behavioral research. Physiological similarity to mammals, ease of genetic manipulations, sensitivity to pharmacological and genetic factors, robust behavior, low cost, and potential for high-throughput screening contribute to the growing utility of zebrafish models in this field. Understanding zebrafish behavioral phenotypes provides important insights into neural pathways, physiological biomarkers, and genetic underpinnings of normal and pathological brain function. Novel zebrafish paradigms continue to appear with an encouraging pace, thus necessitating a consistent terminology and improved understanding of the behavioral repertoire. What can zebrafish 'do', and how does their altered brain function translate into behavioral actions? To help address these questions, we have developed a detailed catalog of zebrafish behaviors (Zebrafish Behavior Catalog, ZBC) that covers both larval and adult models. Representing a beginning of creating a more comprehensive ethogram of zebrafish behavior, this effort will improve interpretation of published findings, foster cross-species behavioral modeling, and encourage new groups to apply zebrafish neurobehavioral paradigms in their research. In addition, this glossary creates a framework for developing a zebrafish neurobehavioral ontology, ultimately to become part of a unified animal neurobehavioral ontology, which collectively will contribute to better integration of biological data within and across species.

  17. Effects of rare earth elements La and Yb on the morphological and functional development of zebrafish embryos

    Institute of Scientific and Technical Information of China (English)

    Jun'an Cui; Zhiyong Zhang; Wei Bai; Ligang Zhang; Xiao He; Yuhui Ma; Yan Liu; Zhifang Chai

    2012-01-01

    In recent years,with the wide applications and mineral exploitation of rare earth elements,their potential environmental and health effects have caused increasing public concern.Effect of rare earth elements La and Yb on the morphological and functional development of zebrafish embryos were studied.The embryos were exposed to La3+ or Yb3+ at 0,0.01,0.1,0.3,0.5 and 1.0 mmol/L,respectively.Early life stage parameters such as egg and embryo mortality,gastrula development,tail detachment,eyes,somite formation,circulatory system,pigmentation,malformations,hatching rate,length of larvae and mortality were investigated.The results showed La3+ and Yb3+ delayed zebrafish embryo and larval development,decreased survival and hatching rates,and caused tail malformation in a concentration-dependent way.Moreover,heavy rare-earth ytterbium led to more severe acute toxicity of zebrafish embryo than light rare-earth lanthanum.

  18. Targeted knock-in of CreER T2 in zebrafish using CRISPR/Cas9.

    Science.gov (United States)

    Kesavan, Gokul; Hammer, Juliane; Hans, Stefan; Brand, Michael

    2018-04-01

    New genome-editing approaches, such as the CRISPR/Cas system, have opened up great opportunities to insert or delete genes at targeted loci and have revolutionized genetics in model organisms like the zebrafish. The Cre-loxp recombination system is widely used to activate or inactivate genes with high spatial and temporal specificity. Using a CRISPR/Cas9-mediated knock-in strategy, we inserted a zebrafish codon-optimized CreER T2 transgene at the otx2 gene locus to generate a conditional Cre-driver line. We chose otx2 as it is a patterning gene of the anterior neural plate that is expressed during early development. By knocking in CreER T2 upstream of the endogenous ATG of otx2, we utilized this gene's native promoter and enhancer elements to perfectly match CreER T2 and endogenous otx2 expression patterns. Next, by combining this novel driver line with a Cre-dependent reporter line, we show that only in the presence of tamoxifen can efficient Cre-loxp-mediated recombination be achieved in the anterior neural plate-derived tissues like the telencephalon, the eye and the optic tectum. Our results imply that the otx2:CreER T2 transgenic fish will be a valuable tool for lineage tracing and conditional mutant studies in larval and adult zebrafish.

  19. Clofibrate and gemfibrozil induce an embryonic malabsorption syndrome in zebrafish

    International Nuclear Information System (INIS)

    Raldua, Demetrio; Andre, Michele; Babin, Patrick J.

    2008-01-01

    Nutrient availability is one of the major non-genetic factors determining embryonic growth and larval or fetal size. Due to the high human consumption of blood lipid regulators, fibrates have recently been reported as pollutants in rivers. Our study investigated the developmental toxicity of fibrates in zebrafish. Treatment with micromolar concentrations of clofibrate or gemfibrozil induced an embryonic malabsorption syndrome (EMS) with very little yolk consumption, resulting in small-sized larvae. This effect was reversible on removing the drug from the water. Clofibrate delayed hatching time and decreased the amount of oil red O lipid staining in the vasculature. It also induced higher density, round-shaped neuromuscular junctions associated with disorganization and less striation of muscular fibers, and pericardial edema, as well as impairing thyroid gland morphogenesis. acox1, apoa1 and mtp hybridization transcript signals were not affected in the yolk syncytial layer (YSL) after clofibrate exposure. Di-(2-ethylhexyl)-phthalate did not slow down yolk resorption, whereas brefeldin A induced EMS. These findings suggest that the inhibition of yolk sac resorption on exposure to fibrate is not at a pre-translational level or peroxisome proliferator-activated receptor alpha dependent and may be due to an inhibition of the YSL constitutive cell secretion. The effects of fibrates and the potential bioconcentration in eggs as well as the additive action of structurally related toxicants warrant an evaluation of the developmental impact of these compounds after long-term exposure at environmentally relevant concentrations. Fibrate-induced EMS in zebrafish seems useful for studying the morphogenetic consequences of impaired nutrient availability during the early stages of vertebrate development

  20. Quantification of birefringence readily measures the level of muscle damage in zebrafish

    Energy Technology Data Exchange (ETDEWEB)

    Berger, Joachim, E-mail: Joachim.Berger@Monash.edu [Australian Regenerative Medicine Institute, EMBL Australia, Monash University, Clayton (Australia); Sztal, Tamar; Currie, Peter D. [Australian Regenerative Medicine Institute, EMBL Australia, Monash University, Clayton (Australia)

    2012-07-13

    Highlights: Black-Right-Pointing-Pointer Report of an unbiased quantification of the birefringence of muscle of fish larvae. Black-Right-Pointing-Pointer Quantification method readily identifies level of overall muscle damage. Black-Right-Pointing-Pointer Compare zebrafish muscle mutants for level of phenotype severity. Black-Right-Pointing-Pointer Proposed tool to survey treatments that aim to ameliorate muscular dystrophy. -- Abstract: Muscular dystrophies are a group of genetic disorders that progressively weaken and degenerate muscle. Many zebrafish models for human muscular dystrophies have been generated and analysed, including dystrophin-deficient zebrafish mutants dmd that model Duchenne Muscular Dystrophy. Under polarised light the zebrafish muscle can be detected as a bright area in an otherwise dark background. This light effect, called birefringence, results from the diffraction of polarised light through the pseudo-crystalline array of the muscle sarcomeres. Muscle damage, as seen in zebrafish models for muscular dystrophies, can readily be detected by a reduction in the birefringence. Therefore, birefringence is a very sensitive indicator of overall muscle integrity within larval zebrafish. Unbiased documentation of the birefringence followed by densitometric measurement enables the quantification of the birefringence of zebrafish larvae. Thereby, the overall level of muscle integrity can be detected, allowing the identification and categorisation of zebrafish muscle mutants. In addition, we propose that the establish protocol can be used to analyse treatments aimed at ameliorating dystrophic zebrafish models.

  1. Quantification of birefringence readily measures the level of muscle damage in zebrafish

    International Nuclear Information System (INIS)

    Berger, Joachim; Sztal, Tamar; Currie, Peter D.

    2012-01-01

    Highlights: ► Report of an unbiased quantification of the birefringence of muscle of fish larvae. ► Quantification method readily identifies level of overall muscle damage. ► Compare zebrafish muscle mutants for level of phenotype severity. ► Proposed tool to survey treatments that aim to ameliorate muscular dystrophy. -- Abstract: Muscular dystrophies are a group of genetic disorders that progressively weaken and degenerate muscle. Many zebrafish models for human muscular dystrophies have been generated and analysed, including dystrophin-deficient zebrafish mutants dmd that model Duchenne Muscular Dystrophy. Under polarised light the zebrafish muscle can be detected as a bright area in an otherwise dark background. This light effect, called birefringence, results from the diffraction of polarised light through the pseudo-crystalline array of the muscle sarcomeres. Muscle damage, as seen in zebrafish models for muscular dystrophies, can readily be detected by a reduction in the birefringence. Therefore, birefringence is a very sensitive indicator of overall muscle integrity within larval zebrafish. Unbiased documentation of the birefringence followed by densitometric measurement enables the quantification of the birefringence of zebrafish larvae. Thereby, the overall level of muscle integrity can be detected, allowing the identification and categorisation of zebrafish muscle mutants. In addition, we propose that the establish protocol can be used to analyse treatments aimed at ameliorating dystrophic zebrafish models.

  2. Zebrafish embryos exposed to alcohol undergo abnormal development of motor neurons and muscle fibers.

    Science.gov (United States)

    Sylvain, Nicole J; Brewster, Daniel L; Ali, Declan W

    2010-01-01

    Children exposed to alcohol in utero have significantly delayed gross and fine motor skills, as well as deficiencies in reflex development. The reasons that underlie the motor deficits caused by ethanol (EtOH) exposure remain to be fully elucidated. The present study was undertaken to investigate the effects of embryonic alcohol exposure (1.5%, 2% and 2.5% EtOH) on motor neuron and muscle fiber morphology in 3 days post fertilization (dpf) larval zebrafish. EtOH treated fish exhibited morphological deformities and fewer bouts of swimming in response to touch, compared with untreated fish. Immunolabelling with anti-acetylated tubulin indicated that fish exposed to 2.5% EtOH had significantly higher rates of motor neuron axon defects. Immunolabelling of primary and secondary motor neurons, using znp-1 and zn-8, revealed that fish exposed to 2% and 2.5% EtOH exhibited significantly higher rates of primary and secondary motor neuron axon defects compared to controls. Examination of red and white muscle fibers revealed that fish exposed to EtOH had significantly smaller fibers compared with controls. These findings indicate that motor neuron and muscle fiber morphology is affected by early alcohol exposure in zebrafish embryos, and that this may be related to deficits in locomotion. Copyright 2010 Elsevier Inc. All rights reserved.

  3. Pregnancy-associated plasma protein-A (PAPP-A) modulates early developmental rate in zebrafish independent of its proteolytic activity

    DEFF Research Database (Denmark)

    Kjær-Sørensen, Kasper; Engholm, Ditte Høyer; Kamei, Hiroyasu

    2013-01-01

    the developmental rate beginning during gastrulation without affecting the normal patterning of the embryo. This phenotype is different from those resulting from deficiency of Igf receptor or ligand in zebrafish, suggesting a function of Papp-a outside the Igf system. Biochemical analysis of recombinant zebrafish...... Papp-a demonstrates conservation of proteolytic activity, specificity, and intrinsic regulatory mechanism. However, in vitro transcribed mRNA, which encodes a proteolytically inactive Papp-a mutant, recues the papp-a knockdown phenotype as efficient as wild-type Papp-a. Thus, the developmental...

  4. Developmental exposure to low concentrations of two brominated flame retardants, BDE-47 and BDE-99, causes life-long behavioral alterations in zebrafish.

    Science.gov (United States)

    Glazer, Lilah; Wells, Corinne N; Drastal, Meghan; Odamah, Kathryn-Ann; Galat, Richard E; Behl, Mamta; Levin, Edward D

    2018-05-01

    Polybrominated diphenyl ethers (PBDEs) were widely used as flame retardants until the early 2000s, mainly in home furnishings and electronics. The persistence of PBDEs in the environment leads to continued ubiquitous exposure to low levels, with infants and children experiencing higher exposures than adults. Accumulating evidence suggest that low-level exposures during early life stages can affect brain development and lead to long-term behavioral impairments. We investigated the effects of zebrafish exposure to low doses of the two prominent PBDEs; 2,2',4,4',5,-Pentabromodiphenyl ether (BDE-99) and 2,2',4,4',-Tetrabromodiphenyl ether (BDE-47), during embryo-development on short- and long-term behavioral endpoints. We included the organophosphate pesticide chlorpyrifos (CPF) due to its well documented neurotoxicity across species from zebrafish to humans. Zebrafish embryos were exposed to the following individual treatments; 0.1% DMSO (vehicle control); 0.3μM CPF; 0.01, 0.03, 0.1, 0.3μM BDE-47; 0.003, 0.03, 0.3, 1, 3, 10, 20μM BDE-99 from 5 until 120h post fertilization (hpf). Low exposure levels were determined as those not causing immediate overt toxicity, and behavior assays were conducted in the low-level range. At 144 hpf the larvae were tested for locomotor activity. At approximately 6 months of age adult zebrafish were tested in a behavioral battery including assays for anxiety-related behavior, sensorimotor response and habituation, social interaction, and predator avoidance. In the short-term, larval locomotor activity was reduced in larvae treated with 0.3μM CPF and 0.1μM BDE-47. BDE-99 treatment caused non-monotonic dose effects, with 0.3μM causing hyperactivity and 1μM or higher causing hypoactivity. In the long-term, adult anxiety-related behavior was reduced in all treatments as measured in both the novel tank dive test and tap test. We show that exposure of zebrafish embryos to low concentrations of the brominated flame retardants BDE-47 and

  5. Afferent connectivity of the zebrafish habenulae

    Directory of Open Access Journals (Sweden)

    Katherine Jane Turner

    2016-04-01

    Full Text Available The habenulae are bilateral nuclei located in the dorsal diencephalon that are conserved across vertebrates.Here we describe the main afferents to the habenulae in larval and adult zebrafish.We observe afferents from the subpallium, nucleus rostrolateralis,posterior tuberculum, posterior hypothalamic lobe, median raphe, olfactory bulb to the right habenula and from the parapineal to the lefthabenula.In addition,we find afferents from a ventrolateral telencephalic nucleus that neurochemical and hodological data identify as the ventral entopeduncular nucleus(vENT,confirming and extending observations of Amo et al.(2014.Fate map and marker studies suggest that vENT originates from the diencephalic prethalamic eminence and extends into the lateral telencephalon from 48 to 120 hpf.No afferents to the habenula were observed from the dorsal entopeduncular nucleus(dENT.Consequently,we confirm that the vENT(and not the dENT should be considered as the entopeduncular nucleus proper in zebrafish.Furthermore,comparison with data in other vertebrates suggests that the vENT is a conserved basal ganglia nucleus,being homologous to the entopeduncular nucleus of mammals(internal segment of the globus pallidus of primates by both embryonic origin and projections,as previously suggested by Amo et al.(2014.Key words: habenula,connections,afferents,entopeduncular nucleus,posterior tuberculum,basal ganglia,zebrafish

  6. Evaluation of visible implant elastomer tags in zebrafish (Danio rerio

    Directory of Open Access Journals (Sweden)

    Claudia Hohn

    2013-11-01

    The use of the visible implant elastomer (VIE tagging system in zebrafish (Danio rerio was examined. Two tag orientations (horizontal and vertical at the dorsal fin base were tested for tag retention, tag fragmentation and whether VIE tags affected growth and survival of juvenile zebrafish (1–4 month post hatch. Six tag locations (abdomen, anal fin base, caudal peduncle, dorsal fin base, pectoral fin base, isthmus and 5 tag colors (yellow, red, pink, orange, blue were evaluated for ease of VIE tag application and tag visibility in adult zebrafish. Long-term retention (1 year and multiple tagging sites (right and left of dorsal fin and pectoral fin base were examined in adult zebrafish. Lastly, survival of recombination activation gene 1−/− (rag1−/− zebrafish was evaluated after VIE tagging. The best tag location was the dorsal fin base, and the most visible tag color was pink. Growth rate of juvenile zebrafish was not affected by VIE tagging. Horizontal tagging is recommended in early stages of fish growth (1–2 months post hatch. VIE tags were retained for 1 year and tagging did not interfere with long-term growth and survival. There was no mortality associated with VIE tagging in rag1−/− zebrafish. The VIE tagging system is highly suitable for small-sized zebrafish. When familiar with the procedure, 120 adult zebrafish can be tagged in one hour. It does not increase mortality in adult zebrafish or interfere with growth in juvenile or adult zebrafish.

  7. Zebrafish as a Model System for Investigating the Compensatory Regulation of Ionic Balance during Metabolic Acidosis

    Directory of Open Access Journals (Sweden)

    Lletta Lewis

    2018-04-01

    Full Text Available Zebrafish (Danio rerio have become an important model for integrative physiological research. Zebrafish inhabit a hypo-osmotic environment; to maintain ionic and acid-base homeostasis, they must actively take up ions and secrete acid to the water. The gills in the adult and the skin at larval stage are the primary sites of ionic regulation in zebrafish. The uptake of ions in zebrafish is mediated by specific ion transporting cells termed ionocytes. Similarly, in mammals, ion reabsorption and acid excretion occur in specific cell types in the terminal region of the renal tubules (distal convoluted tubule and collecting duct. Previous studies have suggested that functional regulation of several ion transporters/channels in the zebrafish ionocytes resembles that in the mammalian renal cells. Additionally, several mechanisms involved in regulating the epithelial ion transport during metabolic acidosis are found to be similar between zebrafish and mammals. In this article, we systemically review the similarities and differences in ionic regulation between zebrafish and mammals during metabolic acidosis. We summarize the available information on the regulation of epithelial ion transporters during acidosis, with a focus on epithelial Na+, Cl− and Ca2+ transporters in zebrafish ionocytes and mammalian renal cells. We also discuss the neuroendocrine responses to acid exposure, and their potential role in ionic compensation. Finally, we identify several knowledge gaps that would benefit from further study.

  8. Patterns of free calcium in zebrafish embryos

    NARCIS (Netherlands)

    Creton, R; Speksnijder, JE; Jaffe, LF

    Direct knowledge of Ca2+ patterns in vertebrate development is largely restricted to early stages, in which they control fertilization, ooplasmic segregation and cleavage. To explore new roles of Ca2+ in vertebrate development, we injected the Ca2+ indicator aequorin into zebrafish eggs and imaged

  9. The search for evolutionary developmental origins of aging in zebrafish: a novel intersection of developmental and senescence biology in the zebrafish model system.

    Science.gov (United States)

    Kishi, Shuji

    2011-09-01

    Senescence may be considered the antithesis of early development, but yet there may be factors and mechanisms in common between these two phenomena during the process of aging. We investigated whether any relationship exists between the regulatory mechanisms that function in early development and in senescence using the zebrafish (Danio rerio), a small freshwater fish and a useful model animal for genetic studies. We conducted experiments to isolate zebrafish mutants expressing an apparent senescence phenotype during embryogenesis (embryonic senescence). Some of the genes we thereby identified had already been associated with cellular senescence and chronological aging in other organisms, but many had not yet been linked to these processes. Complete loss-of-function of developmentally essential genes induce embryonic (or larval) lethality, whereas it seems like their partial loss-of-function (i.e., decrease-of-function by heterozygote or hypomorphic mutations) still remains sufficient to go through the early developmental process because of its adaptive plasticity or rather heterozygote advantage. However, in some cases, such partial loss-of-function of genes compromise normal homeostasis due to haploinsufficiency later in adult life having many environmental stress challenges. By contrast, any heterozygote-advantageous genes might gain a certain benefit(s) (much more fitness) by such partial loss-of-function later in life. Physiological senescence may evolutionarily arise from both genetic and epigenetic drifts as well as from losing adaptive developmental plasticity in face of stress signals from the external environment that interacts with functions of multiple genes rather than effects of only a single gene mutation or defect. Previously uncharacterized developmental genes may thus mediate the aging process and play a pivotal role in senescence. Moreover, unexpected senescence-related genes might also be involved in the early developmental process and

  10. Developmental social isolation affects adult behavior, social interaction, and dopamine metabolite levels in zebrafish.

    Science.gov (United States)

    Shams, Soaleha; Amlani, Shahid; Buske, Christine; Chatterjee, Diptendu; Gerlai, Robert

    2018-01-01

    The zebrafish is a social vertebrate and an excellent translational model for a variety of human disorders. Abnormal social behavior is a hallmark of several human brain disorders. Social behavioral problems can arise as a result of adverse early social environment. Little is known about the effects of early social isolation in adult zebrafish. We compared zebrafish that were isolated for either short (7 days) or long duration (180 days) to socially housed zebrafish, testing their behavior across ontogenesis (ages 10, 30, 60, 90, 120, 180 days), and shoal cohesion and whole-brain monoamines and their metabolites in adulthood. Long social isolation increased locomotion and decreased shoal cohesion and anxiety in the open-field in adult. Additionally, both short and long social isolation reduced dopamine metabolite levels in response to social stimuli. Thus, early social isolation has lasting effects in zebrafish, and may be employed to generate zebrafish models of human neuropsychiatric conditions. © 2017 Wiley Periodicals, Inc.

  11. Assessment of cardiotoxicity and effects of malathion on the early development of zebrafish (Danio rerio) using computer vision for heart rate quantification.

    Science.gov (United States)

    Simoneschi, Daniele; Simoneschi, Francesco; Todd, Nancy E

    2014-06-01

    Malathion, a common organophosphate insecticide, is a proven acetylcholinesterase inhibitor and is the most applied organophosphate insecticide in the United States. The use of zebrafish as a model to study the effects of pesticides on development is an innovative approach yielding relevant implications for determining the potential toxic effects of these pesticides on humans. In this study, a simple noninvasive technique was developed to investigate the cardiotoxicity of malathion on Danio rerio embryos, and to detect and quantify its effect on heart rate. Videos were recorded under a stereomicroscope and examined with our custom-made software (FishBeat) to determine the heart rate of the embryos. The pixel average intensity frequency (PI) of the videos was computed at its maximum probability to indicate the average number of heartbeats per second. Experimental observations successfully demonstrated that this method was able to detect the heart rate of zebrafish embryos as compared with manual stopwatch counting, with no significant difference. Embryos were treated acutely with increasing malathion concentrations (33.3 and 50 μg/mL malathion) at 52, 76, and 96 hpf. Embryos treated with 33.3 μg/mL malathion had significant bradycardia at 52 and 76 hpf, whereas embryos treated with 50 μg/mL malathion presented bradycardia at all hpf. These novel observations confirmed that malathion, acting as an acetylcholinesterase inhibitor, induced heartbeat irregularity in zebrafish embryos.

  12. The Zebrafish Models to Explore Genetic and Epigenetic Impacts on Evolutionary Developmental Origins of Aging

    Science.gov (United States)

    Kishi, Shuji

    2014-01-01

    Can we reset, reprogram, rejuvenate or reverse the organismal aging process? Certain genetic manipulations could at least reset and reprogram epigenetic dynamics beyond phenotypic plasticity and elasticity in cells, which can be further manipulated into organisms. However, in a whole complex aging organism, how can we rejuvenate intrinsic resources and infrastructures in an intact/noninvasive manner? The incidence of diseases increases exponentially with age, accompanied by progressive deteriorations of physiological functions in organisms. Aging-associated diseases are sporadic but essentially inevitable complications arising from senescence. Senescence is often considered the antithesis of early development, but yet there may be factors and mechanisms in common between these two phenomena to rejuvenate over the dynamic process of aging. The association between early development and late-onset disease with advancing age is thought to come from a consequence of developmental plasticity, the phenomenon by which one genotype can give rise to a range of physiologically and/or morphologically adaptive states based on diverse epigenotypes, in response to intrinsic or extrinsic environmental cues and genetic perturbations. We hypothesized that the future aging process can be predictive based on adaptivity during the early developmental period. Modulating the thresholds and windows of plasticity and its robustness by molecular genetic and chemical epigenetic approaches, we have successfully conducted experiments to isolate zebrafish mutants expressing apparently altered senescence phenotypes during their embryonic and/or larval stages (“embryonic/larval senescence”). Subsequently, at least some of these mutant animals were found to show shortened lifespan, while some others would be expected to live longer in adulthoods. We anticipate that previously uncharacterized developmental genes may mediate the aging process and play a pivotal role in senescence. On the other

  13. Statistical Analysis of Zebrafish Locomotor Response.

    Science.gov (United States)

    Liu, Yiwen; Carmer, Robert; Zhang, Gaonan; Venkatraman, Prahatha; Brown, Skye Ashton; Pang, Chi-Pui; Zhang, Mingzhi; Ma, Ping; Leung, Yuk Fai

    2015-01-01

    Zebrafish larvae display rich locomotor behaviour upon external stimulation. The movement can be simultaneously tracked from many larvae arranged in multi-well plates. The resulting time-series locomotor data have been used to reveal new insights into neurobiology and pharmacology. However, the data are of large scale, and the corresponding locomotor behavior is affected by multiple factors. These issues pose a statistical challenge for comparing larval activities. To address this gap, this study has analyzed a visually-driven locomotor behaviour named the visual motor response (VMR) by the Hotelling's T-squared test. This test is congruent with comparing locomotor profiles from a time period. Different wild-type (WT) strains were compared using the test, which shows that they responded differently to light change at different developmental stages. The performance of this test was evaluated by a power analysis, which shows that the test was sensitive for detecting differences between experimental groups with sample numbers that were commonly used in various studies. In addition, this study investigated the effects of various factors that might affect the VMR by multivariate analysis of variance (MANOVA). The results indicate that the larval activity was generally affected by stage, light stimulus, their interaction, and location in the plate. Nonetheless, different factors affected larval activity differently over time, as indicated by a dynamical analysis of the activity at each second. Intriguingly, this analysis also shows that biological and technical repeats had negligible effect on larval activity. This finding is consistent with that from the Hotelling's T-squared test, and suggests that experimental repeats can be combined to enhance statistical power. Together, these investigations have established a statistical framework for analyzing VMR data, a framework that should be generally applicable to other locomotor data with similar structure.

  14. Zebrafish hair cell mechanics and physiology through the lens of noise-induced hair cell death

    Science.gov (United States)

    Coffin, Allison B.; Xu, Jie; Uribe, Phillip M.

    2018-05-01

    Hair cells are exquisitely sensitive to auditory stimuli, but also to damage from a variety of sources including noise trauma and ototoxic drugs. Mammals cannot regenerate cochlear hair cells, while non-mammalian vertebrates exhibit robust regenerative capacity. Our research group uses the lateral line system of larval zebrafish to explore the mechanisms underlying hair cell damage, identify protective therapies, and determine molecular drivers of innate regeneration. The lateral line system contains externally located sensory organs called neuromasts, each composed of ˜8-20 hair cells. Lateral line hair cells are homologous to vertebrate inner ear hair cells and share similar susceptibility to ototoxic damage. In the last decade, the lateral line has emerged as a powerful model system for understanding hair cell death mechanisms and for identifying novel protective compounds. Here we demonstrate that the lateral line is a tractable model for noise-induced hair cell death. We have developed a novel noise damage system capable of inducing over 50% loss of lateral line hair cells, with hair cell death occurring in a dose- and time-dependent manner. Cell death is greatest 72 hours post-exposure. However, early signs of hair cell damage, including changes in membrane integrity and reduced mechanotransduction, are apparent within hours of noise exposure. These features, early signs of damage followed by delayed hair cell death, are consistent with mammalian data, suggesting that noise acts similarly on zebrafish and mammalian hair cells. In our future work we will use our new model system to investigate noise damage events in real time, and to develop protective therapies for future translational research.

  15. Toxic effects of silica nanoparticles on zebrafish embryos and larvae.

    Directory of Open Access Journals (Sweden)

    Junchao Duan

    Full Text Available Silica nanoparticles (SiNPs have been widely used in biomedical and biotechnological applications. Environmental exposure to nanomaterials is inevitable as they become part of our daily life. Therefore, it is necessary to investigate the possible toxic effects of SiNPs exposure. In this study, zebrafish embryos were treated with SiNPs (25, 50, 100, 200 µg/mL during 4-96 hours post fertilization (hpf. Mortality, hatching rate, malformation and whole-embryo cellular death were detected. We also measured the larval behavior to analyze whether SiNPs had adverse effects on larvae locomotor activity. The results showed that as the exposure dosages increasing, the hatching rate of zebrafish embryos was decreased while the mortality and cell death were increased. Exposure to SiNPs caused embryonic malformations, including pericardial edema, yolk sac edema, tail and head malformation. The larval behavior testing showed that the total swimming distance was decreased in a dose-dependent manner. The lower dose (25 and 50 µg/mL SiNPs produced substantial hyperactivity while the higher doses (100 and 200 µg/mL SiNPs elicited remarkably hypoactivity in dark periods. In summary, our data indicated that SiNPs caused embryonic developmental toxicity, resulted in persistent effects on larval behavior.

  16. Swimming Effects on Developing Zebrafish

    NARCIS (Netherlands)

    Kranenbarg, S.; Pelster, B.

    2013-01-01

    Zebrafish represent an important vertebrate model species in developmental biology. This chapter reviews the effects of exercise on the development of the musculoskeletal system, the cardiovascular system, metabolic capacities of developing zebrafish, and regulation of these processes on the gene

  17. Whole plant based treatment of hypercholesterolemia with Crataegus laevigata in a zebrafish model

    Directory of Open Access Journals (Sweden)

    Littleton Robert M

    2012-07-01

    Full Text Available Abstract Background Consumers are increasingly turning to plant-based complementary and alternative medicines to treat hypercholesterolemia. Many of these treatments are untested and their efficacy is unknown. This multitude of potential remedies necessitates a model system amenable to testing large numbers of organisms that maintains similarity to humans in both mode of drug administration and overall physiology. Here we develop the larval zebrafish (4–30 days post fertilization as a vertebrate model of dietary plant-based treatment of hypercholesterolemia and test the effects of Crataegus laevigata in this model. Methods Larval zebrafish were fed high cholesterol diets infused with fluorescent sterols and phytomedicines. Plants were ground with mortar and pestle into a fine powder before addition to food. Fluorescent sterols were utilized to optically quantify relative difference in intravascular cholesterol levels between groups of fish. We utilized the Zeiss 7-Live Duo high-speed confocal platform in order to both quantify intravascular sterol fluorescence and to capture video of the heart beat for determination of cardiac output. Results In this investigation we developed and utilized a larval zebrafish model to investigate dietary plant-based intervention of the pathophysiology of hypercholesterolemia. We found BODIPY-cholesterol effectively labels diet-introduced intravascular cholesterol levels (P t-test. We also established that zebrafish cardiac output declines as cholesterol dose increases (difference between 0.1% and 8% (w/w high cholesterol diet-treated cardiac output significant at P  Conclusions The results of this study demonstrate that the larval zebrafish has the potential to become a powerful model to test plant based dietary intervention of hypercholesterolemia. Using this model we have shown that hawthorn leaves and flowers have the potential to affect cardiac output as well as intravascular cholesterol levels

  18. A tool box for operational mosquito larval control: preliminary results and early lessons from the Urban Malaria Control Programme in Dar es Salaam, Tanzania

    Directory of Open Access Journals (Sweden)

    Govella Nico J

    2008-01-01

    Full Text Available Abstract Background As the population of Africa rapidly urbanizes, large populations could be protected from malaria by controlling aquatic stages of mosquitoes if cost-effective and scalable implementation systems can be designed. Methods A recently initiated Urban Malaria Control Programme in Dar es Salaam delegates responsibility for routine mosquito control and surveillance to modestly-paid community members, known as Community-Owned Resource Persons (CORPs. New vector surveillance, larviciding and management systems were designed and evaluated in 15 city wards to allow timely collection, interpretation and reaction to entomologic monitoring data using practical procedures that rely on minimal technology. After one year of baseline data collection, operational larviciding with Bacillus thuringiensis var. israelensis commenced in March 2006 in three selected wards. Results The procedures and staff management systems described greatly improved standards of larval surveillance relative to that reported at the outset of this programme. In the first year of the programme, over 65,000 potential Anopheles habitats were surveyed by 90 CORPs on a weekly basis. Reaction times to vector surveillance at observations were one day, week and month at ward, municipal and city levels, respectively. One year of community-based larviciding reduced transmission by the primary malaria vector, Anopheles gambiae s.l., by 31% (95% C.I. = 21.6–37.6%; p = 0.04. Conclusion This novel management, monitoring and evaluation system for implementing routine larviciding of malaria vectors in African cities has shown considerable potential for sustained, rapidly responsive, data-driven and affordable application. Nevertheless, the true programmatic value of larviciding in urban Africa can only be established through longer-term programmes which are stably financed and allow the operational teams and management infrastructures to mature by learning from experience.

  19. Imaging Subcellular Structures in the Living Zebrafish Embryo.

    Science.gov (United States)

    Engerer, Peter; Plucinska, Gabriela; Thong, Rachel; Trovò, Laura; Paquet, Dominik; Godinho, Leanne

    2016-04-02

    In vivo imaging provides unprecedented access to the dynamic behavior of cellular and subcellular structures in their natural context. Performing such imaging experiments in higher vertebrates such as mammals generally requires surgical access to the system under study. The optical accessibility of embryonic and larval zebrafish allows such invasive procedures to be circumvented and permits imaging in the intact organism. Indeed the zebrafish is now a well-established model to visualize dynamic cellular behaviors using in vivo microscopy in a wide range of developmental contexts from proliferation to migration and differentiation. A more recent development is the increasing use of zebrafish to study subcellular events including mitochondrial trafficking and centrosome dynamics. The relative ease with which these subcellular structures can be genetically labeled by fluorescent proteins and the use of light microscopy techniques to image them is transforming the zebrafish into an in vivo model of cell biology. Here we describe methods to generate genetic constructs that fluorescently label organelles, highlighting mitochondria and centrosomes as specific examples. We use the bipartite Gal4-UAS system in multiple configurations to restrict expression to specific cell-types and provide protocols to generate transiently expressing and stable transgenic fish. Finally, we provide guidelines for choosing light microscopy methods that are most suitable for imaging subcellular dynamics.

  20. Zebrafish Health Conditions in the China Zebrafish Resource Center and 20 Major Chinese Zebrafish Laboratories.

    Science.gov (United States)

    Liu, Liyue; Pan, Luyuan; Li, Kuoyu; Zhang, Yun; Zhu, Zuoyan; Sun, Yonghua

    2016-07-01

    In China, the use of zebrafish as an experimental animal in the past 15 years has widely expanded. The China Zebrafish Resource Center (CZRC), which was established in 2012, is becoming one of the major resource centers in the global zebrafish community. Large-scale use and regular exchange of zebrafish resources have put forward higher requirements on zebrafish health issues in China. This article reports the current aquatic infrastructure design, animal husbandry, and health-monitoring programs in the CZRC. Meanwhile, through a survey of 20 Chinese zebrafish laboratories, we also describe the current health status of major zebrafish facilities in China. We conclude that it is of great importance to establish a widely accepted health standard and health-monitoring strategy in the Chinese zebrafish research community.

  1. Afferent Connectivity of the Zebrafish Habenulae

    Science.gov (United States)

    Turner, Katherine J.; Hawkins, Thomas A.; Yáñez, Julián; Anadón, Ramón; Wilson, Stephen W.; Folgueira, Mónica

    2016-01-01

    The habenulae are bilateral nuclei located in the dorsal diencephalon that are conserved across vertebrates. Here we describe the main afferents to the habenulae in larval and adult zebrafish. We observe afferents from the subpallium, nucleus rostrolateralis, posterior tuberculum, posterior hypothalamic lobe, median raphe; we also see asymmetric afferents from olfactory bulb to the right habenula, and from the parapineal to the left habenula. In addition, we find afferents from a ventrolateral telencephalic nucleus that neurochemical and hodological data identify as the ventral entopeduncular nucleus (vENT), confirming and extending observations of Amo et al. (2014). Fate map and marker studies suggest that vENT originates from the diencephalic prethalamic eminence and extends into the lateral telencephalon from 48 to 120 hour post-fertilization (hpf). No afferents to the habenula were observed from the dorsal entopeduncular nucleus (dENT). Consequently, we confirm that the vENT (and not the dENT) should be considered as the entopeduncular nucleus “proper” in zebrafish. Furthermore, comparison with data in other vertebrates suggests that the vENT is a conserved basal ganglia nucleus, being homologous to the entopeduncular nucleus of mammals (internal segment of the globus pallidus of primates) by both embryonic origin and projections, as previously suggested by Amo et al. (2014). PMID:27199671

  2. Learning and memory in zebrafish (Danio rerio).

    Science.gov (United States)

    Gerlai, R

    2016-01-01

    Learning and memory are defining features of our own species inherently important to our daily lives and to who we are. Without our memories we cease to exist as a person. Without our ability to learn individuals and collectively our society would cease to function. Diseases of the mind still remain incurable. The interest in understanding of the mechanisms of learning and memory is thus well founded. Given the complexity of such mechanisms, concerted efforts have been made to study them under controlled laboratory conditions, ie, with laboratory model organisms. The zebrafish, although new in this field, is one such model organism. The rapidly developing forward- and reverse genetic methods designed for the zebrafish and the increasing use of pharmacological tools along with numerous neurobiology techniques make this species perhaps the best model for the analysis of the mechanisms of complex central nervous system characteristics. The fact that it is an evolutionarily ancient and simpler vertebrate, but at the same time it possesses numerous conserved features across multiple levels of biological organization makes this species an excellent tool for the analysis of the mechanisms of learning and memory. The bottleneck lies in our understanding of its cognitive and mnemonic features, the topic of this chapter. The current paper builds on a chapter published in the previous edition and continues to focus on associative learning, but now it extends the discussion to other forms of learning and to recent discoveries on memory-related features and findings obtained both in adults and larval zebrafish. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. ESX-5-deficient Mycobacterium marinum is hypervirulent in adult zebrafish

    KAUST Repository

    Weerdenburg, Eveline M.

    2012-02-15

    ESX-5 is a mycobacterial type VII protein secretion system responsible for transport of numerous PE and PPE proteins. It is involved in the induction of host cell death and modulation of the cytokine response in vitro. In this work, we studied the effects of ESX-5 in embryonic and adult zebrafish using Mycobacterium marinum. We found that ESX-5-deficient M.marinum was slightly attenuated in zebrafish embryos. Surprisingly, the same mutant showed highly increased virulence in adult zebrafish, characterized by increased bacterial loads and early onset of granuloma formation with rapid development of necrotic centres. This early onset of granuloma formation was accompanied by an increased expression of pro-inflammatory cytokines and tissue remodelling genes in zebrafish infected with the ESX-5 mutant. Experiments using RAG-1-deficient zebrafish showed that the increased virulence of the ESX-5 mutant was not dependent on the adaptive immune system. Mixed infection experiments with wild-type and ESX-5 mutant bacteria showed that the latter had a specific advantage in adult zebrafish and outcompeted wild-type bacteria. Together our experiments indicate that ESX-5-mediated protein secretion is used by M.marinum to establish a moderate and persistent infection. © 2012 Blackwell Publishing Ltd.

  4. Morphofunctional transformations of the yolk syncytial layer during zebrafish development.

    Science.gov (United States)

    Kondakova, Ekaterina Alexandrovna A; Efremov, Vladimir Ivanovich I

    2014-02-01

    The yolk syncytial layer (YSL) is a provisory extraembryonic structure of teleost fishes and representatives of some other taxa with meroblastic cleavage. The YSL of teleosts is a symplast with polymorphous polyploid nuclei. It is known to perform nutritional, morphogenetic, immune, and, probably, other functions. Data about the YSL organization, functioning and regulation is fragmentary. Although gene expression patterns and other aspects of YSL functioning have been studied in Danio rerio, the morphology of its YSL has not been described in detail. The study of zebrafish YSL structure on sequential developmental stages is necessary to recognize specific features of this important polyfunctional system in this model organism and to extend our knowledge about provisory systems. The thickness of the YSL and the distribution of its nuclei are not uniform on each stage and change during development. During oblong and sphere stages the internal YSL (I-YSL) is filled with yolk inclusions; interphase yolk syncytial nuclei (YSN) and mitotic asters can be seen. During doming and epiboly the external YSL (E-YSL) is thicker than I-YSL. On the subsequent stages the YSL is thickened caudally. The dorsal YSL part is thickened during early segmentation stages and becomes the thinnest YSL region later. The anterior part of the YSL is thin, but enlarges during larval period. The YSN of different size and diverse forms, from regular to lobed, are present and form clusters. The number of irregular-shaped nuclei increases during development. The YSL thickens in the end of endotrophic and in the course of endo-exotrophic period, and its cytoplasm contains numerous yolk inclusions. After yolk exhaustion the YSL is flat. As the YSL degrades, the YSN become pycnotic, and the YSL remnant probably is cleared by phagocytes. Copyright © 2013 Wiley Periodicals, Inc.

  5. Developmental Deltamethrin Exposure Causes Persistent Changes in Dopaminergic Gene Expression, Neurochemistry, and Locomotor Activity in Zebrafish.

    Science.gov (United States)

    Kung, Tiffany S; Richardson, Jason R; Cooper, Keith R; White, Lori A

    2015-08-01

    Pyrethroids are commonly used insecticides that are considered to pose little risk to human health. However, there is an increasing concern that children are more susceptible to the adverse effects of pesticides. We used the zebrafish model to test the hypothesis that developmental exposure to low doses of the pyrethroid deltamethrin results in persistent alterations in dopaminergic gene expression, neurochemistry, and locomotor activity. Zebrafish embryos were treated with deltamethrin (0.25-0.50 μg/l), at concentrations below the LOAEL, during the embryonic period [3-72 h postfertilization (hpf)], after which transferred to fresh water until the larval stage (2-weeks postfertilization). Deltamethrin exposure resulted in decreased transcript levels of the D1 dopamine (DA) receptor (drd1) and increased levels of tyrosine hydroxylase at 72 hpf. The reduction in drd1 transcripts persisted to the larval stage and was associated with decreased D2 dopamine receptor transcripts. Larval fish, exposed developmentally to deltamethrin, had increased levels of homovanillic acid, a DA metabolite. Since the DA system is involved in locomotor activity, we measured the swim activity of larval fish following a transition to darkness. Developmental exposure to deltamethrin significantly increased larval swim activity which was attenuated by concomitant knockdown of the DA transporter. Acute exposure to methylphenidate, a DA transporter inhibitor, increased swim activity in control larva, while reducing swim activity in larva developmentally exposed to deltamethrin. Developmental exposure to deltamethrin causes locomotor deficits in larval zebrafish, which is likely mediated by dopaminergic dysfunction. This highlights the need to understand the persistent effects of low-dose neurotoxicant exposure during development. © The Author 2015. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  6. A zebrafish model of glucocorticoid resistance shows serotonergic modulation of the stress response

    Directory of Open Access Journals (Sweden)

    Brian eGriffiths

    2012-10-01

    Full Text Available One function of glucocorticoids is to restore homeostasis after an acute stress response by providing negative feedback to stress circuits in the brain. Loss of this negative feedback leads to elevated physiological stress and may contribute to depression, anxiety and post-traumatic stress disorder. We investigated the early, developmental effects of glucocorticoid signaling deficits on stress physiology and related behaviors using a mutant zebrafish, grs357, with non-functional glucocorticoid receptors. These mutants are morphologically inconspicuous and adult-viable. A previous study of adult grs357 mutants showed loss of glucocorticoid-mediated negative feedback and elevated physiological and behavioral stress markers. Already at five days post-fertilization, mutant larvae had elevated whole body cortisol, increased expression of pro-opiomelanocortin (POMC, the precursor of adrenocorticotropic hormone (ACTH, and failed to show normal suppression of stress markers after dexamethasone treatment. Mutant larvae had larger auditory-evoked startle responses compared to wildtype sibling controls (grwt, despite having lower spontaneous activity levels. Fluoxetine (Prozac treatment in mutants decreased startle responding and increased spontaneous activity, making them behaviorally similar to wildtype. This result mirrors known effects of selective serotonin reuptake inhibitors (SSRIs in modifying glucocorticoid signaling and alleviating stress disorders in human patients. Our results suggest that larval grs357 zebrafish can be used to study behavioral, physiological and molecular aspects of stress disorders. Most importantly, interactions between glucocorticoid and serotonin signaling appear to be highly conserved among vertebrates, suggesting deep homologies at the neural circuit level and opening up new avenues for research into psychiatric conditions.

  7. Pan-neuronal calcium imaging with cellular resolution in freely swimming zebrafish.

    Science.gov (United States)

    Kim, Dal Hyung; Kim, Jungsoo; Marques, João C; Grama, Abhinav; Hildebrand, David G C; Gu, Wenchao; Li, Jennifer M; Robson, Drew N

    2017-11-01

    Calcium imaging with cellular resolution typically requires an animal to be tethered under a microscope, which substantially restricts the range of behaviors that can be studied. To expand the behavioral repertoire amenable to imaging, we have developed a tracking microscope that enables whole-brain calcium imaging with cellular resolution in freely swimming larval zebrafish. This microscope uses infrared imaging to track a target animal in a behavior arena. On the basis of the predicted trajectory of the animal, we applied optimal control theory to a motorized stage system to cancel brain motion in three dimensions. We combined this motion-cancellation system with differential illumination focal filtering, a variant of HiLo microscopy, which enabled us to image the brain of a freely swimming larval zebrafish for more than an hour. This work expands the repertoire of natural behaviors that can be studied with cellular-resolution calcium imaging to potentially include spatial navigation, social behavior, feeding and reward.

  8. Heart-specific expression of laminopathic mutations in transgenic zebrafish.

    Science.gov (United States)

    Verma, Ajay D; Parnaik, Veena K

    2017-07-01

    Lamins are key determinants of nuclear organization and function in the metazoan nucleus. Mutations in human lamin A cause a spectrum of genetic diseases that affect cardiac muscle and skeletal muscle as well as other tissues. A few laminopathies have been modeled using the mouse. As zebrafish is a well established model for the study of cardiac development and disease, we have investigated the effects of heart-specific lamin A mutations in transgenic zebrafish. We have developed transgenic lines of zebrafish expressing conserved lamin A mutations that cause cardiac dysfunction in humans. Expression of zlamin A mutations Q291P and M368K in the heart was driven by the zebrafish cardiac troponin T2 promoter. Homozygous mutant embryos displayed nuclear abnormalities in cardiomyocyte nuclei. Expression analysis showed the upregulation of genes involved in heart regeneration in transgenic mutant embryos and a cell proliferation marker was increased in adult heart tissue. At the physiological level, there was deviation of up to 20% from normal heart rate in transgenic embryos expressing mutant lamins. Adult homozygous zebrafish were fertile and did not show signs of early mortality. Our results suggest that transgenic zebrafish models of heart-specific laminopathies show cardiac regeneration and moderate deviations in heart rate during embryonic development. © 2017 International Federation for Cell Biology.

  9. Differential modulation of expression of nuclear receptor mediated genes by tris(2-butoxyethyl) phosphate (TBOEP) on early life stages of zebrafish (Danio rerio)

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Zhiyuan, E-mail: zhiyuan_nju@163.com [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Yu, Yijun, E-mail: yjun.yu@gmail.com [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Tang, Song [School of Environment and Sustainability, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Liu, Hongling, E-mail: hlliu@nju.edu.cn [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Su, Guanyong; Xie, Yuwei [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Giesy, John P. [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Toxicology Centre, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Department of Veterinary Biomedical Sciences, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Department of Biology and Chemistry, City University of Hong Kong, Kowloon, Hong Kong Special Administrative Region (Hong Kong); Hecker, Markus [School of Environment and Sustainability, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Toxicology Centre, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Yu, Hongxia [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China)

    2015-12-15

    Highlights: • Effects of TBOEP on expression of genes of several nuclear hormone receptors and their relationship with adverse effect pathways in zebrafish. • TBOEP was neither an agonist nor antagonist of AR or AhR as determined by use of in vitro mammalian cell-based receptor transactivation assays. • Modulation of ER- and MR-dependent pathways allowed for development of feasible receptor-mediated, critical mechanisms of toxic action. - Abstract: As one substitute for phased-out brominated flame retardants (BFRs), tris(2-butoxyethyl) phosphate (TBOEP) is frequently detected in aquatic organisms. However, knowledge about endocrine disrupting mechanisms associated with nuclear receptors caused by TBOEP remained restricted to results from in vitro studies with mammalian cells. In the study, results of which are presented here, embryos/larvae of zebrafish (Danio rerio) were exposed to 0.02, 0.1 or 0.5 μM TBOEP to investigate expression of genes under control of several nuclear hormone receptors (estrogen receptors (ERs), androgen receptor (AR), thyroid hormone receptor alpha (TRα), mineralocorticoid receptor (MR), glucocorticoid receptor (GR), aryl hydrocarbon (AhR), peroxisome proliferator-activated receptor alpha (PPARα), and pregnane × receptor (P × R)) pathways at 120 hpf. Exposure to 0.5 μM TBOEP significantly (p < 0.05, one-way analysis of variance) up-regulated expression of estrogen receptors (ERs, er1, er2a, and er2b) genes and ER-associated genes (vtg4, vtg5, pgr, ncor, and ncoa3), indicating TBOEP modulates the ER pathway. In contrast, expression of most genes (mr, 11βhsd, ube2i,and adrb2b) associated with the mineralocorticoid receptor (MR) pathway were significantly down-regulated. Furthermore, in vitro mammalian cell-based (MDA-kb2 and H4IIE-luc) receptor transactivation assays, were also conducted to investigate possible agonistic or antagonistic effects on AR- and AhR-mediated pathways. In mammalian cells, none of these pathways were

  10. Reducing the Noise in Behavioral Assays: Sex and Age in Adult Zebrafish Locomotion

    OpenAIRE

    Philpott, Catelyn; Donack, Corey J.; Cousin, Margot A.; Pierret, Chris

    2012-01-01

    Many assays are used in animal model systems to measure specific human disease-related behaviors. The use of both adult and larval zebrafish as a behavioral model is gaining popularity. As this work progresses and potentially translates into new treatments, we must do our best to improve the sensitivity of these assays by reducing confounding factors. Scientists who use the mouse model system have demonstrated that sex and age can influence a number of behaviors. As a community, they have mov...

  11. CERKL knockdown causes retinal degeneration in zebrafish.

    Directory of Open Access Journals (Sweden)

    Marina Riera

    Full Text Available The human CERKL gene is responsible for common and severe forms of retinal dystrophies. Despite intense in vitro studies at the molecular and cellular level and in vivo analyses of the retina of murine knockout models, CERKL function remains unknown. In this study, we aimed to approach the developmental and functional features of cerkl in Danio rerio within an Evo-Devo framework. We show that gene expression increases from early developmental stages until the formation of the retina in the optic cup. Unlike the high mRNA-CERKL isoform multiplicity shown in mammals, the moderate transcriptional complexity in fish facilitates phenotypic studies derived from gene silencing. Moreover, of relevance to pathogenicity, teleost CERKL shares the two main human protein isoforms. Morpholino injection has been used to generate a cerkl knockdown zebrafish model. The morphant phenotype results in abnormal eye development with lamination defects, failure to develop photoreceptor outer segments, increased apoptosis of retinal cells and small eyes. Our data support that zebrafish Cerkl does not interfere with proliferation and neural differentiation during early developmental stages but is relevant for survival and protection of the retinal tissue. Overall, we propose that this zebrafish model is a powerful tool to unveil CERKL contribution to human retinal degeneration.

  12. Using fluorescent lipids in live zebrafish larvae: From imaging whole animal physiology to subcellular lipid trafficking.

    Science.gov (United States)

    Anderson, J L; Carten, J D; Farber, S A

    2016-01-01

    Lipids serve essential functions in cells as signaling molecules, membrane components, and sources of energy. Defects in lipid metabolism are implicated in a number of pandemic human diseases, including diabetes, obesity, and hypercholesterolemia. Many aspects of how fatty acids and cholesterol are absorbed and processed by intestinal cells remain unclear and present a hurdle to developing approaches for disease prevention and treatment. Numerous studies have shown that the zebrafish is an excellent model for vertebrate lipid metabolism. In this chapter, we review commercially available fluorescent lipids that can be deployed in live zebrafish to better understand lipid signaling and metabolism. In this chapter, we present criteria one should consider when selecting specific fluorescent lipids for the study of digestive physiology or lipid metabolism in larval zebrafish. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Expression of miRNA-122 Induced by Liver Toxicants in Zebrafish

    Directory of Open Access Journals (Sweden)

    Hyun-Sik Nam

    2016-01-01

    Full Text Available MicroRNA-122 (miRNA-122, also known as liver-specific miRNA, has recently been shown to be a potent biomarker in response to liver injury in mammals. The objective of this study was to examine its expression in response to toxicant treatment and acute liver damage, using the zebrafish system as an alternative model organism. For the hepatotoxicity assay, larval zebrafish were arrayed in 24-well plates. Adult zebrafish were also tested and arrayed in 200 mL cages. Animals were exposed to liver toxicants (tamoxifen or acetaminophen at various doses, and miRNA-122 expression levels were analyzed using qRT-PCR in dissected liver, brain, heart, and intestine, separately. Our results showed no significant changes in miRNA-122 expression level in tamoxifen-treated larvae; however, miRNA-122 expression was highly induced in tamoxifen-treated adults in a tissue-specific manner. In addition, we observed a histological change in adult liver (0.5 μM and cell death in larval liver (5 μM at different doses of tamoxifen. These results indicated that miRNA-122 may be utilized as a liver-specific biomarker for acute liver toxicity in zebrafish.

  14. Characterization of zebrafish larvae suction feeding flow using μPIV and optical coherence tomography

    Science.gov (United States)

    Pekkan, Kerem; Chang, Brian; Uslu, Fazil; Mani, Karthick; Chen, Chia-Yuan; Holzman, Roi

    2016-07-01

    The hydrodynamics of suction feeding is critical for the survival of fish larvae; failure to capture food during the onset of autonomous feeding can rapidly lead to starvation and mortality. Fluid mechanics experiments that investigate the suction feeding of suspended particles are limited to adult fishes, which operate at large Reynolds numbers. This manuscript presents the first literature results in which the external velocity fields generated during suction feeding of early zebrafish larvae (2500-20,000 μm total length) are reported using time-resolved microscopic particle image velocimetry. For the larval stages studied, the maximum peak suction velocity of the inflow bolus is measured at a finite distance from the mouth tip and ranges from 1 to 8 mm/s. The average pressure gradient and the velocity profile proximal to the buccal (mouth) cavity are calculated, and two distinct trends are identified. External recirculation regions and reverse flow feeding cycles are also observed and quantified. One of the unresolved questions in fish suction feeding is the shape and dynamics of the buccal cavity during suction feeding; optical coherence tomography imaging is found to be useful for reconstructing the mouth kinematics. The projected area of the mouth cavity during the feeding cycle varies up to 160 and 22 % for the transverse and mid-sagittal planes, respectively. These findings can inspire novel hydrodynamically efficient biomedical and microfluidic devices.

  15. Effects of probiotic administration on zebrafish development and reproduction.

    Science.gov (United States)

    Carnevali, O; Avella, M A; Gioacchini, G

    2013-07-01

    As the consumption of probiotics increases worldwide, scientists focus on identifying bacterial strains able to improve human life quality and evidence the biological pathways affected by probiotic treatment. In this review, some recent observations on the effects of changes of microbiota on zebrafish metabolism were discussed. In addition, the effects of Lactobacillus rhamnosus - a component of the human gut microflora - as a diet supplement on Danio rerio were presented. When administered chronically, L. rhamnosus may affect larval development and the physiology of reproductive system in the zebrafish model. It was hypothesized exogenous L. rhamnosus accelerates larval growth and backbone development by acting on insulin-like growth factors-I (igfI) and -II (igfII), peroxisome proliferator activated receptors-α and -β, (pparα,β) vitamin D receptor-α (vdrα) and retinoic acid receptor-γ (rarγ). Gonadal differentiation was anticipated at 6weeks together with a higher expression of gnrh3 at the larval stage when L. rhamnosus was administered throughout development. Moreover, brood stock alimented with a L. rhamnosus-supplemented diet showed better reproductive performances as per follicles development, ovulated oocytes quantification and embryos quality. A plausible involvement of factors such as leptin, and kiss1 and 2 in the improvements was concluded. The observations made on the physiology of female reproduction were correlated with the gene expression of a gigantic number of factors as the aromatase cytochrome p 19 (cyp19a), the vitellogenin (vtg) and the α isoform of the E2 receptor (erα), luteinizing hormone receptor (lhr), 20-β hydroxysteroid dehydrogenase (20β-hsd), membrane progesterone receptors α and β, cyclin B, activinβA1, smad2, transforming growth factor β1 (tgfβ1), growth differentiation factor9 (gdf9) and bone morphogenetic protein15 (bmp15.) A model in which the exogenous L. rhamnosus in the digestive tract of zebrafish from the

  16. Analyzing the structure and function of neuronal circuits in zebrafish

    Directory of Open Access Journals (Sweden)

    Rainer eFriedrich

    2013-04-01

    Full Text Available The clever choice of animal models has been instrumental for many breakthrough discoveries in life sciences. One of the outstanding challenges in neuroscience is the in-depth analysis of neuronal circuits to understand how interactions between large numbers of neurons give rise to the computational power of the brain. A promising model organism to address this challenge is the zebrafish, not only because it is cheap, transparent and accessible to sophisticated genetic manipulations but also because it offers unique advantages for quantitative analyses of circuit structure and function. One of the most important advantages of zebrafish is its small brain size, both at larval and adult stages. Small brains enable exhaustive measurements of neuronal activity patterns by optical imaging and facilitate large-scale reconstructions of wiring diagrams by electron microscopic approaches. Such information is important, and probably essential, to obtain mechanistic insights into neuronal computations underlying higher brain functions and dysfunctions. This review provides a brief overview over current methods and motivations for dense reconstructions of neuronal activity and connectivity patterns. It then discusses selective advantages of zebrafish and provides examples how these advantages are exploited to study neuronal computations in the olfactory bulb.

  17. Developmental Toxicity of Dextromethorphan in Zebrafish Embryos/Larvae

    Science.gov (United States)

    Xu, Zheng; Williams, Frederick E.; Liu, Ming-Cheh

    2012-01-01

    Dextromethorphan is widely used in over-the-counter cough and cold medications. Its efficacy and safety for infants and young children remains to be clarified. The present study was designed to use the zebrafish as a model to investigate the potential toxicity of dextromethorphan during the embryonic and larval development. Three sets of zebrafish embryos/larvae were exposed to dextromethorphan at 24 hours post fertilization (hpf), 48 hpf, and 72 hpf, respectively, during the embryonic/larval development. Compared with the 48 and 72 hpf exposure sets, the embryos/larvae in the 24 hpf exposure set showed much higher mortality rates which increased in a dose-dependent manner. Bradycardia and reduced blood flow were observed for the embryos/larvae treated with increasing concentrations of dextromethorphan. Morphological effects of dextromethorphan exposure, including yolk sac and cardiac edema, craniofacial malformation, lordosis, non-inflated swim bladder, and missing gill, were also more frequent and severe among zebrafish embryos/larvae exposed to dextromethorphan at 24 hpf. Whether the more frequent and severe developmental toxicity of dextromethorphan observed among the embryos/larvae in the 24 hpf exposure set, as compared with the 48 and 72 hpf exposure sets, is due to the developmental expression of the Phase I and Phase II enzymes involved in the metabolism of dextromethorphan remains to be clarified. A reverse transcription-polymerase chain reaction (RT-PCR) analysis, nevertheless, revealed developmental stage-dependent expression of mRNAs encoding SULT3 ST1 and SULT3 ST3, two enzymes previously shown to be capable of sulfating dextrorphan, an active metabolite of dextromethorphan. PMID:20737414

  18. Feature Binding in Zebrafish

    Directory of Open Access Journals (Sweden)

    P Neri

    2012-07-01

    Full Text Available Binding operations are primarily ascribed to cortex or similarly complex avian structures. My experiments show that the zebrafish, a lower vertebrate lacking cortex, supports visual feature binding of form and motion for the purpose of social behavior. These results challenge the notion that feature binding may require highly evolved neural structures and demonstrate that the nervous system of lower vertebrates can afford unexpectedly complex computations.

  19. Expression of prostaglandin synthases (pgds and pges) during zebrafish gonadal differentiation

    DEFF Research Database (Denmark)

    Jørgensen, Anne; Nielsen, John E; Nielsen, Betina Frydenlund

    2010-01-01

    The present study aimed at elucidating whether the expression pattern of the membrane bound form of prostaglandin E2 synthase (pges) and especially the lipocalin-type prostaglandin D2 synthase (pgds) indicates involvement in gonadal sex differentiation in zebrafish as has previously been found....... In this study, a sexually dimorphic expression of pgds was found in gonads of adult zebrafish with expression in testis but not in ovaries. To determine whether the sex-specific expression pattern of pgds was present in gonads of juvenile zebrafish and therefore could be an early marker of sex in zebrafish, we...... microdissected gonads from four randomly selected individual zebrafish for every second day in the period 2-20 days post hatch (dph) and 0-1 dph. The temporal expression of pgds and pges was investigated in the microdissected gonads, however, no differential expression that could indicate sex-specific difference...

  20. Capturing tissue repair in zebrafish larvae with time-lapse brightfield stereomicroscopy.

    Science.gov (United States)

    Lisse, Thomas S; Brochu, Elizabeth A; Rieger, Sandra

    2015-01-31

    The zebrafish larval tail fin is ideal for studying tissue regeneration due to the simple architecture of the larval fin-fold, which comprises of two layers of skin that enclose undifferentiated mesenchyme, and because the larval tail fin regenerates rapidly within 2-3 days. Using this system, we demonstrate a method for capturing the repair dynamics of the amputated tail fin with time-lapse video brightfield stereomicroscopy. We demonstrate that fin amputation triggers a contraction of the amputation wound and extrusion of cells around the wound margin, leading to their subsequent clearance. Fin regeneration proceeds from proximal to distal direction after a short delay. In addition, developmental growth of the larva can be observed during all stages. The presented method provides an opportunity for observing and analyzing whole tissue-scale behaviors such as fin development and growth in a simple microscope setting, which is easily adaptable to any stereomicroscope with time-lapse capabilities.

  1. Characterization of Zebrafish von Willebrand Factor Reveals Conservation of Domain Structure, Multimerization, and Intracellular Storage

    Directory of Open Access Journals (Sweden)

    Arunima Ghosh

    2012-01-01

    Full Text Available von Willebrand disease (VWD is the most common inherited human bleeding disorder and is caused by quantitative or qualitative defects in von Willebrand factor (VWF. VWF is a secreted glycoprotein that circulates as large multimers. While reduced VWF is associated with bleeding, elevations in overall level or multimer size are implicated in thrombosis. The zebrafish is a powerful genetic model in which the hemostatic system is well conserved with mammals. The ability of this organism to generate thousands of offspring and its optical transparency make it unique and complementary to mammalian models of hemostasis. Previously, partial clones of zebrafish vwf have been identified, and some functional conservation has been demonstrated. In this paper we clone the complete zebrafish vwf cDNA and show that there is conservation of domain structure. Recombinant zebrafish Vwf forms large multimers and pseudo-Weibel-Palade bodies (WPBs in cell culture. Larval expression is in the pharyngeal arches, yolk sac, and intestinal epithelium. These results provide a foundation for continued study of zebrafish Vwf that may further our understanding of the mechanisms of VWD.

  2. Paclitaxel-induced epithelial damage and ectopic MMP-13 expression promotes neurotoxicity in zebrafish.

    Science.gov (United States)

    Lisse, Thomas S; Middleton, Leah J; Pellegrini, Adriana D; Martin, Paige B; Spaulding, Emily L; Lopes, Olivia; Brochu, Elizabeth A; Carter, Erin V; Waldron, Ashley; Rieger, Sandra

    2016-04-12

    Paclitaxel is a microtubule-stabilizing chemotherapeutic agent that is widely used in cancer treatment and in a number of curative and palliative regimens. Despite its beneficial effects on cancer, paclitaxel also damages healthy tissues, most prominently the peripheral sensory nervous system. The mechanisms leading to paclitaxel-induced peripheral neuropathy remain elusive, and therapies that prevent or alleviate this condition are not available. We established a zebrafish in vivo model to study the underlying mechanisms and to identify pharmacological agents that may be developed into therapeutics. Both adult and larval zebrafish displayed signs of paclitaxel neurotoxicity, including sensory axon degeneration and the loss of touch response in the distal caudal fin. Intriguingly, studies in zebrafish larvae showed that paclitaxel rapidly promotes epithelial damage and decreased mechanical stress resistance of the skin before induction of axon degeneration. Moreover, injured paclitaxel-treated zebrafish skin and scratch-wounded human keratinocytes (HEK001) display reduced healing capacity. Epithelial damage correlated with rapid accumulation of fluorescein-conjugated paclitaxel in epidermal basal keratinocytes, but not axons, and up-regulation of matrix-metalloproteinase 13 (MMP-13, collagenase 3) in the skin. Pharmacological inhibition of MMP-13, in contrast, largely rescued paclitaxel-induced epithelial damage and neurotoxicity, whereas MMP-13 overexpression in zebrafish embryos rendered the skin vulnerable to injury under mechanical stress conditions. Thus, our studies provide evidence that the epidermis plays a critical role in this condition, and we provide a previously unidentified candidate for therapeutic interventions.

  3. Fog1 is required for cardiac looping in zebrafish

    OpenAIRE

    Walton, R. Zaak; Bruce, Ashley E.E.; Olivey, Harold E.; Najib, Khalid; Johnson, Vanitha; Earley, Judy U.; Ho, Robert K.; Svensson, Eric C.

    2006-01-01

    To further our understanding of FOG gene function during cardiac development, we utilized zebrafish to examine FOG’s role in the early steps of heart morphogenesis. We identified fragments of three fog genes in the zebrafish genomic database and isolated full-length coding sequences for each of these genes by using a combination of RT-PCR and 5′-RACE. One gene was similar to murine FOG-1 (fog1), while the remaining two were similar to murine FOG-2 (fog2a and fog2b). All Fog proteins were able...

  4. Biotransformation of ginsenosides F4 and Rg6 in zebrafish.

    Science.gov (United States)

    Shen, Wen-Wen; Zhang, Hai-Xia; Qiu, Shou-Bei; Wei, Ying-Jie; Zhu, Fen-Xia; Wang, Jing; Wang, Dan-Dan; Jia, Xiao-Bin; Tang, Dao-Quan; Chen, Bin

    2017-03-28

    Ginsenosides F 4 and Rg 6 (GF 4 and GRg 6 ), two main active components of steamed notoginseng or red ginseng, are dehydrated disaccharide saponins. In this work, biotransformation of ginsenosides F 4 and Rg 6 in zebrafish was investigated by qualitatively identifying their metabolites and then proposing their possible metabolic pathways. The prediction of possible metabolism of ginsenosides F 4 and Rg 6 using zebrafish model which can effectively simulate existing mammals model was early and quickly performed. Metabolites of ginsenosides F 4 and Rg 6 after exposing to zebrafish for 24 h were identified by Ultraperformance Liquid Chromatography/Quadrupole-Time-of-Flight Mass Spectrometry. A total of 8 and 6 metabolites of ginsenosides F 4 and Rg 6 were identified in zebrafish, respectively. Of these, 7 and 5, including M1, M3-M5, M7-M9 and N1 (N5), N2, N4 (N9), N7-N8 were reported for the first time as far as we know. The mechanisms of their biotransformation involved were further deduced to be desugarization, glucuronidation, sulfation, dehydroxylation, loss of C-17 and/or C-23 residue pathways. It was concluded that loss of rhamnose at position C-6 and glucuronidation at position C-3 in zebrafish were considered as the main physiologic and metabolic processes of ginsenosides F 4 and ginsenosides Rg 6 , respectively.

  5. Influence of carbon nanotube length on toxicity to zebrafish embryos

    Directory of Open Access Journals (Sweden)

    Cheng J

    2012-07-01

    Full Text Available Jinping Cheng,1,2 Shuk Han Cheng11Department of Biology and Chemistry, City University of Hong Kong, Hong Kong; 2State Key Laboratory of Estuarine and Coastal Research, East China Normal University, Shanghai, ChinaAbstract: There is currently a large difference of opinion in nanotoxicology studies of nanomaterials. There is concern about why some studies have indicated that there is strong toxicity, while others have not. In this study, the length of carbon nanotubes greatly affected their toxicity in zebrafish embryos. Multiwalled carbon nanotubes (MWCNTs were sonicated in a nitric acid solution for 24 hours and 48 hours. The modified MWCNTs were tested in early developing zebrafish embryo. MWCNTs prepared with the longer sonication time resulted in severe developmental toxicity; however, the shorter sonication time did not induce any obvious toxicity in the tested developing zebrafish embryos. The cellular and molecular changes of the affected zebrafish embryos were studied and the observed phenotypes scored. This study suggests that length plays an important role in the in vivo toxicity of functionalized CNTs. This study will help in furthering the understanding on current differences in toxicity studies of nanomaterials.Keywords: length, carbon nanotubes, sonication, developmental toxicity, zebrafish

  6. The transcriptomics of glucocorticoid receptor signaling in developing zebrafish.

    Directory of Open Access Journals (Sweden)

    Dinushan Nesan

    Full Text Available Cortisol is the primary corticosteroid in teleosts that is released in response to stressor activation of the hypothalamus-pituitary-interrenal axis. The target tissue action of this hormone is primarily mediated by the intracellular glucocorticoid receptor (GR, a ligand-bound transcription factor. In developing zebrafish (Danio rerio embryos, GR transcripts and cortisol are maternally deposited into the oocyte prior to fertilization and influence early embryogenesis. To better understand of the molecular mechanisms involved, we investigated changes in the developmental transcriptome prior to hatch, in response to morpholino oligonucleotide knockdown of GR using the Agilent zebrafish microarray platform. A total of 1313 and 836 mRNA transcripts were significantly changed at 24 and 36 hours post fertilization (hpf, respectively. Functional analysis revealed numerous developmental processes under GR regulation, including neurogenesis, eye development, skeletal and cardiac muscle formation. Together, this study underscores a critical role for glucocorticoid signaling in programming molecular events essential for zebrafish development.

  7. Characterisation of Culex quinquefasciatus (Diptera: Culicidae larval habitats at ground level and temporal fluctuations of larval abundance in Córdoba, Argentina

    Directory of Open Access Journals (Sweden)

    Marta Grech

    2013-09-01

    Full Text Available The aims of this study were to characterise the ground-level larval habitats of the mosquito Culex quinquefasciatus, to determine the relationships between habitat characteristics and larval abundance and to examine seasonal larval-stage variations in Córdoba city. Every two weeks for two years, 15 larval habitats (natural and artificial water bodies, including shallow wells, drains, retention ponds, canals and ditches were visited and sampled for larval mosquitoes. Data regarding the water depth, temperature and pH, permanence, the presence of aquatic vegetation and the density of collected mosquito larvae were recorded. Data on the average air temperatures and accumulated precipitation during the 15 days prior to each sampling date were also obtained. Cx. quinquefasciatus larvae were collected throughout the study period and were generally most abundant in the summer season. Generalised linear mixed models indicated the average air temperature and presence of dicotyledonous aquatic vegetation as variables that served as important predictors of larval densities. Additionally, permanent breeding sites supported high larval densities. In Córdoba city and possibly in other highly populated cities at the same latitude with the same environmental conditions, control programs should focus on permanent larval habitats with aquatic vegetation during the early spring, when the Cx. quinquefasciatus population begins to increase.

  8. Mitochondrial Dysfunction, Disruption of F-Actin Polymerization, and Transcriptomic Alterations in Zebrafish Larvae Exposed to Trichloroethylene.

    Science.gov (United States)

    Wirbisky, Sara E; Damayanti, Nur P; Mahapatra, Cecon T; Sepúlveda, Maria S; Irudayaraj, Joseph; Freeman, Jennifer L

    2016-02-15

    Trichloroethylene (TCE) is primarily used as an industrial degreasing agent and has been in use since the 1940s. TCE is released into the soil, surface, and groundwater. From an environmental and regulatory standpoint, more than half of Superfund hazardous waste sites on the National Priority List are contaminated with TCE. Occupational exposure to TCE occurs primarily via inhalation, while environmental TCE exposure also occurs through ingestion of contaminated drinking water. Current literature links TCE exposure to various adverse health effects including cardiovascular toxicity. Current studies aiming to address developmental cardiovascular toxicity utilized rodent and avian models, with the majority of studies using relatively higher parts per million (mg/L) doses. In this study, to further investigate developmental cardiotoxicity of TCE, zebrafish embryos were treated with 0, 10, 100, or 500 parts per billion (ppb; μg/L) TCE during embryogenesis and/or through early larval stages. After the appropriate exposure period, angiogenesis, F-actin, and mitochondrial function were assessed. A significant dose-response decrease in angiogenesis, F-actin, and mitochondrial function was observed. To further complement this data, a transcriptomic profile of zebrafish larvae was completed to identify gene alterations associated with the 10 ppb TCE exposure. Results from the transcriptomic data revealed that embryonic TCE exposure caused significant changes in genes associated with cardiovascular disease, cancer, and organismal injury and abnormalities with a number of targets in the FAK signaling pathway. Overall, results from our study support TCE as a developmental cardiovascular toxicant, provide molecular targets and pathways for investigation in future studies, and indicate a need for continued priority for environmental regulation.

  9. Paraquat affects mitochondrial bioenergetics, dopamine system expression, and locomotor activity in zebrafish (Danio rerio).

    Science.gov (United States)

    Wang, Xiao H; Souders, Christopher L; Zhao, Yuan H; Martyniuk, Christopher J

    2018-01-01

    The dipyridyl herbicide paraquat induces oxidative stress in cells and is implicated in adult neurodegenerative diseases. However, less is known about paraquat toxicity in early stages of vertebrate development. To address this gap, zebrafish (Danio rerio) embryos were exposed to 1, 10 and 100 μM paraquat for 96 h. Paraquat did not induce significant mortality nor deformity in embryos and larvae, but it did accelerate time to hatch. To evaluate whether mitochondrial respiration was related to earlier hatch times, oxygen consumption rate was measured in whole embryos. Maximal respiration of embryos exposed to 100 μM paraquat for 24 h was reduced by more than 70%, suggesting that paraquat negatively impacts mitochondrial bioenergetics in early development. Based upon this evidence for mitochondrial dysfunction, transcriptional responses of oxidative stress- and apoptosis-related genes were measured. Fish exposed to 1 μM paraquat showed higher expression levels of superoxide dismutase 2, heat shock protein 70, Bcl-2-associated X protein, and B-cell CLL/lymphoma 2a compared to control fish. No differences among groups were detected in larvae exposed to 10 and 100 μM paraquat, suggesting a non-monotonic response. We also measured endpoints related to larval behavior and dopaminergic signaling as paraquat is associated with degeneration of dopamine neurons. Locomotor activity was stimulated with 100 μM paraquat and dopamine transporter and dopamine receptor 3 mRNA levels were increased in larvae exposed to 1 μM paraquat, interpreted to be a compensatory response at lower concentrations. This study improves mechanistic understanding into the toxic actions of paraquat on early developmental stages. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Disruption of the folate pathway in zebrafish causes developmental defects

    Directory of Open Access Journals (Sweden)

    Lee Marina S

    2012-04-01

    Full Text Available Abstract Background Folic acid supplementation reduces the risk of neural tube defects and congenital heart defects. The biological mechanisms through which folate prevents birth defects are not well understood. We explore the use of zebrafish as a model system to investigate the role of folate metabolism during development. Results We first identified zebrafish orthologs of 12 human folate metabolic genes. RT-PCR and in situ analysis indicated maternal transcripts supply the embryo with mRNA so that the embryo has an intact folate pathway. To perturb folate metabolism we exposed zebrafish embryos to methotrexate (MTX, a potent inhibitor of dihydrofolate reductase (Dhfr an essential enzyme in the folate metabolic pathway. Embryos exposed to high doses of MTX exhibited developmental arrest prior to early segmentation. Lower doses of MTX resulted in embryos with a shortened anterior-posterior axis and cardiac defects: linear heart tubes or incomplete cardiac looping. Inhibition of dhfr mRNA with antisense morpholino oligonucleotides resulted in embryonic lethality. One function of the folate pathway is to provide essential one-carbon units for dTMP synthesis, a rate-limiting step of DNA synthesis. After 24 hours of exposure to high levels of MTX, mutant embryos continue to incorporate the thymidine analog BrdU. However, additional experiments indicate that these embryos have fewer mitotic cells, as assayed with phospho-histone H3 antibodies, and that treated embryos have perturbed cell cycles. Conclusions Our studies demonstrate that human and zebrafish utilize similar one-carbon pathways. Our data indicate that folate metabolism is essential for early zebrafish development. Zebrafish studies of the folate pathway and its deficiencies could provide insight into the underlying etiology of human birth defects and the natural role of folate in development.

  11. Kisspeptins modulate the biology of multiple populations of gonadotropin-releasing hormone neurons during embryogenesis and adulthood in zebrafish (Danio rerio).

    Science.gov (United States)

    Zhao, Yali; Lin, Meng-Chin A; Mock, Allan; Yang, Ming; Wayne, Nancy L

    2014-01-01

    Kisspeptin1 (product of the Kiss1 gene) is the key neuropeptide that gates puberty and maintains fertility by regulating the gonadotropin-releasing hormone (GnRH) neuronal system in mammals. Inactivating mutations in Kiss1 and the kisspeptin receptor (GPR54/Kiss1r) are associated with pubertal failure and infertility. Kiss2, a paralogous gene for kiss1, has been recently identified in several vertebrates including zebrafish. Using our transgenic zebrafish model system in which the GnRH3 promoter drives expression of emerald green fluorescent protein, we investigated the effects of kisspeptins on development of the GnRH neuronal system during embryogenesis and on electrical activity during adulthood. Quantitative PCR showed detectable levels of kiss1 and kiss2 mRNA by 1 day post fertilization, increasing throughout embryonic and larval development. Early treatment with Kiss1 or Kiss2 showed that both kisspeptins stimulated proliferation of trigeminal GnRH3 neurons located in the peripheral nervous system. However, only Kiss1, but not Kiss2, stimulated proliferation of terminal nerve and hypothalamic populations of GnRH3 neurons in the central nervous system. Immunohistochemical analysis of synaptic vesicle protein 2 suggested that Kiss1, but not Kiss2, increased synaptic contacts on the cell body and along the terminal nerve-GnRH3 neuronal processes during embryogenesis. In intact brain of adult zebrafish, whole-cell patch clamp recordings of GnRH3 neurons from the preoptic area and hypothalamus revealed opposite effects of Kiss1 and Kiss2 on spontaneous action potential firing frequency and membrane potential. Kiss1 increased spike frequency and depolarized membrane potential, whereas Kiss2 suppressed spike frequency and hyperpolarized membrane potential. We conclude that in zebrafish, Kiss1 is the primary stimulator of GnRH3 neuronal development in the embryo and an activator of stimulating hypophysiotropic neuron activities in the adult, while Kiss2 plays an

  12. Kisspeptins modulate the biology of multiple populations of gonadotropin-releasing hormone neurons during embryogenesis and adulthood in zebrafish (Danio rerio.

    Directory of Open Access Journals (Sweden)

    Yali Zhao

    Full Text Available Kisspeptin1 (product of the Kiss1 gene is the key neuropeptide that gates puberty and maintains fertility by regulating the gonadotropin-releasing hormone (GnRH neuronal system in mammals. Inactivating mutations in Kiss1 and the kisspeptin receptor (GPR54/Kiss1r are associated with pubertal failure and infertility. Kiss2, a paralogous gene for kiss1, has been recently identified in several vertebrates including zebrafish. Using our transgenic zebrafish model system in which the GnRH3 promoter drives expression of emerald green fluorescent protein, we investigated the effects of kisspeptins on development of the GnRH neuronal system during embryogenesis and on electrical activity during adulthood. Quantitative PCR showed detectable levels of kiss1 and kiss2 mRNA by 1 day post fertilization, increasing throughout embryonic and larval development. Early treatment with Kiss1 or Kiss2 showed that both kisspeptins stimulated proliferation of trigeminal GnRH3 neurons located in the peripheral nervous system. However, only Kiss1, but not Kiss2, stimulated proliferation of terminal nerve and hypothalamic populations of GnRH3 neurons in the central nervous system. Immunohistochemical analysis of synaptic vesicle protein 2 suggested that Kiss1, but not Kiss2, increased synaptic contacts on the cell body and along the terminal nerve-GnRH3 neuronal processes during embryogenesis. In intact brain of adult zebrafish, whole-cell patch clamp recordings of GnRH3 neurons from the preoptic area and hypothalamus revealed opposite effects of Kiss1 and Kiss2 on spontaneous action potential firing frequency and membrane potential. Kiss1 increased spike frequency and depolarized membrane potential, whereas Kiss2 suppressed spike frequency and hyperpolarized membrane potential. We conclude that in zebrafish, Kiss1 is the primary stimulator of GnRH3 neuronal development in the embryo and an activator of stimulating hypophysiotropic neuron activities in the adult, while

  13. Effects of maternal exposure to estrogen and PCB on different life stages of Zebrafish (Danio rerio)

    Energy Technology Data Exchange (ETDEWEB)

    Olsson, Per-Erik; Westerlund, L; Billsson, K; Berg, A H [Umeaa Univ. (Sweden). Dept. of Cellular and Developmental Biology; Teh, S J; Hinton, D E [California Univ., Davis, CA (United States). Dept. of Anatomy, Physiology and Cell Biology; Tysklind, M [Umeaa Univ., (Sweden). Dept. of Environmental Chemistry; Nilsson, Jan; Eriksson, Lars-Ove [Swedish Univ. of Agricultural Sciences, Umeaa (Sweden). Dept. of Aquaculture

    1999-02-01

    PCBs have been found to impair both reproduction and development in fish. We have investigated the effects of 3 PCB congeners, 2,3,3`,4,4`,5,6-HpCB (PCB-190); 2,3,4,4`-TeCB (PCB-60); and 2,2`,4,6,6`-PeCB (PCB-104), and the estrogenic hormone 17{beta}-estradiol on fecundity, early life-stage mortality, gross morphology and histology of zebrafish (Danio rerio). While none of the studied substances reduced fecundity, they increased embryo and larval mortality. The most severe effects on viability were observed following treatment with 17{beta}-estradiol or the weakly estrogenic PCB-104. Following 17{beta}-estradiol or PCB-104 exposure, mortality continued through the yolksac absorption phase. PCB-60, on the other hand, resulted in mortality between the 30% epiboly stage and 75% epiboly stage. At the same time as embryos started to die, embryo development and hatching were delayed. PCB-190 showed only moderate effects on early-life stage mortality. The fish were reared until sexual maturation where after they were subjected to gross morphological and histological analyses. Changes in morphology were observed following PCB-104 and PCB-190 treatment. Both substances gave rise to craniofacial malformations while PCB-104 also led to lordosis in females and scoliosis in fish of both sexes. From histological analysis it was found that PCB-104 and 17{beta}-estradiol resulted in karyorrhexis and karyolysis in the kidney. Possible signs of bile stasis were observed following 17{beta}-estradiol and PCB-190 treatment. Some effects were observed on the gonads, including areas in the ovary showing atresia and limited failure of testicular spermatogenesis in 17{beta}-estradiol, PCB-104, and PCB-60 treated fish. While all studied substances resulted in effects on offspring, the observation that estrogenic substances are highly embryotoxic, raises concern that endocrine disrupting substances may severely reduce fish populations in polluted areas

  14. Multiple zebrafish atoh1 genes specify a diversity of neuronal types in the zebrafish cerebellum.

    Science.gov (United States)

    Kidwell, Chelsea U; Su, Chen-Ying; Hibi, Masahiko; Moens, Cecilia B

    2018-06-01

    A single Atoh1 basic-helix-loop-helix transcription factor specifies multiple neuron types in the mammalian cerebellum and anterior hindbrain. The zebrafish genome encodes three paralagous atoh1 genes whose functions in cerebellum and anterior hindbrain development we explore here. With use of a transgenic reporter, we report that zebrafish atoh1c-expressing cells are organized in two distinct domains that are separated both by space and developmental time. An early isthmic expression domain gives rise to an extracerebellar population in rhombomere 1 and an upper rhombic lip domain gives rise to granule cell progenitors that migrate to populate all four granule cell territories of the fish cerebellum. Using genetic mutants we find that of the three zebrafish atoh1 paralogs, atoh1c and atoh1a are required for the full complement of granule neurons. Surprisingly, the two genes are expressed in non-overlapping granule cell progenitor populations, indicating that fish use duplicate atoh1 genes to generate granule cell diversity that is not detected in mammals. Finally, live imaging of granule cell migration in wildtype and atoh1c mutant embryos reveals that while atoh1c is not required for granule cell specification per se, it is required for granule cells to delaminate and migrate away from the rhombic lip. Copyright © 2018 Elsevier Inc. All rights reserved.

  15. Larval helminths in intermediate hosts

    DEFF Research Database (Denmark)

    Fredensborg, Brian Lund; Poulin, R

    2005-01-01

    Density-dependent effects on parasite fitness have been documented from adult helminths in their definitive hosts. There have, however, been no studies on the cost of sharing an intermediate host with other parasites in terms of reduced adult parasite fecundity. Even if larval parasites suffer a ...

  16. Larval outbreaks in West Greenland

    DEFF Research Database (Denmark)

    Lund, Magnus; Raundrup, Katrine; Westergaard-Nielsen, Andreas

    2017-01-01

    effects of a larval outbreak in 2011 on vegetation productivity and CO2 exchange. We estimate a decreased carbon (C) sink strength in the order of 118–143 g C m−2, corresponding to 1210–1470 tonnes C at the Kobbefjord catchment scale. The decreased C sink was, however, counteracted the following years...

  17. Kauri seeds and larval somersaults

    DEFF Research Database (Denmark)

    Dupont, Steen Thorleif

    2012-01-01

    The trunk morphology of the larvae of the kauri pine (Agathis) seed infesting moth Agathiphaga is described using conventional, polarization, and scanning electron microscopy. The pine seed chamber formed by the larva is also described and commented on. The simple larval chaetotaxy includes more ...

  18. Modeling tuberculous meningitis in zebrafish using Mycobacterium marinum

    Directory of Open Access Journals (Sweden)

    Lisanne M. van Leeuwen

    2014-09-01

    Full Text Available Tuberculous meningitis (TBM is one of the most severe extrapulmonary manifestations of tuberculosis, with a high morbidity and mortality. Characteristic pathological features of TBM are Rich foci, i.e. brain- and spinal-cord-specific granulomas formed after hematogenous spread of pulmonary tuberculosis. Little is known about the early pathogenesis of TBM and the role of Rich foci. We have adapted the zebrafish model of Mycobacterium marinum infection (zebrafish–M. marinum model to study TBM. First, we analyzed whether TBM occurs in adult zebrafish and showed that intraperitoneal infection resulted in granuloma formation in the meninges in 20% of the cases, with occasional brain parenchyma involvement. In zebrafish embryos, bacterial infiltration and clustering of infected phagocytes was observed after infection at three different inoculation sites: parenchyma, hindbrain ventricle and caudal vein. Infection via the bloodstream resulted in the formation of early granulomas in brain tissue in 70% of the cases. In these zebrafish embryos, infiltrates were located in the proximity of blood vessels. Interestingly, no differences were observed when embryos were infected before or after early formation of the blood-brain barrier (BBB, indicating that bacteria are able to cross this barrier with relatively high efficiency. In agreement with this observation, infected zebrafish larvae also showed infiltration of the brain tissue. Upon infection of embryos with an M. marinum ESX-1 mutant, only small clusters and scattered isolated phagocytes with high bacterial loads were present in the brain tissue. In conclusion, our adapted zebrafish–M. marinum infection model for studying granuloma formation in the brain will allow for the detailed analysis of both bacterial and host factors involved in TBM. It will help solve longstanding questions on the role of Rich foci and potentially contribute to the development of better diagnostic tools and therapeutics.

  19. Early life exposure to environmental levels of the aromatase inhibitor tributyltin causes masculinisation and irreversible sperm damage in zebrafish (Danio rerio).

    Science.gov (United States)

    McAllister, Brian G; Kime, David E

    2003-11-19

    To determine whether early life exposure to tributyltin (TBT), an aromatase inhibitor, impaired reproductive function in fish, Danio rerio were exposed to environmentally realistic levels (0.01-100 ng l(-1)) of TBT from 0 to 30, 30 to 60, and 0 to 70 days post-hatch, and the sex ratio and sperm motility of the adults examined 3-5 months after cessation of exposure. Fish exposed for 70 days to 0.1 ng l(-1) of TBT, a concentration presently below the detection limit in water, showed a male biased population which produced a high incidence of sperm lacking flagella. At 1 ng l(-1), the motility of sperm was significantly lower than that of control fish, while at 10 ng l(-1), all sperm lacked flagella and, at 100 ng l(-1), milt volume had increased. The effect of exposure on sex ratio was similar after exposure from 0 to 70 and 0 to 30 days, but even 100 ng l(-1) gave only 65% males after exposure from 30 to 60 days. Effects on sperm motility and morphology and on milt volume were less pronounced after 30 day than 70 day exposure. Our data suggest that screening for aromatase inhibiting activity and assessment of its risks in early life to human and wildlife fertility needs to be urgently addressed, and that the reproductive toxicity of TBT may presently be underestimated.

  20. Remotely Sensing Larval Population Dynamics of Rice Field Anophelines

    Science.gov (United States)

    Beck, Louisa R.; Dister, Sheri W.; Wood, Byron L.; Washino, Robert K.

    1997-01-01

    The primary objective of both studies was to determine if RS and GIS techniques could be used to distinguish between high and low larval-producing rice fields in California. Results of the first study suggested that early-season green-up and proximity to livestock pastures were positively correlated with high larval abundance. Based on the early-season spectral differences between high and low larval-producing fields, it appeared that canopy development and tillering influenced mosquito habitat quality. At that time, rice fields consisted of a mixture of plants and water, a combination that allowed An. freeborni females to lay eggs in partial sunlight, protected from both predators and wind. This established a population earlier in the season than in other, 'less-green' fields where tillering and plant emergence was too minimal for ovipositioning. The study also indicated the importance of the distance that a mosquito would have to fly in order to take a bloodmeal prior to ovipositing. These associations were fully explored in an expanded study two years later. The second study confirmed the positive relationship between early season canopy development and larval abundance, and also demonstrated the relationship between abundance and distance-to-pasture. The association between greenness (as measured using NDVI), distance-to-pasture, and abundance is illustrated. The second study also indicated the siginificance of the landscape context of rice fields for larval production. Fields that included opportunities for feeding and resting within the flight range of the mosquito had higher abundances than did fields that were in a homogeneous rice area.

  1. Developmental toxicity of CdTe QDs in zebrafish embryos and larvae

    Energy Technology Data Exchange (ETDEWEB)

    Duan, Junchao; Yu, Yongbo [School of Public Health, Capital Medical University (China); Li, Yang [School of Public Health, Jilin University (China); Yu, Yang; Li, Yanbo; Huang, Peili; Zhou, Xianqing [School of Public Health, Capital Medical University (China); Peng, Shuangqing, E-mail: pengsq@hotmail.com [Institute of Disease Control and Prevention, Academy of Military Medical Sciences, Evaluation and Research Centre for Toxicology (China); Sun, Zhiwei, E-mail: zwsun@ccmu.edu.cn [School of Public Health, Capital Medical University (China)

    2013-07-15

    Quantum dots (QDs) have widely been used in biomedical and biotechnological applications. However, few studies focus on the assessing toxicity of QDs exposure in vivo. In this study, zebrafish embryos were treated with CdTe QDs (4 nm) during 4-96 h post-fertilization (hpf). Mortality, hatching rate, malformation, heart rate, and QDs uptake were detected. We also measured the larval behavior to analyze whether QDs had persistent effects on larvae locomotor activity at 144 hpf. The results showed that as the exposure dosages increased, the hatching rate and heart rate of zebrafish embryos were decreased, while the mortality increased. Exposure to QDs caused embryonic malformations, including head malformation, pericardial edema, yolk sac edema, bent spine, and yolk not depleted. QDs fluorescence was mainly localized in the intestines region. The larval behavior testing showed that the total swimming distance was decreased in a dose-dependent manner. The lowest dose (2.5 nM QDs) produced substantial hyperactivity while the higher doses groups (5, 10, and 20 nM QDs) elicited remarkably hypoactivity in dark periods. In summary, the data of this article indicated that QDs caused embryonic developmental toxicity, resulted in persistent effects on larval behavior.

  2. Developmental toxicity of CdTe QDs in zebrafish embryos and larvae

    International Nuclear Information System (INIS)

    Duan, Junchao; Yu, Yongbo; Li, Yang; Yu, Yang; Li, Yanbo; Huang, Peili; Zhou, Xianqing; Peng, Shuangqing; Sun, Zhiwei

    2013-01-01

    Quantum dots (QDs) have widely been used in biomedical and biotechnological applications. However, few studies focus on the assessing toxicity of QDs exposure in vivo. In this study, zebrafish embryos were treated with CdTe QDs (4 nm) during 4–96 h post-fertilization (hpf). Mortality, hatching rate, malformation, heart rate, and QDs uptake were detected. We also measured the larval behavior to analyze whether QDs had persistent effects on larvae locomotor activity at 144 hpf. The results showed that as the exposure dosages increased, the hatching rate and heart rate of zebrafish embryos were decreased, while the mortality increased. Exposure to QDs caused embryonic malformations, including head malformation, pericardial edema, yolk sac edema, bent spine, and yolk not depleted. QDs fluorescence was mainly localized in the intestines region. The larval behavior testing showed that the total swimming distance was decreased in a dose-dependent manner. The lowest dose (2.5 nM QDs) produced substantial hyperactivity while the higher doses groups (5, 10, and 20 nM QDs) elicited remarkably hypoactivity in dark periods. In summary, the data of this article indicated that QDs caused embryonic developmental toxicity, resulted in persistent effects on larval behavior

  3. Developmental toxicity of CdTe QDs in zebrafish embryos and larvae

    Science.gov (United States)

    Duan, Junchao; Yu, Yongbo; Li, Yang; Yu, Yang; Li, Yanbo; Huang, Peili; Zhou, Xianqing; Peng, Shuangqing; Sun, Zhiwei

    2013-07-01

    Quantum dots (QDs) have widely been used in biomedical and biotechnological applications. However, few studies focus on the assessing toxicity of QDs exposure in vivo. In this study, zebrafish embryos were treated with CdTe QDs (4 nm) during 4-96 h post-fertilization (hpf). Mortality, hatching rate, malformation, heart rate, and QDs uptake were detected. We also measured the larval behavior to analyze whether QDs had persistent effects on larvae locomotor activity at 144 hpf. The results showed that as the exposure dosages increased, the hatching rate and heart rate of zebrafish embryos were decreased, while the mortality increased. Exposure to QDs caused embryonic malformations, including head malformation, pericardial edema, yolk sac edema, bent spine, and yolk not depleted. QDs fluorescence was mainly localized in the intestines region. The larval behavior testing showed that the total swimming distance was decreased in a dose-dependent manner. The lowest dose (2.5 nM QDs) produced substantial hyperactivity while the higher doses groups (5, 10, and 20 nM QDs) elicited remarkably hypoactivity in dark periods. In summary, the data of this article indicated that QDs caused embryonic developmental toxicity, resulted in persistent effects on larval behavior.

  4. The dlx5a/dlx6a genes play essential roles in the early development of zebrafish median fin and pectoral structures.

    Directory of Open Access Journals (Sweden)

    Eglantine Heude

    Full Text Available The Dlx5 and Dlx6 genes encode homeodomain transcription factors essential for the proper development of limbs in mammalian species. However, the role of their teleost counterparts in fin development has received little attention. Here, we show that dlx5a is an early marker of apical ectodermal cells of the pectoral fin buds and of the median fin fold, but also of cleithrum precursor cells during pectoral girdle development. We propose that early median fin fold establishment results from the medial convergence of dlx5a-expressing cells at the lateral edges of the neural keel. Expression analysis also shows involvement of dlx5a during appendage skeletogenesis. Using morpholino-mediated knock down, we demonstrate that disrupted dlx5a/6a function results in pectoral fin agenesis associated with misexpression of bmp4, fgf8a, and1 and msx genes. In contrast, the median fin fold presents defects in mesenchymal cell migration and actinotrichia formation, whereas the initial specification seems to occur normally. Our results demonstrate that the dlx5a/6a genes are essential for the induction of pectoral fin outgrowth, but are not required during median fin fold specification. The dlx5a/6a knock down also causes a failure of cleithrum formation associated with a drastic loss of runx2b and col10a1 expression. The data indicate distinct requirements for dlx5a/6a during median and pectoral fin development suggesting that initiation of unpaired and paired fin formation are not directed through the same molecular mechanisms. Our results refocus arguments on the mechanistic basis of paired appendage genesis during vertebrate evolution.

  5. The dlx5a/dlx6a Genes Play Essential Roles in the Early Development of Zebrafish Median Fin and Pectoral Structures

    Science.gov (United States)

    Heude, Églantine; Shaikho, Sarah; Ekker, Marc

    2014-01-01

    The Dlx5 and Dlx6 genes encode homeodomain transcription factors essential for the proper development of limbs in mammalian species. However, the role of their teleost counterparts in fin development has received little attention. Here, we show that dlx5a is an early marker of apical ectodermal cells of the pectoral fin buds and of the median fin fold, but also of cleithrum precursor cells during pectoral girdle development. We propose that early median fin fold establishment results from the medial convergence of dlx5a-expressing cells at the lateral edges of the neural keel. Expression analysis also shows involvement of dlx5a during appendage skeletogenesis. Using morpholino-mediated knock down, we demonstrate that disrupted dlx5a/6a function results in pectoral fin agenesis associated with misexpression of bmp4, fgf8a, and1 and msx genes. In contrast, the median fin fold presents defects in mesenchymal cell migration and actinotrichia formation, whereas the initial specification seems to occur normally. Our results demonstrate that the dlx5a/6a genes are essential for the induction of pectoral fin outgrowth, but are not required during median fin fold specification. The dlx5a/6a knock down also causes a failure of cleithrum formation associated with a drastic loss of runx2b and col10a1 expression. The data indicate distinct requirements for dlx5a/6a during median and pectoral fin development suggesting that initiation of unpaired and paired fin formation are not directed through the same molecular mechanisms. Our results refocus arguments on the mechanistic basis of paired appendage genesis during vertebrate evolution. PMID:24858471

  6. The Zebrafish Model Organism Database (ZFIN)

    Data.gov (United States)

    U.S. Department of Health & Human Services — ZFIN serves as the zebrafish model organism database. It aims to: a) be the community database resource for the laboratory use of zebrafish, b) develop and support...

  7. Brain-wide neuronal dynamics during motor adaptation in zebrafish.

    Science.gov (United States)

    Ahrens, Misha B; Li, Jennifer M; Orger, Michael B; Robson, Drew N; Schier, Alexander F; Engert, Florian; Portugues, Ruben

    2012-05-09

    A fundamental question in neuroscience is how entire neural circuits generate behaviour and adapt it to changes in sensory feedback. Here we use two-photon calcium imaging to record the activity of large populations of neurons at the cellular level, throughout the brain of larval zebrafish expressing a genetically encoded calcium sensor, while the paralysed animals interact fictively with a virtual environment and rapidly adapt their motor output to changes in visual feedback. We decompose the network dynamics involved in adaptive locomotion into four types of neuronal response properties, and provide anatomical maps of the corresponding sites. A subset of these signals occurred during behavioural adjustments and are candidates for the functional elements that drive motor learning. Lesions to the inferior olive indicate a specific functional role for olivocerebellar circuitry in adaptive locomotion. This study enables the analysis of brain-wide dynamics at single-cell resolution during behaviour.

  8. Premature aging in telomerase-deficient zebrafish

    Directory of Open Access Journals (Sweden)

    Monique Anchelin

    2013-09-01

    The study of telomere biology is crucial to the understanding of aging and cancer. In the pursuit of greater knowledge in the field of human telomere biology, the mouse has been used extensively as a model. However, there are fundamental differences between mouse and human cells. Therefore, additional models are required. In light of this, we have characterized telomerase-deficient zebrafish (Danio rerio as the second vertebrate model for human telomerase-driven diseases. We found that telomerase-deficient zebrafish show p53-dependent premature aging and reduced lifespan in the first generation, as occurs in humans but not in mice, probably reflecting the similar telomere length in fish and humans. Among these aging symptoms, spinal curvature, liver and retina degeneration, and infertility were the most remarkable. Although the second-generation embryos died in early developmental stages, restoration of telomerase activity rescued telomere length and survival, indicating that telomerase dosage is crucial. Importantly, this model also reproduces the disease anticipation observed in humans with dyskeratosis congenita (DC. Thus, telomerase haploinsufficiency leads to anticipation phenomenon in longevity, which is related to telomere shortening and, specifically, with the proportion of short telomeres. Furthermore, p53 was induced by telomere attrition, leading to growth arrest and apoptosis. Importantly, genetic inhibition of p53 rescued the adverse effects of telomere loss, indicating that the molecular mechanisms induced by telomere shortening are conserved from fish to mammals. The partial rescue of telomere length and longevity by restoration of telomerase activity, together with the feasibility of the zebrafish for high-throughput chemical screening, both point to the usefulness of this model for the discovery of new drugs able to reactivate telomerase in individuals with DC.

  9. Zebrafish yolk lipid processing: a tractable tool for the study of vertebrate lipid transport and metabolism

    Directory of Open Access Journals (Sweden)

    Rosa L. Miyares

    2014-07-01

    Full Text Available Dyslipidemias are a major cause of morbidity and mortality in the world, particularly in developed nations. Investigating lipid and lipoprotein metabolism in experimentally tractable animal models is a crucial step towards understanding and treating human dyslipidemias. The zebrafish, a well-established embryological model, is emerging as a notable system for studies of lipid metabolism. Here, we describe the value of the lecithotrophic, or yolk-metabolizing, stages of the zebrafish as a model for studying lipid metabolism and lipoprotein transport. We demonstrate methods to assay yolk lipid metabolism in embryonic and larval zebrafish. Injection of labeled fatty acids into the zebrafish yolk promotes efficient uptake into the circulation and rapid metabolism. Using a genetic model for abetalipoproteinemia, we show that the uptake of labeled fatty acids into the circulation is dependent on lipoprotein production. Furthermore, we examine the metabolic fate of exogenously delivered fatty acids by assaying their incorporation into complex lipids. Moreover, we demonstrate that this technique is amenable to genetic and pharmacologic studies.

  10. Estrogenic effects of several BPA analogs in the developing zebrafish brain

    Directory of Open Access Journals (Sweden)

    Joel eCano-Nicolau

    2016-03-01

    Full Text Available Important set of studies have demonstrated the endocrine disrupting activity of Bisphenol A (BPA. The present work aimed at defining estrogenic-like activity of several BPA structural analogs, including BPS, BPF, BPAF, and BPAP, on 4-day or 7-day post-fertilization (dpf zebrafish larva as an in vivo model. We measured the induction level of the estrogen-sensitive marker cyp19a1b gene (Aromatase B, expressed in the brain, using three different in situ/in vivo strategies: 1 Quantification of cyp19a1b transcripts using RT-qPCR in wild type 7-dpf larva brains exposed to bisphenols ; 2 Detection and distribution of cyp19a1b transcripts using in situ hybridization on 7-dpf brain sections (hypothalamus; and 3 Quantification of the cyp19a1b promoter activity in live cyp19a1b-GFP transgenic zebrafish (EASZY assay at 4-dpf larval stage. These three different experimental approaches demonstrated that BPS, BPF or BPAF exposure, similarly to BPA, significantly activates the expression of the estrogenic marker in the brain of developing zebrafish. In vitro experiments using both reporter gene assay in a glial cell context and competitive ligand binding assays strongly suggested that up-regulation of cyp19a1b is largely mediated by the zebrafish estrogen nuclear receptor alpha (zfERα. Importantly, and in contrast to other tested bisphenol A analogs, the bisphenol AP (BPAP did not show estrogenic activity in our model.

  11. The Complete and Updated "Rotifer Polyculture Method" for Rearing First Feeding Zebrafish

    Science.gov (United States)

    Lawrence, Christian; Best, Jason; Cockington, Jason; Henry, Eric C.; Hurley, Shane; James, Althea; Lapointe, Christopher; Maloney, Kara; Sanders, Erik

    2016-01-01

    The zebrafish (Danio rerio) is a model organism of increasing importance in many fields of science. One of the most demanding technical aspects of culture of this species in the laboratory is rearing first-feeding larvae to the juvenile stage with high rates of growth and survival. The central management challenge of this developmental period revolves around delivering highly nutritious feed items to the fish on a nearly continuous basis without compromising water quality. Because larval zebrafish are well-adapted to feed on small zooplankton in the water column, live prey items such as brachionid rotifers, Artemia, and Paramecium are widely recognized as the feeds of choice, at least until the fish reach the juvenile stage and are able to efficiently feed on processed diets. This protocol describes a method whereby newly hatched zebrafish larvae are cultured together with live saltwater rotifers (Brachionus plicatilis) in the same system. This polyculture approach provides fish with an "on-demand", nutrient-rich live food source without producing chemical waste at levels that would otherwise limit performance. Importantly, because the system harnesses both the natural high productivity of the rotifers and the behavioral preferences of the fish, the labor involved with maintenance is low. The following protocol details an updated, step-by-step procedure that incorporates rotifer production (scalable to any desired level) for use in a polyculture of zebrafish larvae and rotifers that promotes maximal performance during the first 5 days of exogenous feeding. PMID:26863035

  12. Analysis of a Gene Regulatory Cascade Mediating Circadian Rhythm in Zebrafish

    Science.gov (United States)

    Wang, Haifang; Du, Jiulin; Yan, Jun

    2013-01-01

    In the study of circadian rhythms, it has been a puzzle how a limited number of circadian clock genes can control diverse aspects of physiology. Here we investigate circadian gene expression genome-wide using larval zebrafish as a model system. We made use of a spatial gene expression atlas to investigate the expression of circadian genes in various tissues and cell types. Comparison of genome-wide circadian gene expression data between zebrafish and mouse revealed a nearly anti-phase relationship and allowed us to detect novel evolutionarily conserved circadian genes in vertebrates. We identified three groups of zebrafish genes with distinct responses to light entrainment: fast light-induced genes, slow light-induced genes, and dark-induced genes. Our computational analysis of the circadian gene regulatory network revealed several transcription factors (TFs) involved in diverse aspects of circadian physiology through transcriptional cascade. Of these, microphthalmia-associated transcription factor a (mitfa), a dark-induced TF, mediates a circadian rhythm of melanin synthesis, which may be involved in zebrafish's adaptation to daily light cycling. Our study describes a systematic method to discover previously unidentified TFs involved in circadian physiology in complex organisms. PMID:23468616

  13. A zebrafish model of chordoma initiated by notochord-driven expression of HRASV12.

    Science.gov (United States)

    Burger, Alexa; Vasilyev, Aleksandr; Tomar, Ritu; Selig, Martin K; Nielsen, G Petur; Peterson, Randall T; Drummond, Iain A; Haber, Daniel A

    2014-07-01

    Chordoma is a malignant tumor thought to arise from remnants of the embryonic notochord, with its origin in the bones of the axial skeleton. Surgical resection is the standard treatment, usually in combination with radiation therapy, but neither chemotherapeutic nor targeted therapeutic approaches have demonstrated success. No animal model and only few chordoma cell lines are available for preclinical drug testing, and, although no druggable genetic drivers have been identified, activation of EGFR and downstream AKT-PI3K pathways have been described. Here, we report a zebrafish model of chordoma, based on stable transgene-driven expression of HRASV12 in notochord cells during development. Extensive intra-notochordal tumor formation is evident within days of transgene expression, ultimately leading to larval death. The zebrafish tumors share characteristics of human chordoma as demonstrated by immunohistochemistry and electron microscopy. The mTORC1 inhibitor rapamycin, which has some demonstrated activity in a chordoma cell line, delays the onset of tumor formation in our zebrafish model, and improves survival of tumor-bearing fish. Consequently, the HRASV12-driven zebrafish model of chordoma could enable high-throughput screening of potential therapeutic agents for the treatment of this refractory cancer. © 2014. Published by The Company of Biologists Ltd.

  14. A zebrafish model of chordoma initiated by notochord-driven expression of HRASV12

    Directory of Open Access Journals (Sweden)

    Alexa Burger

    2014-07-01

    Full Text Available Chordoma is a malignant tumor thought to arise from remnants of the embryonic notochord, with its origin in the bones of the axial skeleton. Surgical resection is the standard treatment, usually in combination with radiation therapy, but neither chemotherapeutic nor targeted therapeutic approaches have demonstrated success. No animal model and only few chordoma cell lines are available for preclinical drug testing, and, although no druggable genetic drivers have been identified, activation of EGFR and downstream AKT-PI3K pathways have been described. Here, we report a zebrafish model of chordoma, based on stable transgene-driven expression of HRASV12 in notochord cells during development. Extensive intra-notochordal tumor formation is evident within days of transgene expression, ultimately leading to larval death. The zebrafish tumors share characteristics of human chordoma as demonstrated by immunohistochemistry and electron microscopy. The mTORC1 inhibitor rapamycin, which has some demonstrated activity in a chordoma cell line, delays the onset of tumor formation in our zebrafish model, and improves survival of tumor-bearing fish. Consequently, the HRASV12-driven zebrafish model of chordoma could enable high-throughput screening of potential therapeutic agents for the treatment of this refractory cancer.

  15. Automated processing of zebrafish imaging data: a survey.

    Science.gov (United States)

    Mikut, Ralf; Dickmeis, Thomas; Driever, Wolfgang; Geurts, Pierre; Hamprecht, Fred A; Kausler, Bernhard X; Ledesma-Carbayo, María J; Marée, Raphaël; Mikula, Karol; Pantazis, Periklis; Ronneberger, Olaf; Santos, Andres; Stotzka, Rainer; Strähle, Uwe; Peyriéras, Nadine

    2013-09-01

    Due to the relative transparency of its embryos and larvae, the zebrafish is an ideal model organism for bioimaging approaches in vertebrates. Novel microscope technologies allow the imaging of developmental processes in unprecedented detail, and they enable the use of complex image-based read-outs for high-throughput/high-content screening. Such applications can easily generate Terabytes of image data, the handling and analysis of which becomes a major bottleneck in extracting the targeted information. Here, we describe the current state of the art in computational image analysis in the zebrafish system. We discuss the challenges encountered when handling high-content image data, especially with regard to data quality, annotation, and storage. We survey methods for preprocessing image data for further analysis, and describe selected examples of automated image analysis, including the tracking of cells during embryogenesis, heartbeat detection, identification of dead embryos, recognition of tissues and anatomical landmarks, and quantification of behavioral patterns of adult fish. We review recent examples for applications using such methods, such as the comprehensive analysis of cell lineages during early development, the generation of a three-dimensional brain atlas of zebrafish larvae, and high-throughput drug screens based on movement patterns. Finally, we identify future challenges for the zebrafish image analysis community, notably those concerning the compatibility of algorithms and data formats for the assembly of modular analysis pipelines.

  16. Automated Processing of Zebrafish Imaging Data: A Survey

    Science.gov (United States)

    Dickmeis, Thomas; Driever, Wolfgang; Geurts, Pierre; Hamprecht, Fred A.; Kausler, Bernhard X.; Ledesma-Carbayo, María J.; Marée, Raphaël; Mikula, Karol; Pantazis, Periklis; Ronneberger, Olaf; Santos, Andres; Stotzka, Rainer; Strähle, Uwe; Peyriéras, Nadine

    2013-01-01

    Abstract Due to the relative transparency of its embryos and larvae, the zebrafish is an ideal model organism for bioimaging approaches in vertebrates. Novel microscope technologies allow the imaging of developmental processes in unprecedented detail, and they enable the use of complex image-based read-outs for high-throughput/high-content screening. Such applications can easily generate Terabytes of image data, the handling and analysis of which becomes a major bottleneck in extracting the targeted information. Here, we describe the current state of the art in computational image analysis in the zebrafish system. We discuss the challenges encountered when handling high-content image data, especially with regard to data quality, annotation, and storage. We survey methods for preprocessing image data for further analysis, and describe selected examples of automated image analysis, including the tracking of cells during embryogenesis, heartbeat detection, identification of dead embryos, recognition of tissues and anatomical landmarks, and quantification of behavioral patterns of adult fish. We review recent examples for applications using such methods, such as the comprehensive analysis of cell lineages during early development, the generation of a three-dimensional brain atlas of zebrafish larvae, and high-throughput drug screens based on movement patterns. Finally, we identify future challenges for the zebrafish image analysis community, notably those concerning the compatibility of algorithms and data formats for the assembly of modular analysis pipelines. PMID:23758125

  17. First feeding of larval herring

    DEFF Research Database (Denmark)

    Kiørboe, Thomas; Munk, Peter; Støttrup, Josianne

    1985-01-01

    The transition period from endogenous to exogenous feeding by larval herring was investigated in the laboratory for four herring stocks in order to evaluate the chances of survival at the time of fiest feeding. Observations on larval activity, feeding and growth were related to amount of yolk......, visual experience with potential prey organisms prior to first feeding and prey density. Herring larvae did not initiate exogenous feeding until around the time of yolk resorption. The timing of first feeding was not influenced by prior exposure to potential prey organisms during the yolk sac stage....... In the light of these observations, the ecological significance of the yolk sac stage is discussed. Initiation of exogenous feeding was delayed by 1-4 days at a low (7.5 nauplii .cntdot. l-1) compared to a high (120 nauplii .cntdot. l-1) prey density, but even at prey densities corresponding to the lower end...

  18. The genetics of hair-cell function in zebrafish.

    Science.gov (United States)

    Nicolson, Teresa

    2017-09-01

    Our ears are remarkable sensory organs, providing the important senses of balance and hearing. The complex structure of the inner ear, or 'labyrinth', along with the assorted neuroepithelia, have evolved to detect head movements and sounds with impressive sensitivity. The rub is that the inner ear is highly vulnerable to genetic lesions and environmental insults. According to National Institute of Health estimates, hearing loss is one of the most commonly inherited or acquired sensorineural diseases. To understand the causes of deafness and balance disorders, it is imperative to understand the underlying biology of the inner ear, especially the inner workings of the sensory receptors. These receptors, which are termed hair cells, are particularly susceptible to genetic mutations - more than two dozen genes are associated with defects in this cell type in humans. Over the past decade, a substantial amount of progress has been made in working out the molecular basis of hair-cell function using vertebrate animal models. Given the transparency of the inner ear and the genetic tools that are available, zebrafish have become an increasingly popular animal model for the study of deafness and vestibular dysfunction. Mutagenesis screens for larval defects in hearing and balance have been fruitful in finding key components, many of which have been implicated in human deafness. This review will focus on the genes that are required for hair-cell function in zebrafish, with a particular emphasis on mechanotransduction. In addition, the generation of new tools available for the characterization of zebrafish hair-cell mutants will be discussed.

  19. Tools for automating the imaging of zebrafish larvae.

    Science.gov (United States)

    Pulak, Rock

    2016-03-01

    The VAST BioImager system is a set of tools developed for zebrafish researchers who require the collection of images from a large number of 2-7 dpf zebrafish larvae. The VAST BioImager automates larval handling, positioning and orientation tasks. Color images at about 10 μm resolution are collected from the on-board camera of the system. If images of greater resolution and detail are required, this system is mounted on an upright microscope, such as a confocal or fluorescence microscope, to utilize their capabilities. The system loads a larvae, positions it in view of the camera, determines orientation using pattern recognition analysis, and then more precisely positions to user-defined orientation for optimal imaging of any desired tissue or organ system. Multiple images of the same larva can be collected. The specific part of each larva and the desired orientation and position is identified by the researcher and an experiment defining the settings and a series of steps can be saved and repeated for imaging of subsequent larvae. The system captures images, then ejects and loads another larva from either a bulk reservoir, a well of a 96 well plate using the LP Sampler, or individually targeted larvae from a Petri dish or other container using the VAST Pipettor. Alternative manual protocols for handling larvae for image collection are tedious and time consuming. The VAST BioImager automates these steps to allow for greater throughput of assays and screens requiring high-content image collection of zebrafish larvae such as might be used in drug discovery and toxicology studies. Copyright © 2015 The Author. Published by Elsevier Inc. All rights reserved.

  20. Expression of CALR mutants causes mpl-dependent thrombocytosis in zebrafish.

    Science.gov (United States)

    Lim, K-H; Chang, Y-C; Chiang, Y-H; Lin, H-C; Chang, C-Y; Lin, C-S; Huang, L; Wang, W-T; Gon-Shen Chen, C; Chou, W-C; Kuo, Y-Y

    2016-10-07

    CALR mutations are identified in about 30% of JAK2/MPL-unmutated myeloproliferative neoplasms (MPNs) including essential thrombocythemia (ET) and primary myelofibrosis. Although the molecular pathogenesis of CALR mutations leading to MPNs has been studied using in vitro cell lines models, how mutant CALR may affect developmental hematopoiesis remains unknown. Here we took advantage of the zebrafish model to examine the effects of mutant CALR on early hematopoiesis and model human CALR-mutated MPNs. We identified three zebrafish genes orthologous to human CALR, referred to as calr, calr3a and calr3b. The expression of CALR-del52 and CALR-ins5 mutants caused an increase in the hematopoietic stem/progenitor cells followed by thrombocytosis without affecting normal angiogenesis. The expression of CALR mutants also perturbed early developmental hematopoiesis in zebrafish. Importantly, morpholino knockdown of mpl but not epor or csf3r could significantly attenuate the effects of mutant CALR. Furthermore, the expression of mutant CALR caused jak-stat signaling activation in zebrafish that could be blocked by JAK inhibitors (ruxolitinib and fedratinib). These findings showed that mutant CALR activates jak-stat signaling through an mpl-dependent mechanism to mediate pathogenic thrombopoiesis in zebrafish, and illustrated that the signaling machinery related to mutant CALR tumorigenesis are conserved between human and zebrafish.

  1. 'Peer pressure' in larval Drosophila?

    Science.gov (United States)

    Niewalda, Thomas; Jeske, Ines; Michels, Birgit; Gerber, Bertram

    2014-06-06

    Understanding social behaviour requires a study case that is simple enough to be tractable, yet complex enough to remain interesting. Do larval Drosophila meet these requirements? In a broad sense, this question can refer to effects of the mere presence of other larvae on the behaviour of a target individual. Here we focused in a more strict sense on 'peer pressure', that is on the question of whether the behaviour of a target individual larva is affected by what a surrounding group of larvae is doing. We found that innate olfactory preference of a target individual was neither affected (i) by the level of innate olfactory preference in the surrounding group nor (ii) by the expression of learned olfactory preference in the group. Likewise, learned olfactory preference of a target individual was neither affected (iii) by the level of innate olfactory preference of the surrounding group nor (iv) by the learned olfactory preference the group was expressing. We conclude that larval Drosophila thus do not take note of specifically what surrounding larvae are doing. This implies that in a strict sense, and to the extent tested, there is no social interaction between larvae. These results validate widely used en mass approaches to the behaviour of larval Drosophila. © 2014. Published by The Company of Biologists Ltd.

  2. Transgenic zebrafish reveal tissue-specific differences in estrogen signaling in response to environmental water samples.

    Science.gov (United States)

    Gorelick, Daniel A; Iwanowicz, Luke R; Hung, Alice L; Blazer, Vicki S; Halpern, Marnie E

    2014-04-01

    Environmental endocrine disruptors (EEDs) are exogenous chemicals that mimic endogenous hormones such as estrogens. Previous studies using a zebrafish transgenic reporter demonstrated that the EEDs bisphenol A and genistein preferentially activate estrogen receptors (ERs) in the larval heart compared with the liver. However, it was not known whether the transgenic zebrafish reporter was sensitive enough to detect estrogens from environmental samples, whether environmental estrogens would exhibit tissue-specific effects similar to those of BPA and genistein, or why some compounds preferentially target receptors in the heart. We tested surface water samples using a transgenic zebrafish reporter with tandem estrogen response elements driving green fluorescent protein expression (5xERE:GFP). Reporter activation was colocalized with tissue-specific expression of ER genes by RNA in situ hybridization. We observed selective patterns of ER activation in transgenic fish exposed to river water samples from the Mid-Atlantic United States, with several samples preferentially activating receptors in embryonic and larval heart valves. We discovered that tissue specificity in ER activation was due to differences in the expression of ER subtypes. ERα was expressed in developing heart valves but not in the liver, whereas ERβ2 had the opposite profile. Accordingly, subtype-specific ER agonists activated the reporter in either the heart valves or the liver. The use of 5xERE:GFP transgenic zebrafish revealed an unexpected tissue-specific difference in the response to environmentally relevant estrogenic compounds. Exposure to estrogenic EEDs in utero was associated with adverse health effects, with the potentially unanticipated consequence of targeting developing heart valves.

  3. Reprimo tissue-specific expression pattern is conserved between zebrafish and human.

    Directory of Open Access Journals (Sweden)

    Ricardo J Figueroa

    Full Text Available Reprimo (RPRM, a member of the RPRM gene family, is a tumor-suppressor gene involved in the regulation of the p53-mediated cell cycle arrest at G2/M. RPRM has been associated with malignant tumor progression and proposed as a potential biomarker for early cancer detection. However, the expression and role of RPRM, as well as its family, are poorly understood and their physiology is as yet unstudied. In this scenario, a model system like the zebrafish could serve to dissect the role of the RPRM family members in vivo. Phylogenetic analysis reveals that RPRM and RPRML have been differentially retained by most species throughout vertebrate evolution, yet RPRM3 has been retained only in a small group of distantly related species, including zebrafish. Herein, we characterized the spatiotemporal expression of RPRM (present in zebrafish as an infraclass duplication rprma/rprmb, RPRML and RPRM3 in the zebrafish. By whole-mount in situ hybridization (WISH and fluorescent in situ hybridization (FISH, we demonstrate that rprm (rprma/rprmb and rprml show a similar spatiotemporal expression profile during zebrafish development. At early developmental stages rprmb is expressed in somites. After one day post-fertilization, rprm (rprma/rprmb and rprml are expressed in the notochord, brain, blood vessels and digestive tube. On the other hand, rprm3 shows the most unique expression profile, being expressed only in the central nervous system (CNS. We assessed the expression patterns of RPRM gene transcripts in adult zebrafish and human RPRM protein product in tissue samples by RT-qPCR and immunohistochemistry (IHC staining, respectively. Strikingly, tissue-specific expression patterns of the RPRM transcripts and protein are conserved between zebrafish and humans. We propose the zebrafish as a powerful tool to elucidate the both physiological and pathological roles of the RPRM gene family.

  4. Zebrafish as a visual and dynamic model to study the transport of nanosized drug delivery systems across the biological barriers.

    Science.gov (United States)

    Li, Ye; Miao, Xiaoqing; Chen, Tongkai; Yi, Xiang; Wang, Ruibing; Zhao, Haitao; Lee, Simon Ming-Yuen; Wang, Xueqing; Zheng, Ying

    2017-08-01

    With the wide application of nanotechnology to drug delivery systems, a simple, dynamic and visual in vivo model for high-throughput screening of novel formulations with fluorescence markers across biological barriers is desperately needed. In vitro cell culture models have been widely used, although they are far from a complimentary in vivo system. Mammalian animal models are common predictive models to study transport, but they are costly and time consuming. Zebrafish (Danio rerio), a small vertebrate model, have the potential to be developed as an "intermediate" model for quick evaluations. Based on our previously established coumarin 6 nanocrystals (C6-NCs), which have two different sizes, the present study investigates the transportation of C6-NCs across four biological barriers, including the chorion, blood brain barrier (BBB), blood retinal barrier (BRB) and gastrointestinal (GI) barrier, using zebrafish embryos and larvae as in vivo models. The biodistribution and elimination of C6 from different organs were quantified in adult zebrafish. The results showed that compared to 200nm C6-NCs, 70nm C6-NCs showed better permeability across these biological barriers. A FRET study suggested that intact C6-NCs together with the free dissolved form of C6 were absorbed into the larval zebrafish. More C6 was accumulated in different organs after incubation with small sized NCs via lipid raft-mediated endocytosis in adult zebrafish, which is consistent with the findings from in vitro cell monolayers and the zebrafish larvae model. C6-NCs could be gradually eliminated in each organ over time. This study demonstrated the successful application of zebrafish as a simple and dynamic model to simultaneously assess the transport of nanosized drug delivery systems across several biological barriers and biodistribution in different organs, especially in the brain, which could be used for central nervous system (CNS) drug and delivery system screening. Copyright © 2017 Elsevier B

  5. Carbendazim has the potential to induce oxidative stress, apoptosis, immunotoxicity and endocrine disruption during zebrafish larvae development.

    Science.gov (United States)

    Jiang, Jinhua; Wu, Shenggan; Wang, Yanhua; An, Xuehua; Cai, Leiming; Zhao, Xueping; Wu, Changxing

    2015-10-01

    Increasing evidence have suggested deleterious effects of carbendazim on reproduction, apoptosis, immunotoxicity and endocrine disruption in mice and rats, however, the developmental toxicity of carbendazim to aquatic organisms remains obscure. In the present study, we utilized zebrafish as an environmental monitoring model to characterize the effects of carbendazim on expression of genes related to oxidative stress, apoptosis, immunotoxicity and endocrine disruption during larval development. Different trends in gene expression were observed upon exposing the larvae to 4, 20, 100, and 500 μg/L carbendazim for 4 and 8d. The mRNA levels of catalase, glutathione peroxidase and manganese superoxide dismutase (CAT, GPX, and Mn/SOD) were up-regulated after exposure to different concentrations of carbendazim for 4 or 8d. The up-regulation of p53, Apaf1, Cas8 and the down-regulation of Bcl2, Mdm2, Cas3 in the apoptosis pathway, as well as the increased expression of cytokines and chemokines, including CXCL-C1C, CCL1, IL-1b, IFN, IL-8, and TNFα, suggested carbendazim might trigger apoptosis and immune response during zebrafish larval development. In addition, the alteration of mRNA expression of VTG, ERα, ERβ1, ERβ2, TRα, TRβ, Dio1, and Dio2 indicated the potential of carbendazim to induce endocrine disruption in zebrafish larvae. These data suggested that carbendazim could simultaneously induce multiple responses during zebrafish larval development, and bidirectional interactions among oxidative stress, apoptosis pathway, immune and endocrine systems might be present. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Object recognition memory in zebrafish.

    Science.gov (United States)

    May, Zacnicte; Morrill, Adam; Holcombe, Adam; Johnston, Travis; Gallup, Joshua; Fouad, Karim; Schalomon, Melike; Hamilton, Trevor James

    2016-01-01

    The novel object recognition, or novel-object preference (NOP) test is employed to assess recognition memory in a variety of organisms. The subject is exposed to two identical objects, then after a delay, it is placed back in the original environment containing one of the original objects and a novel object. If the subject spends more time exploring one object, this can be interpreted as memory retention. To date, this test has not been fully explored in zebrafish (Danio rerio). Zebrafish possess recognition memory for simple 2- and 3-dimensional geometrical shapes, yet it is unknown if this translates to complex 3-dimensional objects. In this study we evaluated recognition memory in zebrafish using complex objects of different sizes. Contrary to rodents, zebrafish preferentially explored familiar over novel objects. Familiarity preference disappeared after delays of 5 mins. Leopard danios, another strain of D. rerio, also preferred the familiar object after a 1 min delay. Object preference could be re-established in zebra danios by administration of nicotine tartrate salt (50mg/L) prior to stimuli presentation, suggesting a memory-enhancing effect of nicotine. Additionally, exploration biases were present only when the objects were of intermediate size (2 × 5 cm). Our results demonstrate zebra and leopard danios have recognition memory, and that low nicotine doses can improve this memory type in zebra danios. However, exploration biases, from which memory is inferred, depend on object size. These findings suggest zebrafish ecology might influence object preference, as zebrafish neophobia could reflect natural anti-predatory behaviour. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Glyphosate and Roundup® alter morphology and behavior in zebrafish.

    Science.gov (United States)

    Bridi, Daiane; Altenhofen, Stefani; Gonzalez, Jonas Brum; Reolon, Gustavo Kellermann; Bonan, Carla Denise

    2017-12-01

    Glyphosate has become the most widely used herbicide in the world, due to the wide scale adoption of transgenic glyphosate resistant crops after its introduction in 1996. Glyphosate may be used alone, but it is commonly applied as an active ingredient of the herbicide Roundup ® . This pesticide contains several adjuvants, which may promote an unknown toxicity. The indiscriminate application poses numerous problems, both for the health of the applicators and consumers, and for the environment, contaminating the soil, water and leading to the death of plants and animals. Zebrafish (Danio rerio) is quickly gaining popularity in behavioral research, because of physiological similarity to mammals, sensitivity to pharmacological factors, robust performance, low cost, short spawning intervals, external fertilization, transparency of embryos through larval stages, and rapid development. The aim of this study was evaluate the effects of glyphosate and Roundup ® on behavioral and morphological parameters in zebrafish larvae and adults. Zebrafish larvae at 3days post-fertilization and adults were exposed to glyphosate (0.01, 0.065, and 0.5mg/L) or Roundup ® (0.01, 0.065, and 0.5mg/L) for 96h. Immediately after the exposure, we performed the analysis of locomotor activity, aversive behavior, and morphology for the larvae and exploratory behavior, aggression and inhibitory avoidance memory for adult zebrafish. In zebrafish larvae, there were significant differences in the locomotor activity and aversive behavior after glyphosate or Roundup ® exposure when compared to the control group. Our findings demonstrated that exposure to glyphosate at the concentration of 0.5mg/L, Roundup ® at 0.065 or 0.5mg/L reduced the distance traveled, the mean speed and the line crossings in adult zebrafish. A decreased ocular distance was observed for larvae exposed at 0.5mg/L of glyphosate. We verified that at 0.5mg/L of Roundup ® -treated adult zebrafish demonstrated a significant

  8. Arsenic transport by zebrafish aquaglyceroporins

    Directory of Open Access Journals (Sweden)

    Landfear Scott M

    2009-11-01

    Full Text Available Abstract Background Arsenic is one of the most ubiquitous toxins and endangers the health of tens of millions of humans worldwide. It is a mainly a water-borne contaminant. Inorganic trivalent arsenic (AsIII is one of the major species that exists environmentally. The transport of AsIII has been studied in microbes, plants and mammals. Members of the aquaglyceroporin family have been shown to actively conduct AsIII and its organic metabolite, monomethylarsenite (MAsIII. However, the transport of AsIII and MAsIII in in any fish species has not been characterized. Results In this study, five members of the aquaglyceroporin family from zebrafish (Danio rerio were cloned, and their ability to transport water, glycerol, and trivalent arsenicals (AsIII and MAsIII and antimonite (SbIII was investigated. Genes for at least seven aquaglyceroporins have been annotated in the zebrafish genome project. Here, five genes which are close homologues to human AQP3, AQP9 and AQP10 were cloned from a zebrafish cDNA preparation. These genes were named aqp3, aqp3l, aqp9a, aqp9b and aqp10 according to their similarities to the corresponding human AQPs. Expression of aqp9a, aqp9b, aqp3, aqp3l and aqp10 in multiple zebrafish organs were examined by RT-PCR. Our results demonstrated that these aquaglyceroporins exhibited different tissue expression. They are all detected in more than one tissue. The ability of these five aquaglyceroporins to transport water, glycerol and the metalloids arsenic and antimony was examined following expression in oocytes from Xenopus leavis. Each of these channels showed substantial glycerol transport at equivalent rates. These aquaglyceroporins also facilitate uptake of inorganic AsIII, MAsIII and SbIII. Arsenic accumulation in fish larvae and in different tissues from adult zebrafish was studied following short-term arsenic exposure. The results showed that liver is the major organ of arsenic accumulation; other tissues such as gill, eye

  9. Early gonad development in zebrafish (Danio rerio)

    African Journals Online (AJOL)

    SAM

    2014-08-13

    Aug 13, 2014 ... as a relatively larger proportion of proliferating "gonial" germ cells in the latter gonads was considered ... solution in 100 ml tank water) as described by Westerfield (1993). They were ... 90°C hot plate with toluidine blue and thereafter examined and ... morphological sex differentiation, sex differences could.

  10. Identification of estrogen target genes during zebrafish embryonic development through transcriptomic analysis.

    Directory of Open Access Journals (Sweden)

    Ruixin Hao

    Full Text Available Estrogen signaling is important for vertebrate embryonic development. Here we have used zebrafish (Danio rerio as a vertebrate model to analyze estrogen signaling during development. Zebrafish embryos were exposed to 1 µM 17β-estradiol (E2 or vehicle from 3 hours to 4 days post fertilization (dpf, harvested at 1, 2, 3 and 4 dpf, and subjected to RNA extraction for transcriptome analysis using microarrays. Differentially expressed genes by E2-treatment were analyzed with hierarchical clustering followed by biological process and tissue enrichment analysis. Markedly distinct sets of genes were up and down-regulated by E2 at the four different time points. Among these genes, only the well-known estrogenic marker vtg1 was co-regulated at all time points. Despite this, the biological functional categories targeted by E2 were relatively similar throughout zebrafish development. According to knowledge-based tissue enrichment, estrogen responsive genes were clustered mainly in the liver, pancreas and brain. This was in line with the developmental dynamics of estrogen-target tissues that were visualized using transgenic zebrafish containing estrogen responsive elements driving the expression of GFP (Tg(5xERE:GFP. Finally, the identified embryonic estrogen-responsive genes were compared to already published estrogen-responsive genes identified in male adult zebrafish (Gene Expression Omnibus database. The expressions of a few genes were co-regulated by E2 in both embryonic and adult zebrafish. These could potentially be used as estrogenic biomarkers for exposure to estrogens or estrogenic endocrine disruptors in zebrafish. In conclusion, our data suggests that estrogen effects on early embryonic zebrafish development are stage- and tissue- specific.

  11. The Fanconi anemia/BRCA gene network in zebrafish: Embryonic expression and comparative genomics

    Energy Technology Data Exchange (ETDEWEB)

    Titus, Tom A.; Yan Yilin; Wilson, Catherine; Starks, Amber M.; Frohnmayer, Jonathan D.; Bremiller, Ruth A.; Canestro, Cristian; Rodriguez-Mari, Adriana; He Xinjun [Institute of Neuroscience, University of Oregon, 1425 E. 13th Avenue, Eugene, OR 97403 (United States); Postlethwait, John H., E-mail: jpostle@uoneuro.uoregon.edu [Institute of Neuroscience, University of Oregon, 1425 E. 13th Avenue, Eugene, OR 97403 (United States)

    2009-07-31

    Fanconi anemia (FA) is a genetic disease resulting in bone marrow failure, high cancer risks, and infertility, and developmental anomalies including microphthalmia, microcephaly, hypoplastic radius and thumb. Here we present cDNA sequences, genetic mapping, and genomic analyses for the four previously undescribed zebrafish FA genes (fanci, fancj, fancm, and fancn), and show that they reverted to single copy after the teleost genome duplication. We tested the hypothesis that FA genes are expressed during embryonic development in tissues that are disrupted in human patients by investigating fanc gene expression patterns. We found fanc gene maternal message, which can provide Fanc proteins to repair DNA damage encountered in rapid cleavage divisions. Zygotic expression was broad but especially strong in eyes, central nervous system and hematopoietic tissues. In the pectoral fin bud at hatching, fanc genes were expressed specifically in the apical ectodermal ridge, a signaling center for fin/limb development that may be relevant to the radius/thumb anomaly of FA patients. Hatching embryos expressed fanc genes strongly in the oral epithelium, a site of squamous cell carcinomas in FA patients. Larval and adult zebrafish expressed fanc genes in proliferative regions of the brain, which may be related to microcephaly in FA. Mature ovaries and testes expressed fanc genes in specific stages of oocyte and spermatocyte development, which may be related to DNA repair during homologous recombination in meiosis and to infertility in human patients. The intestine strongly expressed some fanc genes specifically in proliferative zones. Our results show that zebrafish has a complete complement of fanc genes in single copy and that these genes are expressed in zebrafish embryos and adults in proliferative tissues that are often affected in FA patients. These results support the notion that zebrafish offers an attractive experimental system to help unravel mechanisms relevant not only

  12. The Fanconi anemia/BRCA gene network in zebrafish: embryonic expression and comparative genomics.

    Science.gov (United States)

    Titus, Tom A; Yan, Yi-Lin; Wilson, Catherine; Starks, Amber M; Frohnmayer, Jonathan D; Bremiller, Ruth A; Cañestro, Cristian; Rodriguez-Mari, Adriana; He, Xinjun; Postlethwait, John H

    2009-07-31

    Fanconi anemia (FA) is a genetic disease resulting in bone marrow failure, high cancer risks, and infertility, and developmental anomalies including microphthalmia, microcephaly, hypoplastic radius and thumb. Here we present cDNA sequences, genetic mapping, and genomic analyses for the four previously undescribed zebrafish FA genes (fanci, fancj, fancm, and fancn), and show that they reverted to single copy after the teleost genome duplication. We tested the hypothesis that FA genes are expressed during embryonic development in tissues that are disrupted in human patients by investigating fanc gene expression patterns. We found fanc gene maternal message, which can provide Fanc proteins to repair DNA damage encountered in rapid cleavage divisions. Zygotic expression was broad but especially strong in eyes, central nervous system and hematopoietic tissues. In the pectoral fin bud at hatching, fanc genes were expressed specifically in the apical ectodermal ridge, a signaling center for fin/limb development that may be relevant to the radius/thumb anomaly of FA patients. Hatching embryos expressed fanc genes strongly in the oral epithelium, a site of squamous cell carcinomas in FA patients. Larval and adult zebrafish expressed fanc genes in proliferative regions of the brain, which may be related to microcephaly in FA. Mature ovaries and testes expressed fanc genes in specific stages of oocyte and spermatocyte development, which may be related to DNA repair during homologous recombination in meiosis and to infertility in human patients. The intestine strongly expressed some fanc genes specifically in proliferative zones. Our results show that zebrafish has a complete complement of fanc genes in single copy and that these genes are expressed in zebrafish embryos and adults in proliferative tissues that are often affected in FA patients. These results support the notion that zebrafish offers an attractive experimental system to help unravel mechanisms relevant not only

  13. The Fanconi anemia/BRCA gene network in zebrafish: Embryonic expression and comparative genomics

    International Nuclear Information System (INIS)

    Titus, Tom A.; Yan Yilin; Wilson, Catherine; Starks, Amber M.; Frohnmayer, Jonathan D.; Bremiller, Ruth A.; Canestro, Cristian; Rodriguez-Mari, Adriana; He Xinjun; Postlethwait, John H.

    2009-01-01

    Fanconi anemia (FA) is a genetic disease resulting in bone marrow failure, high cancer risks, and infertility, and developmental anomalies including microphthalmia, microcephaly, hypoplastic radius and thumb. Here we present cDNA sequences, genetic mapping, and genomic analyses for the four previously undescribed zebrafish FA genes (fanci, fancj, fancm, and fancn), and show that they reverted to single copy after the teleost genome duplication. We tested the hypothesis that FA genes are expressed during embryonic development in tissues that are disrupted in human patients by investigating fanc gene expression patterns. We found fanc gene maternal message, which can provide Fanc proteins to repair DNA damage encountered in rapid cleavage divisions. Zygotic expression was broad but especially strong in eyes, central nervous system and hematopoietic tissues. In the pectoral fin bud at hatching, fanc genes were expressed specifically in the apical ectodermal ridge, a signaling center for fin/limb development that may be relevant to the radius/thumb anomaly of FA patients. Hatching embryos expressed fanc genes strongly in the oral epithelium, a site of squamous cell carcinomas in FA patients. Larval and adult zebrafish expressed fanc genes in proliferative regions of the brain, which may be related to microcephaly in FA. Mature ovaries and testes expressed fanc genes in specific stages of oocyte and spermatocyte development, which may be related to DNA repair during homologous recombination in meiosis and to infertility in human patients. The intestine strongly expressed some fanc genes specifically in proliferative zones. Our results show that zebrafish has a complete complement of fanc genes in single copy and that these genes are expressed in zebrafish embryos and adults in proliferative tissues that are often affected in FA patients. These results support the notion that zebrafish offers an attractive experimental system to help unravel mechanisms relevant not only

  14. Role of Active Contraction and Tropomodulins in Regulating Actin Filament Length and Sarcomere Structure in Developing Zebrafish Skeletal Muscle.

    Science.gov (United States)

    Mazelet, Lise; Parker, Matthew O; Li, Mei; Arner, Anders; Ashworth, Rachel

    2016-01-01

    Whilst it is recognized that contraction plays an important part in maintaining the structure and function of mature skeletal muscle, its role during development remains undefined. In this study the role of movement in skeletal muscle maturation was investigated in intact zebrafish embryos using a combination of genetic and pharmacological approaches. An immotile mutant line (cacnb1 (ts25) ) which lacks functional voltage-gated calcium channels (dihydropyridine receptors) in the muscle and pharmacological immobilization of embryos with a reversible anesthetic (Tricaine), allowed the study of paralysis (in mutants and anesthetized fish) and recovery of movement (reversal of anesthetic treatment). The effect of paralysis in early embryos (aged between 17 and 24 hours post-fertilization, hpf) on skeletal muscle structure at both myofibrillar and myofilament level was determined using both immunostaining with confocal microscopy and small angle X-ray diffraction. The consequences of paralysis and subsequent recovery on the localization of the actin capping proteins Tropomodulin 1 & 4 (Tmod) in fish aged from 17 hpf until 42 hpf was also assessed. The functional consequences of early paralysis were investigated by examining the mechanical properties of the larval muscle. The length-force relationship, active and passive tension, was measured in immotile, recovered and control skeletal muscle at 5 and 7 day post-fertilization (dpf). Recovery of muscle function was also assessed by examining swimming patterns in recovered and control fish. Inhibition of the initial embryonic movements (up to 24 hpf) resulted in an increase in myofibril length and a decrease in width followed by almost complete recovery in both moving and paralyzed fish by 42 hpf. In conclusion, myofibril organization is regulated by a dual mechanism involving movement-dependent and movement-independent processes. The initial contractile event itself drives the localization of Tmod1 to its sarcomeric

  15. Role of active contraction and tropomodulins in regulating actin filament length and sarcomere structure in developing zebrafish skeletal muscle

    Directory of Open Access Journals (Sweden)

    Lise eMazelet

    2016-03-01

    Full Text Available Whilst it is recognised that contraction plays an important part in maintaining the structure and function of mature skeletal muscle, its role during development remains undefined. In this study the role of movement in skeletal muscle maturation was investigated in intact zebrafish embryos using a combination of genetic and pharmacological approaches. An immotile mutant line (cacnb1ts25 which lacks functional voltage-gated calcium channels (dihydropyridine receptors in the muscle and pharmacological immobilisation of embryos with a reversible anaesthetic (Tricaine, allowed the study of paralysis (in mutants and anaesthetised fish and recovery of movement (reversal of anaesthetic treatment. The effect of paralysis in early embryos (aged between 17-24 hours post fertilisation, hpf on skeletal muscle structure at both myofibrillar and myofilament level was determined using both immunostaining with confocal microscopy and small angle X-ray diffraction. The consequences of paralysis and subsequent recovery on the localisation of the actin capping proteins Tropomodulin 1 &4 (Tmod in fish aged from 17hpf until 42hpf was also assessed. The functional consequences of early paralysis were investigated by examining the mechanical properties of the larval muscle. The length-force relationship, active and passive tension, was measured in immotile, recovered and control skeletal muscle at 5 and 7 day post fertilisation (dpf. Recovery of muscle function was also assessed by examining swimming patterns in recovered and control fish. Inhibition of the initial embryonic movements (up to 24 hpf resulted in an increase in myofibril length and a decrease in width followed by almost complete recovery in both moving and paralysed fish by 42hpf. In conclusion, myofibril organisation is regulated by a dual mechanism involving movement-dependent and movement-independent processes. The initial contractile event itself drives the localisation of Tmod1 to its sarcomeric

  16. Hepassocin is required for hepatic outgrowth during zebrafish hepatogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Ming [Tianjin University, Department of Pharmaceutical Engineering, Tianjin 300072 (China); Beijing Institute of Radiation Medicine, Beijing 100850 (China); Yan, Hui [Beijing Institute of Pharmacology and Toxicology, Beijing 100850 (China); Yin, Rong-Hua [Beijing Institute of Radiation Medicine, Beijing 100850 (China); State Key Laboratory of Proteomics, Beijing 100850 (China); Wang, Qiang [Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China); Zhan, Yi-Qun; Yu, Miao; Ge, Chang-Hui; Li, Chang-Yan; Wang, Xiao-Hui [Beijing Institute of Radiation Medicine, Beijing 100850 (China); State Key Laboratory of Proteomics, Beijing 100850 (China); Ge, Zhi-Qiang [Tianjin University, Department of Pharmaceutical Engineering, Tianjin 300072 (China); Yang, Xiao-Ming, E-mail: xiaomingyang@sina.com [Tianjin University, Department of Pharmaceutical Engineering, Tianjin 300072 (China); Beijing Institute of Radiation Medicine, Beijing 100850 (China); State Key Laboratory of Proteomics, Beijing 100850 (China)

    2015-07-31

    Background & aims: Hepassocin (HPS) is a hepatotrophic growth factor that specifically stimulates hepatocyte proliferation and promotes liver regeneration after liver damage. In this paper, zebrafish were used to investigate the role of HPS in liver development. Methods and results: During zebrafish development, HPS expression is enriched in liver throughout hepatogenesis. Knockdown of HPS using its specific morpholino leads to a smaller liver phenotype. Further results showed that the HPS knockdown has no effect on the expression of the early endoderm marker gata6 and early hepatic marker hhex. In addition, results showed that the smaller-liver phenotype in HPS morphants was caused by suppression of cell proliferation, not induction of cell apoptosis. Conclusions: Current findings indicated that HPS is essential to the later stages of development in vertebrate liver organogenesis. - Highlights: • HPS is enriched in zebrafish liver and has strong similarities with other species. • Knocking down HPS with MOs results in small liver phenotype. • HPS depletion regulates liver outgrowth but not liver specification and budding. • HPS depletion causes hepatocyte proliferation arrest but not apoptosis induction.

  17. Hepassocin is required for hepatic outgrowth during zebrafish hepatogenesis

    International Nuclear Information System (INIS)

    Gao, Ming; Yan, Hui; Yin, Rong-Hua; Wang, Qiang; Zhan, Yi-Qun; Yu, Miao; Ge, Chang-Hui; Li, Chang-Yan; Wang, Xiao-Hui; Ge, Zhi-Qiang; Yang, Xiao-Ming

    2015-01-01

    Background & aims: Hepassocin (HPS) is a hepatotrophic growth factor that specifically stimulates hepatocyte proliferation and promotes liver regeneration after liver damage. In this paper, zebrafish were used to investigate the role of HPS in liver development. Methods and results: During zebrafish development, HPS expression is enriched in liver throughout hepatogenesis. Knockdown of HPS using its specific morpholino leads to a smaller liver phenotype. Further results showed that the HPS knockdown has no effect on the expression of the early endoderm marker gata6 and early hepatic marker hhex. In addition, results showed that the smaller-liver phenotype in HPS morphants was caused by suppression of cell proliferation, not induction of cell apoptosis. Conclusions: Current findings indicated that HPS is essential to the later stages of development in vertebrate liver organogenesis. - Highlights: • HPS is enriched in zebrafish liver and has strong similarities with other species. • Knocking down HPS with MOs results in small liver phenotype. • HPS depletion regulates liver outgrowth but not liver specification and budding. • HPS depletion causes hepatocyte proliferation arrest but not apoptosis induction

  18. Hypoxia-induced metastasis model in embryonic zebrafish

    DEFF Research Database (Denmark)

    Rouhi, Pegah; Jensen, Lasse D.; Cao, Ziquan

    2010-01-01

    Hypoxia facilitates tumor invasion and metastasis by promoting neovascularization and co-option of tumor cells in the peritumoral vasculature, leading to dissemination of tumor cells into the circulation. However, until recently, animal models and imaging technology did not enable monitoring...... of the early events of tumor cell invasion and dissemination in living animals. We recently developed a zebrafish metastasis model to dissect the detailed events of hypoxia-induced tumor cell invasion and metastasis in association with angiogenesis at the single-cell level. In this model, fluorescent Di......I-labeled human or mouse tumor cells are implanted into the perivitelline cavity of 48-h-old zebrafish embryos, which are subsequently placed in hypoxic water for 3 d. Tumor cell invasion, metastasis and pathological angiogenesis are detected under fluorescent microscopy in the living fish. The average...

  19. Optogenetics: a new enlightenment age for zebrafish neurobiology.

    Science.gov (United States)

    Del Bene, Filippo; Wyart, Claire

    2012-03-01

    Zebrafish became a model of choice for neurobiology because of the transparency of its brain and because of its amenability to genetic manipulation. In particular, at early stages of development the intact larva is an ideal system to apply optical techniques for deep imaging in the nervous system, as well as genetically encoded tools for targeting subsets of neurons and monitoring and manipulating their activity. For these applications,new genetically encoded optical tools, fluorescent sensors, and light-gated channels have been generated,creating the field of "optogenetics." It is now possible to monitor and control neuronal activity with minimal perturbation and unprecedented spatio-temporal resolution.We describe here the main achievements that have occurred in the last decade in imaging and manipulating neuronal activity in intact zebrafish larvae. We provide also examples of functional dissection of neuronal circuits achieved with the applications of these techniques in the visual and locomotor systems.

  20. Models of prey capture in larval fish

    NARCIS (Netherlands)

    Drost, M.R.

    1986-01-01

    The food uptake of larval carp and pike is described from high speed movies with synchronous lateral and ventral views.

    During prey intake by larval fishes the velocities of the created suction flow are high relative to their own size: 0.3 m/s for carp larvae of 6

  1. High environmental ammonia exposure has developmental-stage specific and long-term consequences on the cortisol stress response in zebrafish.

    Science.gov (United States)

    Williams, Tegan A; Bonham, Luke A; Bernier, Nicholas J

    2017-12-01

    The capacity for early life environmental stressors to induce programming effects on the endocrine stress response in fish is largely unknown. In this study we determined the effects of high environmental ammonia (HEA) exposure on the stress response in larval zebrafish, assessed the tolerance of embryonic and larval stages to HEA, and evaluated whether early life HEA exposure has long-term consequences on the cortisol response to a novel stressor. Exposure to 500-2000μM NH 4 Cl for 16h did not affect the gene expression of corticotropin-releasing factor (CRF) system components in 1day post-fertilization (dpf) embryos, but differentially increased crfa, crfb and CRF binding protein (crfbp) expression and stimulated both dose- and time-dependent increases in the whole body cortisol of 5dpf larvae. Pre-acclimation to HEA at 1dpf did not affect the cortisol response to a subsequent NH 4 Cl exposure at 5dpf. In contrast, pre-acclimation to HEA at 5dpf caused a small but significant reduction in the cortisol response to a second NH 4 Cl exposure at 10dpf. While continuous exposure to 500-2000μM NH 4 Cl between 0 and 5dpf had a modest effect on mean survival time, exposure to 400-1000μM NH 4 Cl between 10 and 14dpf decreased mean survival time in a dose-dependent manner. Moreover, pre-acclimation to HEA at 5dpf significantly decreased the risk of mortality to continuous NH 4 Cl exposure between 10 and 14dpf. Finally, while HEA at 1dpf did not affect the cortisol stress response to a novel vortex stressor at 5dpf, the same HEA treatment at 5dpf abolished vortex stressor-induced increases in whole body cortisol at 10 and 60dpf. Together these results show that the impact of HEA on the cortisol stress response during development is life-stage specific and closely linked to ammonia tolerance. Further, we demonstrate that HEA exposure at the larval stage can have persistent effects on the capacity to respond to stressors in later life. Copyright © 2017 Elsevier Inc. All

  2. [Canine peritoneal larval cestodosis caused by Mesocestoides spp. larval stages].

    Science.gov (United States)

    Häußler, T C; Peppler, C; Schmitz, S; Bauer, C; Hirzmann, J; Kramer, M

    2016-01-01

    In a female dog with unspecific clinical symptoms, sonography detected a hyperechoic mass in the middle abdomen and blood analysis a middle grade systemic inflammatory reaction. Laparotomy revealed a peritoneal larval cestodosis (PLC). The diagnosis of an infection with tetrathyridia of Mesocestoides spp. was confirmed by parasitological examination and molecularbiological analysis. Reduction of the intra-abdominal parasitic load as well as a high dose administration of fenbendazole over 3 months led to a successful treatment which could be documented sonographically and by decreased concentrations of C-reactive protein (CRP). Seven months after discontinuation of fenbendazole administration, PLC recurred, pre-empted by an elevation of serum CRP values. According to the literature a life-long fenbendazole treatment was initiated. In cases of unclear chronic granulomatous inflammations in the abdominal cavity in dogs, PLC should be considered. CRP concentration and sonographic examinations are suitable to control for treatment success and a possibly occurring relapse.

  3. Neurobehavioral impairments caused by developmental imidacloprid exposure in zebrafish.

    Science.gov (United States)

    Crosby, Emily B; Bailey, Jordan M; Oliveri, Anthony N; Levin, Edward D

    2015-01-01

    Neonicotinoid insecticides are becoming more widely applied as organophosphate (OP) insecticides are decreasing in use. Because of their relative specificity to insect nicotinic receptors, they are thought to have reduced risk of neurotoxicity in vertebrates. However, there is scant published literature concerning the neurobehavioral effects of developmental exposure of vertebrates to neonicotinoids. Using zebrafish, we investigated the neurobehavioral effects of developmental exposure to imidacloprid, a prototypic neonicotinoid pesticide. Nicotine was also administered for comparison. Zebrafish were exposed via immersion in aqueous solutions containing 45 μM or 60 μM of imidacloprid or nicotine (or vehicle control) from 4h to 5d post fertilization. The functional effects of developmental exposure to both imidacloprid and nicotine were assessed in larvae using an activity assay and during adolescence and adulthood using a battery of neurobehavioral assays, including assessment of sensorimotor response and habituation in a tactile startle test, novel tank swimming, and shoaling behavior. In larvae, developmental imidacloprid exposure at both doses significantly decreased swimming activity. The 5D strains of zebrafish were more sensitive to both nicotine and imidacloprid than the AB* strain. In adolescent and adult fish, developmental exposure to imidacloprid significantly decreased novel tank exploration and increased sensorimotor response to startle stimuli. While nicotine did not affect novel tank swimming, it increased sensorimotor response to startle stimuli at the low dose. No effects of either compound were found on shoaling behavior or habituation to a startling stimulus. Early developmental exposure to imidacloprid has both early-life and persisting effects on neurobehavioral function in zebrafish. Its developmental neurotoxicity should be further investigated. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. The role of Sox6 in zebrafish muscle fiber type specification.

    Science.gov (United States)

    Jackson, Harriet E; Ono, Yosuke; Wang, Xingang; Elworthy, Stone; Cunliffe, Vincent T; Ingham, Philip W

    2015-01-01

    The transcription factor Sox6 has been implicated in regulating muscle fiber type-specific gene expression in mammals. In zebrafish, loss of function of the transcription factor Prdm1a results in a slow to fast-twitch fiber type transformation presaged by ectopic expression of sox6 in slow-twitch progenitors. Morpholino-mediated Sox6 knockdown can suppress this transformation but causes ectopic expression of only one of three slow-twitch specific genes assayed. Here, we use gain and loss of function analysis to analyse further the role of Sox6 in zebrafish muscle fiber type specification. The GAL4 binary misexpression system was used to express Sox6 ectopically in zebrafish embryos. Cis-regulatory elements were characterized using transgenic fish. Zinc finger nuclease mediated targeted mutagenesis was used to analyse the effects of loss of Sox6 function in embryonic, larval and adult zebrafish. Zebrafish transgenic for the GCaMP3 Calcium reporter were used to assay Ca2+ transients in wild-type and mutant muscle fibres. Ectopic Sox6 expression is sufficient to downregulate slow-twitch specific gene expression in zebrafish embryos. Cis-regulatory elements upstream of the slow myosin heavy chain 1 (smyhc1) and slow troponin c (tnnc1b) genes contain putative Sox6 binding sites required for repression of the former but not the latter. Embryos homozygous for sox6 null alleles expressed tnnc1b throughout the fast-twitch muscle whereas other slow-specific muscle genes, including smyhc1, were expressed ectopically in only a subset of fast-twitch fibers. Ca2+ transients in sox6 mutant fast-twitch fibers were intermediate in their speed and amplitude between those of wild-type slow- and fast-twitch fibers. sox6 homozygotes survived to adulthood and exhibited continued misexpression of tnnc1b as well as smaller slow-twitch fibers. They also exhibited a striking curvature of the spine. The Sox6 transcription factor is a key regulator of fast-twitch muscle fiber differentiation

  5. Developmental Toxicity of Zinc Oxide Nanoparticles to Zebrafish (Danio rerio: A Transcriptomic Analysis.

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    Jin Soo Choi

    Full Text Available Zinc oxide nanoparticles (ZnO NPs are being utilized in an increasing number of fields and commercial applications. While their general toxicity and associated oxidative stress have been extensively studied, the toxicological pathways that they induce in developmental stages are still largely unknown. In this study, the developmental toxicity of ZnO NPs to embryonic/larval zebrafish was investigated. The transcriptional expression profiles induced by ZnO NPs were also investigated to ascertain novel genomic responses related to their specific toxicity pathway. Zebrafish embryos were exposed to 0.01, 0.1, 1, and 10 mg/L ZnO NPs for 96 h post-fertilization. The toxicity of ZnO NPs, based on their Zn concentration, was quite similar to that in embryonic/larval zebrafish exposed to corresponding ZnSO4 concentrations. Pericardial edema and yolk-sac edema were the principal malformations induced by ZnO NPs. Gene-expression profiling using microarrays demonstrated 689 genes that were differentially regulated (fold change >1.5 following exposure to ZnO NPs (498 upregulated, 191 downregulated. Several genes that were differentially regulated following ZnO NP exposure shared similar biological pathways with those observed with ZnSO4 exposure, but six genes (aicda, cyb5d1, edar, intl2, ogfrl2 and tnfsf13b associated with inflammation and the immune system responded specifically to ZnO NPs (either in the opposite direction or were unchanged in ZnSO4 exposure. Real-time reverse-transcription quantitative polymerase chain reaction confirmed that the responses of these genes to ZnO NPs were significantly different from their response to ZnSO4 exposure. ZnO NPs may affect genes related to inflammation and the immune system, resulting in yolk-sac edema and pericardia edema in embryonic/larval developmental stages. These results will assist in elucidating the mechanisms of toxicity of ZnO NPs during development of zebrafish.

  6. Subdivisions of the adult zebrafish pallium based on molecular marker analysis [version 2; referees: 2 approved, 1 approved with reservations

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    Julia Ganz

    2015-11-01

    Full Text Available Background: The telencephalon shows a remarkable structural diversity among vertebrates. In particular, the everted telencephalon of ray-finned fishes has a markedly different morphology compared to the evaginated telencephalon of all other vertebrates. This difference in development has hampered the comparison between different areas of the pallium of ray-finned fishes and the pallial nuclei of all other vertebrates. Various models of homology between pallial subdivisions in ray-finned fishes and the pallial nuclei in tetrapods have been proposed based on connectional, neurochemical, gene expression and functional data. However, no consensus has been reached so far. In recent years, the analysis of conserved developmental marker genes has assisted the identification of homologies for different parts of the telencephalon among several tetrapod species. Results: We have investigated the gene expression pattern of conserved marker genes in the adult zebrafish (Danio rerio pallium to identify pallial subdivisions and their homology to pallial nuclei in tetrapods. Combinatorial expression analysis of ascl1a, eomesa, emx1, emx2, emx3, and Prox1 identifies four main divisions in the adult zebrafish pallium. Within these subdivisions, we propose that Dm is homologous to the pallial amygdala in tetrapods and that the dorsal subdivision of Dl is homologous to part of the hippocampal formation in mouse. We have complemented this analysis be examining the gene expression of emx1, emx2 and emx3 in the zebrafish larval brain. Conclusions: Based on our gene expression data, we propose a new model of subdivisions in the adult zebrafish pallium and their putative homologies to pallial nuclei in tetrapods. Pallial nuclei control sensory, motor, and cognitive functions, like memory, learning and emotion. The identification of pallial subdivisions in the adult zebrafish and their homologies to pallial nuclei in tetrapods will contribute to the use of the zebrafish

  7. Antigen Uptake during Different Life Stages of Zebrafish (Danio rerio Using a GFP-Tagged Yersinia ruckeri.

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    Rozalia Korbut

    Full Text Available Immersion-vaccines (bacterins are routinely used for aquacultured rainbow trout to protect against Yersinia ruckeri (Yr. During immersion vaccination, rainbow trout take up and process the antigens, which induce protection. The zebrafish was used as a model organism to study uptake mechanisms and subsequent antigen transport in fish. A genetically modified Yr was developed to constitutively express green fluorescent protein (GFP and was used for bacterin production. Larval, juvenile and adult transparent zebrafish (tra:nac mutant received a bath in the bacterin for up to 30 minutes. Samples were taken after 1 min, 15 min, 30 min, 2 h, 12 h and 24 h. At each sampling point fish were used for live imaging of the uptake using a fluorescence stereomicroscope and for immunohistochemistry (IHC. In adult fish, the bacterin could be traced within 30 min in scale pockets, skin, oesophagus, intestine and fins. Within two hours post bath (pb Yr-antigens were visible in the spleen and at 24 h in liver and kidney. Bacteria were associated with the gills, but uptake at this location was limited. Antigens were rarely detected in the blood and never in the nares. In juvenile fish uptake of the bacterin was seen in the intestine 30 min pb and in the nares 2 hpb but never in scale pockets. Antigens were detected in the spleen 12 hpb. Zebrafish larvae exhibited major Yr uptake only in the mid-intestine enterocytes 24 hpb. The different life stages of zebrafish varied with regard to uptake locations, however the gut was consistently a major uptake site. Zebrafish and rainbow trout tend to have similar uptake mechanisms following immersion or bath vaccination, which points towards zebrafish as a suitable model organism for this aquacultured species.

  8. Developmental toxicity of cartap on zebrafish embryos.

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    Zhou, Shengli; Dong, Qiaoxiang; Li, Shaonan; Guo, Jiangfeng; Wang, Xingxing; Zhu, Guonian

    2009-12-13

    Cartap is a widely used insecticide which belongs to a member of nereistoxin derivatives and acts on nicotinic acetylcholine receptor site. Its effects on aquatic species are of grave concern. To explore the potential developmental toxicity of cartap, zebrafish embryos were continually exposed, from 0.5 to 144h post-fertilization, to a range of concentrations of 25-1000microg/l. Results of the experiment indicated that cartap concentrations of 100microg/l and above negatively affected embryo survival and hatching success. Morphological analysis uncovered a large suite of abnormalities such as less melanin pigmentation, wavy notochord, crooked trunk, fuzzy somites, neurogenesis defects and vasculature defects. The most sensitive organ was proved to be the notochord which displayed defects at concentrations as low as 25microg/l. Both sensitivity towards exposure and localization of the defect were stage specific. To elucidate mechanisms concerning notochord, pigmentation, and hatching defects, enzyme assay, RT Q-PCR, and different exposure strategies were performed. For embryos with hatching failure, chorion was verified not to be digested, while removing cartap from exposure at early pre-hatching stage could significantly increase the hatching success. However, cartap was proved, via vitro assay, to have no effect on proteolytic activity of hatching enzyme. These findings implied that the secretion of hatching enzyme might be blocked. We also revealed that cartap inhibited the activity of melanogenic enzyme tyrosinase and matrix enzyme lysyl oxidase and induced expression of their genes. These suggested that cartap could impaired melanin pigmentation of zebrafish embryos through inhibiting tyrosinase activity, while inhibition of lysyl oxidase activity was responsible for notochord undulation, which subsequently caused somite defect, and at least partially responsible for defects in vasculature and neurogenesis.

  9. Innate Color Preference of Zebrafish and Its Use in Behavioral Analyses.

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    Park, Jong-Su; Ryu, Jae-Ho; Choi, Tae-Ik; Bae, Young-Ki; Lee, Suman; Kang, Hae Jin; Kim, Cheol-Hee

    2016-10-01

    Although innate color preference of motile organisms may provide clues to behavioral biases, it has remained a longstanding question. In this study, we investigated innate color preference of zebrafish larvae. A cross maze with different color sleeves around each arm was used for the color preference test (R; red, G; green, B; blue, Y; yellow). The findings showed that 5 dpf zebrafish larvae preferred blue over other colors (B > R > G > Y). To study innate color recognition further, tyrosinase mutants were generated using CRISPR/Cas9 system. As a model for oculocutaneous albinism (OCA) and color vision impairment, tyrosinase mutants demonstrated diminished color sensation, indicated mainly by hypopigmentation of the retinal pigment epithelium (RPE). Due to its relative simplicity and ease, color preference screening using zebrafish larvae is suitable for high-throughput screening applications. This system may potentially be applied to the analysis of drug effects on larval behavior or the detection of sensory deficits in neurological disorder models, such as autism-related disorders, using mutant larvae generated by the CRISPR/Cas9 technique.

  10. Toxicity of zinc oxide nanoparticles to zebrafish embryo: a physicochemical study of toxicity mechanism

    International Nuclear Information System (INIS)

    Bai Wei; Zhang Zhiyong; Tian Wenjing; He Xiao; Ma Yuhui; Zhao Yuliang; Chai Zhifang

    2010-01-01

    The biological impact of engineered nanomaterials released into the aquatic environment is a major concern. In this work, the properties of ZnO nanoparticles (nano-ZnO, 30 nm) were characterized in a water suspension (E3 medium), and a zebrafish 96-h post fertilization (hpf) embryo-larval test was performed to assess the toxicity of nano-ZnO suspension. Nano-ZnO was found to readily form aggregates with different sizes; small aggregates (142.4-517.7 nm) were still suspended in E3 medium, but large aggregates (>1 μm) quickly deposited on the bottom of 24-well plates; nano-ZnO was partially dissolved to Zn species (Zn (dis) ) in E3 medium. In the nano-ZnO suspension, small aggregates, Zn (dis) , and large aggregates might jointly exert influence on the development of zebrafish embryos. The embryo toxicity test revealed that nano-ZnO killed zebrafish embryos (50 and 100 mg/L), retarded the embryo hatching (1-25 mg/L), reduced the body length of larvae, and caused tail malformation after the 96 hpf exposure. Zn (dis) only partially contributed to the toxicity of nano-ZnO. This research highlights the need to further investigate the ecotoxicity of nano-ZnO in the water environment.

  11. Forward Genetic Screening Using Behavioral Tests in Zebrafish: A Proof of Concept Analysis of Mutants.

    Science.gov (United States)

    Gerlai, Robert; Poshusta, Tanya L; Rampersad, Mindy; Fernandes, Yohaan; Greenwood, Tammy M; Cousin, Margot A; Klee, Eric W; Clark, Karl J

    2017-01-01

    The zebrafish enjoys several advantages over other model organisms. It is small, easy to maintain, prolific, and numerous genetic tools are available for it. For example, forward genetic screens have allowed investigators to identify important genes potentially involved in a variety of functions from embryogenesis to cancer. However, despite its sophisticated behavioral repertoire, behavioral methods have rarely been utilized in forward genetic screens. Here, we employ a two-tiered strategy, a proof of concept study, to explore the feasibility of behavioral screens. We generated mutant lines using transposon-based insertional mutagenesis, allowing us to bias mutant selection with target genes expressed within the brain. Furthermore, we employed an efficient and fast behavioral pre-selection in which we investigated the locomotory response of 5-day post-fertilization old larval fish to hyperosmotic shock. Based on this assay, we selected five lines for our lower throughput secondary adult behavioral screen. The latter screen utilized tests in which computer animated image presentation and video-tracking-based automated quantification of behavior allowed us to compare heterozygous zebrafish with their wild-type siblings on their responses to a variety of stimuli. We found significant mutation induced adult behavioral alterations in 4 out of the 5 lines analyzed, including changes in response to social or fear inducing stimuli, to handling and novelty, or in habituation to novelty. We discuss the pros and cons of behavioral phenotyping and of the use of different forward genetic methods in biomedical research with zebrafish.

  12. Behavioural screening of zebrafish using neuroactive traditional Chinese medicine prescriptions and biological targets

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    Wang, Ya-Nan; Hou, Yuan-Yuan; Sun, Ming-Zhu; Zhang, Chun-Yang; Bai, Gang; Zhao, Xin; Feng, Xi-Zeng

    2014-06-01

    The mechanism of the therapeutic action of antidepressants remains uncertain in traditional Chinese medicine (TCM). In this study, we selected 7 classical TCM prescriptions and utilised an automatic video-tracking system to monitor the rest/wake behaviour of larval zebrafish at 4 days post-fertilisation (dpf) for 48 hours. We found that the curative effects of the prescriptions were dose-dependent. K-means clustering was performed according to the shared behavioural phenotypes of the zebrafish. The results revealed that the rest/wake behavioural profiles induced by the same class of prescriptions were similar. A correlation analysis was conducted between the TCM prescriptions and the known compounds. The results showed that the TCM prescriptions correlated well with some well-known compounds. Therefore, we predicted that they may share a similar mechanism of action. This paper describes the first study to combine TCM research with zebrafish rest/wake behaviour in vivo and presents a powerful approach for the discovery of the mechanism of action of TCM prescriptions.

  13. Usefulness of zebrafish larvae to evaluate drug-induced functional and morphological renal tubular alterations.

    Science.gov (United States)

    Gorgulho, Rita; Jacinto, Raquel; Lopes, Susana S; Pereira, Sofia A; Tranfield, Erin M; Martins, Gabriel G; Gualda, Emilio J; Derks, Rico J E; Correia, Ana C; Steenvoorden, Evelyne; Pintado, Petra; Mayboroda, Oleg A; Monteiro, Emilia C; Morello, Judit

    2018-01-01

    Prediction and management of drug-induced renal injury (DIRI) rely on the knowledge of the mechanisms of drug insult and on the availability of appropriate animal models to explore it. Zebrafish (Danio rerio) offers unique advantages for assessing DIRI because the larval pronephric kidney has a high homology with its human counterpart and it is fully mature at 3.5 days post-fertilization. Herein, we aimed to evaluate the usefulness of zebrafish larvae as a model of renal tubular toxicity through a comprehensive analysis of the renal alterations induced by the lethal concentrations for 10% of the larvae for gentamicin, paracetamol and tenofovir. We evaluated drug metabolic profile by mass spectrometry, renal function with the inulin clearance assay, the 3D morphology of the proximal convoluted tubule by two-photon microscopy and the ultrastructure of proximal convoluted tubule mitochondria by transmission electron microscopy. Paracetamol was metabolized by conjugation and oxidation with further detoxification with glutathione. Renal clearance was reduced with gentamicin and paracetamol. Proximal tubules were enlarged with paracetamol and tenofovir. All drugs induced mitochondrial alterations including dysmorphic shapes ("donuts", "pancakes" and "rods"), mitochondrial swelling, cristae disruption and/or loss of matrix granules. These results are in agreement with the tubular effects of gentamicin, paracetamol and tenofovir in man and demonstrate that zebrafish larvae might be a good model to assess functional and structural damage associated with DIRI.

  14. Bioenergetics models to estimate numbers of larval lampreys consumed by smallmouth bass in Elk Creek, Oregon

    Science.gov (United States)

    Schultz, Luke; Heck, Michael; Kowalski, Brandon M; Eagles-Smith, Collin A.; Coates, Kelly C.; Dunham, Jason B.

    2017-01-01

    Nonnative fishes have been increasingly implicated in the decline of native fishes in the Pacific Northwest. Smallmouth Bass Micropterus dolomieu were introduced into the Umpqua River in southwest Oregon in the early 1960s. The spread of Smallmouth Bass throughout the basin coincided with a decline in counts of upstream-migrating Pacific Lampreys Entosphenus tridentatus. This suggested the potential for ecological interactions between Smallmouth Bass and Pacific Lampreys, as well as freshwater-resident Western Brook Lampreys Lampetra richardsoni. To evaluate the potential effects of Smallmouth Bass on lampreys, we sampled diets of Smallmouth Bass and used bioenergetics models to estimate consumption of larval lampreys in a segment of Elk Creek, a tributary to the lower Umpqua River. We captured 303 unique Smallmouth Bass (mean: 197 mm and 136 g) via angling in July and September. We combined information on Smallmouth Bass diet and energy density with other variables (temperature, body size, growth, prey energy density) in a bioenergetics model to estimate consumption of larval lampreys. Larval lampreys were found in 6.2% of diet samples, and model estimates indicated that the Smallmouth Bass we captured consumed 925 larval lampreys in this 2-month study period. When extrapolated to a population estimate of Smallmouth Bass in this segment, we estimated 1,911 larval lampreys were consumed between July and September. Although the precision of these estimates was low, this magnitude of consumption suggests that Smallmouth Bass may negatively affect larval lamprey populations.

  15. Modeling Myeloid Malignancies Using Zebrafish

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    Kathryn S. Potts

    2017-12-01

    Full Text Available Human myeloid malignancies represent a substantial disease burden to individuals, with significant morbidity and death. The genetic underpinnings of disease formation and progression remain incompletely understood. Large-scale human population studies have identified a high frequency of potential driver mutations in spliceosomal and epigenetic regulators that contribute to malignancies, such as myelodysplastic syndromes (MDS and leukemias. The high conservation of cell types and genes between humans and model organisms permits the investigation of the underlying mechanisms of leukemic development and potential therapeutic testing in genetically pliable pre-clinical systems. Due to the many technical advantages, such as large-scale screening, lineage-tracing studies, tumor transplantation, and high-throughput drug screening approaches, zebrafish is emerging as a model system for myeloid malignancies. In this review, we discuss recent advances in MDS and leukemia using the zebrafish model.

  16. The inflammatory bowel disease (IBD susceptibility genes NOD1 and NOD2 have conserved anti-bacterial roles in zebrafish

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    Stefan H. Oehlers

    2011-11-01

    Inflammatory bowel disease (IBD, in the form of Crohn’s disease (CD or ulcerative colitis (UC, is a debilitating chronic immune disorder of the intestine. A complex etiology resulting from dysfunctional interactions between the intestinal immune system and its microflora, influenced by host genetic susceptibility, makes disease modeling challenging. Mutations in NOD2 have the highest disease-specific risk association for CD, and a related gene, NOD1, is associated with UC. NOD1 and NOD2 encode intracellular bacterial sensor proteins acting as innate immune triggers, and represent promising therapeutic targets. The zebrafish has the potential to aid in modeling genetic and environmental aspects of IBD pathogenesis. Here, we report the characterization of the Nod signaling components in the zebrafish larval intestine. The nod1 and nod2 genes are expressed in intestinal epithelial cells and neutrophils together with the Nod signaling pathway genes ripk2, a20, aamp, cd147, centaurin b1, erbin and grim-19. Using a zebrafish embryo Salmonella infection model, morpholino-mediated depletion of Nod1 or Nod2 reduced the ability of embryos to control systemic infection. Depletion of Nod1 or Nod2 decreased expression of dual oxidase in the intestinal epithelium and impaired the ability of larvae to reduce intracellular bacterial burden. This work highlights the potential use of zebrafish larvae in the study of components of IBD pathogenesis.

  17. The Zebrafish as a New Model for the In Vivo Study of Shigella flexneri Interaction with Phagocytes and Bacterial Autophagy

    Science.gov (United States)

    Mostowy, Serge; Boucontet, Laurent; Mazon Moya, Maria J.; Sirianni, Andrea; Boudinot, Pierre; Hollinshead, Michael; Cossart, Pascale; Herbomel, Philippe; Levraud, Jean-Pierre; Colucci-Guyon, Emma

    2013-01-01

    Autophagy, an ancient and highly conserved intracellular degradation process, is viewed as a critical component of innate immunity because of its ability to deliver cytosolic bacteria to the lysosome. However, the role of bacterial autophagy in vivo remains poorly understood. The zebrafish (Danio rerio) has emerged as a vertebrate model for the study of infections because it is optically accessible at the larval stages when the innate immune system is already functional. Here, we have characterized the susceptibility of zebrafish larvae to Shigella flexneri, a paradigm for bacterial autophagy, and have used this model to study Shigella-phagocyte interactions in vivo. Depending on the dose, S. flexneri injected in zebrafish larvae were either cleared in a few days or resulted in a progressive and ultimately fatal infection. Using high resolution live imaging, we found that S. flexneri were rapidly engulfed by macrophages and neutrophils; moreover we discovered a scavenger role for neutrophils in eliminating infected dead macrophages and non-immune cell types that failed to control Shigella infection. We observed that intracellular S. flexneri could escape to the cytosol, induce septin caging and be targeted to autophagy in vivo. Depletion of p62 (sequestosome 1 or SQSTM1), an adaptor protein critical for bacterial autophagy in vitro, significantly increased bacterial burden and host susceptibility to infection. These results show the zebrafish larva as a new model for the study of S. flexneri interaction with phagocytes, and the manipulation of autophagy for anti-bacterial therapy in vivo. PMID:24039575

  18. Identification and characterization of two novel cytosolic sulfotransferases, SULT1 ST7 and SULT1 ST8, from zebrafish

    Energy Technology Data Exchange (ETDEWEB)

    Liu, T.-A. [Department of Pharmacology, College of Pharmacy, University of Toledo, Toledo, OH 43606 (United States); Department of Biological Science and Technology, College of Biological Science and Technology, National Chiao Tung University, Hsinchu, Taiwan (China); Bhuiyan, Shakhawat [Division of Arts and Sciences, Jarvis Christian College, Hawkins, TX 75765 (United States); Snow, Rhodora [School of Mathematics and Science, J. Sargeant Reynolds Community College, Richmond, VA 23285 (United States); Yasuda, Shin; Yasuda, Tomoko [Department of Pharmacology, College of Pharmacy, University of Toledo, Toledo, OH 43606 (United States); Yang, Y.-S. [Department of Biological Science and Technology, College of Biological Science and Technology, National Chiao Tung University, Hsinchu, Taiwan (China); Williams, Frederick E.; Liu, M.-Y.; Suiko, Masahito [Department of Pharmacology, College of Pharmacy, University of Toledo, Toledo, OH 43606 (United States); Carter, Glendora [School of Mathematics and Science, J. Sargeant Reynolds Community College, Richmond, VA 23285 (United States); Liu, M.-C. [Department of Pharmacology, College of Pharmacy, University of Toledo, Toledo, OH 43606 (United States)], E-mail: ming.liu@utoledo.edu

    2008-08-29

    Cytosolic sulfotransferases (SULTs) constitute a family of Phase II detoxification enzymes that are involved in the protection against potentially harmful xenobiotics as well as the regulation and homeostasis of endogenous compounds. Compared with humans and rodents, the zebrafish serves as an excellent model for studying the role of SULTs in the detoxification of environmental pollutants including environmental estrogens. By searching the expressed sequence tag database, two zebrafish cDNAs encoding putative SULTs were identified. Sequence analysis indicated that these two putative zebrafish SULTs belong to the SULT1 gene family. The recombinant form of these two novel zebrafish SULTs, designated SULT1 ST7 and SULT1 ST8, were expressed using the pGEX-2TK glutathione S-transferase (GST) gene fusion system and purified from transformed BL21 (DE3) cells. Purified GST-fusion protein form of SULT1 ST7 and SULT1 ST8 exhibited strong sulfating activities toward environmental estrogens, particularly hydroxylated polychlorinated biphenyls (PCBs), among various endogenous and xenobiotic compounds tested as substrates. pH-dependence experiments showed that SULT1 ST7 and SULT1 ST8 displayed pH optima at 6.5 and 8.0, respectively. Kinetic parameters of the two enzymes in catalyzing the sulfation of catechin and chlorogenic acid as well as 3-chloro-4-biphenylol were determined. Developmental expression experiments revealed distinct patterns of expression of SULT1 ST7 and SULT1 ST8 during embryonic development and throughout the larval stage onto maturity.

  19. Histological Characterization of the Dicer1 Mutant Zebrafish Retina

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    Saeed Akhtar

    2015-01-01

    Full Text Available DICER1, a multidomain RNase III endoribonuclease, plays a critical role in microRNA (miRNA and RNA-interference (RNAi functional pathways. Loss of Dicer1 affects different developmental processes. Dicer1 is essential for retinal development and maintenance. DICER1 was recently shown to have another function of silencing the toxicity of Alu RNAs in retinal pigment epithelium (RPE cells, which are involved in the pathogenesis of age related macular degeneration. In this study, we characterized a Dicer1 mutant fish line, which carries a nonsense mutation (W1457Ter induced by N-ethyl-N-nitrosourea mutagenesis. Zebrafish DICER1 protein is highly conserved in the evolution. Zebrafish Dicer1 is expressed at the earliest stages of zebrafish development and persists into late developmental stages; it is widely expressed in adult tissues. Homozygous Dicer1 mutant fish (DICER1W1457Ter/W1457Ter have an arrest in early growth with significantly smaller eyes and are dead at 14–18 dpf. Heterozygous Dicer1 mutant fish have similar retinal structure to that of control fish; the retinal pigment epithelium (RPE cells are normal with no sign of degeneration at the age of 20 months.

  20. Comparative metal oxide nanoparticle toxicity using embryonic zebrafish

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    Leah C. Wehmas

    2015-01-01

    Full Text Available Engineered metal oxide nanoparticles (MO NPs are finding increasing utility in the medical field as anticancer agents. Before validation of in vivo anticancer efficacy can occur, a better understanding of whole-animal toxicity is required. We compared the toxicity of seven widely used semiconductor MO NPs made from zinc oxide (ZnO, titanium dioxide, cerium dioxide and tin dioxide prepared in pure water and in synthetic seawater using a five-day embryonic zebrafish assay. We hypothesized that the toxicity of these engineered MO NPs would depend on physicochemical properties. Significant agglomeration of MO NPs in aqueous solutions is common making it challenging to associate NP characteristics such as size and charge with toxicity. However, data from our agglomerated MO NPs suggests that the elemental composition and dissolution potential are major drivers of toxicity. Only ZnO caused significant adverse effects of all MO particles tested, and only when prepared in pure water (point estimate median lethal concentration = 3.5–9.1 mg/L. This toxicity was life stage dependent. The 24 h toxicity increased greatly (∼22.7 fold when zebrafish exposures started at the larval life stage compared to the 24 h toxicity following embryonic exposure. Investigation into whether dissolution could account for ZnO toxicity revealed high levels of zinc ion (40–89% of total sample were generated. Exposure to zinc ion equivalents revealed dissolved Zn2+ may be a major contributor to ZnO toxicity.

  1. Mitochondrial behavior during oogenesis in zebrafish: a confocal microscopy analysis.

    Science.gov (United States)

    Zhang, Yong-Zhong; Ouyang, Ying-Chun; Hou, Yi; Schatten, Heide; Chen, Da-Yuan; Sun, Qing-Yuan

    2008-03-01

    The behavior of mitochondria during early oogenesis remains largely unknown in zebrafish. We used three mitochondrial probes (Mito Tracker Red CMXRos, Mito Tracker Green FM, and JC-1) to stain early zebrafish oocyte mitochondria, and confocal microscopy to analyze mitochondrial aggregation and distribution. By using fluorescence recovery after photobleaching (FRAP), we traced mitochondrial movement. The microtubule assembly inhibitor nocodazole and microfilament inhibitor cytochalasin B (CB) were used to analyze the role of microtubules and microfilaments on mitochondrial movement. By using the dual emission probe, JC-1, and oxidative phosphorylation uncoupler, carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone (FCCP), we determined the distribution of active and inactive (low-active) mitochondria. Green/red fluorescence ratios of different sublocations in different oocyte groups stained by JC-1 were detected in merged (green and red) images. Our results showed that mitochondria exhibited a unique distribution pattern in early zebrafish oocytes. They tended to aggregate into large clusters in early stage I oocytes, but in a threadlike state in latter stage I oocytes. We detected a lower density mitochondrial area and a higher density mitochondrial area on opposite sides of the germinal vesicle. The green/red fluorescence ratios in different sublocations in normal oocytes were about 1:1. This implies that active mitochondria were distributed in all sublocations. FCCP treatment caused significant increases in the ratios. CB and nocodazole treatment caused an increase of the ratios in clusters and mitochondrial cloud, but not in dispersed areas. Mitochondria in different sublocations underwent fast dynamic movement. Inhibition or disruption of microtubules or microfilaments resulted in even faster mitochondrial free movement.

  2. Toxicological effects of graphene oxide on adult zebrafish (Danio rerio)

    Energy Technology Data Exchange (ETDEWEB)

    Souza, Jaqueline P., E-mail: souza.jaqueline@gmail.com; Baretta, Jéssica F.; Santos, Fabrício; Paino, Ieda M.M.; Zucolotto, Valtencir

    2017-05-15

    Highlights: • Graphene oxide exposure caused apoptotic and necrotic stages in zebrafish gill cells. • Graphene oxide induced reactive oxygen generation in zebrafish gill cells. • Gill and liver tissues suffered injuries after graphene oxide chronic exposure. • Zebrafish blood cells did not present DNA damages after graphene oxide exposure. - Abstract: Graphene exhibits unique physical and chemical properties that facilitate its application in many fields, including electronics and biomedical areas. However, the use of graphene and its derivatives could result in accumulation in aquatic environments, and the risks posed by these compounds for organisms are not completely understood. In this study, we investigated the effects of graphene oxide (GO) on adult zebrafish (Danio rerio). Experimental fish were exposed to 2, 10 or 20 mg L{sup −1} GO, and the cytotoxicity, genotoxicity and oxidative stress were assessed. The morphology of the gills and liver tissues was also analyzed. Graphene oxide exposure led to an increase in the number of gill cells that were in early apoptotic and necrotic stages, but genotoxicity was not observed in blood cells. We also observed the generation of Reactive Oxygen Species (ROS) in gill cells. Structural analysis revealed injuries to gill tissues, including a dilated marginal channel, lamellar fusion, clubbed tips, swollen mucocytes, epithelial lifting, aneurysms, and necrosis. Liver tissues also presented lesions such as peripherally located nuclei. Furthermore, hepatocytes exhibited a non-uniform shape, picnotic nuclei, vacuole formation, cell rupture, and necrosis. Our results showed that sub-lethal doses of graphene oxide could be harmful to fish species and thus represent risks for the aquatic food chain.

  3. The zebrafish spi1 promoter drives myeloid-specific expression in stable transgenic fish

    NARCIS (Netherlands)

    Ward, AC; McPhee, DO; Condron, MM; Varma, S; Cody, SH; Onnebo, SMN; Paw, BH; Zon, LI; Lieschke, GJ

    2003-01-01

    The spi1 (pu.1) gene has recently been identified as a useful marker of early myeloid cells in zebrafish. To enhance the versatility of this organism as a model for studying myeloid development, the promoter of this gene has been isolated and characterized. Transient transgenesis revealed that a 5.3

  4. Effect of corticosterone on larval growth, antipredator behaviour and metamorphosis of Hylarana indica.

    Science.gov (United States)

    Kulkarni, P S; Gramapurohit, N P

    2017-09-15

    Corticosterone (CORT), a principal glucocorticoid in amphibians, is known to regulate diverse physiological processes including growth and metamorphosis of anuran tadpoles. Environmental stressors activate the neuroendocrine stress axis (hypothalamus-pituitary-interrenal axis, HPI) leading to an acute increase in CORT, which in turn, helps in coping with particular stress. However, chronic increase in CORT can negatively affect other physiological processes such as growth and metamorphosis. Herein, we studied the effect of exogenous CORT on larval growth, antipredator behaviour and metamorphic traits of Hylarana indica. Embryonic exposure to 5 or 20μg/L CORT did not affect their development, hatching duration as well as larval growth and metamorphosis. Exposure of tadpoles to 10 or 20μg/L CORT throughout larval development caused slower growth and development leading to increased body mass at stage 37. However, body and tail morphology of tadpoles was not affected. Interestingly, larval exposure to 5, 10 or 20μg/L CORT enhanced their antipredator response against kairomones in a concentration-dependent manner. Further, larval exposure to increasing concentrations of CORT resulted in the emergence of heavier froglets at 10 and 20μg/L while, delaying metamorphosis at all concentrations. Interestingly, the heavier froglets had shorter hindlimbs and consequently shorter jump distances. Tadpoles exposed to 20μg/L CORT during early, mid or late larval stages grew and developed slowly but tadpole morphology was not altered. Interestingly, exposure during early or mid-larval stages resulted in an enhanced antipredator response. These individuals metamorphosed later but at higher body mass while SVL was unaffected. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Embryonic and larval development in the Midas cichlid fish species flock (Amphilophus spp.): a new evo-devo model for the investigation of adaptive novelties and species differences.

    Science.gov (United States)

    Kratochwil, Claudius F; Sefton, Maggie M; Meyer, Axel

    2015-02-26

    Central American crater lake cichlid fish of the Midas species complex (Amphilophus spp.) are a model system for sympatric speciation and fast ecological diversification and specialization. Midas cichlids have been intensively analyzed from an ecological and morphological perspective. Genomic resources such as transcriptomic and genomic data sets, and a high-quality draft genome are available now. Many ecologically relevant species-specific traits and differences such as pigmentation and cranial morphology arise during development. Detailed descriptions of the early development of the Midas cichlid in particular, will help to investigate the ontogeny of species differences and adaptations. We describe the embryonic and larval development of the crater lake cichlid, Amphilophus xiloaensis, until seven days after fertilization. Similar to previous studies on teleost development, we describe six periods of embryogenesis - the zygote, cleavage, blastula, gastrula, segmentation, and post-hatching period. Furthermore, we define homologous stages to well-described teleost models such as medaka and zebrafish, as well as other cichlid species such as the Nile tilapia and the South American cichlid Cichlasoma dimerus. Key morphological differences between the embryos of Midas cichlids and other teleosts are highlighted and discussed, including the presence of adhesive glands and different early chromatophore patterns, as well as variation in developmental timing. The developmental staging of the Midas cichlid will aid researchers in the comparative investigation of teleost ontogenies. It will facilitate comparative developmental biological studies of Neotropical and African cichlid fish in particular. In the past, the species flocks of the African Great Lakes have received the most attention from researchers, but some lineages of the 300-400 species of Central American lakes are fascinating model systems for adaptive radiation and rapid phenotypic evolution. The availability

  6. Soundscapes and Larval Settlement: Larval Bivalve Responses to Habitat-Associated Underwater Sounds.

    Science.gov (United States)

    Eggleston, David B; Lillis, Ashlee; Bohnenstiehl, DelWayne R

    2016-01-01

    We quantified the effects of habitat-associated sounds on the settlement response of two species of bivalves with contrasting habitat preferences: (1) Crassostrea virginicia (oyster), which prefers to settle on other oysters, and (2) Mercenaria mercenaria (clam), which settles on unstructured habitats. Oyster larval settlement in the laboratory was significantly higher when exposed to oyster reef sound compared with either off-reef or no-sound treatments. Clam larval settlement did not vary according to sound treatments. Similar to laboratory results, field experiments showed that oyster larval settlement in "larval housings" suspended above oyster reefs was significantly higher compared with off-reef sites.

  7. Zebrafish kidney phagocytes utilize macropinocytosis and Ca+-dependent endocytic mechanisms.

    Directory of Open Access Journals (Sweden)

    Claudia Hohn

    Full Text Available BACKGROUND: The innate immune response constitutes the first line of defense against invading pathogens and consists of a variety of immune defense mechanisms including active endocytosis by macrophages and granulocytes. Endocytosis can be used as a reliable measure of selective and non-selective mechanisms of antigen uptake in the early phase of an immune response. Numerous assays have been developed to measure this response in a variety of mammalian and fish species. The small size of the zebrafish has prevented the large-scale collection of monocytes/macrophages and granulocytes for these endocytic assays. METHODOLOGY/PRINCIPAL FINDINGS: Pooled zebrafish kidney hematopoietic tissues were used as a source of phagocytic cells for flow-cytometry based endocytic assays. FITC-Dextran, Lucifer Yellow and FITC-Edwardsiella ictaluri were used to evaluate selective and non-selective mechanisms of uptake in zebrafish phagocytes. CONCLUSIONS/SIGNIFICANCE: Zebrafish kidney phagocytes characterized as monocytes/macrophages, neutrophils and lymphocytes utilize macropinocytosis and Ca(2+-dependant endocytosis mechanisms of antigen uptake. These cells do not appear to utilize a mannose receptor. Heat-killed Edwardsiella ictaluri induces cytoskeletal interactions for internalization in zebrafish kidney monocytes/macrophages and granulocytes. The proposed method is easy to implement and should prove especially useful in immunological, toxicological and epidemiological research.

  8. Impact of CdSe/ZnS quantum dots on the development of zebrafish embryos

    Science.gov (United States)

    Lei, Yong; Xiao, Qi; Huang, Shan; Xu, Wansu; Zhang, Zhe; He, Zhike; Liu, Yi; Deng, Fengjiao

    2011-12-01

    Due to their unique fluorescent characteristics, quantum dots (QDs) have been successfully applied in the fields of biotechnology and medicine, but there is very limited information regarding their biodistribution and chronic toxicity in vivo. In this article, the biological behavior and toxic effects of mercaptoacetic acid-CdSe/ZnS QDs (MAA-QDs) in developing zebrafish embryos were investigated by in vivo tests. The MAA-QDs were introduced into zebrafish through microinjection at early stage. The results showed that the MAA-QDs at certain concentrations influenced the survival of zebrafish embryos, but treated embryos without developmental defects were also observed. MAA-QDs injected into the cytoplasm at the one-cell stage were allocated to progeny blastoderm cells during proliferation and almost never entered the yolk. The formation of notochord and primordial germ cells with normal morphologies was detected in the treated embryos by whole-mount in situ hybridization. Furthermore, traces of the element cadmium were mainly discovered in the tissue of liver and kidney of 3-month-old-treated zebrafish by quantitative assessment with inductively coupled plasma mass spectrometry. Thus, we hypothesized that low concentration MAA-QDs have chronic toxicities when they were delivered into zebrafish organs.

  9. Non-invasive imaging of zebrafish with spinal deformities using optical coherence tomography: a preliminary study

    Science.gov (United States)

    Bernstein, Liane; Beaudette, Kathy; Patten, Kessen; Beaulieu-Ouellet, Émilie; Strupler, Mathias; Moldovan, Florina; Boudoux, Caroline

    2013-03-01

    A zebrafish model has recently been introduced to study various genetic mutations that could lead to spinal deformities such as scoliosis. However, current imaging techniques make it difficult to perform longitudinal studies of this condition in zebrafish, especially in the early stages of development. The goal of this project is to determine whether optical coherence tomography (OCT) is a viable non-invasive method to image zebrafish exhibiting spinal deformities. Images of both live and fixed malformed zebrafish (5 to 21 days postfertilization) as well as wild-type fish (5 to 29 days postfertilization) were acquired non-invasively using a commercial SD-OCT system, with a laser source centered at 930nm (λ=100nm), permitting axial and lateral resolutions of 7 and 8μm respectively. Using two-dimensional images and three-dimensional reconstructions, it was possible to identify the malformed notochord as well as deformities in other major organs at different stages of formation. Visualization of the notochord was facilitated with the development of a segmentation algorithm. OCT images were compared to HE histological sections and images obtained by calcein staining. Because of the possibility of performing longitudinal studies on a same fish and reducing image processing time as compared with staining techniques and histology, the use of OCT could facilitate phenotypic characterization in studying genetic factors leading to spinal deformities in zebrafish and could eventually contribute to the identification of the genetic causes of spinal deformities such as scoliosis.

  10. Impact of CdSe/ZnS quantum dots on the development of zebrafish embryos

    International Nuclear Information System (INIS)

    Lei Yong; Xiao Qi; Huang Shan; Xu Wansu; Zhang Zhe; He Zhike; Liu Yi; Den, Fengjiao

    2011-01-01

    Due to their unique fluorescent characteristics, quantum dots (QDs) have been successfully applied in the fields of biotechnology and medicine, but there is very limited information regarding their biodistribution and chronic toxicity in vivo. In this article, the biological behavior and toxic effects of mercaptoacetic acid-CdSe/ZnS QDs (MAA-QDs) in developing zebrafish embryos were investigated by in vivo tests. The MAA-QDs were introduced into zebrafish through microinjection at early stage. The results showed that the MAA-QDs at certain concentrations influenced the survival of zebrafish embryos, but treated embryos without developmental defects were also observed. MAA-QDs injected into the cytoplasm at the one-cell stage were allocated to progeny blastoderm cells during proliferation and almost never entered the yolk. The formation of notochord and primordial germ cells with normal morphologies was detected in the treated embryos by whole-mount in situ hybridization. Furthermore, traces of the element cadmium were mainly discovered in the tissue of liver and kidney of 3-month-old-treated zebrafish by quantitative assessment with inductively coupled plasma mass spectrometry. Thus, we hypothesized that low concentration MAA-QDs have chronic toxicities when they were delivered into zebrafish organs.

  11. Integrated mosquito larval source management reduces larval numbers in two highland villages in western Kenya

    Directory of Open Access Journals (Sweden)

    Imbahale Susan S

    2012-05-01

    Full Text Available Abstract Background In western Kenya, malaria remains one of the major health problems and its control remains an important public health measure. Malaria control is by either use of drugs to treat patients infected with malaria parasites or by controlling the vectors. Vector control may target the free living adult or aquatic (larval stages of mosquito. The most commonly applied control strategies target indoor resting mosquitoes. However, because mosquitoes spend a considerable time in water, targeting the aquatic stages can complement well with existing adult control measures. Methods Larval source management (LSM of malaria vectors was examined in two villages i.e. Fort Ternan and Lunyerere, with the aim of testing strategies that can easily be accessed by the affected communities. Intervention strategies applied include environmental management through source reduction (drainage of canals, land levelling or by filling ditches with soil, habitat manipulation (by provision of shading from arrow root plant, application of Bacillus thuringiensis var israelensis (Bti and the use of predatory fish, Gambusia affinis. The abundance of immature stages of Anopheles and Culex within intervention habitats was compared to that within non-intervention habitats. Results The findings show that in Fort Ternan no significant differences were observed in the abundance of Anopheles early and late instars between intervention and non-intervention habitats. In Lunyerere, the abundance of Anopheles early instars was fifty five times more likely to be present within non-intervention habitats than in habitats under drainage. No differences in early instars abundance were observed between non-intervention and habitats applied with Bti. However, late instars had 89 % and 91 % chance of being sampled from non-intervention rather than habitats under drainage and those applied with Bti respectively. Conclusion Most of these interventions were applied in habitats

  12. Standardized orthotopic xenografts in zebrafish reveal glioma cell-line-specific characteristics and tumor cell heterogeneity

    Directory of Open Access Journals (Sweden)

    Alessandra M. Welker

    2016-02-01

    Full Text Available Glioblastoma (GBM is a deadly brain cancer, for which few effective drug treatments are available. Several studies have used zebrafish models to study GBM, but a standardized approach to modeling GBM in zebrafish was lacking to date, preventing comparison of data across studies. Here, we describe a new, standardized orthotopic xenotransplant model of GBM in zebrafish. Dose-response survival assays were used to define the optimal number of cells for tumor formation. Techniques to measure tumor burden and cell spread within the brain over real time were optimized using mouse neural stem cells as control transplants. Applying this standardized approach, we transplanted two patient-derived GBM cell lines, serum-grown adherent cells and neurospheres, into the midbrain region of embryonic zebrafish and analyzed transplanted larvae over time. Progressive brain tumor growth and premature larval death were observed using both cell lines; however, fewer transplanted neurosphere cells were needed for tumor growth and lethality. Tumors were heterogeneous, containing both cells expressing stem cell markers and cells expressing markers of differentiation. A small proportion of transplanted neurosphere cells expressed glial fibrillary acidic protein (GFAP or vimentin, markers of more differentiated cells, but this number increased significantly during tumor growth, indicating that these cells undergo differentiation in vivo. By contrast, most serum-grown adherent cells expressed GFAP and vimentin at the earliest times examined post-transplant. Both cell types produced brain tumors that contained Sox2+ cells, indicative of tumor stem cells. Transplanted larvae were treated with currently used GBM therapeutics, temozolomide or bortezomib, and this resulted in a reduction in tumor volume in vivo and an increase in survival. The standardized model reported here facilitates robust and reproducible analysis of glioblastoma tumor cells in real time and provides a

  13. Lidocaine Hydrochloride Compared with MS222 for the Euthanasia of Zebrafish (Danio rerio)

    Science.gov (United States)

    Collymore, Chereen; Banks, E Kate; Turner, Patricia V

    2016-01-01

    Despite several shortcomings, MS222 is the most commonly used chemical agent for euthanasia of zebrafish. Although lidocaine hydrochloride has some advantages over MS222, its effectiveness as a euthanasia agent for zebrafish is unknown. Larvae at 9 to 16 d postfertilization were exposed to 250 mg/L MS222 or 400, 500, 600, 700, 800, 900, or 1000 mg/L lidocaine and observed for cessation of heartbeat. Adult zebrafish were exposed to 250 mg/L MS222 or 400, 500, or 600 mg/L lidocaine; times to loss of righting reflex, cessation of opercular movement, and complete recovery; body length; aversive behavior; and gross and microscopic evidence of acute toxicity were evaluated. The heartbeat was not lost from any larvae in any group, regardless of drug or dosage. For adults, time to loss of righting reflex was greatest in the 500-mg/L lidocaine group. Opercular movement ceased earlier in all lidocaine groups compared with the MS222 group. Fish in the 500-mg/L lidocaine group were smaller than those in other groups. Fewer fish in the lidocaine groups displayed aversive behavior (erratic swimming and piping) compared with the MS222 group. No fish in the lidocaine hydrochloride groups (n = 30) recovered from euthanasia, whereas one fish in the MS222 group did (n = 10). Neither the MS222 nor lidocaine groups showed any gross or histologic changes suggestive of acute toxicity. Our results suggest that lidocaine hydrochloride may be an effective alternative chemical euthanasia agent for adult zebrafish but should not be used in larval fish. PMID:27931323

  14. Essential role for fibrillin-2 in zebrafish notochord and vascular morphogenesis.

    Science.gov (United States)

    Gansner, John M; Madsen, Erik C; Mecham, Robert P; Gitlin, Jonathan D

    2008-10-01

    Recent studies demonstrate that lysyl oxidase cuproenzymes are critical for zebrafish notochord formation, but the molecular mechanisms of copper-dependent notochord morphogenesis are incompletely understood. We, therefore, conducted a forward genetic screen for zebrafish mutants that exhibit notochord sensitivity to lysyl oxidase inhibition, yielding a mutant with defects in notochord and vascular morphogenesis, puff daddygw1 (pfdgw1). Meiotic mapping and cloning reveal that the pfdgw1 phenotype results from disruption of the gene encoding the extracellular matrix protein fibrillin-2, and the spatiotemporal expression of fibrillin-2 is consistent with the pfdgw1 phenotype. Furthermore, each aspect of the pfdgw1 phenotype is recapitulated by morpholino knockdown of fibrillin-2. Taken together, the data reveal a genetic interaction between fibrillin-2 and the lysyl oxidases in notochord formation and demonstrate the importance of fibrillin-2 in specific early developmental processes in zebrafish. Copyright (c) 2008 Wiley-Liss, Inc.

  15. Silver Nanopartilces After Zebrafish Development and Larval Behavior: Distinct Roles for Particle Size, Coating and Composition

    Science.gov (United States)

    Background: Silver nanoparticles (AgNPs) act as antibacterials by releasing monovalent silver (Ag+) and are increasingly used in consumer products, thus elevating exposures in human and environmental populations. In vitro models indicate that AgNPs are likely to be developmental ...

  16. A novel mechanism for mechanosensory-based rheotaxis in larval zebrafish

    OpenAIRE

    Oteiza, Pablo; Odstrcil, Iris; Lauder, George; Portugues, Ruben; Engert, Florian

    2017-01-01

    When flying or swimming, animals must adjust their own movement to compensate for displacements induced by the flow of the surrounding air or water1. These flow-induced displacements can most easily be detected as visual whole-field motion with respect to the animal’s frame of reference2. In spite of this, many aquatic animals consistently orient and swim against oncoming flows (a behavior known as rheotaxis) even in the absence of visual cues3,4. How animals achieve this task, and its underl...

  17. Developmental exposure to triclosan alters microbiota community structure and locomotor activity in larval zebrafish

    Science.gov (United States)

    Growing evidence indicates that host-associated microbiota modify the toxicokinetics and/or toxicodynamics of environmental chemicals; however, current risk assessment methods do not consider interactions between microbiota and chemical toxicity. We previously reported that micro...

  18. Investigating phenology of larval fishes in St. Louis River estuary shallow water habitats

    Science.gov (United States)

    As part of the development of an early detection monitoring strategy for non-native fishes, larval fish surveys have been conducted since 2012 in the St. Louis River estuary. Survey data demonstrates there is considerable variability in fish abundance and species assemblages acro...

  19. The importance of Zebrafish in biomedical research.

    Science.gov (United States)

    Tavares, Bárbara; Santos Lopes, Susana

    2013-01-01

    Zebrafish (Danio rerio) is an ideal model organism for the study of vertebrate development. This is due to the large clutches that each couple produces, with up to 200 embryos every 7 days, and to the fact that the embryos and larvae are small, transparent and undergo rapid external development. Using scientific literature research tools available online and the keywords Zebrafish, biomedical research, human disease, and drug screening, we reviewed original studies and reviews indexed in PubMed. In this review we summarized work conducted with this model for the advancement of our knowledge related to several human diseases. We also focused on the biomedical research being performed in Portugal with the zebrafish model. Powerful live imaging and genetic tools are currently available for zebrafish making it a valuable model in biomedical research. The combination of these properties with the optimization of automated systems for drug screening has transformed the zebrafish into "a top model" in biomedical research, drug discovery and toxicity testing. Furthermore, with the optimization of xenografts technology it will be possible to use zebrafish to aide in the choice of the best therapy for each patient. Zebrafish is an excellent model organism in biomedical research, drug development and in clinical therapy.

  20. Sampling uncharted waters: Examining rearing habitat of larval Longfin Smelt (Spirinchus thaleichthys) in the upper San Francisco Estuary

    Science.gov (United States)

    Grimaldo, Lenny; Feyrer, Frederick; Burns, Jillian; Maniscalco, Donna

    2017-01-01

    The southern-most reproducing Longfin Smelt population occurs in the San Francisco Estuary, California, USA. Long-term monitoring of estuarine habitat for this species has generally only considered deep channels, with little known of the role shallow waters play in supporting their early life stage. To address the need for focused research on shallow-water habitat, a targeted study of Longfin Smelt larvae in littoral habitat was conducted to identify potential rearing habitats during 2013 and 2014. Our study objectives were to (1) determine if larval densities vary between littoral habitats (tidal slough vs. open-water shoal), (2) determine how larval densities in littoral habitats vary with physicochemical and biological attributes, (3) determine if larval densities vary between littoral habitats and long-term monitoring channel collections, and (4) determine what factors predict larval rearing distributions from the long-term monitoring channel collections. Larval densities did not vary between littoral habitats but they did vary between years. Water temperature, salinity, and chlorophyll a were found important in predicting larval densities in littoral habitats. Larval densities do not vary between littoral and channel surveys; however, the analysis based on channel data suggests that Longfin Smelt are hatching and rearing in a much broader region and under higher salinities (∼2–12 psu) than previously recognized. Results of this study indicate that conservation efforts should consider how freshwater flow, habitat, climate, and food webs interact as mechanisms that influence Longfin Smelt recruitment in estuarine environments.

  1. 2017 Midwest Zebrafish Meeting Report.

    Science.gov (United States)

    Sandquist, Elizabeth; Petersen, Sarah C; Smith, Cody J

    2017-12-01

    The 2017 Midwest Zebrafish meeting was held from June 16 to 18 at the University of Cincinnati, sponsored by the Cincinnati Children's Hospital Divisions of Developmental Biology, Molecular Cardiovascular Biology, and Gastroenterology, Hepatology, and Nutrition. The meeting, organized by Saulius Sumanas, Joshua Waxman, and Chunyue Yin, hosted >130 attendees from 16 different states. Scientific sessions were focused on morphogenesis, neural development, novel technologies, and disease models, with Steve Ekker, Stephen Potter, and Lila Solnica-Krezel presenting keynote talks. In this article, we highlight the results and emerging themes from the meeting.

  2. Evolution of the osteoblast: skeletogenesis in gar and zebrafish

    Directory of Open Access Journals (Sweden)

    Eames B Frank

    2012-03-01

    surprising finding that the "chondrogenic" transcription factor sox9 is expressed in developing osteoblasts of both zebrafish and gar can help explain the expression of chondrocyte genes in osteoblasts of ray-finned fish. More broadly, our data suggest that the molecular fingerprint of the osteoblast, which largely is constrained among land animals, was not fixed during early vertebrate evolution.

  3. Larval trophodynamics, turbulence, and drift on Georges Bank : A sensitivity analysis of cod and haddock

    DEFF Research Database (Denmark)

    Werner, F.E.; MacKenzie, Brian; Perry, R.I.

    2001-01-01

    Using an individual-based model approach we consider trophodynamic effects on the growth and survival of larval cod (Gadus morhua) and haddock (Melanogrammus aeglefinus) on Georges Bank during late winter/early spring. These studies represent an extension of results described in Werner et al. (1996...... be an important input to larval growth and survival models. The inclusion of turbulence in determining the position of passive larvae in the water column allows the larvae to sample the entire water column, contributing to a decrease in the variance of the size of the larvae over time. The ability of larvae...

  4. Sprouting Buds of Zebrafish Research in Malaysia: First Malaysia Zebrafish Disease Model Workshop.

    Science.gov (United States)

    Okuda, Kazuhide Shaun; Tan, Pei Jean; Patel, Vyomesh

    2016-04-01

    Zebrafish is gaining prominence as an important vertebrate model for investigating various human diseases. Zebrafish provides unique advantages such as optical clarity of embryos, high fecundity rate, and low cost of maintenance, making it a perfect complement to the murine model equivalent in biomedical research. Due to these advantages, researchers in Malaysia are starting to take notice and incorporate the zebrafish model into their research activities. However, zebrafish research in Malaysia is still in its infancy stage and many researchers still remain unaware of the full potential of the zebrafish model or have limited access to related tools and techniques that are widely utilized in many zebrafish laboratories worldwide. To overcome this, we organized the First Malaysia Zebrafish Disease Model Workshop in Malaysia that took place on 11th and 12th of November 2015. In this workshop, we showcased how the zebrafish model is being utilized in the biomedical field in international settings as well as in Malaysia. For this, notable international speakers and those from local universities known to be carrying out impactful research using zebrafish were invited to share some of the cutting edge techniques that are used in their laboratories that may one day be incorporated in the Malaysian scientific community.

  5. A review of monoaminergic neuropsychopharmacology in zebrafish.

    Science.gov (United States)

    Maximino, Caio; Herculano, Anderson Manoel

    2010-12-01

    Monoamine neurotransmitters are the major regulatory mechanisms in the vertebrate brain, involved in the adjustment of motivation, emotion, and cognition. The chemical anatomy of these systems is thought to be highly conserved in the brain of all vertebrates, including zebrafish. Recently, the development of behavioral assays in zebrafish allowed the neuropsychopharmacological investigation of these circuits and its functions. Here we review neuroanatomical, genetic, neurochemical, and psychopharmacological evidence regarding the roles of histaminergic, dopaminergic, noradrenergic, serotonergic, and melatonergic systems in this species. We conclude that, in spite of species differences, zebrafish are suitable for the investigation of neuropsychopharmacology of drugs that affect theses systems; nonetheless, more thorough validation of behavioral methods is still needed.

  6. Whole-brain activity mapping onto a zebrafish brain atlas

    Science.gov (United States)

    Randlett, Owen; Wee, Caroline L.; Naumann, Eva A.; Nnaemeka, Onyeka; Schoppik, David; Fitzgerald, James E.; Portugues, Ruben; Lacoste, Alix M.B.; Riegler, Clemens; Engert, Florian; Schier, Alexander F.

    2015-01-01

    In order to localize the neural circuits involved in generating behaviors, it is necessary to assign activity onto anatomical maps of the nervous system. Using brain registration across hundreds of larval zebrafish, we have built an expandable open source atlas containing molecular labels and anatomical region definitions, the Z-Brain. Using this platform and immunohistochemical detection of phosphorylated-Extracellular signal-regulated kinase (ERK/MAPK) as a readout of neural activity, we have developed a system to create and contextualize whole brain maps of stimulus- and behavior-dependent neural activity. This MAP-Mapping (Mitogen Activated Protein kinase – Mapping) assay is technically simple, fast, inexpensive, and data analysis is completely automated. Since MAP-Mapping is performed on fish that are freely swimming, it is applicable to nearly any stimulus or behavior. We demonstrate the utility of our high-throughput approach using hunting/feeding, pharmacological, visual and noxious stimuli. The resultant maps outline hundreds of areas associated with behaviors. PMID:26778924

  7. Whole-brain activity mapping onto a zebrafish brain atlas.

    Science.gov (United States)

    Randlett, Owen; Wee, Caroline L; Naumann, Eva A; Nnaemeka, Onyeka; Schoppik, David; Fitzgerald, James E; Portugues, Ruben; Lacoste, Alix M B; Riegler, Clemens; Engert, Florian; Schier, Alexander F

    2015-11-01

    In order to localize the neural circuits involved in generating behaviors, it is necessary to assign activity onto anatomical maps of the nervous system. Using brain registration across hundreds of larval zebrafish, we have built an expandable open-source atlas containing molecular labels and definitions of anatomical regions, the Z-Brain. Using this platform and immunohistochemical detection of phosphorylated extracellular signal–regulated kinase (ERK) as a readout of neural activity, we have developed a system to create and contextualize whole-brain maps of stimulus- and behavior-dependent neural activity. This mitogen-activated protein kinase (MAP)-mapping assay is technically simple, and data analysis is completely automated. Because MAP-mapping is performed on freely swimming fish, it is applicable to studies of nearly any stimulus or behavior. Here we demonstrate our high-throughput approach using pharmacological, visual and noxious stimuli, as well as hunting and feeding. The resultant maps outline hundreds of areas associated with behaviors.

  8. On Growth and Form of the Zebrafish Gut Microbiome

    Science.gov (United States)

    Jemielita, Matthew; Taormina, Michael; Rolig, Annah; Burns, Adam; Hampton, Jennifer; Guillemin, Karen; Parthasarathy, Raghuveer

    2014-03-01

    The vertebrate gut is home to a diverse microbial community whose composition has a strong influence on the development and health of the host organism. Researchers can identify the members of the microbiota, yet little is known about the spatial and temporal dynamics of these microbial communities, including the mechanisms guiding their nucleation, growth, and interactions. We address these issues using the larval zebrafish (Danio rerio) as a model organism, which are raised microbe-free and then inoculated with controlled compositions of fluorophore-expressing bacteria. Live imaging using light sheet fluorescence microscopy enables visualization of the gut's entire microbial population over the first 24 hours of colonization. Image analysis allows us to quantify microbial populations that range from a few individuals to tens of thousands of microbes, and analyze the structure and growth kinetics of gut bacterial communities. We find that genetically-identical microbes can show surprisingly different growth rates and colonization abilities depending on their order of arrival. This demonstrates that knowing only the constituents of the gut community is insufficient to determine their dynamics; rather, the history of colonization matters.

  9. Gene expression patterns during the larval development of European sea bass (dicentrarchus labrax) by microarray analysis.

    Science.gov (United States)

    Darias, M J; Zambonino-Infante, J L; Hugot, K; Cahu, C L; Mazurais, D

    2008-01-01

    During the larval period, marine teleosts undergo very fast growth and dramatic changes in morphology, metabolism, and behavior to accomplish their metamorphosis into juvenile fish. Regulation of gene expression is widely thought to be a key mechanism underlying the management of the biological processes required for harmonious development over this phase of life. To provide an overall analysis of gene expression in the whole body during sea bass larval development, we monitored the expression of 6,626 distinct genes at 10 different points in time between 7 and 43 days post-hatching (dph) by using heterologous hybridization of a rainbow trout cDNA microarray. The differentially expressed genes (n = 485) could be grouped into two categories: genes that were generally up-expressed early, between 7 and 23 dph, and genes up-expressed between 25 and 43 dph. Interestingly, among the genes regulated during the larval period, those related to organogenesis, energy pathways, biosynthesis, and digestion were over-represented compared with total set of analyzed genes. We discuss the quantitative regulation of whole-body contents of these specific transcripts with regard to the ontogenesis and maturation of essential functions that take place over larval development. Our study is the first utilization of a transcriptomic approach in sea bass and reveals dynamic changes in gene expression patterns in relation to marine finfish larval development.

  10. Predator-induced larval cloning in the sand dollar Dendraster excentricus: might mothers matter?

    Science.gov (United States)

    Vaughn, Dawn

    2009-10-01

    Predator-induced cloning in echinoid larvae, with reduced size a consequence of cloning, is a dramatic modification of development and a novel response to risks associated with prolonged planktonic development. Recent laboratory studies demonstrate that exposure to stimuli from predators (i.e., fish mucus) induces cloning in the pluteus larvae (plutei) of Dendraster excentricus. However, the timing and incidence of cloning and size reduction of unrelated conspecific plutei differed across experiments. A variable cloning response suggests the effects of such factors as cue quality, egg provisioning, maternal experience, and genetic background, indicating that the potential advantages of cloning as an adaptive response to predators are not available to all larvae. This study tested the hypothesis that cloning in D. excentricus plutei is maternally influenced. Plutei from three half-sibling larval families (different mothers, same father) were exposed to fish mucus for 9 days during early development. Cloning was inferred in a percentage of plutei from each family; however, the rate and success of cloning differed significantly among the larval half-siblings. Unexpectedly, all mucus-treated plutei were smaller and developmentally delayed when compared to all plutei reared in the absence of a mucus stimulus. Thus, while the results from this study support the hypothesis of an influence of mothers on cloning of larval offspring, reduced larval size was a uniform response to fish mucus and did not indicate an effect of mothers. Hypotheses of the developmental effects of fish mucus on larval size with or without successful cloning are discussed.

  11. Anopheline larval habitats seasonality and species distribution: a prerequisite for effective targeted larval habitats control programmes.

    Directory of Open Access Journals (Sweden)

    Eliningaya J Kweka

    Full Text Available Larval control is of paramount importance in the reduction of malaria vector abundance and subsequent disease transmission reduction. Understanding larval habitat succession and its ecology in different land use managements and cropping systems can give an insight for effective larval source management practices. This study investigated larval habitat succession and ecological parameters which influence larval abundance in malaria epidemic prone areas of western Kenya.A total of 51 aquatic habitats positive for anopheline larvae were surveyed and visited once a week for a period of 85 weeks in succession. Habitats were selected and identified. Mosquito larval species, physico-chemical parameters, habitat size, grass cover, crop cycle and distance to nearest house were recorded. Polymerase chain reaction revealed that An. gambiae s.l was the most dominant vector species comprised of An.gambiae s.s (77.60% and An.arabiensis (18.34%, the remaining 4.06% had no amplification by polymerase chain reaction. Physico-chemical parameters and habitat size significantly influenced abundance of An. gambiae s.s (P = 0.024 and An. arabiensis (P = 0.002 larvae. Further, larval species abundance was influenced by crop cycle (P≤0.001, grass cover (P≤0.001, while distance to nearest houses significantly influenced the abundance of mosquito species larvae (r = 0.920;P≤0.001. The number of predator species influenced mosquito larval abundance in different habitat types. Crop weeding significantly influenced with the abundance of An.gambiae s.l (P≤0.001 when preceded with fertilizer application. Significantly higher anopheline larval abundance was recorded in habitats in pasture compared to farmland (P = 0.002. When habitat stability and habitat types were considered, hoof print were the most productive followed by disused goldmines.These findings suggest that implementation of effective larval control programme should be targeted with larval

  12. Screening on the differentially expressed miRNAs in zebrafish (Danio rerio) exposed to trace β-diketone antibiotics and their related functions

    International Nuclear Information System (INIS)

    Li, Jieyi; Liu, Jinfeng; Zhang, Yuhuan; Wang, Xuedong; Li, Weijun; Zhang, Hongqin; Wang, Huili

    2016-01-01

    Highlights: • DKAs possessed toxic effect transfer relation across larval and adult zebrafish. • 215 mature miRNAs were differentially expressed in three comparison groups. • A regulatory network for 4 positive miRNA genes (miR-10, −96, −92, −184) was plotted. • Expression of miR-184, −96, −10 and −92 was proved with miRNA-seq, qRT-PCR and ISH. • DKA exposure induced severe histopathological changes in zebrafish tissues. - Abstract: The toxicity of β-diketone antibiotics (DKAs) to larval and adult zebrafish (Danio rerio) was investigated by miRNA sequencing and bioinformatics analyses. In control and DKA-exposed groups, 215 differentially expressed miRNAs were screened, and 4076 differential target genes were predicted. Among 51 co-differentially expressed genes, 45 were annotated in KOG functional classification, and 34 in KEGG pathway analysis. The homology analysis of 20 miRNAs with human hsa-miRNAs demonstrated 17 high homologous sequences. The expression levels of 12 miRNAs by qRT-PCR were consistent with those by sRNA-seq. A regulatory network for 4 positive miRNA genes (dre-miR-10, −96, −92 and −184) was plotted, and the high-degree of connectivity between miRNA-gene pairs suggests that these miRNAs play critical roles during zebrafish development. The consistent expression of dre-miR-184 and dre-miR-96 was proved in 120-hpf zebrafish brain, gill, otoliths and lateral line neuromast by qRT-PCR, miRNA-seq, W-ISH and ISH. DKA-exposure led to vacuolation of interstitial cells, reduced number of neurons, glial cell proliferation and formation of glial scar, and the obvious abnormality of cell structure might result from abnormal expression of differentially expressed miRNAs. In general, chronic DKA-exposure resulted in comprehensively toxic effects on larval and adult zebrafish tissues, especially for nervous system.

  13. Screening on the differentially expressed miRNAs in zebrafish (Danio rerio) exposed to trace β-diketone antibiotics and their related functions

    Energy Technology Data Exchange (ETDEWEB)

    Li, Jieyi; Liu, Jinfeng; Zhang, Yuhuan [College of Life Sciences, Wenzhou Medical University, Wenzhou 325035 (China); Wang, Xuedong [Key Laboratory of Watershed Sciences and Health of Zhejiang Province, Wenzhou Medical University, Wenzhou 325035 (China); Li, Weijun [Puyang People’s Hospital of Henan Province, Puyang 457000 (China); Zhang, Hongqin [College of Life Sciences, Wenzhou Medical University, Wenzhou 325035 (China); Wang, Huili, E-mail: wxdong@wzmc.edu.cn [College of Life Sciences, Wenzhou Medical University, Wenzhou 325035 (China)

    2016-09-15

    Highlights: • DKAs possessed toxic effect transfer relation across larval and adult zebrafish. • 215 mature miRNAs were differentially expressed in three comparison groups. • A regulatory network for 4 positive miRNA genes (miR-10, −96, −92, −184) was plotted. • Expression of miR-184, −96, −10 and −92 was proved with miRNA-seq, qRT-PCR and ISH. • DKA exposure induced severe histopathological changes in zebrafish tissues. - Abstract: The toxicity of β-diketone antibiotics (DKAs) to larval and adult zebrafish (Danio rerio) was investigated by miRNA sequencing and bioinformatics analyses. In control and DKA-exposed groups, 215 differentially expressed miRNAs were screened, and 4076 differential target genes were predicted. Among 51 co-differentially expressed genes, 45 were annotated in KOG functional classification, and 34 in KEGG pathway analysis. The homology analysis of 20 miRNAs with human hsa-miRNAs demonstrated 17 high homologous sequences. The expression levels of 12 miRNAs by qRT-PCR were consistent with those by sRNA-seq. A regulatory network for 4 positive miRNA genes (dre-miR-10, −96, −92 and −184) was plotted, and the high-degree of connectivity between miRNA-gene pairs suggests that these miRNAs play critical roles during zebrafish development. The consistent expression of dre-miR-184 and dre-miR-96 was proved in 120-hpf zebrafish brain, gill, otoliths and lateral line neuromast by qRT-PCR, miRNA-seq, W-ISH and ISH. DKA-exposure led to vacuolation of interstitial cells, reduced number of neurons, glial cell proliferation and formation of glial scar, and the obvious abnormality of cell structure might result from abnormal expression of differentially expressed miRNAs. In general, chronic DKA-exposure resulted in comprehensively toxic effects on larval and adult zebrafish tissues, especially for nervous system.

  14. Development of molecular markers for zebrafish (Danio rerio) ovarian follicle growth assessment following in-vitro culture in cryopreservation studies.

    Science.gov (United States)

    Anil, Siji; Rawson, David; Zhang, Tiantian

    2018-05-29

    Development of in vitro culture protocol for early stage ovarian follicles of zebrafish is important since cryopreserved early stage ovarian follicles would need to be matured in vitro following cryopreservation before they can be fertilised. Development of molecular markers for zebrafish (Danio rerio) ovarian follicle growth assessment following in vitro culture of early stage zebrafish ovarian follicles in ovarian tissue fragments is reported here for the first time although some work has been reported for in vitro culture of isolated early stage zebrafish ovarian follicles. The main aim of the present study was to develop molecular markers in an optimised in vitro culture protocol for stage I and stage II zebrafish ovarian follicles in ovarian tissue fragments. The effect of concentration of the hormones human chorionic gonadotropin and follicle stimulating hormones, and additives such as Foetal Bovine Serum and Bovine Serum Albumin were studied. The results showed that early stage zebrafish ovarian fragments containing stage I and stage II follicles which are cultured in vitro for 24 h in 20% FBS and 100mIU/ml FSH in 90% L-15 medium at 28 °C can grow to the size of stage II and stage III ovarian follicles respectively. More importantly the follicle growth from stage I to stage II and from stage II to stage III were confirmed using molecular markers such as cyp19a1a (also known as P450aromA) and vtg1 genes respectively. However, no follicle growth was observed following cryopreservation and in vitro culture. Copyright © 2018 Elsevier Inc. All rights reserved.

  15. Episodic-like memory in zebrafish.

    Science.gov (United States)

    Hamilton, Trevor J; Myggland, Allison; Duperreault, Erika; May, Zacnicte; Gallup, Joshua; Powell, Russell A; Schalomon, Melike; Digweed, Shannon M

    2016-11-01

    Episodic-like memory tests often aid in determining an animal's ability to recall the what, where, and which (context) of an event. To date, this type of memory has been demonstrated in humans, wild chacma baboons, corvids (Scrub jays), humming birds, mice, rats, Yucatan minipigs, and cuttlefish. The potential for this type of memory in zebrafish remains unexplored even though they are quickly becoming an essential model organism for the study of a variety of human cognitive and mental disorders. Here we explore the episodic-like capabilities of zebrafish (Danio rerio) in a previously established mammalian memory paradigm. We demonstrate that when zebrafish were presented with a familiar object in a familiar context but a novel location within that context, they spend more time in the novel quadrant. Thus, zebrafish display episodic-like memory as they remember what object they saw, where they saw it (quadrant location), and on which occasion (yellow or blue walls) it was presented.

  16. The Macrophage-Specific Promoter mfap4 Allows Live, Long-Term Analysis of Macrophage Behavior during Mycobacterial Infection in Zebrafish.

    Directory of Open Access Journals (Sweden)

    Eric M Walton

    Full Text Available Transgenic labeling of innate immune cell lineages within the larval zebrafish allows for real-time, in vivo analyses of microbial pathogenesis within a vertebrate host. To date, labeling of zebrafish macrophages has been relatively limited, with the most specific expression coming from the mpeg1 promoter. However, mpeg1 transcription at both endogenous and transgenic loci becomes attenuated in the presence of intracellular pathogens, including Salmonella typhimurium and Mycobacterium marinum. Here, we describe mfap4 as a macrophage-specific promoter capable of producing transgenic lines in which transgene expression within larval macrophages remains stable throughout several days of infection. Additionally, we have developed a novel macrophage-specific Cre transgenic line under the control of mfap4, enabling macrophage-specific expression using existing floxed transgenic lines. These tools enrich the repertoire of transgenic lines and promoters available for studying zebrafish macrophage dynamics during infection and inflammation and add flexibility to the design of future macrophage-specific transgenic lines.

  17. Extraocular muscle regeneration in zebrafish requires late signals from Insulin-like growth factors.

    Science.gov (United States)

    Saera-Vila, Alfonso; Louie, Ke'ale W; Sha, Cuilee; Kelly, Ryan M; Kish, Phillip E; Kahana, Alon

    2018-01-01

    Insulin-like growth factors (Igfs) are key regulators of key biological processes such as embryonic development, growth, and tissue repair and regeneration. The role of Igf in myogenesis is well documented and, in zebrafish, promotes fin and heart regeneration. However, the mechanism of action of Igf in muscle repair and regeneration is not well understood. Using adult zebrafish extraocular muscle (EOM) regeneration as an experimental model, we show that Igf1 receptor blockage using either chemical inhibitors (BMS754807 and NVP-AEW541) or translation-blocking morpholino oligonucleotides (MOs) reduced EOM regeneration. Zebrafish EOMs regeneration depends on myocyte dedifferentiation, which is driven by early epigenetic reprogramming and requires autophagy activation and cell cycle reentry. Inhibition of Igf signaling had no effect on either autophagy activation or cell proliferation, indicating that Igf signaling was not involved in the early reprogramming steps of regeneration. Instead, blocking Igf signaling produced hypercellularity of regenerating EOMs and diminished myosin expression, resulting in lack of mature differentiated muscle fibers even many days after injury, indicating that Igf was involved in late re-differentiation steps. Although it is considered the main mediator of myogenic Igf actions, Akt activation decreased in regenerating EOMs, suggesting that alternative signaling pathways mediate Igf activity in muscle regeneration. In conclusion, Igf signaling is critical for re-differentiation of reprogrammed myoblasts during late steps of zebrafish EOM regeneration, suggesting a regulatory mechanism for determining regenerated muscle size and timing of differentiation, and a potential target for regenerative therapy.

  18. Duplication and diversification of the hypoxia-inducible IGFBP-1 gene in zebrafish.

    Directory of Open Access Journals (Sweden)

    Hiroyasu Kamei

    2008-08-01

    Full Text Available Gene duplication is the primary force of new gene evolution. Deciphering whether a pair of duplicated genes has evolved divergent functions is often challenging. The zebrafish is uniquely positioned to provide insight into the process of functional gene evolution due to its amenability to genetic and experimental manipulation and because it possess a large number of duplicated genes.We report the identification and characterization of two hypoxia-inducible genes in zebrafish that are co-ortholgs of human IGF binding protein-1 (IGFBP-1. IGFBP-1 is a secreted protein that binds to IGF and modulates IGF actions in somatic growth, development, and aging. Like their human and mouse counterparts, in adult zebrafish igfbp-1a and igfbp-1b are exclusively expressed in the liver. During embryogenesis, the two genes are expressed in overlapping spatial domains but with distinct temporal patterns. While zebrafish IGFBP-1a mRNA was easily detected throughout embryogenesis, IGFBP-1b mRNA was detectable only in advanced stages. Hypoxia induces igfbp-1a expression in early embryogenesis, but induces the igfbp-1b expression later in embryogenesis. Both IGFBP-1a and -b are capable of IGF binding, but IGFBP-1b has much lower affinities for IGF-I and -II because of greater dissociation rates. Overexpression of IGFBP-1a and -1b in zebrafish embryos caused significant decreases in growth and developmental rates. When tested in cultured zebrafish embryonic cells, IGFBP-1a and -1b both inhibited IGF-1-induced cell proliferation but the activity of IGFBP-1b was significantly weaker.These results indicate subfunction partitioning of the duplicated IGFBP-1 genes at the levels of gene expression, physiological regulation, protein structure, and biological actions. The duplicated IGFBP-1 may provide additional flexibility in fine-tuning IGF signaling activities under hypoxia and other catabolic conditions.

  19. Rehydration of forensically important larval Diptera specimens.

    Science.gov (United States)

    Sanford, Michelle R; Pechal, Jennifer L; Tomberlin, Jeffery K

    2011-01-01

    Established procedures for collecting and preserving evidence are essential for all forensic disciplines to be accepted in court and by the forensic community at large. Entomological evidence, such as Diptera larvae, are primarily preserved in ethanol, which can evaporate over time, resulting in the dehydration of specimens. In this study, methods used for rehydrating specimens were compared. The changes in larval specimens with respect to larval length and weight for three forensically important blow fly (Diptera: Calliphoridae) species in North America were quantified. Phormia regina (Meigen), Cochliomyia macellaria (F.), and Chrysomya rufifacies (Macquart) third-instar larvae were collected from various decomposing animals and preserved with three preservation methods (80% ethanol, 70% isopropyl alcohol, and hot-water kill then 80% ethanol). Preservative solutions were allowed to evaporate. Rehydration was attempted with either of the following: 80% ethanol, commercial trisodium phosphate substitute solution, or 0.5% trisodium phosphate solution. All three methods partially restored weight and length of specimens recorded before preservation. Analysis of variance results indicated that effects of preservation, rehydration treatment, and collection animal were different in each species. The interaction between preservative method and rehydration treatment had a significant effect on both P. regina and C. macellaria larval length and weight. In addition, there was a significant interaction effect of collection animal on larval C. macellaria measurements. No significant effect was observed in C. rufifacies larval length or weight among the preservatives or treatments. These methods could be used to establish a standard operating procedure for dealing with dehydrated larval specimens in forensic investigations.

  20. Do effects of mercury in larval amphibians persist after metamorphosis?

    Science.gov (United States)

    Todd, Brian D; Willson, John D; Bergeron, Christine M; Hopkins, William A

    2012-01-01

    Despite widespread concern about the role of environmental contaminants in global amphibian declines, and evidence that post-metamorphic life stages contribute disproportionately to amphibian population dynamics, most studies in amphibian ecotoxicology focus on larval life stages. Studies that focus solely on early life stages may miss important effects of contaminant exposure, such as latent effects that manifest some time after previous exposure. Moreover, it is often assumed that effects observed in amphibian larvae will persist to affect survival or reproduction later in life. We used terrestrial enclosures to determine whether exposure to mercury (Hg) through maternal transfer and/or larval diet had any adverse effects in post-metamorphic American toads (Bufo americanus). We found a 5% difference in size at metamorphosis that was attributed to maternal Hg exposure persisted for 1 year in the terrestrial environment, resulting in a 7% difference at the conclusion of the study. Although patterns of survival differed among treatments through time, we found no overall difference in survival after 1 year. We also found no evidence of emergent latent effects in the terrestrial toads that could be attributed to earlier exposure. Our results indicate that adverse effects of maternal Hg exposure that were observed in larval amphibians may persist to affect later terrestrial life stages but that no novel adverse effects developed when animals were raised in a semi-natural environment. Moreover, we found no evidence of persistent effects of dietary Hg exposure in larvae, highlighting a need for greater focus on maternal effects in amphibian ecotoxicology. Finally, we suggest an increase in the use of longitudinal studies to better understand contaminant impacts to amphibian populations via effects in both aquatic and terrestrial life stages.

  1. Soundscapes and Larval Settlement: Characterizing the Stimulus from a Larval Perspective.

    Science.gov (United States)

    Lillis, Ashlee; Eggleston, David B; Bohnenstiehl, DelWayne R

    2016-01-01

    There is growing evidence that underwater sounds serve as a cue for the larvae of marine organisms to locate suitable settlement habitats; however, the relevant spatiotemporal scales of variability in habitat-related sounds and how this variation scales with larval settlement processes remain largely uncharacterized, particularly in estuarine habitats. Here, we provide an overview of the approaches we have developed to characterize an estuarine soundscape as it relates to larval processes, and a conceptual framework is provided for how habitat-related sounds may influence larval settlement, using oyster reef soundscapes as an example.

  2. Identification and characterization of zebrafish thrombocytes.

    Science.gov (United States)

    Jagadeeswaran, P; Sheehan, J P; Craig, F E; Troyer, D

    1999-12-01

    To analyse primary haemostasis in the zebrafish we have identified and characterized the zebrafish thrombocyte by morphologic, immunologic and functional approaches. Novel methods were developed for harvesting zebrafish blood with preservation of thrombocytes, and assaying whole blood adhesion/aggregation responses in microtitre plates. Light and electron microscopy of the thrombocyte illustrated morphological characteristics including the formation of aggregates, pseudopodia, and surface-connected vesicles analagous to the platelet canalicular system. Immunostaining with polyclonal antisera versus human platelet glycoproteins demonstrated the presence of glycoprotein Ib and IIb/IIIa-like complexes on the thrombocyte surface. Whole blood assays for adhesion/aggregation and ATP release showed ristocetin-induced adhesion without ATP release, and platelet agonist (collagen, arachidonic acid) induced aggregation with ATP release. Blood harvested from zebrafish treated with aspirin demonstrated inhibition of arachidonic acid induced aggregation and agonist induced ATP release, consistent with at least partial dependence on an intact cyclo oxygenase pathway. The combined morphologic immunologic and functional evidence suggest that the zebrafish thrombocyte is the haemostatic homologue of the mammalian platelet. Conservation of major haemostatic pathways involved in platelet function and coagulation suggests that the zebrafish is a relevant model for mammalian haemostasis and thrombosis.

  3. Genomic Organization of Zebrafish microRNAs

    Directory of Open Access Journals (Sweden)

    Paydar Ima

    2008-05-01

    Full Text Available Abstract Background microRNAs (miRNAs are small (~22 nt non-coding RNAs that regulate cell movement, specification, and development. Expression of miRNAs is highly regulated, both spatially and temporally. Based on direct cloning, sequence conservation, and predicted secondary structures, a large number of miRNAs have been identified in higher eukaryotic genomes but whether these RNAs are simply a subset of a much larger number of noncoding RNA families is unknown. This is especially true in zebrafish where genome sequencing and annotation is not yet complete. Results We analyzed the zebrafish genome to identify the number and location of proven and predicted miRNAs resulting in the identification of 35 new miRNAs. We then grouped all 415 zebrafish miRNAs into families based on seed sequence identity as a means to identify possible functional redundancy. Based on genomic location and expression analysis, we also identified those miRNAs that are likely to be encoded as part of polycistronic transcripts. Lastly, as a resource, we compiled existing zebrafish miRNA expression data and, where possible, listed all experimentally proven mRNA targets. Conclusion Current analysis indicates the zebrafish genome encodes 415 miRNAs which can be grouped into 44 families. The largest of these families (the miR-430 family contains 72 members largely clustered in two main locations along chromosome 4. Thus far, most zebrafish miRNAs exhibit tissue specific patterns of expression.

  4. Genetic diversity, classification and comparative study on the larval ...

    African Journals Online (AJOL)

    Genetic diversity, classification and comparative study on the larval phenotypic ... B. mori showed different performance based on larval phenotypic data. The analysis of variance regarding the studied traits showed that different strains have ...

  5. R-spondin 3 regulates dorsoventral and anteroposterior patterning by antagonizing Wnt/β-catenin signaling in zebrafish embryos.

    Directory of Open Access Journals (Sweden)

    Xiaozhi Rong

    Full Text Available The Wnt/β-catenin or canonical Wnt signaling pathway plays fundamental roles in early development and in maintaining adult tissue homeostasis. R-spondin 3 (Rspo3 is a secreted protein that has been implicated in activating the Wnt/β-catenin signaling in amphibians and mammals. Here we report that zebrafish Rspo3 plays a negative role in regulating the zygotic Wnt/β-catenin signaling. Zebrafish Rspo3 has a unique domain structure. It contains a third furin-like (FU3 domain. This FU3 is present in other four ray-finned fish species studied but not in elephant shark. In zebrafish, rspo3 mRNA is maternally deposited and has a ubiquitous expression in early embryonic stages. After 12 hpf, its expression becomes tissue-specific. Forced expression of rspo3 promotes dorsoanterior patterning and increases the expression of dorsal and anterior marker genes. Knockdown of rspo3 increases ventral-posterior development and stimulates ventral and posterior marker genes expression. Forced expression of rspo3 abolishes exogenous Wnt3a action and reduces the endogenous Wnt signaling activity. Knockdown of rspo3 results in increased Wnt/β-catenin signaling activity. Further analyses indicate that Rspo3 does not promote maternal Wnt signaling. Human RSPO3 has similar action when tested in zebrafish embryos. These results suggest that Rspo3 regulates dorsoventral and anteroposterior patterning by negatively regulating the zygotic Wnt/β-catenin signaling in zebrafish embryos.

  6. Zebrafish neurobehavioral phenomics for aquatic neuropharmacology and toxicology research.

    Science.gov (United States)

    Kalueff, Allan V; Echevarria, David J; Homechaudhuri, Sumit; Stewart, Adam Michael; Collier, Adam D; Kaluyeva, Aleksandra A; Li, Shaomin; Liu, Yingcong; Chen, Peirong; Wang, JiaJia; Yang, Lei; Mitra, Anisa; Pal, Subharthi; Chaudhuri, Adwitiya; Roy, Anwesha; Biswas, Missidona; Roy, Dola; Podder, Anupam; Poudel, Manoj K; Katare, Deepshikha P; Mani, Ruchi J; Kyzar, Evan J; Gaikwad, Siddharth; Nguyen, Michael; Song, Cai

    2016-01-01

    Zebrafish (Danio rerio) are rapidly emerging as an important model organism for aquatic neuropharmacology and toxicology research. The behavioral/phenotypic complexity of zebrafish allows for thorough dissection of complex human brain disorders and drug-evoked pathological states. As numerous zebrafish models become available with a wide spectrum of behavioral, genetic, and environmental methods to test novel drugs, here we discuss recent zebrafish phenomics methods to facilitate drug discovery, particularly in the field of biological psychiatry. Additionally, behavioral, neurological, and endocrine endpoints are becoming increasingly well-characterized in zebrafish, making them an inexpensive, robust and effective model for toxicology research and pharmacological screening. We also discuss zebrafish behavioral phenotypes, experimental considerations, pharmacological candidates and relevance of zebrafish neurophenomics to other 'omics' (e.g., genomic, proteomic) approaches. Finally, we critically evaluate the limitations of utilizing this model organism, and outline future strategies of research in the field of zebrafish phenomics. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Biophysical models of larval dispersal in the Benguela Current ...

    African Journals Online (AJOL)

    We synthesise and update results from the suite of biophysical, larval-dispersal models developed in the Benguela Current ecosystem. Biophysical models of larval dispersal use outputs of physical hydrodynamic models as inputs to individual-based models in which biological processes acting during the larval life are ...

  8. Primary Spinal OPC Culture System from Adult Zebrafish to Study Oligodendrocyte Differentiation In Vitro

    Directory of Open Access Journals (Sweden)

    Volker Kroehne

    2017-09-01

    Full Text Available Endogenous oligodendrocyte progenitor cells (OPCs are a promising target to improve functional recovery after spinal cord injury (SCI by remyelinating denuded, and therefore vulnerable, axons. Demyelination is the result of a primary insult and secondary injury, leading to conduction blocks and long-term degeneration of the axons, which subsequently can lead to the loss of their neurons. In response to SCI, dormant OPCs can be activated and subsequently start to proliferate and differentiate into mature myelinating oligodendrocytes (OLs. Therefore, researchers strive to control OPC responses, and utilize small molecule screening approaches in order to identify mechanisms of OPC activation, proliferation, migration and differentiation. In zebrafish, OPCs remyelinate axons of the optic tract after lysophosphatidylcholine (LPC-induced demyelination back to full thickness myelin sheaths. In contrast to zebrafish, mammalian OPCs are highly vulnerable to excitotoxic stress, a cause of secondary injury, and remyelination remains insufficient. Generally, injury induced remyelination leads to shorter internodes and thinner myelin sheaths in mammals. In this study, we show that myelin sheaths are lost early after a complete spinal transection injury, but are re-established within 14 days after lesion. We introduce a novel, easy-to-use, inexpensive and highly reproducible OPC culture system based on dormant spinal OPCs from adult zebrafish that enables in vitro analysis. Zebrafish OPCs are robust, can easily be purified with high viability and taken into cell culture. This method enables to examine why zebrafish OPCs remyelinate better than their mammalian counterparts, identify cell intrinsic responses, which could lead to pro-proliferating or pro-differentiating strategies, and to test small molecule approaches. In this methodology paper, we show efficient isolation of OPCs from adult zebrafish spinal cord and describe culture conditions that enable

  9. Evaluating human cancer cell metastasis in zebrafish

    International Nuclear Information System (INIS)

    Teng, Yong; Xie, Xiayang; Walker, Steven; White, David T; Mumm, Jeff S; Cowell, John K

    2013-01-01

    In vivo metastasis assays have traditionally been performed in mice, but the process is inefficient and costly. However, since zebrafish do not develop an adaptive immune system until 14 days post-fertilization, human cancer cells can survive and metastasize when transplanted into zebrafish larvae. Despite isolated reports, there has been no systematic evaluation of the robustness of this system to date. Individual cell lines were stained with CM-Dil and injected into the perivitelline space of 2-day old zebrafish larvae. After 2-4 days fish were imaged using confocal microscopy and the number of metastatic cells was determined using Fiji software. To determine whether zebrafish can faithfully report metastatic potential in human cancer cells, we injected a series of cells with different metastatic potential into the perivitelline space of 2 day old embryos. Using cells from breast, prostate, colon and pancreas we demonstrated that the degree of cell metastasis in fish is proportional to their invasion potential in vitro. Highly metastatic cells such as MDA231, DU145, SW620 and ASPC-1 are seen in the vasculature and throughout the body of the fish after only 24–48 hours. Importantly, cells that are not invasive in vitro such as T47D, LNCaP and HT29 do not metastasize in fish. Inactivation of JAK1/2 in fibrosarcoma cells leads to loss of invasion in vitro and metastasis in vivo, and in zebrafish these cells show limited spread throughout the zebrafish body compared with the highly metastatic parental cells. Further, knockdown of WASF3 in DU145 cells which leads to loss of invasion in vitro and metastasis in vivo also results in suppression of metastasis in zebrafish. In a cancer progression model involving normal MCF10A breast epithelial cells, the degree of invasion/metastasis in vitro and in mice is mirrored in zebrafish. Using a modified version of Fiji software, it is possible to quantify individual metastatic cells in the transparent larvae to correlate with

  10. Biosecurity and Health Monitoring at the Zebrafish International Resource Center

    OpenAIRE

    Murray, Katrina N.; Varga, Zolt?n M.; Kent, Michael L.

    2016-01-01

    The Zebrafish International Resource Center (ZIRC) is a repository and distribution center for mutant, transgenic, and wild-type zebrafish. In recent years annual imports of new zebrafish lines to ZIRC have increased tremendously. In addition, after 15 years of research, we have identified some of the most virulent pathogens affecting zebrafish that should be avoided in large production facilities, such as ZIRC. Therefore, while importing a high volume of new lines we prioritize safeguarding ...

  11. Polybrominated diphenyl ethers affect the reproduction and development, and alter the sex ratio of zebrafish (Danio rerio)

    International Nuclear Information System (INIS)

    Han, X.B.; Yuen, Karen W.Y.; Wu, Rudolf S.S.

    2013-01-01

    Polybrominated diphenyl ethers (PBDEs) have been commonly used as flame retardants and now become ubiquitous in the global environment. Using zebrafish as a model, we tested the hypothesis that PBDEs may affect the reproduction and development of fish. Zebrafish were exposed to environmentally relevant concentrations of DE-71 (a congener of PBDE commonly found in the environment) throughout their whole life cycle, and the effects of DE-71 on gonadal development, gamete quality, fertilization success, hatching success, embryonic development and sex ratio were investigated. Despite gonadal development was enhanced, reductions in spawning, fertilization success, hatching success and larval survival rate were evident, while significant increases in malformation and percentage of male were also observed in the F1 generation. Our laboratory results suggest that PBDEs may pose a risk to reproductive success and alter the sex ratio of fish in environments highly contaminated with PBDEs. -- Highlights: •Zebrafish were exposed to PBDE from eggs to adults. •An increase in Gonadal-Somatic Index and enhanced gonadal development was enhanced. •Fertilization and hatching successes were reduced, while malformation was increased. •PBDE alters sex differentiation, leading to a male biased F1 population. •Environmental relevant concentrations of PBDE threaten natural fish populations. -- PBDE reduces fertilization and hatching successes, causes malformation and leads to a male biased F1 generation in fish

  12. ngs (notochord granular surface) gene encodes a novel type of intermediate filament family protein essential for notochord maintenance in zebrafish.

    Science.gov (United States)

    Tong, Xiangjun; Xia, Zhidan; Zu, Yao; Telfer, Helena; Hu, Jing; Yu, Jingyi; Liu, Huan; Zhang, Quan; Sodmergen; Lin, Shuo; Zhang, Bo

    2013-01-25

    The notochord is an important organ involved in embryonic patterning and locomotion. In zebrafish, the mature notochord consists of a single stack of fully differentiated, large vacuolated cells called chordocytes, surrounded by a single layer of less differentiated notochordal epithelial cells called chordoblasts. Through genetic analysis of zebrafish lines carrying pseudo-typed retroviral insertions, a mutant exhibiting a defective notochord with a granular appearance was isolated, and the corresponding gene was identified as ngs (notochord granular surface), which was specifically expressed in the notochord. In the mutants, the notochord started to degenerate from 32 hours post-fertilization, and the chordocytes were then gradually replaced by smaller cells derived from chordoblasts. The granular notochord phenotype was alleviated by anesthetizing the mutant embryos with tricaine to prevent muscle contraction and locomotion. Phylogenetic analysis showed that ngs encodes a new type of intermediate filament (IF) family protein, which we named chordostatin based on its function. Under the transmission electron microcopy, bundles of 10-nm-thick IF-like filaments were enriched in the chordocytes of wild-type zebrafish embryos, whereas the chordocytes in ngs mutants lacked IF-like structures. Furthermore, chordostatin-enhanced GFP (EGFP) fusion protein assembled into a filamentous network specifically in chordocytes. Taken together, our work demonstrates that ngs encodes a novel type of IF protein and functions to maintain notochord integrity for larval development and locomotion. Our work sheds light on the mechanisms of notochord structural maintenance, as well as the evolution and biological function of IF family proteins.

  13. Strategic larval decision-making in a bivoltine butterfly.

    Science.gov (United States)

    Friberg, Magne; Dahlerus, Josefin; Wiklund, Christer

    2012-07-01

    In temperate areas, insect larvae must decide between entering winter diapause or developing directly and reproducing in the same season. Long daylength and high temperature promote direct development, which is generally associated with a higher growth rate. In this work, we investigated whether the larval pathway decision precedes the adjustment of growth rate (state-independent), or whether the pathway decision is conditional on the individual's growth rate (state-dependent), in the butterfly Pieris napi. This species typically makes the pathway decision in the penultimate instar. We measured growth rate throughout larval development under two daylengths: slightly shorter and slightly longer than the critical daylength. Results indicate that the pathway decision can be both state-independent and state-dependent; under the shorter daylength condition, most larvae entered diapause, and direct development was chosen exclusively by a small subset of larvae showing the highest growth rates already in the early instars; under the longer daylength condition, most larvae developed directly, and the diapause pathway was chosen exclusively by a small subset of slow-growing individuals. Among the remainder, the choice of pathway was independent of the early growth rate; larvae entering diapause under the short daylength grew as fast as or faster than the direct developers under the longer daylength in the early instars, whereas the direct developers grew faster than the diapausers only in the ultimate instar. Hence, the pathway decision was state-dependent in a subset with a very high or very low growth rate, whereas the decision was state-independent in the majority of the larvae, which made the growth rate adjustment downstream from the pathway decision.

  14. Zebrafish neurofibromatosis type 1 genes have redundant functions in tumorigenesis and embryonic development

    Directory of Open Access Journals (Sweden)

    Jimann Shin

    2012-11-01

    Neurofibromatosis type 1 (NF1 is a common, dominantly inherited genetic disorder that results from mutations in the neurofibromin 1 (NF1 gene. Affected individuals demonstrate abnormalities in neural-crest-derived tissues that include hyperpigmented skin lesions and benign peripheral nerve sheath tumors. NF1 patients also have a predisposition to malignancies including juvenile myelomonocytic leukemia (JMML, optic glioma, glioblastoma, schwannoma and malignant peripheral nerve sheath tumors (MPNSTs. In an effort to better define the molecular and cellular determinants of NF1 disease pathogenesis in vivo, we employed targeted mutagenesis strategies to generate zebrafish harboring stable germline mutations in nf1a and nf1b, orthologues of NF1. Animals homozygous for loss-of-function alleles of nf1a or nf1b alone are phenotypically normal and viable. Homozygous loss of both alleles in combination generates larval phenotypes that resemble aspects of the human disease and results in larval lethality between 7 and 10 days post fertilization. nf1-null larvae demonstrate significant central and peripheral nervous system defects. These include aberrant proliferation and differentiation of oligodendrocyte progenitor cells (OPCs, dysmorphic myelin sheaths and hyperplasia of Schwann cells. Loss of nf1 contributes to tumorigenesis as demonstrated by an accelerated onset and increased penetrance of high-grade gliomas and MPNSTs in adult nf1a+/−; nf1b−/−; p53e7/e7 animals. nf1-null larvae also demonstrate significant motor and learning defects. Importantly, we identify and quantitatively analyze a novel melanophore phenotype in nf1-null larvae, providing the first animal model of the pathognomonic pigmentation lesions of NF1. Together, these findings support a role for nf1a and nf1b as potent tumor suppressor genes that also function in the development of both central and peripheral glial cells as well as melanophores in zebrafish.

  15. Genome-wide survey and developmental expression mapping of zebrafish SET domain-containing genes.

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    Xiao-Jian Sun

    Full Text Available SET domain-containing proteins represent an evolutionarily conserved family of epigenetic regulators, which are responsible for most histone lysine methylation. Since some of these genes have been revealed to be essential for embryonic development, we propose that the zebrafish, a vertebrate model organism possessing many advantages for developmental studies, can be utilized to study the biological functions of these genes and the related epigenetic mechanisms during early development. To this end, we have performed a genome-wide survey of zebrafish SET domain genes. 58 genes total have been identified. Although gene duplication events give rise to several lineage-specific paralogs, clear reciprocal orthologous relationship reveals high conservation between zebrafish and human SET domain genes. These data were further subject to an evolutionary analysis ranging from yeast to human, leading to the identification of putative clusters of orthologous groups (COGs of this gene family. By means of whole-mount mRNA in situ hybridization strategy, we have also carried out a developmental expression mapping of these genes. A group of maternal SET domain genes, which are implicated in the programming of histone modification states in early development, have been identified and predicted to be responsible for all known sites of SET domain-mediated histone methylation. Furthermore, some genes show specific expression patterns in certain tissues at certain stages, suggesting the involvement of epigenetic mechanisms in the development of these systems. These results provide a global view of zebrafish SET domain histone methyltransferases in evolutionary and developmental dimensions and pave the way for using zebrafish to systematically study the roles of these genes during development.

  16. Developmental mechanisms of arsenite toxicity in zebrafish (Danio rerio) embryos

    International Nuclear Information System (INIS)

    Li Dan; Lu Cailing; Wang Ju; Hu Wei; Cao Zongfu; Sun Daguang; Xia Hongfei; Ma Xu

    2009-01-01

    Arsenic usually accumulates in soil, water and airborne particles, from which it is taken up by various organisms. Exposure to arsenic through food and drinking water is a major public health problem affecting some countries. At present there are limited laboratory data on the effects of arsenic exposure on early embryonic development and the mechanisms behind its toxicity. In this study, we used zebrafish as a model system to investigate the effects of arsenite on early development. Zebrafish embryos were exposed to a range of sodium arsenite concentrations (0-10.0 mM) between 4 and 120 h post-fertilization (hpf). Survival and early development of the embryos were not obviously influenced by arsenite concentrations below 0.5 mM. However, embryos exposed to higher concentrations (0.5-10.0 mM) displayed reduced survival and abnormal development including delayed hatching, retarded growth and changed morphology. Alterations in neural development included weak tactile responses to light (2.0-5.0 mM, 30 hpf), malformation of the spinal cord and disordered motor axon projections (2.0 mM, 48 hpf). Abnormal cardiac function was observed as bradycardia (0.5-2.0 mM, 60 hpf) and altered ventricular shape (2.0 mM, 48 hpf). Furthermore, altered cell proliferation (2.0 mM, 24 hpf) and apoptosis status (2.0 mM, 24 and 48 hpf), as well as abnormal genomic DNA methylation patterning (2.0 mM, 24 and 48 hpf) were detected in the arsenite-treated embryos. All of these indicate a possible relationship between arsenic exposure and developmental failure in early embryogenesis. Our studies suggest that the negative effects of arsenic on vertebrate embryogenesis are substantial

  17. Otolith tethering in the zebrafish otic vesicle requires Otogelin and α-Tectorin.

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    Stooke-Vaughan, Georgina A; Obholzer, Nikolaus D; Baxendale, Sarah; Megason, Sean G; Whitfield, Tanya T

    2015-03-15

    Otoliths are biomineralised structures important for balance and hearing in fish. Their counterparts in the mammalian inner ear, otoconia, have a primarily vestibular function. Otoliths and otoconia form over sensory maculae and are attached to the otolithic membrane, a gelatinous extracellular matrix that provides a physical coupling between the otolith and the underlying sensory epithelium. In this study, we have identified two proteins required for otolith tethering in the zebrafish ear, and propose that there are at least two stages to this process: seeding and maintenance. The initial seeding step, in which otolith precursor particles tether directly to the tips of hair cell kinocilia, fails to occur in the einstein (eis) mutant. The gene disrupted in eis is otogelin (otog); mutations in the human OTOG gene have recently been identified as causative for deafness and vestibular dysfunction (DFNB18B). At later larval stages, maintenance of otolith tethering to the saccular macula is dependent on tectorin alpha (tecta) function, which is disrupted in the rolling stones (rst) mutant. α-Tectorin (Tecta) is a major constituent of the tectorial membrane in the mammalian cochlea. Mutations in the human TECTA gene can cause either dominant (DFNA8/12) or recessive (DFNB21) forms of deafness. Our findings indicate that the composition of extracellular otic membranes is highly conserved between mammals and fish, reinforcing the view that the zebrafish is an excellent model system for the study of deafness and vestibular disease. © 2015. Published by The Company of Biologists Ltd.

  18. Genomic Amplification of an Endogenous Retrovirus in Zebrafish T-Cell Malignancies

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    J. Kimble Frazer

    2012-01-01

    Full Text Available Genomic instability plays a crucial role in oncogenesis. Somatically acquired mutations can disable some genes and inappropriately activate others. In addition, chromosomal rearrangements can amplify, delete, or even fuse genes, altering their functions and contributing to malignant phenotypes. Using array comparative genomic hybridization (aCGH, a technique to detect numeric variations between different DNA samples, we examined genomes from zebrafish (Danio rerio T-cell leukemias of three cancer-prone lines. In all malignancies tested, we identified recurring amplifications of a zebrafish endogenous retrovirus. This retrovirus, ZFERV, was first identified due to high expression of proviral transcripts in thymic tissue from larval and adult fish. We confirmed ZFERV amplifications by quantitative PCR analyses of DNA from wild-type fish tissue and normal and malignant D. rerio T cells. We also quantified ZFERV RNA expression and found that normal and neoplastic T cells both produce retrovirally encoded transcripts, but most cancers show dramatically increased transcription. In aggregate, these data imply that ZFERV amplification and transcription may be related to T-cell leukemogenesis. Based on these data and ZFERV’s phylogenetic relation to viruses of the murine-leukemia-related virus class of gammaretroviridae, we posit that ZFERV may be oncogenic via an insertional mutagenesis mechanism.

  19. Distinct innate immune phagocyte responses to Aspergillus fumigatus conidia and hyphae in zebrafish larvae.

    Science.gov (United States)

    Knox, Benjamin P; Deng, Qing; Rood, Mary; Eickhoff, Jens C; Keller, Nancy P; Huttenlocher, Anna

    2014-10-01

    Aspergillus fumigatus is the most common filamentous fungal pathogen of immunocompromised hosts, resulting in invasive aspergillosis (IA) and high mortality rates. Innate immunity is known to be the predominant host defense against A. fumigatus; however, innate phagocyte responses to A. fumigatus in an intact host and their contributions to host survival remain unclear. Here, we describe a larval zebrafish A. fumigatus infection model amenable to real-time imaging of host-fungal interactions in live animals. Following infection with A. fumigatus, innate phagocyte populations exhibit clear preferences for different fungal morphologies: macrophages rapidly phagocytose conidia and form aggregates around hyphae, while the neutrophil response is dependent upon the presence of hyphae. Depletion of macrophages rendered host larvae susceptible to invasive disease. Moreover, a zebrafish model of human leukocyte adhesion deficiency with impaired neutrophil function also resulted in invasive disease and impaired host survival. In contrast, macrophage-deficient but not neutrophil-deficient larvae exhibited attenuated disease following challenge with a less virulent (ΔlaeA) strain of A. fumigatus, which has defects in secondary metabolite production. Taking these results together, we have established a new vertebrate model for studying innate immune responses to A. fumigatus that reveals distinct roles for neutrophils and macrophages in mediating host defense against IA. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  20. Natural bizbenzoquinoline derivatives protect zebrafish lateral line sensory hair cells from aminoglycoside toxicity

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    Matthew eKruger

    2016-03-01

    Full Text Available Moderate to severe hearing loss affects 360 million people worldwide and most often results from damage to sensory hair cells. Hair cell damage can result from aging, genetic mutations, excess noise exposure, and certain medications including aminoglycoside antibiotics. Aminoglycosides are effective at treating infections associated with cystic fibrosis and other life-threatening conditions such as sepsis, but cause hearing loss in 20-30% of patients. It is therefore imperative to develop new therapies to combat hearing loss and allow safe use of these potent antibiotics. We approach this drug discovery question using the larval zebrafish lateral line because zebrafish hair cells are structurally and functionally similar to mammalian inner ear hair cells and respond similarly to toxins. We screened a library of 502 natural compounds in order to identify novel hair cell protectants. Our screen identified four bisbenzylisoquinoline derivatives: berbamine, E6 berbamine, hernandezine, and isotetrandrine, each of which robustly protected hair cells from aminoglycoside-induced damage. Using fluorescence microscopy and electrophysiology, we demonstrated that the natural compounds confer protection by reducing antibiotic uptake into hair cells and showed that hair cells remain functional during and after incubation in E6 berbamine. We also determined that these natural compounds do not reduce antibiotic efficacy. Together, these natural compounds represent a novel source of possible otoprotective drugs that may offer therapeutic options for patients receiving aminoglycoside treatment.

  1. Alternative splicing of sept9a and sept9b in zebrafish produces multiple mRNA transcripts expressed throughout development.

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    Megan L Landsverk

    2010-05-01

    Full Text Available Septins are involved in a number of cellular processes including cytokinesis and organization of the cytoskeleton. Alterations in human septin-9 (SEPT9 levels have been linked to multiple cancers, whereas mutations in SEPT9 cause the episodic neuropathy, hereditary neuralgic amyotrophy (HNA. Despite its important function in human health, the in vivo role of SEPT9 is unknown.Here we utilize zebrafish to study the role of SEPT9 in early development. We show that zebrafish possess two genes, sept9a and sept9b that, like humans, express multiple transcripts. Knockdown or overexpression of sept9a transcripts results in specific developmental alterations including circulation defects and aberrant epidermal development.Our work demonstrates that sept9 plays an important role in zebrafish development, and establishes zebrafish as a valuable model organism for the study of SEPT9.

  2. Dissection and lateral mounting of zebrafish embryos: analysis of spinal cord development.

    Science.gov (United States)

    Beck, Aaron P; Watt, Roland M; Bonner, Jennifer

    2014-02-28

    The zebrafish spinal cord is an effective investigative model for nervous system research for several reasons. First, genetic, transgenic and gene knockdown approaches can be utilized to examine the molecular mechanisms underlying nervous system development. Second, large clutches of developmentally synchronized embryos provide large experimental sample sizes. Third, the optical clarity of the zebrafish embryo permits researchers to visualize progenitor, glial, and neuronal populations. Although zebrafish embryos are transparent, specimen thickness can impede effective microscopic visualization. One reason for this is the tandem development of the spinal cord and overlying somite tissue. Another reason is the large yolk ball, which is still present during periods of early neurogenesis. In this article, we demonstrate microdissection and removal of the yolk in fixed embryos, which allows microscopic visualization while preserving surrounding somite tissue. We also demonstrate semipermanent mounting of zebrafish embryos. This permits observation of neurodevelopment in the dorso-ventral and anterior-posterior axes, as it preserves the three-dimensionality of the tissue.

  3. In Vivo Testing of MicroRNA-Mediated Gene Knockdown in Zebrafish

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    Ivone Un San Leong

    2012-01-01

    Full Text Available The zebrafish (Danio rerio has become an attractive model for human disease modeling as there are a large number of orthologous genes that encode similar proteins to those found in humans. The number of tools available to manipulate the zebrafish genome is limited and many currently used techniques are only effective during early development (such as morpholino-based antisense technology or it is phenotypically driven and does not offer targeted gene knockdown (such as chemical mutagenesis. The use of RNA interference has been met with controversy as off-target effects can make interpreting phenotypic outcomes difficult; however, this has been resolved by creating zebrafish lines that contain stably integrated miRNA constructs that target the desired gene of interest. In this study, we show that a commercially available miRNA vector system with a mouse-derived miRNA backbone is functional in zebrafish and is effective in causing eGFP knockdown in a transient in vivo eGFP sensor assay system. We chose to apply this system to the knockdown of transcripts that are implicated in the human cardiac disorder, Long QT syndrome.

  4. Control over the morphology and segregation of Zebrafish germ cell granules during embryonic development

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    Nakkrasae La-Iad

    2008-05-01

    Full Text Available Abstract Background Zebrafish germ cells contain granular-like structures, organized around the cell nucleus. These structures share common features with polar granules in Drosophila, germinal granules in Xenopus and chromatoid bodies in mice germ cells, such as the localization of the zebrafish Vasa, Piwi and Nanos proteins, among others. Little is known about the structure of these granules as well as their segregation in mitosis during early germ-cell development. Results Using transgenic fish expressing a fluorescently labeled novel component of Zebrafish germ cell granules termed Granulito, we followed the morphology and distribution of the granules. We show that whereas these granules initially exhibit a wide size variation, by the end of the first day of development they become a homogeneous population of medium size granules. We investigated this resizing event and demonstrated the role of microtubules and the minus-end microtubule dependent motor protein Dynein in the process. Last, we show that the function of the germ cell granule resident protein the Tudor domain containing protein-7 (Tdrd7 is required for determination of granule morphology and number. Conclusion Our results suggest that Zebrafish germ cell granules undergo a transformation process, which involves germ cell specific proteins as well as the microtubular network.

  5. Developmental sub-chronic exposure to chlorpyrifos reduces anxiety-related behavior in zebrafish larvae

    Science.gov (United States)

    Richendrfer, Holly; Pelkowski, Sean D.; Colwill, Ruth M.; Créton, Robbert

    2013-01-01

    Neurobehavioral disorders such as anxiety, autism, and attention deficit hyperactivity disorders are typically influenced by genetic and environmental factors. Although several genetic risk factors have been identified in recent years, little is known about the environmental factors that either cause neurobehavioral disorders or contribute to their progression in genetically predisposed individuals. One environmental factor that has raised concerns is chlorpyrifos, an organophosphate pesticide that is widely used in agriculture and is found ubiquitously in the environment. In the present study, we examined the effects of sub-chronic chlorpyrifos exposure on anxiety-related behavior during development using zebrafish larvae. We found that sub-chronic exposure to 0.01 or 0.1 μM chlorpyrifos during development induces specific behavioral defects in 7-day-old zebrafish larvae. The larvae displayed decreases in swim speed and thigmotaxis, yet no changes in avoidance behavior were seen. Exposure to 0.001 μM chlorpyrifos did not affect swimming, thigmotaxis, or avoidance behavior and exposure to 1 μM chlorpyrifos induced behavioral defects, but also induced defects in larval morphology. Since thigmotaxis, a preference for the edge, is an anxiety-related behavior in zebrafish larvae, we propose that sub-chronic chlorpyrifos exposure interferes with the development of anxiety-related behaviors. The results of this study provide a good starting point for examination of the molecular, cellular, developmental, and neural mechanisms that are affected by environmentally relevant concentrations of organophosphate pesticides. A more detailed understanding of these mechanisms is important for the development of predictive models and refined health policies to prevent toxicant-induced neurobehavioral disorders. PMID:22579535

  6. Joint toxic effects of triazophos and imidacloprid on zebrafish (Danio rerio).

    Science.gov (United States)

    Wu, Shenggan; Li, Xinfang; Liu, Xinju; Yang, Guiling; An, Xuehua; Wang, Qiang; Wang, Yanhua

    2018-04-01

    Pesticide contamination is more often found as a mixture of different pesticides in water bodies rather than individual compounds. However, regulatory risk evaluation is mostly based on the effects of individual pesticides. In the present study, we aimed to investigate the individual and joint toxicities of triazophos (TRI) and imidacloprid (IMI) to the zebrafish (Danio rerio) using acute indices and various sublethal endpoints. Results from 96-h semi-static test indicated that the LC 50 values of TRI to D. rerio at multiple life stages (embryonic, larval, juvenile and adult stages) ranged from 0.49 (0.36-0.71) to 4.99 (2.06-6.81) mg a.i. L -1 , which were higher than those of IMI ranging from 26.39 (19.04-38.01) to 128.9 (68.47-173.6) mg a.i. L -1 . Pesticide mixtures of TRI and IMI displayed synergistic response to zebrafish embryos. Activities of carboxylesterase (CarE) and catalase (CAT) were significantly changed in most of the individual and joint exposures of pesticides compared with the control group. The expressions of 26 genes related to oxidative stress, cellular apoptosis, immune system, hypothalamic-pituitary-thyroid and hypothalamic-pituitary-gonadal axis at the mRNA level revealed that zebrafish embryos were affected by the individual or joint pesticides, and greater changes in the expressions of six genes (Mn-sod, CXCL-CIC, Dio1, Dio2, tsh and vtg1) were observed when exposed to joint pesticides compared with their individual pesticides. Taken together, the synergistic effects indicated that it was highly important to incorporate joint toxicity studies, especially at low concentrations, when assessing the risk of pesticides. Copyright © 2018 Elsevier Ltd. All rights reserved.

  7. Transcriptome analysis of zebrafish embryogenesis using microarrays.

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    Sinnakaruppan Mathavan

    2005-08-01

    Full Text Available Zebrafish (Danio rerio is a well-recognized model for the study of vertebrate developmental genetics, yet at the same time little is known about the transcriptional events that underlie zebrafish embryogenesis. Here we have employed microarray analysis to study the temporal activity of developmentally regulated genes during zebrafish embryogenesis. Transcriptome analysis at 12 different embryonic time points covering five different developmental stages (maternal, blastula, gastrula, segmentation, and pharyngula revealed a highly dynamic transcriptional profile. Hierarchical clustering, stage-specific clustering, and algorithms to detect onset and peak of gene expression revealed clearly demarcated transcript clusters with maximum gene activity at distinct developmental stages as well as co-regulated expression of gene groups involved in dedicated functions such as organogenesis. Our study also revealed a previously unidentified cohort of genes that are transcribed prior to the mid-blastula transition, a time point earlier than when the zygotic genome was traditionally thought to become active. Here we provide, for the first time to our knowledge, a comprehensive list of developmentally regulated zebrafish genes and their expression profiles during embryogenesis, including novel information on the temporal expression of several thousand previously uncharacterized genes. The expression data generated from this study are accessible to all interested scientists from our institute resource database (http://giscompute.gis.a-star.edu.sg/~govind/zebrafish/data_download.html.

  8. Effects of uranium on the metabolism of zebrafish, Danio rerio

    International Nuclear Information System (INIS)

    Augustine, Starrlight; Gagnaire, Béatrice; Adam-Guillermin, Christelle; Kooijman, Sebastiaan A.L.M.

    2012-01-01

    The increasing demand for nuclear energy results in heightened levels of uranium (U) in aquatic systems which present a potential health hazard to resident organisms. The aim of this study was to mechanistically assess how chronic exposure to environmentally relevant concentrations of U perturbs the complex interplay between feeding, growth, maintenance, maturation and reproduction throughout the life-cycle of an individual. To this end we analysed literature-based and original zebrafish toxicity data within a same mass and energy balancing conceptual framework. U was found to increase somatic maintenance leading to inhibition of spawning as well as increase hazard rate and costs for growth during the early life stages. The fish's initial conditions and elimination through reproduction greatly affected toxico-kinetics and effects. We demonstrate that growth and reproduction should be measured on specific individuals since mean values were hardly interpretable. The mean food level differed between experiments, conditions and individuals. This last ‘detail’ contributed substantially to the observed variability by its combined effect on metabolism, toxic effects and toxico-kinetics. The significance of this work is that we address exactly how these issues are related and derive conclusions which are independent of experimental protocol and coherent with a very large body of literature on zebrafish eco-physiology.

  9. Effects of uranium on the metabolism of zebrafish, Danio rerio

    Energy Technology Data Exchange (ETDEWEB)

    Augustine, Starrlight, E-mail: starr-light.augustine@irsn.fr [Laboratory of Radionuclide Ecotoxicology, PRP-ENV/SERIS/LECO, Institute of Radioprotection and Nuclear Safety (IRSN), Caradache, Building 186, BP3, 13115 St-Paul-lez-Durance Cedex (France); Gagnaire, Beatrice, E-mail: beatrice.gagnaire@irsn.fr [Laboratory of Radionuclide Ecotoxicology, PRP-ENV/SERIS/LECO, Institute of Radioprotection and Nuclear Safety (IRSN), Caradache, Building 186, BP3, 13115 St-Paul-lez-Durance Cedex (France); Adam-Guillermin, Christelle, E-mail: christelle.adam-guillermin@irsn.fr [Laboratory of Radionuclide Ecotoxicology, PRP-ENV/SERIS/LECO, Institute of Radioprotection and Nuclear Safety (IRSN), Caradache, Building 186, BP3, 13115 St-Paul-lez-Durance Cedex (France); Kooijman, Sebastiaan A.L.M., E-mail: bas.kooijman@vu.nl [Department of Theoretical Biology, Vrije Universiteit, de Boelelaan 1087, 1081 HV Amsterdam (Netherlands)

    2012-08-15

    The increasing demand for nuclear energy results in heightened levels of uranium (U) in aquatic systems which present a potential health hazard to resident organisms. The aim of this study was to mechanistically assess how chronic exposure to environmentally relevant concentrations of U perturbs the complex interplay between feeding, growth, maintenance, maturation and reproduction throughout the life-cycle of an individual. To this end we analysed literature-based and original zebrafish toxicity data within a same mass and energy balancing conceptual framework. U was found to increase somatic maintenance leading to inhibition of spawning as well as increase hazard rate and costs for growth during the early life stages. The fish's initial conditions and elimination through reproduction greatly affected toxico-kinetics and effects. We demonstrate that growth and reproduction should be measured on specific individuals since mean values were hardly interpretable. The mean food level differed between experiments, conditions and individuals. This last 'detail' contributed substantially to the observed variability by its combined effect on metabolism, toxic effects and toxico-kinetics. The significance of this work is that we address exactly how these issues are related and derive conclusions which are independent of experimental protocol and coherent with a very large body of literature on zebrafish eco-physiology.

  10. Aqueous Hg(2+) associates with TiO2 nanoparticles according to particle size, changes particle agglomeration, and becomes less bioavailable to zebrafish.

    Science.gov (United States)

    Boran, Halis; Boyle, David; Altinok, Ilhan; Patsiou, Danae; Henry, Theodore B

    2016-05-01

    Engineered nanoparticles (NPs) have unique physicochemistry and potential to interact with other substances in the aqueous phase. Here, gene [metallothionein 2 (mt2)] expression changes in larval zebrafish were used to evaluate the association between aqueous Hg(2+) and TiO2 (NPs and bulk particle size control) to investigate the relationship between changes in Hg(2+) behavior and TiO2 size. During 24h exposures, TiO2 agglomerates increased in size and in the presence of 25μg Hg(2+)/L, greater increases in size were observed. The concentration of Hg(2+) in suspension also decreased in the presence of TiO2-NPs. Mercury increased expression of mt2 in larval zebrafish, but this response was lessened when zebrafish were exposed to Hg(2+) in the presence of TiO2-NPs, and which suggests that TiO2-NPs alter the bioavailability of Hg(2+) to zebrafish larvae. This ameliorative effect of TiO2 was also likely due to surface binding of Hg(2+) because a greater decrease in mt2 expression was observed in the presence of 1mg/L TiO2-NPs than 1mg/L TiO2-bulk. In conclusion, the results show that Hg(2+) will associate with TiO2-NPs, TiO2-NPs that have associated Hg(2+) will settle out of the aqueous phase more rapidly, and agglomerates will deliver associated Hg(2+) to sediment surfaces. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. The zebrafish genome: a review and msx gene case study.

    Science.gov (United States)

    Postlethwait, J H

    2006-01-01

    Zebrafish is one of several important teleost models for understanding principles of vertebrate developmental, molecular, organismal, genetic, evolutionary, and genomic biology. Efficient investigation of the molecular genetic basis of induced mutations depends on knowledge of the zebrafish genome. Principles of zebrafish genomic analysis, including gene mapping, ortholog identification, conservation of syntenies, genome duplication, and evolution of duplicate gene function are discussed here using as a case study the zebrafish msxa, msxb, msxc, msxd, and msxe genes, which together constitute zebrafish orthologs of tetrapod Msx1, Msx2, and Msx3. Genomic analysis suggests orthologs for this difficult to understand group of paralogs.

  12. Live Cell Imaging of Butterfly Pupal and Larval Wings In Vivo.

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    Yoshikazu Ohno

    Full Text Available Butterfly wing color patterns are determined during the late larval and early pupal stages. Characterization of wing epithelial cells at these stages is thus critical to understand how wing structures, including color patterns, are determined. Previously, we successfully recorded real-time in vivo images of developing butterfly wings over time at the tissue level. In this study, we employed similar in vivo fluorescent imaging techniques to visualize developing wing epithelial cells in the late larval and early pupal stages 1 hour post-pupation. Both larval and pupal epithelial cells were rich in mitochondria and intracellular networks of endoplasmic reticulum, suggesting high metabolic activities, likely in preparation for cellular division, polyploidization, and differentiation. Larval epithelial cells in the wing imaginal disk were relatively large horizontally and tightly packed, whereas pupal epithelial cells were smaller and relatively loosely packed. Furthermore, larval cells were flat, whereas pupal cells were vertically elongated as deep as 130 μm. In pupal cells, many endosome-like or autophagosome-like structures were present in the cellular periphery down to approximately 10 μm in depth, and extensive epidermal feet or filopodia-like processes were observed a few micrometers deep from the cellular surface. Cells were clustered or bundled from approximately 50 μm in depth to deeper levels. From 60 μm to 80 μm in depth, horizontal connections between these clusters were observed. The prospective eyespot and marginal focus areas were resistant to fluorescent dyes, likely because of their non-flat cone-like structures with a relatively thick cuticle. These in vivo images provide important information with which to understand processes of epithelial cell differentiation and color pattern determination in butterfly wings.

  13. Live Cell Imaging of Butterfly Pupal and Larval Wings In Vivo.

    Science.gov (United States)

    Ohno, Yoshikazu; Otaki, Joji M

    2015-01-01

    Butterfly wing color patterns are determined during the late larval and early pupal stages. Characterization of wing epithelial cells at these stages is thus critical to understand how wing structures, including color patterns, are determined. Previously, we successfully recorded real-time in vivo images of developing butterfly wings over time at the tissue level. In this study, we employed similar in vivo fluorescent imaging techniques to visualize developing wing epithelial cells in the late larval and early pupal stages 1 hour post-pupation. Both larval and pupal epithelial cells were rich in mitochondria and intracellular networks of endoplasmic reticulum, suggesting high metabolic activities, likely in preparation for cellular division, polyploidization, and differentiation. Larval epithelial cells in the wing imaginal disk were relatively large horizontally and tightly packed, whereas pupal epithelial cells were smaller and relatively loosely packed. Furthermore, larval cells were flat, whereas pupal cells were vertically elongated as deep as 130 μm. In pupal cells, many endosome-like or autophagosome-like structures were present in the cellular periphery down to approximately 10 μm in depth, and extensive epidermal feet or filopodia-like processes were observed a few micrometers deep from the cellular surface. Cells were clustered or bundled from approximately 50 μm in depth to deeper levels. From 60 μm to 80 μm in depth, horizontal connections between these clusters were observed. The prospective eyespot and marginal focus areas were resistant to fluorescent dyes, likely because of their non-flat cone-like structures with a relatively thick cuticle. These in vivo images provide important information with which to understand processes of epithelial cell differentiation and color pattern determination in butterfly wings.

  14. DDT exposure of zebrafish embryos enhances seizure susceptibility: relationship to fetal p,p'-DDE burden and domoic acid exposure of California sea lions.

    Science.gov (United States)

    Tiedeken, Jessica A; Ramsdell, John S

    2009-01-01

    California sea lions have a large body burden of organochlorine pesticides, and over the last decade they have also been subject to domoic acid poisoning. Domoic acid poisoning, previously recognized in adult animals, is now viewed as a major cause of prenatal mortality. The appearance of a chronic juvenile domoic acid disease in the sea lions, characterized by behavioral abnormalities and epilepsy, is consistent with early life poisoning and may be potentiated by organochlorine burden. We investigated the interactive effect of DDT (dichlorodiphenyltrichloroethane) on neurodevelopment using a zebrafish (Danio rerio) model for seizure behavior to examine the susceptibility to domoic acid-induced seizures after completion of neurodevelopment. Embryos were exposed (6-30 hr postfertilization) to either o,p'-DDT or p,p'-DDE (dichlorodiphenyldichloroethylene) during neurodevelopment via a 0.1% dimethyl sulfoxide solution. These larval (7 days postfertilization) fish were then exposed to either the seizure-inducing drug pentylenetetrazol (PTZ) or domoic acid; resulting seizure behavior was monitored and analyzed for changes using cameras and behavioral tracking software. Embryonic exposure to DDTs enhanced PTZ seizures and caused distinct and increased seizure behaviors to domoic acid, most notably a type of head-shaking behavior. These studies demonstrate that embryonic exposure to DDTs leads to asymptomatic animals at completion of neurodevelopment with greater sensitivity to domoic acid-induced seizures. The body burden levels of p,p'-DDE are close to the range recently found in fetal California sea lions and suggest a potential interactive effect of p,p'-DDE embryonic poisoning and domoic acid toxicity.

  15. Larval Population Density Alters Adult Sleep in Wild-Type Drosophila melanogaster but Not in Amnesiac Mutant Flies

    Directory of Open Access Journals (Sweden)

    Michael W. Chi

    2014-08-01

    Full Text Available Sleep has many important biological functions, but how sleep is regulated remains poorly understood. In humans, social isolation and other stressors early in life can disrupt adult sleep. In fruit flies housed at different population densities during early adulthood, social enrichment was shown to increase subsequent sleep, but it is unknown if population density during early development can also influence adult sleep. To answer this question, we maintained Drosophila larvae at a range of population densities throughout larval development, kept them isolated during early adulthood, and then tested their sleep patterns. Our findings reveal that flies that had been isolated as larvae had more fragmented sleep than those that had been raised at higher population densities. This effect was more prominent in females than in males. Larval population density did not affect sleep in female flies that were mutant for amnesiac, which has been shown to be required for normal memory consolidation, adult sleep regulation, and brain development. In contrast, larval population density effects on sleep persisted in female flies lacking the olfactory receptor or83b, suggesting that olfactory signals are not required for the effects of larval population density on adult sleep. These findings show that population density during early development can alter sleep behavior in adulthood, suggesting that genetic and/or structural changes are induced by this developmental manipulation that persist through metamorphosis.

  16. Scrambled eggs: Proteomic portraits and novel biomarkers of egg quality in zebrafish (Danio rerio)

    OpenAIRE

    Yilmaz, Ozlem; Patinote, Amélie; Nguyen, Thao Vi; Com, Emmanuelle; Lavigne, Regis; Pineau, Charles; Sullivan, Craig V.; Bobe, Julien

    2017-01-01

    Egg quality is a complex biological trait and a major determinant of reproductive fitness in all animals. This study delivered the first proteomic portraits of egg quality in zebrafish, a leading biomedical model for early development. Egg batches of good and poor quality, evidenced by embryo survival for 24 h, were sampled immediately after spawning and used to create pooled or replicated sample sets whose protein extracts were subjected to different levels of fractionation before liquid chr...

  17. Characterization of zebrafish larvae suction feeding flow using μPIV and optical coherence tomography

    OpenAIRE

    Pekkan, Kerem; Uslu, Fazıl E.; Chang, Brain; Mani, Karthick; Chen, Chia-Yuan; Holzman, Roi

    2016-01-01

    The hydrodynamics of suction feeding is critical for the survival of fish larvae; failure to capture food during the onset of autonomous feeding can rapidly lead to starvation and mortality. Fluid mechanics experiments that investigate the suction feeding of suspended particles are limited to adult fishes, which operate at large Reynolds numbers. This manuscript presents the first literature results in which the external velocity fields generated during suction feeding of early zebrafish larv...

  18. New tides: using zebrafish to study renal regeneration.

    Science.gov (United States)

    McCampbell, Kristen K; Wingert, Rebecca A

    2014-02-01

    Over the past several decades, the zebrafish has become one of the major vertebrate model organisms used in biomedical research. In this arena, the zebrafish has emerged as an applicable system for the study of kidney diseases and renal regeneration. The relevance of the zebrafish model for nephrology research has been increasingly appreciated as the understanding of zebrafish kidney structure, ontogeny, and the response to damage has steadily expanded. Recent studies have documented the amazing regenerative characteristics of the zebrafish kidney, which include the ability to replace epithelial populations after acute injury and to grow new renal functional units, termed nephrons. Here we discuss how nephron composition is conserved between zebrafish and mammals, and highlight how recent findings from zebrafish studies utilizing transgenic technologies and chemical genetics can complement traditional murine approaches in the effort to dissect how the kidney responds to acute damage and identify therapeutics that enhance human renal regeneration. Copyright © 2014 Mosby, Inc. All rights reserved.

  19. Morphological and Physiological Interactions Between GnRH3 and Hypocretin/Orexin Neuronal Systems in Zebrafish (Danio rerio).

    Science.gov (United States)

    Zhao, Yali; Singh, Chanpreet; Prober, David A; Wayne, Nancy L

    2016-10-01

    GnRH neurons integrate internal and external cues to control sexual maturation and fertility. Homeostasis of energy balance and food intake correlates strongly with the status of reproduction. Neuropeptides secreted by the hypothalamus involved in modulating energy balance and feeding may play additional roles in the regulation of reproduction. Hypocretin (Hcrt) (also known as orexin) is one such peptide, primarily controlling sleep/wakefulness, food intake, and reward processing. There is a growing body of evidence indicating that Hcrt/orexin (Hcrt) modulates reproduction through interacting with the hypothalamo-pituitary-gonadal axis in mammals. To explore potential morphological and functional interactions between the GnRH and Hcrt neuronal systems, we employed a variety of experimental approaches including confocal imaging, immunohistochemistry, and electrophysiology in transgenic zebrafish, in which fluorescent proteins are genetically expressed in GnRH3 and Hcrt neurons. Our imaging data revealed close apposition and direct connection between GnRH3 and Hcrt neuronal systems in the hypothalamus during larval development through adulthood. Furthermore, the Hcrt receptor (HcrtR) is expressed in GnRH3 neurons. Electrophysiological data revealed a reversible inhibitory effect of Hcrt on GnRH3 neuron electrical activity, which was blocked by the HcrtR antagonist almorexant. In addition, Hcrt had no effect on the electrical activity of GnRH3 neurons in the HcrtR null mutant zebrafish (HcrtR -/- ). Our findings demonstrate a close anatomical and functional relationship between Hcrt and GnRH neuronal systems in zebrafish. It is the first demonstration of a link between neuronal circuits controlling sleeping/arousal/feeding and reproduction in zebrafish, an important animal model for investigating the molecular genetics of development.

  20. Zebrafish-Based Discovery of Antiseizure Compounds from the Red Sea: Pseurotin A2 and Azaspirofuran A.

    Science.gov (United States)

    Copmans, Daniëlle; Rateb, Mostafa; Tabudravu, Jioji N; Pérez-Bonilla, Mercedes; Dirkx, Nina; Vallorani, Riccardo; Diaz, Caridad; Pérez Del Palacio, José; Smith, Alan J; Ebel, Rainer; Reyes, Fernando; Jaspars, Marcel; de Witte, Peter A M

    2018-04-19

    In search for novel antiseizure drugs (ASDs), the European FP7-funded PharmaSea project used zebrafish embryos and larvae as a drug discovery platform to screen marine natural products to identify promising antiseizure hits in vivo for further development. Within the framework of this project, seven known heterospirocyclic γ-lactams, namely, pseurotin A, pseurotin A 2 , pseurotin F1, 11- O-methylpseurotin A, pseurotin D, azaspirofuran A, and azaspirofuran B, were isolated from the bioactive marine fungus Aspergillus fumigatus, and their antiseizure activity was evaluated in the larval zebrafish pentylenetetrazole (PTZ) seizure model. Pseurotin A 2 and azaspirofuran A were identified as antiseizure hits, while their close chemical analogues were inactive. Besides, electrophysiological analysis from the zebrafish midbrain demonstrated that pseurotin A 2 and azaspirofuran A also ameliorate PTZ-induced epileptiform discharges. Next, to determine whether these findings translate to mammalians, both compounds were analyzed in the mouse 6 Hz (44 mA) psychomotor seizure model. They lowered the seizure duration dose-dependently, thereby confirming their antiseizure properties and suggesting activity against drug-resistant seizures. Finally, in a thorough ADMET assessment, pseurotin A 2 and azaspirofuran A were found to be drug-like. Based on the prominent antiseizure activity in both species and the drug-likeness, we propose pseurotin A 2 and azaspirofuran A as lead compounds that are worth further investigation for the treatment of epileptic seizures. This study not only provides the first evidence of antiseizure activity of pseurotins and azaspirofurans, but also demonstrates the value of the zebrafish model in (marine) natural product drug discovery in general, and for ASD discovery in particular.

  1. Dietary β-glucan enhances the contents of complement component 3 and factor B in eggs of zebrafish.

    Science.gov (United States)

    Jiang, Chengyan; Wang, Peng; Li, Mengyang; Liu, Shousheng; Zhang, Shicui

    2016-12-01

    β-glucan has been shown to increase non-specific immunity and resistance against infections or pathogenic bacteria in several fish species, but no information is available regarding its trans-generational effects to date. Here we clearly demonstrated that β-glucan enhanced the contents of immune-relevant molecules C3 and Bf in eggs of zebrafish, and the embryos derived from β-1,3 glucan-treated zebrafish were more resistant to bacterial challenge than control embryos. Moreover, the transferred C3 and Bf were directly associated with the antimicrobial defense of early embryos. In addition, feeding female zebrafish with β-glucan had little detrimental effects on the number of spawned eggs and their embryonic development. Collectively, these data show for the first time that β-glucan can be safely used to promote the non-specific immunity in offspring of fishes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Neutrophil Reverse Migration Becomes Transparent with Zebrafish

    Directory of Open Access Journals (Sweden)

    Taylor W. Starnes

    2012-01-01

    Full Text Available The precise control of neutrophil-mediated inflammation is critical for both host defense and the prevention of immunopathology. In vivo imaging studies in zebrafish, and more recently in mice, have made the novel observation that neutrophils leave a site of inflammation through a process called neutrophil reverse migration. The application of advanced imaging techniques to the genetically tractable, optically transparent zebrafish larvae was critical for these advances. Still, the mechanisms underlying neutrophil reverse migration and its effects on the resolution or priming of immune responses remain unclear. Here, we review the current knowledge of neutrophil reverse migration, its potential roles in host immunity, and the live imaging tools that make zebrafish a valuable model for increasing our knowledge of neutrophil behavior in vivo.

  3. Tributyltin and Zebrafish: Swimming in Dangerous Water

    Directory of Open Access Journals (Sweden)

    Clemilson Berto-Júnior

    2018-04-01

    Full Text Available Zebrafish has been established as a reliable biological model with important insertion in academy (morphologic, biochemical, and pathophysiological studies and pharmaceutical industry (toxicology and drug development due to its molecular complexity and similar systems biology that recapitulate those from other organisms. Considering the toxicological aspects, many efforts using zebrafish models are being done in order to elucidate the effects of endocrine disruptors, and some of them are focused on tributyltin (TBT and its mechanism of action. TBT is an antifouling agent applied in ship’s hull that is constantly released into the water and absorbed by marine organisms, leading to bioaccumulation and biomagnification effects. Thus, several findings of malformations and changes in the normal biochemical and physiologic aspects of these marine animals have been related to TBT contamination. In the present review, we have compiled the most significant studies related to TBT effects in zebrafish, also taking into consideration the effects found in other study models.

  4. Tributyltin and Zebrafish: Swimming in Dangerous Water

    Science.gov (United States)

    Berto-Júnior, Clemilson; de Carvalho, Denise Pires; Soares, Paula; Miranda-Alves, Leandro

    2018-01-01

    Zebrafish has been established as a reliable biological model with important insertion in academy (morphologic, biochemical, and pathophysiological studies) and pharmaceutical industry (toxicology and drug development) due to its molecular complexity and similar systems biology that recapitulate those from other organisms. Considering the toxicological aspects, many efforts using zebrafish models are being done in order to elucidate the effects of endocrine disruptors, and some of them are focused on tributyltin (TBT) and its mechanism of action. TBT is an antifouling agent applied in ship’s hull that is constantly released into the water and absorbed by marine organisms, leading to bioaccumulation and biomagnification effects. Thus, several findings of malformations and changes in the normal biochemical and physiologic aspects of these marine animals have been related to TBT contamination. In the present review, we have compiled the most significant studies related to TBT effects in zebrafish, also taking into consideration the effects found in other study models. PMID:29692757

  5. Dynamic nucleosome organization at hox promoters during zebrafish embryogenesis.

    Directory of Open Access Journals (Sweden)

    Steven E Weicksel

    Full Text Available Nucleosome organization at promoter regions plays an important role in regulating gene activity. Genome-wide studies in yeast, flies, worms, mammalian embryonic stem cells and transformed cell lines have found well-positioned nucleosomes flanking a nucleosome depleted region (NDR at transcription start sites. This nucleosome arrangement depends on DNA sequence (cis-elements as well as DNA binding factors and ATP-dependent chromatin modifiers (trans-factors. However, little is understood about how the nascent embryonic genome positions nucleosomes during development. This is particularly intriguing since the embryonic genome must undergo a broad reprogramming event upon fusion of sperm and oocyte. Using four stages of early embryonic zebrafish development, we map nucleosome positions at the promoter region of 37 zebrafish hox genes. We find that nucleosome arrangement at the hox promoters is a progressive process that takes place over several stages. At stages immediately after fertilization, nucleosomes appear to be largely disordered at hox promoter regions. At stages after activation of the embryonic genome, nucleosomes are detectable at hox promoters, with positions becoming more uniform and more highly occupied. Since the genomic sequence is invariant during embryogenesis, this progressive change in nucleosome arrangement suggests that trans-factors play an important role in organizing nucleosomes during embryogenesis. Separating hox genes into expressed and non-expressed groups shows that expressed promoters have better positioned and occupied nucleosomes, as well as distinct NDRs, than non-expressed promoters. Finally, by blocking the retinoic acid-signaling pathway, we disrupt early hox gene transcription, but observe no effect on nucleosome positions, suggesting that active hox transcription is not a driving force behind the arrangement of nucleosomes at the promoters of hox genes during early development.

  6. Effect of massing on larval growth rate.

    Science.gov (United States)

    Johnson, Aidan P; Wallman, James F

    2014-08-01

    Estimation of minimum postmortem interval commonly relies on predicting the age of blowfly larvae based on their size and an estimate of the temperatures to which they have been exposed throughout their development. The majority of larval growth rate data have been developed using small larval masses in order to avoid excess heat generation. The current study collected growth rate data for larvae at different mass volumes, and assessed the temperature production of these masses, for two forensically important blow fly species, Chrysomya rufifacies and Calliphora vicina. The growth rate of larvae in a small mass, exposed to the higher temperatures equivalent to those experienced by large masses, was also assessed to determine if observed differences were due to the known temperature effects of maggot masses. The results showed that temperature production increased with increasing mass volume, with temperature increases of 11 °C observed in the large Ch. rufifacies masses and increases of 5 °C in the large C. vicina masses. Similarly, the growth rate of the larvae was affected by mass size. The larvae from small masses grown at the higher temperatures experienced by large masses displayed an initial delay in growth, but then grew at a similar rate to those larvae at a constant 23 °C. Since these larvae from masses of equivalent sizes displayed similar patterns of growth rate, despite differing temperatures, and these growth rates differed from larger masses exposed to the same temperatures, it can be concluded that larval growth rate within a mass may be affected by additional factors other than temperature. Overall, this study highlights the importance of understanding the role of massing in larval development and provides initial developmental data for mass sizes of two forensically important blowfly species commonly encountered in Australian forensic casework. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  7. Microbial gut diversity of Africanized and European honey bee larval instars.

    Directory of Open Access Journals (Sweden)

    Svjetlana Vojvodic

    Full Text Available The first step in understanding gut microbial ecology is determining the presence and potential niche breadth of associated microbes. While the core gut bacteria of adult honey bees is becoming increasingly apparent, there is very little and inconsistent information concerning symbiotic bacterial communities in honey bee larvae. The larval gut is the target of highly pathogenic bacteria and fungi, highlighting the need to understand interactions between typical larval gut flora, nutrition and disease progression. Here we show that the larval gut is colonized by a handful of bacterial groups previously described from guts of adult honey bees or other pollinators. First and second larval instars contained almost exclusively Alpha 2.2, a core Acetobacteraceae, while later instars were dominated by one of two very different Lactobacillus spp., depending on the sampled site. Royal jelly inhibition assays revealed that of seven bacteria occurring in larvae, only one Neisseriaceae and one Lactobacillus sp. were inhibited. We found both core and environmentally vectored bacteria with putatively beneficial functions. Our results suggest that early inoculation by Acetobacteraceae may be important for microbial succession in larvae. This assay is a starting point for more sophisticated in vitro models of nutrition and disease resistance in honey bee larvae.

  8. Culturable gut microbiota diversity in zebrafish.

    Science.gov (United States)

    Cantas, Leon; Sørby, Jan Roger Torp; Aleström, Peter; Sørum, Henning

    2012-03-01

    The zebrafish (Danio rerio) is an increasingly used laboratory animal model in basic biology and biomedicine, novel drug development, and toxicology. The wide use has increased the demand for optimized husbandry protocols to ensure animal health care and welfare. The knowledge about the correlation between culturable zebrafish intestinal microbiota and health in relation to environmental factors and management procedures is very limited. A semi-quantitative level of growth of individual types of bacteria was determined and associated with sampling points. A total of 72 TAB line zebrafish from four laboratories (Labs A-D) in the Zebrafish Network Norway were used. Diagnostic was based on traditional bacterial culture methods and biochemical characterization using commercial kits, followed by 16S rDNA gene sequencing from pure subcultures. Also selected Gram-negative isolates were analyzed for antibiotic susceptibility to 8 different antibiotics. A total of 13 morphologically different bacterial species were the most prevalent: Aeromonas hydrophila, Aeromonas sobria, Vibrio parahaemolyticus, Photobacterium damselae, Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas luteola, Comamonas testosteroni, Ochrobactrum anthropi, Staphylococcus cohnii, Staphylococcus epidermidis, Staphylococcus capitis, and Staphylococcus warneri. Only Lab B had significantly higher levels of total bacterial growth (OR=2.03), whereas numbers from Lab C (OR=1.01) and Lab D (OR=1.12) were found to be similar to the baseline Lab A. Sexually immature individuals had a significantly higher level of harvested total bacterial growth than mature fish (OR=0.82), no statistically significant differences were found between male and female fish (OR=1.01), and the posterior intestinal segment demonstrated a higher degree of culturable bacteria than the anterior segment (OR=4.1). Multiple antibiotic (>3) resistance was observed in 17% of the strains. We propose that a rapid conventional

  9. The microcephaly gene aspm is involved in brain development in zebrafish

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Hyun-Taek; Lee, Mi-Sun; Choi, Jung-Hwa [Department of Biology and GRAST, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Jung, Ju-Yeon [Department of Biotechnology, Konkuk University, Chungju 380-701 (Korea, Republic of); Ahn, Dae-Gwon [Department of Biology and GRAST, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Yeo, Sang-Yeob [Department of Biotechnology, Division of Applied Chemistry and Biotechnology, Hanbat National University, Daejeon 305-719 (Korea, Republic of); Choi, Dong-Kug, E-mail: choidk@kku.ac.kr [Department of Biotechnology, Konkuk University, Chungju 380-701 (Korea, Republic of); Kim, Cheol-Hee, E-mail: zebrakim@cnu.ac.kr [Department of Biology and GRAST, Chungnam National University, Daejeon 305-764 (Korea, Republic of)

    2011-06-17

    Highlights: {yields} We identified a zebrafish aspm/mcph5 gene that is expressed in proliferating cells in the CNS during early development. {yields} Embryos injected with the aspm MO consistently showed a reduced head and eye size but were otherwise grossly normal, closely mimicking the known phenotypes of human microcephaly patients. {yields} Knock-down of aspm causes cell cycle arrest and apoptotic cell death during early development. -- Abstract: MCPH is a neurodevelopmental disorder characterized by a global reduction in cerebral cortical volume. Homozygous mutation of the MCPH5 gene, also known as ASPM, is the most common cause of the MCPH phenotype. To elucidate the roles of ASPM during embryonic development, the zebrafish aspm was identified, which is specifically expressed in proliferating cells in the CNS. Morpholino-mediated knock-down of aspm resulted in a significant reduction in head size. Furthermore, aspm-deficient embryos exhibited a mitotic arrest during early development. These findings suggest that the reduction in brain size in MCPH might be caused by lack of aspm function in the mitotic cell cycle and demonstrate that the zebrafish can provide a model system for congenital diseases of the human nervous system.

  10. The microcephaly gene aspm is involved in brain development in zebrafish

    International Nuclear Information System (INIS)

    Kim, Hyun-Taek; Lee, Mi-Sun; Choi, Jung-Hwa; Jung, Ju-Yeon; Ahn, Dae-Gwon; Yeo, Sang-Yeob; Choi, Dong-Kug; Kim, Cheol-Hee

    2011-01-01

    Highlights: → We identified a zebrafish aspm/mcph5 gene that is expressed in proliferating cells in the CNS during early development. → Embryos injected with the aspm MO consistently showed a reduced head and eye size but were otherwise grossly normal, closely mimicking the known phenotypes of human microcephaly patients. → Knock-down of aspm causes cell cycle arrest and apoptotic cell death during early development. -- Abstract: MCPH is a neurodevelopmental disorder characterized by a global reduction in cerebral cortical volume. Homozygous mutation of the MCPH5 gene, also known as ASPM, is the most common cause of the MCPH phenotype. To elucidate the roles of ASPM during embryonic development, the zebrafish aspm was identified, which is specifically expressed in proliferating cells in the CNS. Morpholino-mediated knock-down of aspm resulted in a significant reduction in head size. Furthermore, aspm-deficient embryos exhibited a mitotic arrest during early development. These findings suggest that the reduction in brain size in MCPH might be caused by lack of aspm function in the mitotic cell cycle and demonstrate that the zebrafish can provide a model system for congenital diseases of the human nervous system.

  11. Granulomatous responses in larval taeniid infections.

    Science.gov (United States)

    Díaz, Á; Sagasti, C; Casaravilla, C

    2018-05-01

    Granulomas are responses to persistent nonliving bodies or pathogens, centrally featuring specialized macrophage forms called epithelioid and multinucleated giant cells. The larval stages of the cestode parasites of the Taeniidae family (Taenia, Echinococcus) develop for years in fixed tissue sites in mammals. In consequence, they are targets of granulomatous responses. The information on tissue responses to larval taeniids is fragmented among host and parasite species and scattered over many decades. We attempt to draw an integrated picture of these responses in solid tissues. The intensity of inflammation around live parasites spans a spectrum from minimal to high, parasite vitality correlating with low inflammation. The low end of the inflammatory spectrum features collagen capsules proximal to the parasites and moderate distal infiltration. The middle of the spectrum is dominated by classical granulomatous responses, whereas the high end features massive eosinophil invasions. Across the range of parasite species, much observational evidence suggests that eosinophils are highly effective at killing larval taeniids in solid tissues, before and during chronic granulomatous responses. The evidence available also suggests that these parasites are adapted to inhibit host granulomatous responses, in part through the exacerbation of host regulatory mechanisms including regulatory T cells and TGF-β. © 2018 John Wiley & Sons Ltd.

  12. Generation and characterization of gsuα:EGFP transgenic zebrafish for evaluating endocrine-disrupting effects

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Xiaoxia [Key Laboratory of Aquatic Biodiversity and Conservation of the Chinese Academy of Sciences, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei (China); University of Chinese Academy of Sciences, Beijing (China); Chen, Xiaowen; Jin, Xia; He, Jiangyan [Key Laboratory of Aquatic Biodiversity and Conservation of the Chinese Academy of Sciences, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei (China); Yin, Zhan, E-mail: zyin@ihb.ac.cn [Key Laboratory of Aquatic Biodiversity and Conservation of the Chinese Academy of Sciences, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei (China); Ningbo Laboratory, State Key Laboratory of Freshwater Ecology and Biotechnology (China)

    2014-07-01

    The glycoprotein subunit α (gsuα) gene encodes the shared α subunit of the three pituitary heterodimeric glycoprotein hormones: follicle-stimulating hormone β (Fshβ), luteinizing hormone β (Lhβ) and thyroid stimulating hormone β (Tshβ). In our current study, we identified and characterized the promoter region of zebrafish gsuα and generated a stable gsuα:EGFP transgenic line, which recapitulated the endogenous gsuα expression in the early developing pituitary gland. A relatively conserved regulatory element set is presented in the promoter regions of zebrafish and three other known mammalian gsuα promoters. Our results also demonstrated that the expression patterns of the gsuα:EGFP transgene were all identical to those expression patterns of the endogenous gsuα expression in the pituitary tissue when our transgenic fish were treated with various endocrine chemicals, including forskolin (FSK), SP600125, trichostatin A (TSA), KClO{sub 4}, dexamethasone (Dex), β-estradiol and progesterone. Thus, this gsuα:EGFP transgenic fish reporter line provides another valuable tool for investigating the lineage development of gsuα-expressing gonadotrophins and the coordinated regulation of various glycoprotein hormone subunit genes. These reporter fish can serve as a novel platform to perform screenings of endocrine-disrupting chemicals (EDCs) in vivo as well. - Highlights: • Identification of the promoter of zebrafish glycoprotein subunit α (gsuα) gene • Generation of stable transmission gsuα:EGFP transgenic zebrafish reporter • Demonstration of the recapitulation of the gsuα:EGFP and endogenous gsuα expression • Suggestion of the gsuα:EGFP transgenic zebrafish as a novel platform for EDC study.

  13. Generation and characterization of gsuα:EGFP transgenic zebrafish for evaluating endocrine-disrupting effects

    International Nuclear Information System (INIS)

    Cheng, Xiaoxia; Chen, Xiaowen; Jin, Xia; He, Jiangyan; Yin, Zhan

    2014-01-01

    The glycoprotein subunit α (gsuα) gene encodes the shared α subunit of the three pituitary heterodimeric glycoprotein hormones: follicle-stimulating hormone β (Fshβ), luteinizing hormone β (Lhβ) and thyroid stimulating hormone β (Tshβ). In our current study, we identified and characterized the promoter region of zebrafish gsuα and generated a stable gsuα:EGFP transgenic line, which recapitulated the endogenous gsuα expression in the early developing pituitary gland. A relatively conserved regulatory element set is presented in the promoter regions of zebrafish and three other known mammalian gsuα promoters. Our results also demonstrated that the expression patterns of the gsuα:EGFP transgene were all identical to those expression patterns of the endogenous gsuα expression in the pituitary tissue when our transgenic fish were treated with various endocrine chemicals, including forskolin (FSK), SP600125, trichostatin A (TSA), KClO 4 , dexamethasone (Dex), β-estradiol and progesterone. Thus, this gsuα:EGFP transgenic fish reporter line provides another valuable tool for investigating the lineage development of gsuα-expressing gonadotrophins and the coordinated regulation of various glycoprotein hormone subunit genes. These reporter fish can serve as a novel platform to perform screenings of endocrine-disrupting chemicals (EDCs) in vivo as well. - Highlights: • Identification of the promoter of zebrafish glycoprotein subunit α (gsuα) gene • Generation of stable transmission gsuα:EGFP transgenic zebrafish reporter • Demonstration of the recapitulation of the gsuα:EGFP and endogenous gsuα expression • Suggestion of the gsuα:EGFP transgenic zebrafish as a novel platform for EDC study

  14. Cytochrome P450 20A1 in zebrafish: Cloning, regulation and potential involvement in hyperactivity disorders

    International Nuclear Information System (INIS)

    Lemaire, Benjamin; Kubota, Akira; O'Meara, Conor M.; Lamb, David C.; Tanguay, Robert L.; Goldstone, Jared V.; Stegeman, John J.

    2016-01-01

    Cytochrome P450 (CYP) enzymes for which there is no functional information are considered “orphan” CYPs. Previous studies showed that CYP20A1, an orphan, is expressed in human hippocampus and substantia nigra, and in zebrafish (Danio rerio) CYP20A1 maternal transcript occurs in eggs, suggesting involvement in brain and in early development. Moreover, hyperactivity is reported in humans with chromosome 2 microdeletions including CYP20A1. We examined CYP20A1 in zebrafish, including impacts of chemical exposure on expression. Zebrafish CYP20A1 cDNA was cloned, sequenced, and aligned with cloned human CYP20A1 and predicted vertebrate orthologs. CYP20A1s share a highly conserved N-terminal region and unusual sequences in the I-helix and the heme-binding CYP signature motifs. CYP20A1 mRNA expression was observed in adult zebrafish organs including the liver, heart, gonads, spleen and brain, as well as the eye and optic nerve. Putative binding sites in proximal promoter regions of CYP20A1s, and response of zebrafish CYP20A1 to selected nuclear and xenobiotic receptor agonists, point to up-regulation by agents involved in steroid hormone response, cholesterol and lipid metabolism. There also was a dose-dependent reduction of CYP20A1 expression in embryos exposed to environmentally relevant levels of methylmercury. Morpholino knockdown of CYP20A1 in developing zebrafish resulted in behavioral effects, including hyperactivity and a slowing of the optomotor response in larvae. The results suggest that altered expression of CYP20A1 might be part of a mechanism linking methylmercury exposure to neurobehavioral deficits. The expanded information on CYP20A1 brings us closer to “deorphanization”, that is, identifying CYP20A1 functions and its roles in health and disease. - Highlights: • The “orphan” CYP20A1 was cloned from zebrafish and its sequence analyzed. • Knockdown of CYP20A1 reduced an optomotor response and elicited bursts of activity. • Effects of

  15. Cytochrome P450 20A1 in zebrafish: Cloning, regulation and potential involvement in hyperactivity disorders

    Energy Technology Data Exchange (ETDEWEB)

    Lemaire, Benjamin; Kubota, Akira; O' Meara, Conor M. [Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA (United States); Lamb, David C. [Institute of Life Science, Medical School, Swansea University, Swansea (United Kingdom); Tanguay, Robert L. [Department of Environmental and Molecular Toxicology, Oregon State University, Corvallis, OR (United States); Goldstone, Jared V. [Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA (United States); Stegeman, John J., E-mail: jstegeman@whoi.edu [Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA (United States)

    2016-04-01

    Cytochrome P450 (CYP) enzymes for which there is no functional information are considered “orphan” CYPs. Previous studies showed that CYP20A1, an orphan, is expressed in human hippocampus and substantia nigra, and in zebrafish (Danio rerio) CYP20A1 maternal transcript occurs in eggs, suggesting involvement in brain and in early development. Moreover, hyperactivity is reported in humans with chromosome 2 microdeletions including CYP20A1. We examined CYP20A1 in zebrafish, including impacts of chemical exposure on expression. Zebrafish CYP20A1 cDNA was cloned, sequenced, and aligned with cloned human CYP20A1 and predicted vertebrate orthologs. CYP20A1s share a highly conserved N-terminal region and unusual sequences in the I-helix and the heme-binding CYP signature motifs. CYP20A1 mRNA expression was observed in adult zebrafish organs including the liver, heart, gonads, spleen and brain, as well as the eye and optic nerve. Putative binding sites in proximal promoter regions of CYP20A1s, and response of zebrafish CYP20A1 to selected nuclear and xenobiotic receptor agonists, point to up-regulation by agents involved in steroid hormone response, cholesterol and lipid metabolism. There also was a dose-dependent reduction of CYP20A1 expression in embryos exposed to environmentally relevant levels of methylmercury. Morpholino knockdown of CYP20A1 in developing zebrafish resulted in behavioral effects, including hyperactivity and a slowing of the optomotor response in larvae. The results suggest that altered expression of CYP20A1 might be part of a mechanism linking methylmercury exposure to neurobehavioral deficits. The expanded information on CYP20A1 brings us closer to “deorphanization”, that is, identifying CYP20A1 functions and its roles in health and disease. - Highlights: • The “orphan” CYP20A1 was cloned from zebrafish and its sequence analyzed. • Knockdown of CYP20A1 reduced an optomotor response and elicited bursts of activity. • Effects of

  16. Exposure to waterborne Cu inhibits cutaneous Na⁺ uptake in post-hatch larval rainbow trout (Oncorhynchus mykiss).

    Science.gov (United States)

    Zimmer, Alex M; Brauner, Colin J; Wood, Chris M

    2014-05-01

    In freshwater rainbow trout (Oncorhynchus mykiss), two common responses to acute waterborne copper (Cu) exposure are reductions in ammonia excretion and Na(+) uptake at the gills, with the latter representing the likely lethal mechanism of action for Cu in adult fish. Larval fish, however, lack a functional gill following hatch and rely predominantly on cutaneous exchange, yet represent the most Cu-sensitive life stage. It is not known if Cu toxicity in larval fish occurs via the skin or gills. The present study utilized divided chambers to assess cutaneous and branchial Cu toxicity over larval development, using disruptions in ammonia excretion (Jamm) and Na(+) uptake (Jin(Na)) as toxicological endpoints. Early in development (early; 3 days post-hatch; dph), approximately 95% of Jamm and 78% of Jin(Na) occurred cutaneously, while in the late developmental stage (late; 25 dph), the gills were the dominant site of exchange (83 and 87% of Jamm and Jin(Na), respectively). Exposure to 50 μg/l Cu led to a 49% inhibition of Jamm in the late developmental stage only, while in the early and middle developmental (mid; 17 dph) stages, Cu had no effect on Jamm. Jin(Na), however, was significantly inhibited by Cu exposure at the early (53% reduction) and late (47% reduction) stages. Inhibition at the early stage of development was mediated by a reduction in cutaneous uptake, representing the first evidence of cutaneous metal toxicity in an intact aquatic organism. The inhibitions of both Jamm and Jin(Na) in the late developmental stage occurred via a reduction in branchial exchange only. The differential responses of the skin and gills to Cu exposure suggest that the mechanisms of Jamm and Jin(Na) and/or Cu toxicity differ between these tissues. Exposure to 20μg/l Cu revealed that Jamm is the more Cu-sensitive process. The results presented here have important implications in predicting metal toxicity in larval fish. The Biotic Ligand Model (BLM) is currently used to predict

  17. Zooplankton diversity and the predatory impact by larval and small juvenile fish at the Fisher Banks in the North Sea

    DEFF Research Database (Denmark)

    Nielsen, Torkel Gissel; Munk, Peter

    1998-01-01

    The biomass and diversity of the mesozooplankton and fish larvae community were investigated across a frontal zone in the central North Sea in the early summer, to investigate whether larval fish predation is a regulator of mesozooplankton production. Pronounced changes in the mesozooplankton com...

  18. Zebrafish swimming in the flow: a particle image velocimetry study

    Directory of Open Access Journals (Sweden)

    Violet Mwaffo

    2017-11-01

    Full Text Available Zebrafish is emerging as a species of choice for the study of a number of biomechanics problems, including balance development, schooling, and neuromuscular transmission. The precise quantification of the flow physics around swimming zebrafish is critical toward a mechanistic understanding of the complex swimming style of this fresh-water species. Although previous studies have elucidated the vortical structures in the wake of zebrafish swimming in placid water, the flow physics of zebrafish swimming against a water current remains unexplored. In an effort to illuminate zebrafish swimming in a dynamic environment reminiscent of its natural habitat, we experimentally investigated the locomotion and hydrodynamics of a single zebrafish swimming in a miniature water tunnel using particle image velocimetry. Our results on zebrafish locomotion detail the role of flow speed on tail beat undulations, heading direction, and swimming speed. Our findings on zebrafish hydrodynamics offer a precise quantification of vortex shedding during zebrafish swimming and demonstrate that locomotory patterns play a central role on the flow physics. This knowledge may help clarify the evolutionary advantage of burst and cruise swimming movements in zebrafish.

  19. Fish larval transport in the coastal waters through ecological modelling

    Digital Repository Service at National Institute of Oceanography (India)

    George, G.

    are as follows: (i) to find out the influence of environmental parameters on the biology of the given ecosystem (ii) to track larval transport and biological abundance in relation to environmental vari- ables (iii) to compare biological abundance and fish larval... include the following investigations: (i) analysis of satellite chlorophyll data along the southwest coastal waters of India to derive a biological calender for sardine (ii) tracking the larval survival and establish a link between food and sardine inter...

  20. Thyroid endocrine disruption and external body morphology of Zebrafish

    Science.gov (United States)

    Sharma, Prakash; Grabowski, Timothy B.; Patino, Reynaldo

    2016-01-01

    This study examined the effects thyroid-active compounds during early development on body morphology of Zebrafish (Danio rerio). Three-day postfertilization (dpf) larvae were exposed to goitrogen [methimazole (MZ, 0.15 mM)], combination of MZ (0.15 mM) and thyroxine (T4, 2 nM), T4 (2 nM), or control (reconstituted water) treatments until 33 dpf and subsequently maintained in reconstituted water until 45 dpf. Samples were taken at 33 and 45 dpf for multivariate analysis of geometric distances between selected homologous landmarks placed on digital images of fish, and for histological assessment of thyrocytes. Body mass, standard length, and pectoral fin length were separately measured on remaining fish at 45 dpf. Histological analysis confirmed the hypothyroid effect (increased thyrocyte height) of MZ and rescue effect of T4 co-administration. Geometric distance analysis showed that pectoral and pelvic fins shifted backward along the rostrocaudal axis under hypothyroid conditions at 45 dpf and that T4 co-treatment prevented this shift. Pectoral fin length at 45 dpf was reduced by exposure to MZ and rescued by co-administration of T4, but it was not associated with standard length. Methimazole caused a reduction in body mass and length at 45 dpf that could not be rescued by T4 co-administration, and non-thyroidal effects of MZ on body shape were also recognized at 33 and 45 dpf. Alterations in the length and position of paired fins caused by exposure to thyroid-disrupting chemicals during early development, as shown here for Zebrafish, could affect physical aspects of locomotion and consequently other important organismal functions such as foraging, predator avoidance, and ultimately survival and recruitment into the adult population. Results of this study also suggest the need to include rescue treatments in endocrine disruption studies that rely on goitrogens as reference for thyroid-mediated effects.

  1. Development of the acoustically evoked behavioral response in larval plainfin midshipman fish, Porichthys notatus.

    Directory of Open Access Journals (Sweden)

    Peter W Alderks

    Full Text Available The ontogeny of hearing in fishes has become a major interest among bioacoustics researchers studying fish behavior and sensory ecology. Most fish begin to detect acoustic stimuli during the larval stage which can be important for navigation, predator avoidance and settlement, however relatively little is known about the hearing capabilities of larval fishes. We characterized the acoustically evoked behavioral response (AEBR in the plainfin midshipman fish, Porichthys notatus, and used this innate startle-like response to characterize this species' auditory capability during larval development. Age and size of larval midshipman were highly correlated (r(2 = 0.92. The AEBR was first observed in larvae at 1.4 cm TL. At a size ≥ 1.8 cm TL, all larvae responded to a broadband stimulus of 154 dB re1 µPa or -15.2 dB re 1 g (z-axis. Lowest AEBR thresholds were 140-150 dB re 1 µPa or -33 to -23 dB re 1 g for frequencies below 225 Hz. Larval fish with size ranges of 1.9-2.4 cm TL had significantly lower best evoked frequencies than the other tested size groups. We also investigated the development of the lateral line organ and its function in mediating the AEBR. The lateral line organ is likely involved in mediating the AEBR but not necessary to evoke the startle-like response. The midshipman auditory and lateral line systems are functional during early development when the larvae are in the nest and the auditory system appears to have similar tuning characteristics throughout all life history stages.

  2. Development of the acoustically evoked behavioral response in larval plainfin midshipman fish, Porichthys notatus.

    Science.gov (United States)

    Alderks, Peter W; Sisneros, Joseph A

    2013-01-01

    The ontogeny of hearing in fishes has become a major interest among bioacoustics researchers studying fish behavior and sensory ecology. Most fish begin to detect acoustic stimuli during the larval stage which can be important for navigation, predator avoidance and settlement, however relatively little is known about the hearing capabilities of larval fishes. We characterized the acoustically evoked behavioral response (AEBR) in the plainfin midshipman fish, Porichthys notatus, and used this innate startle-like response to characterize this species' auditory capability during larval development. Age and size of larval midshipman were highly correlated (r(2) = 0.92). The AEBR was first observed in larvae at 1.4 cm TL. At a size ≥ 1.8 cm TL, all larvae responded to a broadband stimulus of 154 dB re1 µPa or -15.2 dB re 1 g (z-axis). Lowest AEBR thresholds were 140-150 dB re 1 µPa or -33 to -23 dB re 1 g for frequencies below 225 Hz. Larval fish with size ranges of 1.9-2.4 cm TL had significantly lower best evoked frequencies than the other tested size groups. We also investigated the development of the lateral line organ and its function in mediating the AEBR. The lateral line organ is likely involved in mediating the AEBR but not necessary to evoke the startle-like response. The midshipman auditory and lateral line systems are functional during early development when the larvae are in the nest and the auditory system appears to have similar tuning characteristics throughout all life history stages.

  3. Zebrafish in Toxicology and Environmental Health.

    Science.gov (United States)

    Bambino, Kathryn; Chu, Jaime

    2017-01-01

    As manufacturing processes and development of new synthetic compounds increase to keep pace with the expanding global demand, environmental health, and the effects of toxicant exposure are emerging as critical public health concerns. Additionally, chemicals that naturally occur in the environment, such as metals, have profound effects on human and animal health. Many of these compounds are in the news: lead, arsenic, and endocrine disruptors such as bisphenol A have all been widely publicized as causing disease or damage to humans and wildlife in recent years. Despite the widespread appreciation that environmental toxins can be harmful, there is limited understanding of how many toxins cause disease. Zebrafish are at the forefront of toxicology research; this system has been widely used as a tool to detect toxins in water samples and to investigate the mechanisms of action of environmental toxins and their related diseases. The benefits of zebrafish for studying vertebrate development are equally useful for studying teratogens. Here, we review how zebrafish are being used both to detect the presence of some toxins as well as to identify how environmental exposures affect human health and disease. We focus on areas where zebrafish have been most effectively used in ecotoxicology and in environmental health, including investigation of exposures to endocrine disruptors, industrial waste byproducts, and arsenic. © 2017 Elsevier Inc. All rights reserved.

  4. Visualizing infections and immune mechanisms in zebrafish

    DEFF Research Database (Denmark)

    Jørgensen, Louise von Gersdorff; Korbut, Rozalia; Mehrdana, Foojan

    , immunological reactions during e.g. transplant rejections or the spread and pathogenicity of pathogens. We have, in our laboratory, used the zebrafish as a model for aquacultured fish species and their pathogens. We have 1) visualized antigen uptake in vivo following a bath in a soup containing fluorescent...

  5. Highly Efficient ENU Mutagenesis in Zebrafish.

    NARCIS (Netherlands)

    de Bruijn, E.; Cuppen, E.; Feitsma, H.

    2009-01-01

    ENU (N-ethyl-N-nitrosourea) mutagenesis is a widely accepted and proven method to introduce random point mutations in the genome. Because there are no targeted knockout strategies available for zebrafish so far, random mutagenesis is currently the preferred method in both forward and reverse genetic

  6. Molecular genetics of pituitary development in zebrafish.

    Science.gov (United States)

    Pogoda, Hans-Martin; Hammerschmidt, Matthias

    2007-08-01

    The pituitary gland of vertebrates consists of two major parts, the neurohypophysis (NH) and the adenohypophysis (AH). As a central part of the hypothalamo-hypophyseal system (HHS), it constitutes a functional link between the nervous and the endocrine system to regulate basic body functions, such as growth, metabolism and reproduction. The development of the AH has been intensively studied in mouse, serving as a model for organogenesis and differential cell specification. However, given that the AH is a relatively recent evolutionary advance of the chordate phylum, it is also interesting to understand its development in lower chordate systems. In recent years, the zebrafish has emerged as a powerful lower vertebrate system for developmental studies, being amenable for large-scale genetic approaches, embryological manipulations, and in vivo imaging. Here, we present an overview of current knowledge of the mechanisms and genetic control of pituitary formation during zebrafish development. First, we describe the components of the zebrafish HHS, and the different pituitary cell types and hormones, followed by a description of the different steps of normal pituitary development. The central part of the review deals with the genes found to be essential for zebrafish AH development, accompanied by a description of the corresponding mutant phenotypes. Finally, we discuss future directions, with particular focus on evolutionary aspects, and some novel functional aspects with growing medical and social relevance.

  7. A zebrafish model of inflammatory lymphangiogenesis

    Directory of Open Access Journals (Sweden)

    Kazuhide S. Okuda

    2015-10-01

    Full Text Available Inflammatory bowel disease (IBD is a disabling chronic inflammatory disease of the gastrointestinal tract. IBD patients have increased intestinal lymphatic vessel density and recent studies have shown that this may contribute to the resolution of IBD. However, the molecular mechanisms involved in IBD-associated lymphangiogenesis are still unclear. In this study, we established a novel inflammatory lymphangiogenesis model in zebrafish larvae involving colitogenic challenge stimulated by exposure to 2,4,6-trinitrobenzenesulfonic acid (TNBS or dextran sodium sulphate (DSS. Treatment with either TNBS or DSS resulted in vascular endothelial growth factor receptor (Vegfr-dependent lymphangiogenesis in the zebrafish intestine. Reduction of intestinal inflammation by the administration of the IBD therapeutic, 5-aminosalicylic acid, reduced intestinal lymphatic expansion. Zebrafish macrophages express vascular growth factors vegfaa, vegfc and vegfd and chemical ablation of these cells inhibits intestinal lymphatic expansion, suggesting that the recruitment of macrophages to the intestine upon colitogenic challenge is required for intestinal inflammatory lymphangiogenesis. Importantly, this study highlights the potential of zebrafish as an inflammatory lymphangiogenesis model that can be used to investigate the role and mechanism of lymphangiogenesis in inflammatory diseases such as IBD.

  8. Axonal regeneration in zebrafish spinal cord

    Science.gov (United States)

    Hui, Subhra Prakash

    2018-01-01

    Abstract In the present review we discuss two interrelated events—axonal damage and repair—known to occur after spinal cord injury (SCI) in the zebrafish. Adult zebrafish are capable of regenerating axonal tracts and can restore full functionality after SCI. Unlike fish, axon regeneration in the adult mammalian central nervous system is extremely limited. As a consequence of an injury there is very little repair of disengaged axons and therefore functional deficit persists after SCI in adult mammals. In contrast, peripheral nervous system axons readily regenerate following injury and hence allow functional recovery both in mammals and fish. A better mechanistic understanding of these three scenarios could provide a more comprehensive insight into the success or failure of axonal regeneration after SCI. This review summarizes the present understanding of the cellular and molecular basis of axonal regeneration, in both the peripheral nervous system and the central nervous system, and large scale gene expression analysis is used to focus on different events during regeneration. The discovery and identification of genes involved in zebrafish spinal cord regeneration and subsequent functional experimentation will provide more insight into the endogenous mechanism of myelination and remyelination. Furthermore, precise knowledge of the mechanism underlying the extraordinary axonal regeneration process in zebrafish will also allow us to unravel the potential therapeutic strategies to be implemented for enhancing regrowth and remyelination of axons in mammals. PMID:29721326

  9. Effects of alpha particles on zebrafish embryos

    International Nuclear Information System (INIS)

    Yum, E.H.W.; Choi, V.W.Y.; Yu, K.N.; Li, V.W.T.; Cheng, S.H.

    2008-01-01

    Full text: Ionizing radiation such as X-ray and alpha particles can damage cellular macromolecules, which can lead to DNA single- and double-strand breaks. In the present work, we studied the effects of alpha particles on dechorionated zebrafish embryos. Thin polyallyldiglycol carbonate (PADC) films with a thickness of 16 μm were prepared from commercially available PADC films (with thickness of 100 μm) by chemical etching and used as support substrates for holding zebrafish embryos for alpha-particle irradiation. These films recorded alpha-particle hit positions, quantified the number and energy of alpha particles actually incident on the embryo cells, and thus enabled the calculation of the dose absorbed by the embryo cells. Irradiation was made at 1.25 hours post fertilization (hpf) with various absorbed dose. TdT-mediated dUTP Nick-End Labeling (TUNEL) assay was performed on the embryos at different time stages after irradiation. Marked apoptosis was detected only in embryos at earlier time stages. The results showed that DNA double-strand break during zebrafish embryogenesis can be induced by alpha-particle irradiation, which suggests that zebrafish is a potential model for assessing the effects of alpha-particle radiation

  10. Effects of Tail Clipping on Larval Performance and Tail Regeneration Rates in the Near Eastern Fire Salamander, Salamandra infraimmaculata.

    Directory of Open Access Journals (Sweden)

    Ori Segev

    Full Text Available Tail-tip clipping is a common technique for collecting tissue samples from amphibian larvae and adults. Surprisingly, studies of this invasive sampling procedure or of natural tail clipping--i.e., bites inflicted by predators including conspecifics--on the performance and fitness of aquatic larval stages of urodeles are scarce. We conducted two studies in which we assessed the effects of posterior tail clipping (~30 percent of tail on Near Eastern fire salamander (Salamandra infraimmaculata larvae. In a laboratory study, we checked regeneration rates of posterior tail-tip clipping at different ages. Regeneration rates were hump-shaped, peaking at the age of ~30 days and then decreasing. This variation in tail regeneration rates suggests tradeoffs in resource allocation between regeneration and somatic growth during early and advanced development. In an outdoor artificial pond experiment, under constant larval densities, we assessed how tail clipping of newborn larvae affects survival to, time to, and size at metamorphosis. Repeated measures ANOVA on mean larval survival per pond revealed no effect of tail clipping. Tail clipping had correspondingly no effect on larval growth and development expressed in size (mass and snout-vent length at, and time to, metamorphosis. We conclude that despite the given variation in tail regeneration rates throughout larval ontogeny, clipping of 30% percent of the posterior tail area seems to have no adverse effects on larval fitness and survival. We suggest that future use of this imperative tool for the study of amphibian should take into account larval developmental stage during the time of application and not just the relative size of the clipped tail sample.

  11. Comparative analysis of biological effect of corannulene and graphene on developmental and sleep/wake profile of zebrafish larvae.

    Science.gov (United States)

    Li, Xiang; Zhang, Yuan; Li, Xu; Feng, DaoFu; Zhang, ShuHui; Zhao, Xin; Chen, DongYan; Zhang, ZhiXiang; Feng, XiZeng

    2017-06-01

    Little is known about the biological effect of non-planar polycyclic aromatic hydrocarbons (PAH) such as corannulene on organisms. In this study, we compared the effect of corannulene (non-planar PAH) and graphene (planar PAH) on embryonic development and sleep/wake behaviors of larval zebrafish. First, the toxicity of graded doses of corannulene (1, 10, and 50μg/mL) was tested in developing zebrafish embryos. Corannulene showed minimal developmental toxicity only induced an epiboly delay. Further, a significant decrease in locomotion/increase in sleep was observed in larvae treated with the highest dose (50μg/mL) of corannulene while no significant locomotion alterations were induced by graphene. Finally, the effect of corannulene or graphene on the hypocretin (hcrt) system and sleep/wake regulators such as hcrt, hcrt G-protein coupled receptor (hcrtr), and arylalkylamine N-acetyltransferase-2 (aanat2) was evaluated. Corannulene increased sleep and reduced locomotor activity and the expression of hcrt and hcrtr mRNA while graphene did not obviously disturb the sleep behavior and gene expression patterns. These results suggest that the corannulene has the potential to cause hypnosis-like behavior in larvae and provides a fundamental comparative understanding of the effects of corannulene and graphene on biology systems. Little is known about the biological effect of non-planar polycyclic aromatic hydrocarbons (PAH) such as corannulene on organisms. Here, we compare the effect of corannulene (no-planar PAH) and graphene (planar PAH) on embryonic development and sleep/wake behaviors of larval zebrafish. And we aim to investigate the effect of curvature on biological system. First, toxicity of corannulene over the range of doses (1μg/mL, 10μg/mL and 50μg/mL) was tested in developing zebrafish embryos. Corannulene has minimal developmental toxicity, only incurred epiboly delay. Subsequently, a significant decrease in locomotion/increase in sleep at the highest

  12. Evaluating sampling strategies for larval cisco (Coregonus artedi)

    Science.gov (United States)

    Myers, J.T.; Stockwell, J.D.; Yule, D.L.; Black, J.A.

    2008-01-01

    To improve our ability to assess larval cisco (Coregonus artedi) populations in Lake Superior, we conducted a study to compare several sampling strategies. First, we compared density estimates of larval cisco concurrently captured in surface waters with a 2 x 1-m paired neuston net and a 0.5-m (diameter) conical net. Density estimates obtained from the two gear types were not significantly different, suggesting that the conical net is a reasonable alternative to the more cumbersome and costly neuston net. Next, we assessed the effect of tow pattern (sinusoidal versus straight tows) to examine if propeller wash affected larval density. We found no effect of propeller wash on the catchability of larval cisco. Given the availability of global positioning systems, we recommend sampling larval cisco using straight tows to simplify protocols and facilitate straightforward measurements of volume filtered. Finally, we investigated potential trends in larval cisco density estimates by sampling four time periods during the light period of a day at individual sites. Our results indicate no significant trends in larval density estimates during the day. We conclude estimates of larval cisco density across space are not confounded by time at a daily timescale. Well-designed, cost effective surveys of larval cisco abundance will help to further our understanding of this important Great Lakes forage species.

  13. Embryonic Zebrafish Model - A Well-Established Method for Rapidly Assessing the Toxicity of Homeopathic Drugs - Toxicity Evaluation of Homeopathic Drugs Using Zebrafish Embryo Model -

    Directory of Open Access Journals (Sweden)

    Himanshu R Gupta

    2016-12-01

    Full Text Available Objectives: Advancements in nanotechnology have led to nanoparticle (NP use in various fields of medicine. Although the potential of NPs is promising, the lack of documented evidence on the toxicological effects of NPs is concerning. A few studies have documented that homeopathy uses NPs. Unfortunately, very few sound scientific studies have explored the toxic effects of homeopathic drugs. Citing this lack of high-quality scientific evidence, regulatory agencies have been reluctant to endorse homeopathic treatment as an alternative or adjunct treatment. This study aimed to enhance our insight into the impact of commercially-available homeopathic drugs, to study the presence of NPs in those drugs and any deleterious effects they might have, and to determine the distribution pattern of NPs in zebrafish embryos (Danio rerio. Methods: Homeopathic dilutions were studied using high-resolution transmission electron microscopy with selected area electron diffraction (SAED. For the toxicity assessment on Zebrafish, embryos were exposed to a test solution from 4 - 6 hours post-fertilization, and embryos/larvae were assessed up to 5 days post-fertilization (dpf for viability and morphology. Toxicity was recorded in terms of mortality, hatching delay, phenotypic defects and metal accumulation. Around 5 dpf was found to be the optimum developmental stage for evaluation. Results: The present study aimed to conclusively prove the presence of NPs in all high dilutions of homeopathic drugs. Embryonic zebrafish were exposed to three homeopathic drugs with two potencies (30CH, 200CH during early embryogenesis. The resulting morphological and cellular responses were observed. Exposure to these potencies produced no visibly significant malformations, pericardial edema, and mortality and no necrotic and apoptotic cellular death. Conclusion: Our findings clearly demonstrate that no toxic effects were observed for these three homeopathic drugs at the potencies and

  14. Embryonic Zebrafish Model - A Well-Established Method for Rapidly Assessing the Toxicity of Homeopathic Drugs: - Toxicity Evaluation of Homeopathic Drugs Using Zebrafish Embryo Model.

    Science.gov (United States)

    Gupta, Himanshu R; Patil, Yogesh; Singh, Dipty; Thakur, Mansee

    2016-12-01

    Advancements in nanotechnology have led to nanoparticle (NP) use in various fields of medicine. Although the potential of NPs is promising, the lack of documented evidence on the toxicological effects of NPs is concerning. A few studies have documented that homeopathy uses NPs. Unfortunately, very few sound scientific studies have explored the toxic effects of homeopathic drugs. Citing this lack of high-quality scientific evidence, regulatory agencies have been reluctant to endorse homeopathic treatment as an alternative or adjunct treatment. This study aimed to enhance our insight into the impact of commercially-available homeopathic drugs, to study the presence of NPs in those drugs and any deleterious effects they might have, and to determine the distribution pattern of NPs in zebrafish embryos ( Danio rerio ). Homeopathic dilutions were studied using high-resolution transmission electron microscopy with selected area electron diffraction (SAED). For the toxicity assessment on Zebrafish, embryos were exposed to a test solution from 4 - 6 hours post-fertilization, and embryos/larvae were assessed up to 5 days post-fertilization (dpf) for viability and morphology. Toxicity was recorded in terms of mortality, hatching delay, phenotypic defects and metal accumulation. Around 5 dpf was found to be the optimum developmental stage for evaluation. The present study aimed to conclusively prove the presence of NPs in all high dilutions of homeopathic drugs. Embryonic zebrafish were exposed to three homeopathic drugs with two potencies (30CH, 200CH) during early embryogenesis. The resulting morphological and cellular responses were observed. Exposure to these potencies produced no visibly significant malformations, pericardial edema, and mortality and no necrotic and apoptotic cellular death. Our findings clearly demonstrate that no toxic effects were observed for these three homeopathic drugs at the potencies and exposure times used in this study. The embryonic zebrafish

  15. Oxidative stress and DNA damage induced by imidacloprid in zebrafish (Danio rerio).

    Science.gov (United States)

    Ge, Weili; Yan, Saihong; Wang, Jinhua; Zhu, Lusheng; Chen, Aimei; Wang, Jun

    2015-02-18

    Imidacloprid is a neonicotinoid insecticide that can have negative effects on nontarget animals. The present study was conducted to assess the toxicity of various imidacloprid doses (0.3, 1.25, and 5 mg/mL) on zebrafish sampled after 7, 14, 21, and 28 days of exposure. The levels of catalase (CAT), superoxide dismutase (SOD), reactive oxygen species (ROS), glutathione-S-transferase (GST), and malondialdehyde (MDA) and the extent of DNA damage were measured to evaluate the toxicity of imidacloprid on zebrafish. SOD and GST activities were noticeably increased during early exposure but were inhibited toward the end of the exposure period. In addition, the CAT levels decreased to the control level following their elevation during early exposure. High concentrations of imidacloprid (1.25 and 5 mg/L) induced excessive ROS production and markedly increased MDA content on the 21st day of exposure. DNA damage was dose- and time-dependent. In conclusion, the present study showed that imidacloprid can induce oxidative stress and DNA damage in zebrafish.

  16. Discovery, characterization and expression of a novel zebrafish gene, znfr, important for notochord formation.

    Science.gov (United States)

    Xu, Yan; Zou, Peng; Liu, Yao; Deng, Fengjiao

    2010-06-01

    Genes specifically expressed in the notochord may be crucial for proper notochord development. Using the digital differential display program offered by the National Center for Biotechnology Information, we identified a novel EST sequence from a zebrafish ovary library (No. XM_701450). The full-length cDNA of this transcript was cloned by performing 3' and 5'-RACE and was further confirmed by PCR and sequencing. The resulting 614 bp gene was found to encode a novel 94 amino acid protein that did not share significant homology with any other known protein. Characterization of the genomic sequence revealed that the gene spanned 4.9 kb and was composed of four exons and three introns. RT-PCR gene expression analysis revealed that our gene of interest was expressed in ovary, kidney, brain, mature oocytes and during the early stages of embryogenesis. During embryonic development, znfr mRNA was found to be expressed in the embryonic shield, chordamesoderm and the vacuolated notochord cells by in situ hybridization. Based on this information, we hypothesize that this novel gene is an important maternal factor required for zebrafish notochord formation during early embryonic development. We have thus named this gene znfr (zebrafish notochord formation related).

  17. Defects of the Glycinergic Synapse in Zebrafish

    Science.gov (United States)

    Ogino, Kazutoyo; Hirata, Hiromi

    2016-01-01

    Glycine mediates fast inhibitory synaptic transmission. Physiological importance of the glycinergic synapse is well established in the brainstem and the spinal cord. In humans, the loss of glycinergic function in the spinal cord and brainstem leads to hyperekplexia, which is characterized by an excess startle reflex to sudden acoustic or tactile stimulation. In addition, glycinergic synapses in this region are also involved in the regulation of respiration and locomotion, and in the nociceptive processing. The importance of the glycinergic synapse is conserved across vertebrate species. A teleost fish, the zebrafish, offers several advantages as a vertebrate model for research of glycinergic synapse. Mutagenesis screens in zebrafish have isolated two motor defective mutants that have pathogenic mutations in glycinergic synaptic transmission: bandoneon (beo) and shocked (sho). Beo mutants have a loss-of-function mutation of glycine receptor (GlyR) β-subunit b, alternatively, sho mutant is a glycinergic transporter 1 (GlyT1) defective mutant. These mutants are useful animal models for understanding of glycinergic synaptic transmission and for identification of novel therapeutic agents for human diseases arising from defect in glycinergic transmission, such as hyperekplexia or glycine encephalopathy. Recent advances in techniques for genome editing and for imaging and manipulating of a molecule or a physiological process make zebrafish more attractive model. In this review, we describe the glycinergic defective zebrafish mutants and the technical advances in both forward and reverse genetic approaches as well as in vivo visualization and manipulation approaches for the study of the glycinergic synapse in zebrafish. PMID:27445686

  18. Turbulence-enhanced prey encounter rates in larval fish : Effects of spatial scale, larval behaviour and size

    DEFF Research Database (Denmark)

    Kiørboe, Thomas; MacKenzie, Brian

    1995-01-01

    Turbulent water motion has several effects on the feeding ecology of larval fish and other planktivorous predators. In this paper, we consider the appropriate spatial scales for estimating relative velocities between larval fish predators and their prey, and the effect that different choices of s...... in the range in which turbulent intensity has an overall positive effect on larval fish ingestion rate probability. However, experimental data to test the model predictions are lacking. We suggest that the model inputs require further empirical study....

  19. Transcriptome data on maternal RNA of 24 individual zebrafish eggs from five sibling mothers

    Directory of Open Access Journals (Sweden)

    Johanna F.B. Pagano

    2016-09-01

    Full Text Available Maternal mRNA that is present in the mature oocyte plays an important role in the proper development of the early embryo. To elucidate the role of the maternal transcriptome we recently reported a microarray study on individual zebrafish eggs from five different clutches from sibling mothers and showed differences in maternal RNA abundance between and within clutches, “Mother-specific signature in the maternal transcriptome composition of mature, unfertilized Eggs” [1]. Here we provide in detail the applied preprocessing method as well as the R-code to identify expressed and non-expressed genes in the associated transcriptome dataset. Additionally, we provide a website that allows a researcher to search for the expression of their gene of interest in this experiment. Keywords: Zebrafish, Danio rerio, Egg transcriptome, Single egg

  20. Oral exposure of adult zebrafish (Danio rerio) to 2,4,6-tribromophenol affects reproduction

    DEFF Research Database (Denmark)

    Halden, Anna Norman; Nyholm, Jenny Rattfelt; Andersson, Patrik L

    2010-01-01

    The bromophenol 2,4,6-tribromophenol (TBP) is widely used as an industrial chemical, formed by degradation of tetrabromobisphenol-A, and it occurs naturally in marine organisms. Concentrations of TBP in fish have been related to intake via feed, but little is known about effects on fish health...... after oral exposure. In this study, we exposed adult male and female zebrafish (Danio rerio) to TBP via feed in nominal concentrations of 33, 330, and 3300 mu g/g feed (or control feed) for 6 weeks to assess the effects of TBP on reproductive output, gonad morphology, circulatory vitellogenin levels......, and early embryo development. The aim was also to investigate the extent to which TBP was metabolised to 2,4,6-tribromoanisole (IBA) in dietary exposed zebrafish, and the amounts of TBP and TBA found in offspring. After 6 weeks of exposure, we found about 3% of the daily dose of TBP in adult fish...

  1. Microhabitat influence on larval fish assemblages within ...

    Science.gov (United States)

    We examined larval and juvenile fish assemblage structure in relation to microhabitat variables within the St. Louis River estuary, a drowned river mouth of Lake Superior. Fish were sampled in vegetated beds throughout the estuary, across a gradient of vegetation types and densities (including disturbed, preserved and post-restoration sites). Canonical correspondence analysis, relating species abundances to environmental variables revealed that plant species richness, turbidity and aquatic plant cover were most influential in structuring assemblages. Results from this microhabitat analysis at this crucial life stage has potential to inform wetland restoration efforts within the St. Louis River and other Great Lake coastal wetlands. not applicable

  2. Relative importance of temperature and diet to larval development and adult size of the winter stonefly, Soyedina carolinensis (Plecoptera: Nemouridae)

    Energy Technology Data Exchange (ETDEWEB)

    Sweeney, B.W.; Vannote, R.L.; Dodds, P.J.

    1986-02-01

    Soyedina carolinensis Claassen, a leaf shredding stonefly, was reared in a series of three laboratory experiments from early instar to adult on different species of deciduous leaves and at various constant and fluctuating temperature regimes. Experiment 1, which involved rearing larvae on fourteen different leaf diets at ambient stream temperatures, showed that diet significantly affected larval growth and adult size but did not affect overall developmental time. Experiment 2, which involved rearing larvae on five different leaf diets at each of three fluctuating temperature regimes, showed that: adding 6/sup 0/C to the normal temperature regime of WCC was lethal to 99% of the larvae regardless of diet; and warming WCC by 3/sup 0/C did not affect developmental time but did significantly reduce adult size relative to adults reared at WCC temperatures on certain diets. Experiment 3, which involved rearing larvae on five different leaf diets at each of five constant temperatures showed that: temperature significantly affected the mortality, growth, and development time of larvae whereas diet only affected larval growth and mortality; temperatures at or near 10/sup 0/C yielded maximum larval growth and survival for most diets; at 5/sup 0/C, larval mortality was high and growth was low resulting in a few small adults for most diets; larval mortality was at or near 100% at 15/sup 0/C regardless of diet; and no larvae survived at 20 and 25/sup 0/C.

  3. Detection of vitellogenin incorporation into zebrafish oocytes by FITC fluorescence

    Directory of Open Access Journals (Sweden)

    Yokoi Hayato

    2011-04-01

    Full Text Available Abstract Background Large volumes of lymph can be collected from the eye-sacs of bubble-eye goldfish. We attempted to induce vitellogenin (Vtg in the eye-sac lymph of bubble-eye goldfish and develop a method for visualizing Vtg incorporation by zebrafish oocytes using FITC-labeling. Methods Estrogen efficiently induced Vtg in the eye-sac lymph of goldfish. After FITC-labeled Vtg was prepared, it was injected into mature female zebrafish. Results Incorporation of FITC-labeled Vtg by zebrafish oocytes was detected in in vivo and in vitro experiments. The embryos obtained from zebrafish females injected with FITC-labeled Vtg emitted FITC fluorescence from the yolk sac and developed normally. Conclusion This method for achieving Vtg incorporation by zebrafish oocytes could be useful in experiments related to the development and endocrinology of zebrafish oocytes.

  4. Analysis of Lethality and Malformations During Zebrafish (Danio rerio) Development.

    Science.gov (United States)

    Raghunath, Azhwar; Perumal, Ekambaram

    2018-01-01

    The versatility offered by zebrafish (Danio rerio) makes it a powerful and an attractive vertebrate model in developmental toxicity and teratogenicity assays. Apart from the newly introduced chemicals as drugs, xenobiotics also induce abnormal developmental abnormalities and congenital malformations in living organisms. Over the recent decades, zebrafish embryo/larva has emerged as a potential tool to test teratogenicity potential of these chemicals. Zebrafish responds to compounds as mammals do as they share similarities in their development, metabolism, physiology, and signaling pathways with that of mammals. The methodology used by the different scientists varies enormously in the zebrafish embryotoxicity test. In this chapter, we present methods to assess lethality and malformations during zebrafish development. We propose two major malformations scoring systems: binomial and relative morphological scoring systems to assess the malformations in zebrafish embryos/larvae. Based on the scoring of the malformations, the test compound can be classified as a teratogen or a nonteratogen and its teratogenic potential is evaluated.

  5. Biosecurity and Health Monitoring at the Zebrafish International Resource Center.

    Science.gov (United States)

    Murray, Katrina N; Varga, Zoltán M; Kent, Michael L

    2016-07-01

    The Zebrafish International Resource Center (ZIRC) is a repository and distribution center for mutant, transgenic, and wild-type zebrafish. In recent years annual imports of new zebrafish lines to ZIRC have increased tremendously. In addition, after 15 years of research, we have identified some of the most virulent pathogens affecting zebrafish that should be avoided in large production facilities, such as ZIRC. Therefore, while importing a high volume of new lines we prioritize safeguarding the health of our in-house fish colony. Here, we describe the biosecurity and health-monitoring program implemented at ZIRC. This strategy was designed to prevent introduction of new zebrafish pathogens, minimize pathogens already present in the facility, and ensure a healthy zebrafish colony for in-house uses and shipment to customers.

  6. Laser capture microdissection of gonads from juvenile zebrafish

    DEFF Research Database (Denmark)

    Jørgensen, Anne; Nielsen, John; Morthorst, Jane Ebsen

    2009-01-01

    was adjusted and optimised to isolate juvenile zebrafish gonads. Results: The juvenile zebrafish gonad is not morphologically distinguishable when using dehydrated cryosections on membrane slides and a specific staining method is necessary to identify the gonads. The protocol setup in this study allows......Background: Investigating gonadal gene expression is important in attempting to elucidate the molecular mechanism of sex determination and differentiation in the model species zebrafish. However, the small size of juvenile zebrafish and correspondingly their gonads complicates this type...... of investigation. Furthermore, the lack of a genetic sex marker in juvenile zebrafish prevents pooling gonads from several individuals. The aim of this study was to establish a method to isolate the gonads from individual juvenile zebrafish allowing future investigations of gonadal gene expression during sex...

  7. In Vivo Quantitative Study of Sized-Dependent Transport and Toxicity of Single Silver Nanoparticles Using Zebrafish Embryos

    Science.gov (United States)

    Lee, Kerry J.; Browning, Lauren M.; Nallathamby, Prakash D.; Desai, Tanvi; Cherukui, Pavan K.; Xu, Xiao-Hong Nancy

    2012-01-01

    Nanomaterials possess distinctive physicochemical properties (e.g., small sizes, high surface area-to-volume ratios) and promise a wide variety of applications, ranging from design of high quality consumer products to effective disease diagnosis and therapy. These properties can lead to toxic effects, potentially hindering advance in nanotechnology. In this study, we have synthesized and characterized purified and stable (non-aggregation) silver nanoparticles (Ag NPs, 41.6±9.1 nm in average diameters), and utilized early-developing (cleavage-stage) zebrafish embryos (critical aquatic and eco- species) as in vivo model organisms to probe diffusion and toxicity of Ag NPs. We found that single Ag NPs (30–72 nm diameters) passively diffused into the embryos through chorionic pores via random Brownian motion and stayed inside the embryos throughout their entire development (120 hours-post-fertilization, hpf). Dose and size dependent toxic effects of the NPs on embryonic development were observed, showing the possibility of tuning biocompatibility and toxicity of the NPs. At lower concentrations of the NPs (≤ 0.02 nM), 75–91% of embryos developed to normal zebrafish. At the higher concentrations of NPs (≥ 0.20 nM), 100% of embryos became dead. At the concentrations in between (0.02–0.2 nM), embryos developed to various deformed zebrafish. Number and sizes of individual Ag NPs embedded in tissues of normal and deformed zebrafish at 120 hpf were quantitatively analyzed, showing deformed zebrafish with higher number of larger NPs than normal zebrafish, and size-dependent nanotoxicity. By comparing with our previous studies of smaller Ag NPs (11.6±3.5 nm), the results further demonstrate striking size-dependent nanotoxicity that, at the same molar concentration, the larger Ag NPs (41.6±9.1 nm) are more toxic than the smaller Ag NPs (11.6±3.5 nm). PMID:22486336

  8. The paracrine effect of exogenous growth hormone alleviates dysmorphogenesis caused by tbx5 deficiency in zebrafish (Danio rerio embryos

    Directory of Open Access Journals (Sweden)

    Tsai Tzu-Chun

    2012-07-01

    Full Text Available Abstract Background Dysmorphogenesis and multiple organ defects are well known in zebrafish (Danio rerio embryos with T-box transcription factor 5 (tbx5 deficiencies, mimicking human Holt-Oram syndrome. Methods Using an oligonucleotide-based microarray analysis to study the expression of special genes in tbx5 morphants, we demonstrated that GH and some GH-related genes were markedly downregulated. Zebrafish embryos microinjected with tbx5-morpholino (MO antisense RNA and mismatched antisense RNA in the 1-cell stage served as controls, while zebrafish embryos co-injected with exogenous growth hormone (GH concomitant with tbx5-MO comprised the treatment group. Results The attenuating effects of GH in tbx5-MO knockdown embryos were quantified and observed at 24, 30, 48, 72, and 96 h post-fertilization. Though the understanding of mechanisms involving GH in the tbx5 functioning complex is limited, exogenous GH supplied to tbx5 knockdown zebrafish embryos is able to enhance the expression of downstream mediators in the GH and insulin-like growth factor (IGF-1 pathway, including igf1, ghra, and ghrb, and signal transductors (erk1, akt2, and eventually to correct dysmorphogenesis in various organs including the heart and pectoral fins. Supplementary GH also reduced apoptosis as determined by a TUNEL assay and decreased the expression of apoptosis-related genes and proteins (bcl2 and bad according to semiquantitative reverse-transcription polymerase chain reaction and immunohistochemical analysis, respectively, as well as improving cell cycle-related genes (p27 and cdk2 and cardiomyogenetic genes (amhc, vmhc, and cmlc2. Conclusions Based on our results, tbx5 knockdown causes a pseudo GH deficiency in zebrafish during early embryonic stages, and supplementation of exogenous GH can partially restore dysmorphogenesis, apoptosis, cell growth inhibition, and abnormal cardiomyogenesis in tbx5 knockdown zebrafish in a paracrine manner.

  9. Patterning Muscles Using Organizers: Larval Muscle Templates and Adult Myoblasts Actively Interact to Pattern the Dorsal Longitudinal Flight Muscles of Drosophila

    Science.gov (United States)

    Roy, Sudipto; VijayRaghavan, K.

    1998-01-01

    Pattern formation in muscle development is often mediated by special cells called muscle organizers. During metamorphosis in Drosophila, a set of larval muscles function as organizers and provide scaffolding for the development of the dorsal longitudinal flight muscles. These organizers undergo defined morphological changes and dramatically split into templates as adult fibers differentiate during pupation. We have investigated the cellular mechanisms involved in the use of larval fibers as templates. Using molecular markers that label myoblasts and the larval muscles themselves, we show that splitting of the larval muscles is concomitant with invasion by imaginal myoblasts and the onset of differentiation. We show that the Erect wing protein, an early marker of muscle differentiation, is not only expressed in myoblasts just before and after fusion, but also in remnant larval nuclei during muscle differentiation. We also show that interaction between imaginal myoblasts and larval muscles is necessary for transformation of the larval fibers. In the absence of imaginal myoblasts, the earliest steps in metamorphosis, such as the escape of larval muscles from histolysis and changes in their innervation, are normal. However, subsequent events, such as the splitting of these muscles, fail to progress. Finally, we show that in a mutant combination, null for Erect wing function in the mesoderm, the splitting of the larval muscles is aborted. These studies provide a genetic and molecular handle for the understanding of mechanisms underlying the use of muscle organizers in muscle patterning. Since the use of such organizers is a common theme in myogenesis in several organisms, it is likely that many of the processes that we describe are conserved. PMID:9606206

  10. Pentachlorophenol exposure causes Warburg-like effects in zebrafish embryos at gastrulation stage

    International Nuclear Information System (INIS)

    Xu, Ting; Zhao, Jing; Hu, Ping; Dong, Zhangji; Li, Jingyun; Zhang, Hongchang; Yin, Daqiang; Zhao, Qingshun

    2014-01-01

    Pentachlorophenol (PCP) is a prevalent pollutant in the environment and has been demonstrated to be a serious toxicant to humans and animals. However, little is known regarding the molecular mechanism underlying its toxic effects on vertebrate early development. To explore the impacts and underlying mechanisms of PCP on early development, zebrafish (Danio rerio) embryos were exposed to PCP at concentrations of 0, 20 and 50 μg/L, and microscopic observation and cDNA microarray analysis were subsequently conducted at gastrulation stage. The morphological observations revealed that PCP caused a developmental delay of zebrafish embryos in a concentration-dependent manner. Transcriptomic data showed that 50 μg/L PCP treatment resulted in significant changes in gene expression level, and the genes involved in energy metabolism and cell behavior were identified based on gene functional enrichment analysis. The energy production of embryos was influenced by PCP via the activation of glycolysis along with the inhibition of oxidative phosphorylation (OXPHOS). The results suggested that PCP acts as an inhibitor of OXPHOS at 8 hpf (hours postfertilization). Consistent with the activated glycolysis, the cell cycle activity of PCP-treated embryos was higher than the controls. These characteristics are similar to the Warburg effect, which occurs in human tumors. The microinjection of exogenous ATP confirmed that an additional energy supply could rescue PCP-treated embryos from the developmental delay due to the energy deficit. Taken together, our results demonstrated that PCP causes a Warburg-like effect on zebrafish embryos during gastrulation, and the affected embryos had the phenotype of developmental delay. - Highlights: • We treat zebrafish embryos with PCP at gastrula stage. • PCP acts as an oxidative phosphorylation inhibitor, not an uncoupler, in gastrulation. • Exogenous ATP injection will rescue the development of effected embryos. • The transcriptome of PCP

  11. Pentachlorophenol exposure causes Warburg-like effects in zebrafish embryos at gastrulation stage

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Ting; Zhao, Jing [Key Laboratory of Yangtze River Water Environment, Ministry of Education, College of Environmental Science and Technology, Tongji University, Shanghai 200092 (China); Hu, Ping [Key Laboratory of Model Animal for Disease Study, Ministry of Education, Model Animal Research Center, Nanjing University, Nanjing 210061 (China); State Key Laboratory of Reproductive Medicine, Department of Prenatal Diagnosis, Nanjing Maternity and Child Health Care Hospital Affiliated to Nanjing Medical University, Nanjing 210029 (China); Dong, Zhangji; Li, Jingyun [Key Laboratory of Model Animal for Disease Study, Ministry of Education, Model Animal Research Center, Nanjing University, Nanjing 210061 (China); Zhang, Hongchang [Key Laboratory of Yangtze River Water Environment, Ministry of Education, College of Environmental Science and Technology, Tongji University, Shanghai 200092 (China); Yin, Daqiang, E-mail: yindq@tongji.edu.cn [Key Laboratory of Yangtze River Water Environment, Ministry of Education, College of Environmental Science and Technology, Tongji University, Shanghai 200092 (China); Zhao, Qingshun, E-mail: qingshun@nju.edu.cn [Key Laboratory of Model Animal for Disease Study, Ministry of Education, Model Animal Research Center, Nanjing University, Nanjing 210061 (China)

    2014-06-01

    Pentachlorophenol (PCP) is a prevalent pollutant in the environment and has been demonstrated to be a serious toxicant to humans and animals. However, little is known regarding the molecular mechanism underlying its toxic effects on vertebrate early development. To explore the impacts and underlying mechanisms of PCP on early development, zebrafish (Danio rerio) embryos were exposed to PCP at concentrations of 0, 20 and 50 μg/L, and microscopic observation and cDNA microarray analysis were subsequently conducted at gastrulation stage. The morphological observations revealed that PCP caused a developmental delay of zebrafish embryos in a concentration-dependent manner. Transcriptomic data showed that 50 μg/L PCP treatment resulted in significant changes in gene expression level, and the genes involved in energy metabolism and cell behavior were identified based on gene functional enrichment analysis. The energy production of embryos was influenced by PCP via the activation of glycolysis along with the inhibition of oxidative phosphorylation (OXPHOS). The results suggested that PCP acts as an inhibitor of OXPHOS at 8 hpf (hours postfertilization). Consistent with the activated glycolysis, the cell cycle activity of PCP-treated embryos was higher than the controls. These characteristics are similar to the Warburg effect, which occurs in human tumors. The microinjection of exogenous ATP confirmed that an additional energy supply could rescue PCP-treated embryos from the developmental delay due to the energy deficit. Taken together, our results demonstrated that PCP causes a Warburg-like effect on zebrafish embryos during gastrulation, and the affected embryos had the phenotype of developmental delay. - Highlights: • We treat zebrafish embryos with PCP at gastrula stage. • PCP acts as an oxidative phosphorylation inhibitor, not an uncoupler, in gastrulation. • Exogenous ATP injection will rescue the development of effected embryos. • The transcriptome of PCP

  12. Transient exposure to ethanol during zebrafish embryogenesis results in defects in neuronal differentiation: an alternative model system to study FASD.

    Directory of Open Access Journals (Sweden)

    Xavier Joya

    Full Text Available The exposure of the human embryo to ethanol results in a spectrum of disorders involving multiple organ systems, including the impairment of the development of the central nervous system (CNS. In spite of the importance for human health, the molecular basis of prenatal ethanol exposure remains poorly understood, mainly to the difficulty of sample collection. Zebrafish is now emerging as a powerful organism for the modeling and the study of human diseases. In this work, we have assessed the sensitivity of specific subsets of neurons to ethanol exposure during embryogenesis and we have visualized the sensitive embryonic developmental periods for specific neuronal groups by the use of different transgenic zebrafish lines.In order to evaluate the teratogenic effects of acute ethanol exposure, we exposed zebrafish embryos to ethanol in a given time window and analyzed the effects in neurogenesis, neuronal differentiation and brain patterning. Zebrafish larvae exposed to ethanol displayed small eyes and/or a reduction of the body length, phenotypical features similar to the observed in children with prenatal exposure to ethanol. When neuronal populations were analyzed, we observed a clear reduction in the number of differentiated neurons in the spinal cord upon ethanol exposure. There was a decrease in the population of sensory neurons mainly due to a decrease in cell proliferation and subsequent apoptosis during neuronal differentiation, with no effect in motoneuron specification.Our investigation highlights that transient exposure to ethanol during early embryonic development affects neuronal differentiation although does not result in defects in early neurogenesis. These results establish the use of zebrafish embryos as an alternative research model to elucidate the molecular mechanism(s of ethanol-induced developmental toxicity at very early stages of embryonic development.

  13. Transient Exposure to Ethanol during Zebrafish Embryogenesis Results in Defects in Neuronal Differentiation: An Alternative Model System to Study FASD

    Science.gov (United States)

    Joya, Xavier; Garcia-Algar, Oscar; Vall, Oriol; Pujades, Cristina

    2014-01-01

    Background The exposure of the human embryo to ethanol results in a spectrum of disorders involving multiple organ systems, including the impairment of the development of the central nervous system (CNS). In spite of the importance for human health, the molecular basis of prenatal ethanol exposure remains poorly understood, mainly to the difficulty of sample collection. Zebrafish is now emerging as a powerful organism for the modeling and the study of human diseases. In this work, we have assessed the sensitivity of specific subsets of neurons to ethanol exposure during embryogenesis and we have visualized the sensitive embryonic developmental periods for specific neuronal groups by the use of different transgenic zebrafish lines. Methodology/Principal Findings In order to evaluate the teratogenic effects of acute ethanol exposure, we exposed zebrafish embryos to ethanol in a given time window and analyzed the effects in neurogenesis, neuronal differentiation and brain patterning. Zebrafish larvae exposed to ethanol displayed small eyes and/or a reduction of the body length, phenotypical features similar to the observed in children with prenatal exposure to ethanol. When neuronal populations were analyzed, we observed a clear reduction in the number of differentiated neurons in the spinal cord upon ethanol exposure. There was a decrease in the population of sensory neurons mainly due to a decrease in cell proliferation and subsequent apoptosis during neuronal differentiation, with no effect in motoneuron specification. Conclusion Our investigation highlights that transient exposure to ethanol during early embryonic development affects neuronal differentiation although does not result in defects in early neurogenesis. These results establish the use of zebrafish embryos as an alternative research model to elucidate the molecular mechanism(s) of ethanol-induced developmental toxicity at very early stages of embryonic development. PMID:25383948

  14. Larval growth and drift pattern and the separation of herring spawning groups in the North Sea

    DEFF Research Database (Denmark)

    Munk, Peter; Christensen, Villy

    1990-01-01

    Data from two internationally co-ordinated survey programmes are used in an attempt to display the events in the early life that took place during the 'recovery period' of the North Sea herring stock, 1979-1986. During the period, an increasing importance of the spawning grounds off Buchan and off...... Yorkshire is evident from distributions of newly hatched herring larvae. The dramatic changes in relative importance of spawning grounds are traced in the data on both late larvae (6 months old) and juveniles (18 months old). The onset of extended spawning off Buchan and off Yorkshire was followed...... in the relative contributions from spawning grounds apparently influence the overall spatial distribution as well as size composition of larval and juvenile North Sea herring. The findings indicate that groups of larvae retain, to a large extent, separate distributions until metamorphosis, and point to larval...

  15. Larval development and shape variation of the kelpfish Myxodes viridis (Teleostei: Clinidae

    Directory of Open Access Journals (Sweden)

    Francisca Zavala-Muñoz

    2016-03-01

    Full Text Available Larval development and shape ontogeny of the kelpfish Myxodes viridis (Clinidae are described for the first time. A total of 214 individuals ranging between 3.51 and 23.09 mm standard length collected off central Chile were assessed employing classic and geometric morphometrics, illustration with camera lucida and a double-staining technique for cartilaginous and bone structure observation. Based on characteristics such as yolk sac presence and fin formation, six stages of larval development were differentiated: yolk sac, preflexion, flexion, early postflexion, late postflexion and juvenile. Shape changes during development are subtle and occur smoothly, being more significant in the head and preanal length, and ontogenetic allometry accounts for almost 15%. Cartilage formation takes place first at the branchial arches and cranium; then hypural, haemal and neural arches are consecutively formed. Bony structure ossification occurs late in the development. Vertebral centra ossify directly, without cartilaginous matrix replacement.

  16. Drivers of larval fish assemblage shift during the spring-summer transition in the coastal Mediterranean

    Science.gov (United States)

    Álvarez, Itziar; Catalán, Ignacio A.; Jordi, Antoni; Palmer, Miquel; Sabatés, Ana; Basterretxea, Gotzon

    2012-01-01

    The influence of coastal environmental conditions from winter-spring to summer on fish larvae assemblages in a temperate area has suggested a seasonal shift in ecosystem-level variation through which trophic pathways shift from the pelagic to the benthic system. This variation may be related to marked effects in the reproductive strategies in the fishes inhabiting the area and indirectly affect ichthyoplankton assemblages. Larval fish assemblages were sampled fortnightly at three stations located in coastal waters off southern Mallorca (Western Mediterranean) from March to August 2007, covering the main spawning period for the resident coastal fish in this region. The larval fish assemblage showed clear seasonality with higher specific abundance but lower diversity in the spring. Two main assemblages were identified: a spring assemblage, occurring at surface seawater temperatures ichthyoplankton communities occurred in early June, coinciding with the onset of summer hydrographical conditions and the local benthic productivity peak.

  17. Reduced larval feeding rate is a strong evolutionary correlate of ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Genetics; Volume 85; Issue 3. Reduced larval feeding rate is a strong evolutionary correlate of rapid development in Drosophila melanogaster. M. Rajamani N. Raghavendra ... Keywords. life-history evolution; development time; larval feeding rate; competition; tradeoffs; Drosophila melanogaster.

  18. The larval development of the red mangrove crab Sesarma meinerti ...

    African Journals Online (AJOL)

    The larval stages of the red mangrove crab Sesarma meinerti de Man were reared in the laboratory. Larval development consists of five zoeal stages and one megalopa. Zoeal development lasts an average of 25 days at 25°C. The external morphology of larvae is described in detail and their relationship with larvae of.

  19. Molecular and Chemical Genetic Approaches to Developmental Origins of Aging and Disease in Zebrafish

    Science.gov (United States)

    Sasaki, Tomoyuki; Kishi, Shuji

    2013-01-01

    The incidence of diseases increases rapidly with age, accompanied by progressive deteriorations of physiological functions in organisms. Aging-associated diseases are sporadic but mostly inevitable complications arising from senescence. Senescence is often considered the antithesis of early development, but yet there may be factors and mechanisms in common between these two phenomena over the dynamic process of aging. The association between early development and late-onset disease with advancing age is thought to come from a consequence of developmental plasticity, the phenomenon by which one genotype can give rise to a range of physiologically and/or morphologically adaptive states in response to different environmental or genetic perturbations. On the one hand, we hypothesized that the future aging process can be predictive based on adaptivity during the early developmental period. Modulating the thresholds of adaptive plasticity by chemical genetic approaches, we have been investigating whether any relationship exists between the regulatory mechanisms that function in early development and in senescence using the zebrafish (Danio rerio), a small freshwater fish and a useful model animal for genetic studies. We have successfully conducted experiments to isolate zebrafish mutants expressing apparently altered senescence phenotypes during embryogenesis (“embryonic senescence”), subsequently showing shortened lifespan in adulthoods. We anticipate that previously uncharacterized developmental genes may mediate the aging process and play a pivotal role in senescence. On the other hand, unexpected senescence-related genes might also be involved in the early developmental process and regulation. The ease of manipulation using the zebrafish system allows us to conduct an exhaustive exploration of novel genes and small molecular compounds that can be linked to the senescence phenotype, and thereby facilitates searching for the evolutionary and developmental origins

  20. Cloning of aquaporin-1 of the blue crab, Callinectes sapidus: its expression during the larval development in hyposalinity.

    Science.gov (United States)

    Chung, J Sook; Maurer, Leah; Bratcher, Meagan; Pitula, Joseph S; Ogburn, Matthew B

    2012-09-03

    of C. sapidus and the induction of its expression at early larval stages in the exposure of hyposalinity. However, it remains to be determined if the increase in CasAQP-1 expression at later larval stages may have a role in adaptation to hyposalinity.

  1. Chondroitin / dermatan sulfate modification enzymes in zebrafish development.

    Directory of Open Access Journals (Sweden)

    Judith Habicher

    Full Text Available Chondroitin/dermatan sulfate (CS/DS proteoglycans consist of unbranched sulfated polysaccharide chains of repeating GalNAc-GlcA/IdoA disaccharide units, attached to serine residues on specific proteins. The CS/DS proteoglycans are abundant in the extracellular matrix where they have essential functions in tissue development and homeostasis. In this report a phylogenetic analysis of vertebrate genes coding for the enzymes that modify CS/DS is presented. We identify single orthologous genes in the zebrafish genome for the sulfotransferases chst7, chst11, chst13, chst14, chst15 and ust and the epimerase dse. In contrast, two copies were found for mammalian sulfotransferases CHST3 and CHST12 and the epimerase DSEL, named chst3a and chst3b, chst12a and chst12b, dsela and dselb, respectively. Expression of CS/DS modification enzymes is spatially and temporally regulated with a large variation between different genes. We found that CS/DS 4-O-sulfotransferases and 6-O-sulfotransferases as well as CS/DS epimerases show a strong and partly overlapping expression, whereas the expression is restricted for enzymes with ability to synthesize di-sulfated disaccharides. A structural analysis further showed that CS/DS sulfation increases during embryonic development mainly due to synthesis of 4-O-sulfated GalNAc while the proportion of 6-O-sulfated GalNAc increases in later developmental stages. Di-sulfated GalNAc synthesized by Chst15 and 2-O-sulfated GlcA/IdoA synthesized by Ust are rare, in accordance with the restricted expression of these enzymes. We also compared CS/DS composition with that of heparan sulfate (HS. Notably, CS/DS biosynthesis in early zebrafish development is more dynamic than HS biosynthesis. Furthermore, HS contains disaccharides with more than one sulfate group, which are virtually absent in CS/DS.

  2. Linking larval history to juvenile demography in the bicolor damselfish Stegastes partitus (Perciformes:Pomacentridae

    Directory of Open Access Journals (Sweden)

    Richard S Nemeth

    2005-05-01

    Full Text Available Otolith-based reconstructions of daily larval growth increments were used to examine the effect of variation in larval growth on size and age at settlement and post-settlement growth,survival and habitat preferences of juvenile bicolor damselfish (Stegastes partitus Poey.During August 1992 and 1994,newly settled S. partitus were collected from Montastraea coral heads and Porites rubble piles in Tague Bay,St.Croix,U.S. Virgin Islands (17 °45 ’ N,64 °42 ’ W.Daily lapillar otolith increments from each fish were counted and measured with Optimas image analysis software.S.partitus pelagic larval duration was 23.7 d in 1992 (n =70and 24.6 d in 1994 (n =38and larval age at settlement averaged 13.0 mm total length both years.Analysis of daily otolith increments demonstrated that variation in larval growth rates and size and age at settlement had no detectable effect on post-settlement survivorship but that larger larvae showed a preference for Montastraea coral at settlement.Late larval and early juvenile growth rates showed a significant positive relationship indicating that growth patterns established during the planktonic stage can span metamorphosis and continue into the benthic juvenile phase.Larval growth rates during the first two weeks post-hatching also had a strong effect on age to developmental competence (ability to undergo metamorphosisin both 1992 and 1994 with the fastest growing larvae being 8 d younger and 0.8 mm smaller at settlement than the slowest growing larvae.These differential growth rates in early stage larvae established trajectories toward larval developmental competence and may prove important in biogeographical studies of larval dispersal.Reconstruyendo aumentos diarios de otolitos se compará la variación en crecimiento larval sobre el tamaño y la edad de asentamiento,y el crecimiento post-acentamiento, sobrevivencia y preferencia de hábitat,del pez damisela bicolor (Stegastes partitus Poeyjoven.En agosto de 1992

  3. Dissection of vertebrate hematopoiesis using zebrafish thrombopoietin

    Czech Academy of Sciences Publication Activity Database

    Svoboda, Ondřej; Stachura, D.L.; Machoňová, Olga; Pajer, Petr; Brynda, Jiří; Zon, L.I.; Traver, D.; Bartůněk, Petr

    2014-01-01

    Roč. 124, č. 2 (2014), s. 220-228 ISSN 0006-4971 R&D Projects: GA ČR GAP305/10/0953 Grant - others:NIH(US) K01-DK087814-01A1; NIH(US) R01-DK074482 Keywords : Zebrafish * hematopoiesis * progenitors * thrombopoietin * erythropoietin Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 10.452, year: 2014

  4. Ethanol Exposure Causes Muscle Degeneration in Zebrafish

    Directory of Open Access Journals (Sweden)

    Elizabeth C. Coffey

    2018-03-01

    Full Text Available Alcoholic myopathies are characterized by neuromusculoskeletal symptoms such as compromised movement and weakness. Although these symptoms have been attributed to neurological damage, EtOH may also target skeletal muscle. EtOH exposure during zebrafish primary muscle development or adulthood results in smaller muscle fibers. However, the effects of EtOH exposure on skeletal muscle during the growth period that follows primary muscle development are not well understood. We determined the effects of EtOH exposure on muscle during this phase of development. Strikingly, muscle fibers at this stage are acutely sensitive to EtOH treatment: EtOH induces muscle degeneration. The severity of EtOH-induced muscle damage varies but muscle becomes more refractory to EtOH as muscle develops. NF-kB induction in muscle indicates that EtOH triggers a pro-inflammatory response. EtOH-induced muscle damage is p53-independent. Uptake of Evans blue dye shows that EtOH treatment causes sarcolemmal instability before muscle fiber detachment. Dystrophin-null sapje mutant zebrafish also exhibit sarcolemmal instability. We tested whether Trichostatin A (TSA, which reduces muscle degeneration in sapje mutants, would affect EtOH-treated zebrafish. We found that TSA and EtOH are a lethal combination. EtOH does, however, exacerbate muscle degeneration in sapje mutants. EtOH also disrupts adhesion of muscle fibers to their extracellular matrix at the myotendinous junction: some detached muscle fibers retain beta-Dystroglycan indicating failure of muscle end attachments. Overexpression of Paxillin, which reduces muscle degeneration in zebrafish deficient for beta-Dystroglycan, is not sufficient to rescue degeneration. Taken together, our results suggest that EtOH exposure has pleiotropic deleterious effects on skeletal muscle.

  5. Molecular cloning and expression analysis of a zebrafish novel zinc finger protein gene rnf141

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    Wenqian Deng

    2009-01-01

    Full Text Available ZNF230 is a novel zinc finger gene cloned by our laboratory. In order to understand the potential functions of this gene in vertebrate development, we cloned the zebrafish orthologue of human ZNF230, named rnf141. The cDNA fragment of rnf141 was obtained by rapid amplification of cDNA ends (RACE. The open reading frame (ORF encodes a polypeptide of 222 amino acids which shares 75.65% identity with the human ZNF230. RT-PCR analysis in zebrafish embryo and adult tissues revealed that rnf141 transcripts are maternally derived and that rnf141 mRNA has a broad distribution. Zygotic rnf141 message is strongly localized in the central nervous system, as shown by whole-mount in situ hybridization. Knockdown and over expression of rnf141 can induce abnormal phenotypes, including abnormal development of brain, as well as yolk sac and axis extendsion. Marker gene analysis showed that rnf141 may play a role in normal dorsoventral patterning of zebrafish embryos, suggesting that rnf141 may have a broad function during early development of vertebrates.

  6. Scrambled eggs: Proteomic portraits and novel biomarkers of egg quality in zebrafish (Danio rerio).

    Science.gov (United States)

    Yilmaz, Ozlem; Patinote, Amélie; Nguyen, Thao Vi; Com, Emmanuelle; Lavigne, Regis; Pineau, Charles; Sullivan, Craig V; Bobe, Julien

    2017-01-01

    Egg quality is a complex biological trait and a major determinant of reproductive fitness in all animals. This study delivered the first proteomic portraits of egg quality in zebrafish, a leading biomedical model for early development. Egg batches of good and poor quality, evidenced by embryo survival for 24 h, were sampled immediately after spawning and used to create pooled or replicated sample sets whose protein extracts were subjected to different levels of fractionation before liquid chromatography and tandem mass spectrometry. Obtained spectra were searched against a zebrafish proteome database and detected proteins were annotated, categorized and quantified based on normalized spectral counts. Manually curated and automated enrichment analyses revealed poor quality eggs to be deficient of proteins involved in protein synthesis and energy and lipid metabolism, and of some vitellogenin products and lectins, and to have a surfeit of proteins involved in endo-lysosomal activities, autophagy, and apoptosis, and of some oncogene products, lectins and egg envelope proteins. Results of pathway and network analyses suggest that this aberrant proteomic profile results from failure of oocytes giving rise to poor quality eggs to properly transit through final maturation, and implicated Wnt signaling in the etiology of this defect. Quantitative comparisons of abundant proteins in good versus poor quality eggs revealed 17 candidate egg quality markers. Thus, the zebrafish egg proteome is clearly linked to embryo developmental potential, a phenomenon that begs further investigation to elucidate the root causes of poor egg quality, presently a serious and intractable problem in livestock and human reproductive medicine.

  7. The mechanism for primordial germ-cell migration is conserved between Japanese eel and zebrafish.

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    Taiju Saito

    Full Text Available Primordial germ cells (PGCs are segregated and specified from somatic cells during early development. These cells arise elsewhere and have to migrate across the embryo to reach developing gonadal precursors. Several molecules associated with PGC migration (i.e. dead-end, nanos1, and cxcr4 are highly conserved across phylum boundaries. However, since cell migration is a complicated process that is regulated spatially and temporally by multiple adaptors and signal effectors, the process is unlikely to be explained by these known genes only. Indeed, it has been shown that there are variations in PGC migration pattern during development among teleost species. However, it is still unclear whether the actual mechanism of PGC migration is conserved among species. In this study, we studied the migration of PGCs in Japanese eel (Anguilla japonica embryos and tested the migration mechanism between Japanese eel and zebrafish (Danio rerio for conservation, by transplanting eel PGCs into zebrafish embryos. The experiments showed that eel PGCs can migrate toward the gonadal region of zebrafish embryos along with endogenous PGCs, even though the migration patterns, behaviors, and settlements of PGCs are somewhat different between these species. Our results demonstrate that the migration mechanism of PGCs during embryonic development is highly conserved between these two distantly related species (belonging to different teleost orders.

  8. Apoc2 loss-of-function zebrafish mutant as a genetic model of hyperlipidemia

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    Chao Liu

    2015-08-01

    Full Text Available Apolipoprotein C-II (APOC2 is an obligatory activator of lipoprotein lipase. Human patients with APOC2 deficiency display severe hypertriglyceridemia while consuming a normal diet, often manifesting xanthomas, lipemia retinalis and pancreatitis. Hypertriglyceridemia is also an important risk factor for development of cardiovascular disease. Animal models to study hypertriglyceridemia are limited, with no Apoc2-knockout mouse reported. To develop a genetic model of hypertriglyceridemia, we generated an apoc2 mutant zebrafish characterized by the loss of Apoc2 function. apoc2 mutants show decreased plasma lipase activity and display chylomicronemia and severe hypertriglyceridemia, which closely resemble the phenotype observed in human patients with APOC2 deficiency. The hypertriglyceridemia in apoc2 mutants is rescued by injection of plasma from wild-type zebrafish or by injection of a human APOC2 mimetic peptide. Consistent with a previous report of a transient apoc2 knockdown, apoc2 mutant larvae have a minor delay in yolk consumption and angiogenesis. Furthermore, apoc2 mutants fed a normal diet accumulate lipid and lipid-laden macrophages in the vasculature, which resemble early events in the development of human atherosclerotic lesions. In addition, apoc2 mutant embryos show ectopic overgrowth of pancreas. Taken together, our data suggest that the apoc2 mutant zebrafish is a robust and versatile animal model to study hypertriglyceridemia and the mechanisms involved in the pathogenesis of associated human diseases.

  9. Use of zebrafish and knockdown technology to define proprotein convertase activity.

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    Chitramuthu, Babykumari P; Bennett, Hugh P J

    2011-01-01

    The Zebrafish (Danio rerio) is a powerful and well-established tool used extensively for the study of early vertebrate development and as a model of human diseases. Zebrafish genes orthologous to their mammalian counterparts generally share conserved biological function. Protein knockdown or overexpression can be effectively achieved by microinjection of morpholino antisense oligonucleotides (MOs) or mRNA, respectively, into developing embryos at the one- to two-cell stage. Correlating gene expression patterns with the characterizing of phenotypes resulting from over- or underexpression can reveal the function of a particular protein. The microinjection technique is simple and results are reproducible. We defined the expression pattern of the proprotein convertase PCSK5 within the lateral line neuromasts and various organs including the liver, gut and otic vesicle by whole-mount in situ hybridization (ISH) and immunofluorescence (IF). MO-mediated knockdown of zebrafish PCSK5 expression generated embryos that display abnormal neuromast deposition within the lateral line system resulting in uncoordinated patterns of swimming.

  10. Aryl hydrocarbon receptor 2 mediates the toxicity of Paclobutrazol on the digestive system of zebrafish embryos.

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    Wang, Wen-Der; Chen, Guan-Ting; Hsu, Hwei-Jan; Wu, Chang-Yi

    2015-02-01

    Paclobutrazol (PBZ), a trazole-containing fungicide and plant growth retardant, has been widely used for over 30 years to regulate plant growth and promote early fruit setting. Long-term usage of PBZ in agriculture and natural environments has resulted in residual PBZ in the soil and water. Chronic exposure to waterborne PBZ can cause various physiological effects in fish, including hepatic steatosis, antioxidant activity, and disruption of spermatogenesis. We have previously shown that PBZ also affects the rates of zebrafish embryonic survival and hatching, and causes developmental failure of the head skeleton and eyes; here, we further show that PBZ has embryonic toxic effects on digestive organs of zebrafish, and describe the underlying mechanisms. PBZ treatment of embryos resulted in dose-dependent morphological and functional abnormalities of the digestive organs. Real-time RT-PCR and in situ hybridization were used to show that PBZ strongly induces cyp1a1 expression in the digestive system, and slightly induces ahr2 expression in zebrafish embryos. Knockdown of ahr2 with morpholino oligonucleotides prevents PBZ toxicity. Thus, the toxic effect of PBZ on digestive organs is mediated by AhR2, as was previously reported for retene and TCDD. These findings have implications for understanding the potential toxicity of PBZ during embryogenesis, and thus the potential impact of fungicides on public health and the environment. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Uncoupling nicotine mediated motoneuron axonal pathfinding errors and muscle degeneration in zebrafish

    International Nuclear Information System (INIS)

    Welsh, Lillian; Tanguay, Robert L.; Svoboda, Kurt R.

    2009-01-01

    Zebrafish embryos offer a unique opportunity to investigate the mechanisms by which nicotine exposure impacts early vertebrate development. Embryos exposed to nicotine become functionally paralyzed by 42 hpf suggesting that the neuromuscular system is compromised in exposed embryos. We previously demonstrated that secondary spinal motoneurons in nicotine-exposed embryos were delayed in development and that their axons made pathfinding errors (Svoboda, K.R., Vijayaraghaven, S., Tanguay, R.L., 2002. Nicotinic receptors mediate changes in spinal motoneuron development and axonal pathfinding in embryonic zebrafish exposed to nicotine. J. Neurosci. 22, 10731-10741). In that study, we did not consider the potential role that altered skeletal muscle development caused by nicotine exposure could play in contributing to the errors in spinal motoneuron axon pathfinding. In this study, we show that an alteration in skeletal muscle development occurs in tandem with alterations in spinal motoneuron development upon exposure to nicotine. The alteration in the muscle involves the binding of nicotine to the muscle-specific AChRs. The nicotine-induced alteration in muscle development does not occur in the zebrafish mutant (sofa potato, [sop]), which lacks muscle-specific AChRs. Even though muscle development is unaffected by nicotine exposure in sop mutants, motoneuron axonal pathfinding errors still occur in these mutants, indicating a direct effect of nicotine exposure on nervous system development.

  12. Znrg, a novel gene expressed mainly in the developing notochord of zebrafish.

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    Zhou, Yaping; Xu, Yan; Li, Jianzhen; Liu, Yao; Zhang, Zhe; Deng, Fengjiao

    2010-06-01

    The notochord, a defining characteristic of the chordate embryo is a critical midline structure required for axial skeletal formation in vertebrates, and acts as a signaling center throughout embryonic development. We utilized the digital differential display program of the National Center for Biotechnology Information, and identified a contig of expressed sequence tags (no. Dr. 83747) from the zebrafish ovary library in Genbank. Full-length cDNA of the identified gene was cloned by 5'- and 3'- RACE, and the resulting sequence was confirmed by polymerase chain reaction and sequencing. The cDNA clone contains 2,505 base pairs and encodes a novel protein of 707 amino acids that shares no significant homology with any known proteins. This gene was expressed in mature oocytes and at the one-cell stage, and persisted until the 5th day of development, as determined by RT-PCR. Transcripts were detected by whole-mount RNA in situ hybridization from the two-cell stage to 72 h of embryonic development. This gene was uniformly distributed from the cleavage stage up to the blastula stage. During early gastrulation, it was present in the dorsal region, and became restricted to the notochord and pectoral fin at 48 and 72 h of embryonic development. Based on its abundance in the notochord, we hypothesized that the novel gene may play an important role in notochord development in zebrafish; we named this gene, zebrafish notochord-related gene, or znrg.

  13. Clonal origins of cells in the pigmented retina of the zebrafish eye

    International Nuclear Information System (INIS)

    Streisinger, G.; Coale, F.; Taggart, C.; Walker, C.; Grunwald, D.J.

    1989-01-01

    Mosaic analysis has been used to study the clonal basis of the development of the pigmented retina of the zebrafish, Brachydanio rerio. Zebrafish embryos heterozygous for a recessive mutation at the gol-1 locus were exposed to gamma-irradiation at various developmental stages to create mosaic individuals consisting of wild-type pigmented cells and a clone of pigmentless (golden) cells in the eye. The contribution of individual embryonic cells to the pigmented retina was measured and the total number of cells in the embryo that contributed descendants to this tissue was determined. Until the 32-cell stage, almost every blastomere has some descendants that participate in the formation of the pigmented retina of the zebrafish. During subsequent cell divisions, up to the several thousand-cell stage, the number of ancestral cells is constant: approximately 40 cells are present that will give rise to progeny in the pigmented retina. Analysis of the size of clones in the pigmented retina indicates that the cells of this tissue do not arise through a rigid series of cell divisions originating in the early embryo. The findings that each cleavage stage cell contributes to the pigmented retina and yet the contribution of such cells is highly variable are consistent with the interpretation that clonal descendants of different blastomeres normally intermix extensively prior to formation of the pigmented retina

  14. Disease modeling in genetic kidney diseases: zebrafish.

    Science.gov (United States)

    Schenk, Heiko; Müller-Deile, Janina; Kinast, Mark; Schiffer, Mario

    2017-07-01

    Growing numbers of translational genomics studies are based on the highly efficient and versatile zebrafish (Danio rerio) vertebrate model. The increasing types of zebrafish models have improved our understanding of inherited kidney diseases, since they not only display pathophysiological changes but also give us the opportunity to develop and test novel treatment options in a high-throughput manner. New paradigms in inherited kidney diseases have been developed on the basis of the distinct genome conservation of approximately 70 % between zebrafish and humans in terms of existing gene orthologs. Several options are available to determine the functional role of a specific gene or gene sets. Permanent genome editing can be induced via complete gene knockout by using the CRISPR/Cas-system, among others, or via transient modification by using various morpholino techniques. Cross-species rescues succeeding knockdown techniques are employed to determine the functional significance of a target gene or a specific mutation. This article summarizes the current techniques and discusses their perspectives.

  15. Social dominance modulates eavesdropping in zebrafish

    Science.gov (United States)

    Abril-de-Abreu, Rodrigo; Cruz, Ana S.; Oliveira, Rui F.

    2015-01-01

    Group living animals may eavesdrop on signalling interactions between conspecifics and integrate it with their own past social experience in order to optimize the use of relevant information from others. However, little is known about this interplay between public (eavesdropped) and private social information. To investigate it, we first manipulated the dominance status of bystander zebrafish. Next, we either allowed or prevented bystanders from observing a fight. Finally, we assessed their behaviour towards the winners and losers of the interaction, using a custom-made video-tracking system and directional analysis. We found that only dominant bystanders who had seen the fight revealed a significant increase in directional focus (a measure of attention) towards the losers of the fights. Furthermore, our results indicate that information about the fighters' acquired status was collected from the signalling interaction itself and not from post-interaction status cues, which implies the existence of individual recognition in zebrafish. Thus, we show for the first time that zebrafish, a highly social model organism, eavesdrop on conspecific agonistic interactions and that this process is modulated by the eavesdroppers' dominance status. We suggest that this type of integration of public and private information may be ubiquitous in social learning processes. PMID:26361550

  16. Use of zebrafish to study Shigella infection

    Science.gov (United States)

    Duggan, Gina M.

    2018-01-01

    ABSTRACT Shigella is a leading cause of dysentery worldwide, responsible for up to 165 million cases of shigellosis each year. Shigella is also recognised as an exceptional model pathogen to study key issues in cell biology and innate immunity. Several infection models have been useful to explore Shigella biology; however, we still lack information regarding the events taking place during the Shigella infection process in vivo. Here, we discuss a selection of mechanistic insights recently gained from studying Shigella infection of zebrafish (Danio rerio), with a focus on cytoskeleton rearrangements and cellular immunity. We also discuss how infection of zebrafish can be used to investigate new concepts underlying infection control, including emergency granulopoiesis and the use of predatory bacteria to combat antimicrobial resistance. Collectively, these insights illustrate how Shigella infection of zebrafish can provide fundamental advances in our understanding of bacterial pathogenesis and vertebrate host defence. This information should also provide vital clues for the discovery of new therapeutic strategies against infectious disease in humans. PMID:29590642

  17. Use of zebrafish to study Shigella infection

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    Gina M. Duggan

    2018-02-01

    Full Text Available Shigella is a leading cause of dysentery worldwide, responsible for up to 165 million cases of shigellosis each year. Shigella is also recognised as an exceptional model pathogen to study key issues in cell biology and innate immunity. Several infection models have been useful to explore Shigella biology; however, we still lack information regarding the events taking place during the Shigella infection process in vivo. Here, we discuss a selection of mechanistic insights recently gained from studying Shigella infection of zebrafish (Danio rerio, with a focus on cytoskeleton rearrangements and cellular immunity. We also discuss how infection of zebrafish can be used to investigate new concepts underlying infection control, including emergency granulopoiesis and the use of predatory bacteria to combat antimicrobial resistance. Collectively, these insights illustrate how Shigella infection of zebrafish can provide fundamental advances in our understanding of bacterial pathogenesis and vertebrate host defence. This information should also provide vital clues for the discovery of new therapeutic strategies against infectious disease in humans.

  18. Cell migration during heart regeneration in zebrafish.

    Science.gov (United States)

    Tahara, Naoyuki; Brush, Michael; Kawakami, Yasuhiko

    2016-07-01

    Zebrafish possess the remarkable ability to regenerate injured hearts as adults, which contrasts the very limited ability in mammals. Although very limited, mammalian hearts do in fact have measurable levels of cardiomyocyte regeneration. Therefore, elucidating mechanisms of zebrafish heart regeneration would provide information of naturally occurring regeneration to potentially apply to mammalian studies, in addition to addressing this biologically interesting phenomenon in itself. Studies over the past 13 years have identified processes and mechanisms of heart regeneration in zebrafish. After heart injury, pre-existing cardiomyocytes dedifferentiate, enter the cell cycle, and repair the injured myocardium. This process requires interaction with epicardial cells, endocardial cells, and vascular endothelial cells. Epicardial cells envelope the heart, while endocardial cells make up the inner lining of the heart. They provide paracrine signals to cardiomyocytes to regenerate the injured myocardium, which is vascularized during heart regeneration. In addition, accumulating results suggest that local migration of these major cardiac cell types have roles in heart regeneration. In this review, we summarize the characteristics of various heart injury methods used in the research community and regeneration of the major cardiac cell types. Then, we discuss local migration of these cardiac cell types and immune cells during heart regeneration. Developmental Dynamics 245:774-787, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  19. Comparative transcriptomic profiling of hydrogen peroxide signaling networks in zebrafish and human keratinocytes: Implications toward conservation, migration and wound healing.

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    Lisse, Thomas S; King, Benjamin L; Rieger, Sandra

    2016-02-05

    Skin wounds need to be repaired rapidly after injury to restore proper skin barrier function. Hydrogen peroxide (H2O2) is a conserved signaling factor that has been shown to promote a variety of skin wound repair processes, including immune cell migration, angiogenesis and sensory axon repair. Despite growing research on H2O2 functions in wound repair, the downstream signaling pathways activated by this reactive oxygen species in the context of injury remain largely unknown. The goal of this study was to provide a comprehensive analysis of gene expression changes in the epidermis upon exposure to H2O2 concentrations known to promote wound repair. Comparative transcriptome analysis using RNA-seq data from larval zebrafish and previously reported microarray data from a human epidermal keratinocyte line shows that H2O2 activates conserved cell migration, adhesion, cytoprotective and anti-apoptotic programs in both zebrafish and human keratinocytes. Further assessment of expression characteristics and signaling pathways revealed the activation of three major H2O2-dependent pathways, EGF, FOXO1, and IKKα. This study expands on our current understanding of the clinical potential of low-level H2O2 for the promotion of epidermal wound repair and provides potential candidates in the treatment of wound healing deficits.

  20. Single and joint toxicity assessment of four currently used pesticides to zebrafish (Danio rerio) using traditional and molecular endpoints.

    Science.gov (United States)

    Wang, Yanhua; Wu, Shenggan; Chen, Jine; Zhang, Changpeng; Xu, Zhenlan; Li, Gang; Cai, Leiming; Shen, Weifeng; Wang, Qiang

    2018-02-01

    Pesticides usually present in mixtures in surface waters, although they are traditionally regulated on an individual basis in aquatic ecosystems. In this study, we aimed to investigate the lethal and transcriptional responses of individual and combined pesticides (iprodione, pyrimethanil, pyraclostrobin and acetamiprid) on zebrafish (Danio rerio). Semi-static toxicity test indicated that the greatest toxicity to the four life stages (embryonic, larval, juvenile and adult stages) of D. rerio was detected from pyraclostrobin, followed by iprodione and pyrimethanil. In contrast, the lowest toxicity to the organisms was found from acetamiprid. Most of the selected pesticides exerted greater toxicities to D. rerio of embryonic stage compared with other life stages. Synergistic responses were observed from all binary mixtures of iprodione in combination with pyrimethanil or acetamiprid and ternary mixtures of iprodione+pyraclostrobin in combination with pyrimethanil or acetamiprid. The expressions of 16 genes related to cell apoptosis pathway, oxidative stress response, innate immunity and endocrine disruption at the mRNA level showed that zebrafish embryos were affected by the individual or combined pesticides. The expressions of P53, Tnf, TRβ, Tsh and Cyp19a exhibited greater changes upon exposure to combined pesticides compared with individual pesticides. Taken together, increased toxicity might be triggered by the simultaneous presence of several pesticides in the aquatic environment, which seriously damaged the non-target organisms. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Characterization of genetically targeted neuron types in the zebrafish optic tectum

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    Estuardo eRobles

    2011-02-01

    Full Text Available The optically transparent larval zebrafish is ideally suited for in vivo analyses of neural circuitry controlling visually guided behaviors. However, there is a lack of information regarding specific cell types in the major retinorecipient brain region of the fish, the optic tectum. Here we report the characterization of three previously unidentified tectal cell types that are specifically labeled by dlx5/6 enhancer elements. In vivo laser scanning microscopy in conjunction with ex vivo array tomography revealed that these neurons differ in their morphologies, synaptic connectivity, and neurotransmitter phenotypes. The first type is an excitatory bistratified periventricular interneuron (bsPVIN that forms a dendritic arbor in the retinorecipient stratum fibrosum griseum et superficiale (SFGS and an axonal arbor in the stratum griseum centrale (SGC. The second type, a GABAergic nonstratified periventricular interneuron (nsPVIN, extends a bushy arbor containing both dendrites and axons into the SGC and the deepest sublayers of the SFGS. The third type is a GABAergic periventricular projection neuron (PVPN that extends a dendritic arbor into the SGC and a long axon to the torus semicircularis, medulla oblongata, and anterior hindbrain. Interestingly, the same axons form en passant synapses within the deepest neuropil layer of the tectum, the stratum album centrale. This approach revealed several novel aspects of tectal circuitry, including: (1 a glutamatergic mode of transmission from the superficial, retinorecipient neuropil layers to the deeper, output layers, (2 the presence of interneurons with mixed dendrite/axon arbors likely involved in local processing, and (3 a heretofore unknown GABAergic tectofugal projection to midbrain and hindbrain. These observations establish a framework for studying the morphological and functional differentiation of neural circuits in the zebrafish visual system.

  2. β-catenin/Wnt signaling controls progenitor fate in the developing and regenerating zebrafish retina

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    Meyers Jason R

    2012-08-01

    Full Text Available Abstract Background The zebrafish retina maintains two populations of stem cells: first, the germinal zone or ciliary marginal zone (CMZ contains multipotent retinal progenitors that add cells to the retinal periphery as the fish continue to grow; second, radial glia (Müller cells occasionally divide asymmetrically to generate committed progenitors that differentiate into rod photoreceptors, which are added interstitially throughout the retina with growth. Retinal injury stimulates Müller glia to dedifferentiate, re-enter the cell cycle, and generate multipotent retinal progenitors similar to those in the CMZ to replace missing neurons. The specific signals that maintain these two distinct populations of endogenous retinal stem cells are not understood. Results We used genetic and pharmacological manipulation of the β-catenin/Wnt signaling pathway to show that it is required to maintain proliferation in the CMZ and that hyperstimulation of β-catenin/Wnt signaling inhibits normal retinal differentiation and expands the population of proliferative retinal progenitors. To test whether similar effects occur during regeneration, we developed a method for making rapid, selective photoreceptor ablations in larval zebrafish with intense light. We found that dephosphorylated β-catenin accumulates in Müller glia as they re-enter the cell cycle following injury, but not in Müller glia that remain quiescent. Activation of Wnt signaling is required for regenerative proliferation, and hyperstimulation results in loss of Müller glia from the INL as all proliferative cells move into the ONL. Conclusions β-catenin/Wnt signaling is thus required for the maintenance of retinal progenitors during both initial development and lesion-induced regeneration, and is sufficient to prevent differentiation of those progenitors and maintain them in a proliferative state. This suggests that the β-catenin/Wnt cascade is part of the shared molecular circuitry that

  3. The heartstrings mutation in zebrafish causes heart/fin Tbx5 deficiency syndrome.

    Science.gov (United States)

    Garrity, Deborah M; Childs, Sarah; Fishman, Mark C

    2002-10-01

    Holt-Oram syndrome is one of the autosomal dominant human "heart-hand" disorders, with a combination of upper limb malformations and cardiac defects. Holt-Oram syndrome is caused by mutations in the TBX5 gene, a member of a large family of T-box transcription factors that play important roles in cell-type specification and morphogenesis. In a screen for mutations affecting zebrafish cardiac function, we isolated the recessive lethal mutant heartstrings, which lacks pectoral fins and exhibits severe cardiac dysfunction, beginning with a slow heart rate and progressing to a stretched, non-functional heart. We mapped and cloned the heartstrings mutation and find it to encode the zebrafish ortholog of the TBX5 gene. The heartstrings mutation causes premature termination at amino acid 316. Homozygous mutant embryos never develop pectoral fin buds and do not express several markers of early fin differentiation. The total absence of any fin bud differentiation distinguishes heartstrings from most other mutations that affect zebrafish fin development, suggesting that Tbx5 functions very early in the pectoral fin induction pathway. Moderate reduction of Tbx5 by morpholino causes fin malformations, revealing an additional early requirement for Tbx5 in coordinating the axes of fin outgrowth. The heart of heartstrings mutant embryos appears to form and function normally through the early heart tube stage, manifesting only a slight bradycardia compared with wild-type siblings. However, the heart fails to loop and then progressively deteriorates, a process affecting the ventricle as well as the atrium. Relative to mammals, fish require lower levels of Tbx5 to produce malformed appendages and display whole-heart rather than atrial-predominant cardiac defects. However, the syndromic deficiencies of tbx5 mutation are remarkably well retained between fish and mammals.

  4. Zebrafish tissue injury causes upregulation of interleukin-1 and caspase-dependent amplification of the inflammatory response

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    Nikolay V. Ogryzko

    2014-02-01

    Full Text Available Interleukin-1 (IL-1, the ‘gatekeeper’ of inflammation, is the apical cytokine in a signalling cascade that drives the early response to injury or infection. Expression, processing and secretion of IL-1 are tightly controlled, and dysregulated IL-1 signalling has been implicated in a number of pathologies ranging from atherosclerosis to complications of infection. Our understanding of these processes comes from in vitro monocytic cell culture models as lines or primary isolates, in which a range and spectra of IL-1 secretion mechanisms have been described. We therefore investigated whether zebrafish embryos provide a suitable in vivo model for studying IL-1-mediated inflammation. Structurally, zebrafish IL-1β shares a β-sheet-rich trefoil structure with its human counterpart. Functionally, leukocyte expression of IL-1β was detectable only following injury, which activated leukocytes throughout zebrafish embryos. Migration of macrophages and neutrophils was attenuated by inhibitors of either caspase-1 or P2X7, which similarly inhibited the activation of NF-κB at the site of injury. Zebrafish offer a new and versatile model to study the IL-1β pathway in inflammatory disease and should offer unique insights into IL-1 biology in vivo.

  5. Zebrafish tissue injury causes upregulation of interleukin-1 and caspase-dependent amplification of the inflammatory response.

    Science.gov (United States)

    Ogryzko, Nikolay V; Hoggett, Emily E; Solaymani-Kohal, Sara; Tazzyman, Simon; Chico, Timothy J A; Renshaw, Stephen A; Wilson, Heather L

    2014-02-01

    Interleukin-1 (IL-1), the 'gatekeeper' of inflammation, is the apical cytokine in a signalling cascade that drives the early response to injury or infection. Expression, processing and secretion of IL-1 are tightly controlled, and dysregulated IL-1 signalling has been implicated in a number of pathologies ranging from atherosclerosis to complications of infection. Our understanding of these processes comes from in vitro monocytic cell culture models as lines or primary isolates, in which a range and spectra of IL-1 secretion mechanisms have been described. We therefore investigated whether zebrafish embryos provide a suitable in vivo model for studying IL-1-mediated inflammation. Structurally, zebrafish IL-1β shares a β-sheet-rich trefoil structure with its human counterpart. Functionally, leukocyte expression of IL-1β was detectable only following injury, which activated leukocytes throughout zebrafish embryos. Migration of macrophages and neutrophils was attenuated by inhibitors of either caspase-1 or P2X7, which similarly inhibited the activation of NF-κB at the site of injury. Zebrafish offer a new and versatile model to study the IL-1β pathway in inflammatory disease and should offer unique insights into IL-1 biology in vivo.

  6. Altered Adipogenesis in Zebrafish Larvae Following High Fat Diet and Chemical Exposure Is Visualised by Stimulated Raman Scattering Microscopy

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    Marjo J. den Broeder

    2017-04-01

    Full Text Available Early life stage exposure to environmental chemicals may play a role in obesity by altering adipogenesis; however, robust in vivo methods to quantify these effects are lacking. The goal of this study was to analyze the effects of developmental exposure to chemicals on adipogenesis in the zebrafish (Danio rerio. We used label-free Stimulated Raman Scattering (SRS microscopy for the first time to image zebrafish adipogenesis at 15 days post fertilization (dpf and compared standard feed conditions (StF to a high fat diet (HFD or high glucose diet (HGD. We also exposed zebrafish embryos to a non-toxic concentration of tributyltin (TBT, 1 nM or Tris(1,3-dichloroisopropylphosphate (TDCiPP, 0.5 µM from 0–6 dpf and reared larvae to 15 dpf under StF. Potential molecular mechanisms of altered adipogenesis were examined by qPCR. Diet-dependent modulation of adipogenesis was observed, with HFD resulting in a threefold increase in larvae with adipocytes, compared to StF and HGD. Developmental exposure to TBT but not TDCiPP significantly increased adipocyte differentiation. The expression of adipogenic genes such as pparda, lxr and lepa was altered in response to HFD or chemicals. This study shows that SRS microscopy can be successfully applied to zebrafish to visualize and quantify adipogenesis, and is a powerful approach for identifying obesogenic chemicals in vivo.

  7. Altered Adipogenesis in Zebrafish Larvae Following High Fat Diet and Chemical Exposure Is Visualised by Stimulated Raman Scattering Microscopy

    Science.gov (United States)

    den Broeder, Marjo J.; Moester, Miriam J. B.; Kamstra, Jorke H.; Cenijn, Peter H.; Davidoiu, Valentina; Kamminga, Leonie M.; Ariese, Freek; de Boer, Johannes F.; Legler, Juliette

    2017-01-01

    Early life stage exposure to environmental chemicals may play a role in obesity by altering adipogenesis; however, robust in vivo methods to quantify these effects are lacking. The goal of this study was to analyze the effects of developmental exposure to chemicals on adipogenesis in the zebrafish (Danio rerio). We used label-free Stimulated Raman Scattering (SRS) microscopy for the first time to image zebrafish adipogenesis at 15 days post fertilization (dpf) and compared standard feed conditions (StF) to a high fat diet (HFD) or high glucose diet (HGD). We also exposed zebrafish embryos to a non-toxic concentration of tributyltin (TBT, 1 nM) or Tris(1,3-dichloroisopropyl)phosphate (TDCiPP, 0.5 µM) from 0–6 dpf and reared larvae to 15 dpf under StF. Potential molecular mechanisms of altered adipogenesis were examined by qPCR. Diet-dependent modulation of adipogenesis was observed, with HFD resulting in a threefold increase in larvae with adipocytes, compared to StF and HGD. Developmental exposure to TBT but not TDCiPP significantly increased adipocyte differentiation. The expression of adipogenic genes such as pparda, lxr and lepa was altered in response to HFD or chemicals. This study shows that SRS microscopy can be successfully applied to zebrafish to visualize and quantify adipogenesis, and is a powerful approach for identifying obesogenic chemicals in vivo. PMID:28441764

  8. Zebrafish embryos as a screen for DNA methylation modifications after compound exposure

    Energy Technology Data Exchange (ETDEWEB)

    Bouwmeester, Manon C.; Ruiter, Sander; Lommelaars, Tobias; Sippel, Josefine; Hodemaekers, Hennie M. [Center for Health Protection, National Institute for Public Health and the Environment (RIVM), PO Box 1, 3720 BA Bilthoven (Netherlands); Brandhof, Evert-Jan van den [Center for Environmental Quality, National Institute for Public Health and the Environment (RIVM), PO Box 1, 3720 BA Bilthoven (Netherlands); Pennings, Jeroen L.A. [Center for Health Protection, National Institute for Public Health and the Environment (RIVM), PO Box 1, 3720 BA Bilthoven (Netherlands); Kamstra, Jorke H. [Institute for Environmental Studies (IVM), VU University, De Boelelaan 1085, 1081 HV Amsterdam (Netherlands); Jelinek, Jaroslav [Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, PA (United States); Issa, Jean-Pierre J. [Fels Institute for Cancer Research and Molecular Biology, Temple University School of Medicine, Philadelphia, PA (United States); Department of Leukemia, The University of Texas MD Anderson Cancer Center, Houston, TX (United States); Legler, Juliette [Institute for Environmental Studies (IVM), VU University, De Boelelaan 1085, 1081 HV Amsterdam (Netherlands); Ven, Leo T.M. van der, E-mail: leo.van.der.ven@rivm.nl [Center for Health Protection, National Institute for Public Health and the Environment (RIVM), PO Box 1, 3720 BA Bilthoven (Netherlands)

    2016-01-15

    Modified epigenetic programming early in life is proposed to underlie the development of an adverse adult phenotype, known as the Developmental Origins of Health and Disease (DOHaD) concept. Several environmental contaminants have been implicated as modifying factors of the developing epigenome. This underlines the need to investigate this newly recognized toxicological risk and systematically screen for the epigenome modifying potential of compounds. In this study, we examined the applicability of the zebrafish embryo as a screening model for DNA methylation modifications. Embryos were exposed from 0 to 72 h post fertilization (hpf) to bisphenol-A (BPA), diethylstilbestrol, 17α-ethynylestradiol, nickel, cadmium, tributyltin, arsenite, perfluoroctanoic acid, valproic acid, flusilazole, 5-azacytidine (5AC) in subtoxic concentrations. Both global and site-specific methylation was examined. Global methylation was only affected by 5AC. Genome wide locus-specific analysis was performed for BPA exposed embryos using Digital Restriction Enzyme Analysis of Methylation (DREAM), which showed minimal wide scale effects on the genome, whereas potential informative markers were not confirmed by pyrosequencing. Site-specific methylation was examined in the promoter regions of three selected genes vasa, vtg1 and cyp19a2, of which vasa (ddx4) was the most responsive. This analysis distinguished estrogenic compounds from metals by direction and sensitivity of the effect compared to embryotoxicity. In conclusion, the zebrafish embryo is a potential screening tool to examine DNA methylation modifications after xenobiotic exposure. The next step is to examine the adult phenotype of exposed embryos and to analyze molecular mechanisms that potentially link epigenetic effects and altered phenotypes, to support the DOHaD hypothesis. - Highlights: • Compound induced effects on DNA methylation in zebrafish embryos • Global methylation not an informative biomarker • Minimal genome

  9. Zebrafish eda and edar mutants reveal conserved and ancestral roles of ectodysplasin signaling in vertebrates.

    Directory of Open Access Journals (Sweden)

    Matthew P Harris

    2008-10-01

    Full Text Available The genetic basis of the development and variation of adult form of vertebrates is not well understood. To address this problem, we performed a mutant screen to identify genes essential for the formation of adult skeletal structures of the zebrafish. Here, we describe the phenotypic and molecular characterization of a set of mutants showing loss of adult structures of the dermal skeleton, such as the rays of the fins and the scales, as well as the pharyngeal teeth. The mutations represent adult-viable, loss of function alleles in the ectodysplasin (eda and ectodysplasin receptor (edar genes. These genes are frequently mutated in the human hereditary disease hypohidrotic ectodermal dysplasia (HED; OMIM 224900, 305100 that affects the development of integumentary appendages such as hair and teeth. We find mutations in zebrafish edar that affect similar residues as mutated in human cases of HED and show similar phenotypic consequences. eda and edar are not required for early zebrafish development, but are rather specific for the development of adult skeletal and dental structures. We find that the defects of the fins and scales are due to the role of Eda signaling in organizing epidermal cells into discrete signaling centers of the scale epidermal placode and fin fold. Our genetic analysis demonstrates dose-sensitive and organ-specific response to alteration in levels of Eda signaling. In addition, we show substantial buffering of the effect of loss of edar function in different genetic backgrounds, suggesting canalization of this developmental system. We uncover a previously unknown role of Eda signaling in teleosts and show conservation of the developmental mechanisms involved in the formation and variation of both integumentary appendages and limbs. Lastly, our findings point to the utility of adult genetic screens in the zebrafish in identifying essential developmental processes involved in human disease and in morphological evolution.

  10. Zebrafish embryos as a screen for DNA methylation modifications after compound exposure

    International Nuclear Information System (INIS)

    Bouwmeester, Manon C.; Ruiter, Sander; Lommelaars, Tobias; Sippel, Josefine; Hodemaekers, Hennie M.; Brandhof, Evert-Jan van den; Pennings, Jeroen L.A.; Kamstra, Jorke H.; Jelinek, Jaroslav; Issa, Jean-Pierre J.; Legler, Juliette; Ven, Leo T.M. van der

    2016-01-01

    Modified epigenetic programming early in life is proposed to underlie the development of an adverse adult phenotype, known as the Developmental Origins of Health and Disease (DOHaD) concept. Several environmental contaminants have been implicated as modifying factors of the developing epigenome. This underlines the need to investigate this newly recognized toxicological risk and systematically screen for the epigenome modifying potential of compounds. In this study, we examined the applicability of the zebrafish embryo as a screening model for DNA methylation modifications. Embryos were exposed from 0 to 72 h post fertilization (hpf) to bisphenol-A (BPA), diethylstilbestrol, 17α-ethynylestradiol, nickel, cadmium, tributyltin, arsenite, perfluoroctanoic acid, valproic acid, flusilazole, 5-azacytidine (5AC) in subtoxic concentrations. Both global and site-specific methylation was examined. Global methylation was only affected by 5AC. Genome wide locus-specific analysis was performed for BPA exposed embryos using Digital Restriction Enzyme Analysis of Methylation (DREAM), which showed minimal wide scale effects on the genome, whereas potential informative markers were not confirmed by pyrosequencing. Site-specific methylation was examined in the promoter regions of three selected genes vasa, vtg1 and cyp19a2, of which vasa (ddx4) was the most responsive. This analysis distinguished estrogenic compounds from metals by direction and sensitivity of the effect compared to embryotoxicity. In conclusion, the zebrafish embryo is a potential screening tool to examine DNA methylation modifications after xenobiotic exposure. The next step is to examine the adult phenotype of exposed embryos and to analyze molecular mechanisms that potentially link epigenetic effects and altered phenotypes, to support the DOHaD hypothesis. - Highlights: • Compound induced effects on DNA methylation in zebrafish embryos • Global methylation not an informative biomarker • Minimal genome

  11. Sex reversal in zebrafish fancl mutants is caused by Tp53-mediated germ cell apoptosis.

    Directory of Open Access Journals (Sweden)

    Adriana Rodríguez-Marí

    2010-07-01

    Full Text Available The molecular genetic mechanisms of sex determination are not known for most vertebrates, including zebrafish. We identified a mutation in the zebrafish fancl gene that causes homozygous mutants to develop as fertile males due to female-to-male sex reversal. Fancl is a member of the Fanconi Anemia/BRCA DNA repair pathway. Experiments showed that zebrafish fancl was expressed in developing germ cells in bipotential gonads at the critical time of sexual fate determination. Caspase-3 immunoassays revealed increased germ cell apoptosis in fancl mutants that compromised oocyte survival. In the absence of oocytes surviving through meiosis, somatic cells of mutant gonads did not maintain expression of the ovary gene cyp19a1a and did not down-regulate expression of the early testis gene amh; consequently, gonads masculinized and became testes. Remarkably, results showed that the introduction of a tp53 (p53 mutation into fancl mutants rescued the sex-reversal phenotype by reducing germ cell apoptosis and, thus, allowed fancl mutants to become fertile females. Our results show that Fancl function is not essential for spermatogonia and oogonia to become sperm or mature oocytes, but instead suggest that Fancl function is involved in the survival of developing oocytes through meiosis. This work reveals that Tp53-mediated germ cell apoptosis induces sex reversal after the mutation of a DNA-repair pathway gene by compromising the survival of oocytes and suggests the existence of an oocyte-derived signal that biases gonad fate towards the female developmental pathway and thereby controls zebrafish sex determination.

  12. Zebrafish eda and edar Mutants Reveal Conserved and Ancestral Roles of Ectodysplasin Signaling in Vertebrates

    Science.gov (United States)

    Harris, Matthew P.; Rohner, Nicolas; Schwarz, Heinz; Perathoner, Simon; Konstantinidis, Peter; Nüsslein-Volhard, Christiane

    2008-01-01

    The genetic basis of the development and variation of adult form of vertebrates is not well understood. To address this problem, we performed a mutant screen to identify genes essential for the formation of adult skeletal structures of the zebrafish. Here, we describe the phenotypic and molecular characterization of a set of mutants showing loss of adult structures of the dermal skeleton, such as the rays of the fins and the scales, as well as the pharyngeal teeth. The mutations represent adult-viable, loss of function alleles in the ectodysplasin (eda) and ectodysplasin receptor (edar) genes. These genes are frequently mutated in the human hereditary disease hypohidrotic ectodermal dysplasia (HED; OMIM 224900, 305100) that affects the development of integumentary appendages such as hair and teeth. We find mutations in zebrafish edar that affect similar residues as mutated in human cases of HED and show similar phenotypic consequences. eda and edar are not required for early zebrafish development, but are rather specific for the development of adult skeletal and dental structures. We find that the defects of the fins and scales are due to the role of Eda signaling in organizing epidermal cells into discrete signaling centers of the scale epidermal placode and fin fold. Our genetic analysis demonstrates dose-sensitive and organ-specific response to alteration in levels of Eda signaling. In addition, we show substantial buffering of the effect of loss of edar function in different genetic backgrounds, suggesting canalization of this developmental system. We uncover a previously unknown role of Eda signaling in teleosts and show conservation of the developmental mechanisms involved in the formation and variation of both integumentary appendages and limbs. Lastly, our findings point to the utility of adult genetic screens in the zebrafish in identifying essential developmental processes involved in human disease and in morphological evolution. PMID:18833299

  13. Embryonic Zebrafish Model - A Well-Established Method for Rapidly Assessing the Toxicity of Homeopathic Drugs

    Science.gov (United States)

    Gupta, Himanshu R; Patil, Yogesh; Singh, Dipty

    2016-01-01

    Objectives: Advancements in nanotechnology have led to nanoparticle (NP) use in various fields of medicine. Although the potential of NPs is promising, the lack of documented evidence on the toxicological effects of NPs is concerning. A few studies have documented that homeopathy uses NPs. Unfortunately, very few sound scientific studies have explored the toxic effects of homeopathic drugs. Citing this lack of high-quality scientific evidence, regulatory agencies have been reluctant to endorse homeopathic treatment as an alternative or adjunct treatment. This study aimed to enhance our insight into the impact of commercially-available homeopathic drugs, to study the presence of NPs in those drugs and any deleterious effects they might have, and to determine the distribution pattern of NPs in zebrafish embryos (Danio rerio). Methods: Homeopathic dilutions were studied using high-resolution transmission electron microscopy with selected area electron diffraction (SAED). For the toxicity assessment on Zebrafish, embryos were exposed to a test solution from 4 - 6 hours post-fertilization, and embryos/larvae were assessed up to 5 days post-fertilization (dpf) for viability and morphology. Toxicity was recorded in terms of mortality, hatching delay, phenotypic defects and metal accumulation. Around 5 dpf was found to be the optimum developmental stage for evaluation. Results: The present study aimed to conclusively prove the presence of NPs in all high dilutions of homeopathic drugs. Embryonic zebrafish were exposed to three homeopathic drugs with two potencies (30CH, 200CH) during early embryogenesis. The resulting morphological and cellular responses were observed. Exposure to these potencies produced no visibly significant malformations, pericardial edema, and mortality and no necrotic and apoptotic cellular death. Conclusion: Our findings clearly demonstrate that no toxic effects were observed for these three homeopathic drugs at the potencies and exposure times used

  14. Establishment of infection models in zebrafish larvae (Danio rerio to study the pathogenesis of Aeromonas hydrophila.

    Directory of Open Access Journals (Sweden)

    Paolo Roberto Saraceni

    2016-08-01

    Full Text Available Aeromonas hydrophila is a Gram-negative opportunistic pathogen of fish and terrestrial animals. In humans, A. hydrophila mainly causes gastroenteritis, septicaemia and tissue infections. The mechanisms of infection, the main virulence factors and the host immune response triggered by A. hydrophila have been studied in detail using murine models and adult fish. However, the great limitation of studying adult animals is that the animal must be sacrificed and its tissues/organs extracted, which prevents the study of the infectious processes in the whole living animal.Zebrafish larvae are being used for the analysis of several infectious diseases, but their use for studying the pathogenesis of A. hydrophila has never been explored. The great advantage of zebrafish larvae is their transparency during the first week after fertilization, which allows detailed descriptions of the infectious processes using in vivo imaging techniques such as differential interferential contrast (DIC and fluorescence microscopy. Moreover, the availability of fluorescent pathogens and transgenic reporter zebrafish lines expressing fluorescent immune cells, immune marker genes or cytokines/chemokines allows the host-pathogen interactions to be characterized.The present study explores the suitability of zebrafish larvae to study the pathogenesis of A. hydrophila and the interaction mechanisms between the bacterium and the innate immune responses through an infection model using different routes for infection. We used an early-embryo infection model at 3 days post-fertilization (dpf through the microinjection of A. hydrophila into the duct of Cuvier, caudal vein, notochord or muscle and two bath infection models using 4 dpf healthy and injured larvae. The latter resembled the natural conditions under which A. hydrophila produces infectious diseases in animals. We compared the cellular processes after infection in each anatomical site by confocal fluorescence imaging and

  15. High-content screening in zebrafish embryos identifies butafenacil as a potent inducer of anemia.

    Directory of Open Access Journals (Sweden)

    Jessica K Leet

    Full Text Available Using transgenic zebrafish (fli1:egfp that stably express enhanced green fluorescent protein (eGFP within vascular endothelial cells, we recently developed and optimized a 384-well high-content screening (HCS assay that enables us to screen and identify chemicals affecting cardiovascular development and function at non-teratogenic concentrations. Within this assay, automated image acquisition procedures and custom image analysis protocols are used to quantify body length, heart rate, circulation, pericardial area, and intersegmental vessel area within individual live embryos exposed from 5 to 72 hours post-fertilization. After ranking developmental toxicity data generated from the U.S. Environmental Protection Agency's (EPA's zebrafish teratogenesis assay, we screened 26 of the most acutely toxic chemicals within EPA's ToxCast Phase-I library in concentration-response format (0.05-50 µM using this HCS assay. Based on this screen, we identified butafenacil as a potent inducer of anemia, as exposure from 0.39 to 3.125 µM butafenacil completely abolished arterial circulation in the absence of effects on all other endpoints evaluated. Butafenacil is an herbicide that inhibits protoporphyrinogen oxidase (PPO--an enzyme necessary for heme production in vertebrates. Using o-dianisidine staining, we then revealed that severe butafenacil-induced anemia in zebrafish was due to a complete loss of hemoglobin following exposure during early development. Therefore, six additional PPO inhibitors within the ToxCast Phase-I library were screened to determine whether anemia represents a common adverse outcome for these herbicides. Embryonic exposure to only one of these PPO inhibitors--flumioxazin--resulted in a similar phenotype as butafenacil, albeit not as severe as butafenacil. Overall, this study highlights the potential utility of this assay for (1 screening chemicals for cardiovascular toxicity and (2 prioritizing chemicals for future hypothesis

  16. Zebrafish model of tuberous sclerosis complex reveals cell-autonomous and non-cell-autonomous functions of mutant tuberin

    Directory of Open Access Journals (Sweden)

    Seok-Hyung Kim

    2011-03-01

    Tuberous sclerosis complex (TSC is an autosomal dominant disease caused by mutations in either the TSC1 (encodes hamartin or TSC2 (encodes tuberin genes. Patients with TSC have hamartomas in various organs throughout the whole body, most notably in the brain, skin, eye, heart, kidney and lung. To study the development of hamartomas, we generated a zebrafish model of TSC featuring a nonsense mutation (vu242 in the tsc2 gene. This tsc2vu242 allele encodes a truncated Tuberin protein lacking the GAP domain, which is required for inhibition of Rheb and of the TOR kinase within TORC1. We show that tsc2vu242 is a recessive larval-lethal mutation that causes increased cell size in the brain and liver. Greatly elevated TORC1 signaling is observed in tsc2vu242/vu242 homozygous zebrafish, and is moderately increased in tsc2vu242/+ heterozygotes. Forebrain neurons are poorly organized in tsc2vu242/vu242 homozygous mutants, which have extensive gray and white matter disorganization and ectopically positioned cells. Genetic mosaic analyses demonstrate that tsc2 limits TORC1 signaling in a cell-autonomous manner. However, in chimeric animals, tsc2vu242/vu242 mutant cells also mislocalize wild-type host cells in the forebrain in a non-cell-autonomous manner. These results demonstrate a highly conserved role of tsc2 in zebrafish and establish a new animal model for studies of TSC. The finding of a non-cell-autonomous function of mutant cells might help explain the formation of brain hamartomas and cortical malformations in human TSC.

  17. ngs (Notochord Granular Surface) Gene Encodes a Novel Type of Intermediate Filament Family Protein Essential for Notochord Maintenance in Zebrafish*

    Science.gov (United States)

    Tong, Xiangjun; Xia, Zhidan; Zu, Yao; Telfer, Helena; Hu, Jing; Yu, Jingyi; Liu, Huan; Zhang, Quan; Sodmergen; Lin, Shuo; Zhang, Bo

    2013-01-01

    The notochord is an important organ involved in embryonic patterning and locomotion. In zebrafish, the mature notochord consists of a single stack of fully differentiated, large vacuolated cells called chordocytes, surrounded by a single layer of less differentiated notochordal epithelial cells called chordoblasts. Through genetic analysis of zebrafish lines carrying pseudo-typed retroviral insertions, a mutant exhibiting a defective notochord with a granular appearance was isolated, and the corresponding gene was identified as ngs (notochord granular surface), which was specifically expressed in the notochord. In the mutants, the notochord started to degenerate from 32 hours post-fertilization, and the chordocytes were then gradually replaced by smaller cells derived from chordoblasts. The granular notochord phenotype was alleviated by anesthetizing the mutant embryos with tricaine to prevent muscle contraction and locomotion. Phylogenetic analysis showed that ngs encodes a new type of intermediate filament (IF) family protein, which we named chordostatin based on its function. Under the transmission electron microcopy, bundles of 10-nm-thick IF-like filaments were enriched in the chordocytes of wild-type zebrafish embryos, whereas the chordocytes in ngs mutants lacked IF-like structures. Furthermore, chordostatin-enhanced GFP (EGFP) fusion protein assembled into a filamentous network specifically in chordocytes. Taken together, our work demonstrates that ngs encodes a novel type of IF protein and functions to maintain notochord integrity for larval development and locomotion. Our work sheds light on the mechanisms of notochord structural maintenance, as well as the evolution and biological function of IF family proteins. PMID:23132861

  18. PEG-b-PCL polymeric nano-micelle inhibits vascular angiogenesis by activating p53-dependent apoptosis in zebrafish.

    Science.gov (United States)

    Zhou, Tian; Dong, Qinglei; Shen, Yang; Wu, Wei; Wu, Haide; Luo, Xianglin; Liao, Xiaoling; Wang, Guixue

    Micro/nanoparticles could cause adverse effects on cardiovascular system and increase the risk for cardiovascular disease-related events. Nanoparticles prepared from poly(ethylene glycol) (PEG)- b -poly( ε -caprolactone) (PCL), namely PEG- b -PCL, a widely studied biodegradable copolymer, are promising carriers for the drug delivery systems. However, it is unknown whether polymeric PEG- b -PCL nano-micelles give rise to potential complications of the cardiovascular system. Zebrafish were used as an in vivo model to evaluate the effects of PEG- b -PCL nano-micelle on cardiovascular development. The results showed that PEG- b -PCL nano-micelle caused embryo mortality as well as embryonic and larval malformations in a dose-dependent manner. To determine PEG- b -PCL nano-micelle effects on embryonic angiogenesis, a critical process in zebrafish cardiovascular development, growth of intersegmental vessels (ISVs) and caudal vessels (CVs) in flk1-GFP transgenic zebrafish embryos using fluorescent stereomicroscopy were examined. The expression of fetal liver kinase 1 (flk1), an angiogenic factor, by real-time quantitative polymerase chain reaction (qPCR) and in situ whole-mount hybridization were also analyzed. PEG- b -PCL nano-micelle decreased growth of ISVs and CVs, as well as reduced flk1 expression in a concentration-dependent manner. Parallel to the inhibitory effects on angiogenesis, PEG- b -PCL nano-micelle exposure upregulated p53 pro-apoptotic pathway and induced cellular apoptosis in angiogenic regions by qPCR and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) apoptosis assay. This study further showed that inhibiting p53 activity, either by pharmacological inhibitor or RNA interference, could abrogate the apoptosis and angiogenic defects caused by PEG- b -PCL nano-micelles, indicating that PEG- b -PCL nano-micelle inhibits angiogenesis by activating p53-mediated apoptosis. This study indicates that polymeric PEG- b -PCL nano-micelle could

  19. Mosquito larval source management for controlling malaria

    Science.gov (United States)

    Tusting, Lucy S; Thwing, Julie; Sinclair, David; Fillinger, Ulrike; Gimnig, John; Bonner, Kimberly E; Bottomley, Christian; Lindsay, Steven W

    2015-01-01

    Background Malaria is an important cause of illness and death in people living in many parts of the world, especially sub-Saharan Africa. Long-lasting insecticide treated bed nets (LLINs) and indoor residual spraying (IRS) reduce malaria transmission by targeting the adult mosquito vector and are key components of malaria control programmes. However, mosquito numbers may also be reduced by larval source management (LSM), which targets mosquito larvae as they mature in aquatic habitats. This is conducted by permanently or temporarily reducing the availability of larval habitats (habitat modification and habitat manipulation), or by adding substances to standing water that either kill or inhibit the development of larvae (larviciding). Objectives To evaluate the effectiveness of mosquito LSM for preventing malaria. Search methods We searched the Cochrane Infectious Diseases Group Specialized Register; Cochrane Central Register of Controlled Trials (CENTRAL); MEDLINE; EMBASE; CABS Abstracts; and LILACS up to 24 October 2012. We handsearched the Tropical Diseases Bulletin from 1900 to 2010, the archives of the World Health Organization (up to 11 February 2011), and the literature database of the Armed Forces Pest Management Board (up to 2 March 2011). We also contacted colleagues in the field for relevant articles. Selection criteria We included cluster randomized controlled trials (cluster-RCTs), controlled before-and-after trials with at least one year of baseline data, and randomized cross-over trials that compared LSM with no LSM for malaria control. We excluded trials that evaluated biological control of anopheline mosquitoes with larvivorous fish. Data collection and analysis At least two authors assessed each trial for eligibility. We extracted data and at least two authors independently determined the risk of bias in the included studies. We resolved all disagreements through discussion with a third author. We analyzed the data using Review Manager 5 software

  20. Characterization of behavioral and endocrine effects of LSD on zebrafish.

    Science.gov (United States)

    Grossman, Leah; Utterback, Eli; Stewart, Adam; Gaikwad, Siddharth; Chung, Kyung Min; Suciu, Christopher; Wong, Keith; Elegante, Marco; Elkhayat, Salem; Tan, Julia; Gilder, Thomas; Wu, Nadine; Dileo, John; Cachat, Jonathan; Kalueff, Allan V

    2010-12-25

    Lysergic acid diethylamide (LSD) is a potent hallucinogenic drug that strongly affects animal and human behavior. Although adult zebrafish (Danio rerio) are emerging as a promising neurobehavioral model, the effects of LSD on zebrafish have not been investigated previously. Several behavioral paradigms (the novel tank, observation cylinder, light-dark box, open field, T-maze, social preference and shoaling tests), as well as modern video-tracking tools and whole-body cortisol assay were used to characterize the effects of acute LSD in zebrafish. While lower doses (5-100 microg/L) did not affect zebrafish behavior, 250 microg/L LSD increased top dwelling and reduced freezing in the novel tank and observation cylinder tests, also affecting spatiotemporal patterns of activity (as assessed by 3D reconstruction of zebrafish traces and ethograms). LSD evoked mild thigmotaxis in the open field test, increased light behavior in the light-dark test, reduced the number of arm entries and freezing in the T-maze and social preference test, without affecting social preference. In contrast, LSD affected zebrafish shoaling (increasing the inter-fish distance in a group), and elevated whole-body cortisol levels. Overall, our findings show sensitivity of zebrafish to LSD action, and support the use of zebrafish models to study hallucinogenic drugs of abuse. Copyright (c) 2010 Elsevier B.V. All rights reserved.

  1. A bioenergetic model for zebrafish Danio rerio (Hamilton)

    Science.gov (United States)

    Chizinski, C.J.; Sharma, Bibek; Pope, K.L.; Patino, R.

    2008-01-01

    A bioenergetics model was developed from observed consumption, respiration and growth rates for zebrafish Danio rerio across a range (18-32?? C) of water temperatures, and evaluated with a 50 day laboratory trial at 28?? C. No significant bias in variable estimates was found during the validation trial; namely, predicted zebrafish mass generally agreed with observed mass. ?? 2008 The Authors.

  2. Contrasting effects of heavy metals and hydrocarbons on larval settlement and juvenile survival in sponges

    International Nuclear Information System (INIS)

    Cebrian, E.; Uriz, M.J.

    2007-01-01

    Metals and polycyclic aromatic hydrocarbons (PAHs) contaminate sediments and waters of coastal areas threatening early stages of invertebrate development. Effects on these stages may largely determine the decline and even disappearance of invertebrate populations in polluted environments. Our study aimed to determine the possible influence of metals (Cu and Cd) and PAHs on larval settlement and consecutive survival of two widespread sponges of the Mediterranean: Crambe crambe and Scopalina lophyropoda. Larvae of both species were exposed to Cu and Cd for a short period during 1 week, and settlement and following (6 months) survival of juvenile were monitored. Short exposures to copper and cadmium at the concentrations used did not affect C. crambe settlement compared with SW control, and no effect on consecutive survival of juvenile