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Sample records for adult zebrafish teleost

  1. Electroporation of adult zebrafish.

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    Rao, N Madhusudhana; Rambabu, K Murali; Rao, S Harinarayana

    2008-01-01

    We generated transient transgenic zebrafish by applying electrical pulses subsequent to injection of DNA into muscle tissue of 3-6-month old adult zebrafish. Electroporation parameters, such as number of pulses, voltage, and amount of plasmid DNA, were optimized and found that 6 pulses of 40 V/cm at 15 mug/fish increased the luciferase expression by 10-fold compared with those in controls. By measuring the expression of luciferase, in vivo by electroporation in adult zebrafish and in vitro using fish cell line (Xiphophorus xiphidium A2 cells), the strength of three promoters (CMV, human EF-1alpha, and Xenopus EF-1alpha) was compared. Subsequent to electroporation after injecting DNA in the mid region of zebrafish, expression of green fluorescent protein was found far away from the site of injection in the head and the tail sections. Thus, electroporation in adult zebrafish provides a rapid way of testing the behavior of gene sequences in the whole organism.

  2. Transducin duplicates in the zebrafish retina and pineal complex: differential specialisation after the teleost tetraploidisation.

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    David Lagman

    Full Text Available Gene duplications provide raw materials that can be selected for functional adaptations by evolutionary mechanisms. We describe here the results of 350 million years of evolution of three functionally related gene families: the alpha, beta and gamma subunits of transducins, the G protein involved in vision. Early vertebrate tetraploidisations resulted in separate transducin heterotrimers: gnat1/gnb1/gngt1 for rods, and gnat2/gnb3/gngt2 for cones. The teleost-specific tetraploidisation generated additional duplicates for gnb1, gnb3 and gngt2. We report here that the duplicates have undergone several types of subfunctionalisation or neofunctionalisation in the zebrafish. We have found that gnb1a and gnb1b are co-expressed at different levels in rods; gnb3a and gnb3b have undergone compartmentalisation restricting gnb3b to the dorsal and medial retina, however, gnb3a expression was detected only at very low levels in both larvae and adult retina; gngt2b expression is restricted to the dorsal and medial retina, whereas gngt2a is expressed ventrally. This dorsoventral distinction could be an adaptation to protect the lower part of the retina from intense light damage. The ontogenetic analysis shows earlier onset of expression in the pineal complex than in the retina, in accordance with its earlier maturation. Additionally, gnb1a but not gnb1b is expressed in the pineal complex, and gnb3b and gngt2b are transiently expressed in the pineal during ontogeny, thus showing partial temporal subfunctionalisation. These retina-pineal distinctions presumably reflect their distinct functional roles in vision and circadian rhythmicity. In summary, this study describes several functional differences between transducin gene duplicates resulting from the teleost-specific tetraploidisation.

  3. Transducin duplicates in the zebrafish retina and pineal complex: differential specialisation after the teleost tetraploidisation.

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    Lagman, David; Callado-Pérez, Amalia; Franzén, Ilkin E; Larhammar, Dan; Abalo, Xesús M

    2015-01-01

    Gene duplications provide raw materials that can be selected for functional adaptations by evolutionary mechanisms. We describe here the results of 350 million years of evolution of three functionally related gene families: the alpha, beta and gamma subunits of transducins, the G protein involved in vision. Early vertebrate tetraploidisations resulted in separate transducin heterotrimers: gnat1/gnb1/gngt1 for rods, and gnat2/gnb3/gngt2 for cones. The teleost-specific tetraploidisation generated additional duplicates for gnb1, gnb3 and gngt2. We report here that the duplicates have undergone several types of subfunctionalisation or neofunctionalisation in the zebrafish. We have found that gnb1a and gnb1b are co-expressed at different levels in rods; gnb3a and gnb3b have undergone compartmentalisation restricting gnb3b to the dorsal and medial retina, however, gnb3a expression was detected only at very low levels in both larvae and adult retina; gngt2b expression is restricted to the dorsal and medial retina, whereas gngt2a is expressed ventrally. This dorsoventral distinction could be an adaptation to protect the lower part of the retina from intense light damage. The ontogenetic analysis shows earlier onset of expression in the pineal complex than in the retina, in accordance with its earlier maturation. Additionally, gnb1a but not gnb1b is expressed in the pineal complex, and gnb3b and gngt2b are transiently expressed in the pineal during ontogeny, thus showing partial temporal subfunctionalisation. These retina-pineal distinctions presumably reflect their distinct functional roles in vision and circadian rhythmicity. In summary, this study describes several functional differences between transducin gene duplicates resulting from the teleost-specific tetraploidisation.

  4. Structural and functional characterization of neuromedin S in the teleost fish, zebrafish (Danio rerio).

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    Chen, Huapu; Huang, Hongxin; Chen, Xinggui; Deng, Siping; Zhu, Chunhua; Huang, Hai; Li, Guangli

    2016-01-01

    Neuromedin S (NMS) has been demonstrated to have important roles in many vertebrate physiological processes. However, the function of NMS in teleost fishes remains unclear. We explored the physiological roles of the NMS gene in the zebrafish model. An NMS cDNA was cloned from zebrafish brain tissue, and the full-length cDNA sequence was 521 bp in length and encoded a precursor of 110 amino acid residues. Interestingly, fish prepro-NMS is predicted to generate a short 34-residue peptide, designated as NMS-related peptide (NMSRP). Zebrafish prepro-NMS does not contain the NMS peptide which is found in the NMS precursors of mammals, and just retains the MNSRP peptide. A multiple-species sequence alignment showed that NMSRPs are conserved among the other sampled vertebrates. Zebrafish NMS mRNA was detected by RT-PCR revealing a tissue-specific distribution with high levels of expression in the brain, spleen, ovary, pituitary, and muscle. Furthermore, the locations of NMS-expressing cells in the zebrafish brain were detected by in situ hybridization in the parvocellular preoptic nucleus (PPa), the ventral zone of the periventricular hypothalamus (Hv), and lateral hypothalamic nucleus (LH). The levels of NMS mRNA in the hypothalamus were significantly increased after three days of food deprivation. Administration of zebrafish NMSRP by intraperitoneal injection significantly promoted the expression of neuropeptide Y (NPY) and orexin, suggesting an orexigenic role for NMSRP in zebrafish. The present study offers a new understanding of the NMS gene in vertebrates and increases our knowledge of the neuroendocrine regulation of feeding.

  5. Cerebroventricular microinjection (CVMI into adult zebrafish brain is an efficient misexpression method for forebrain ventricular cells.

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    Caghan Kizil

    Full Text Available The teleost fish Danio rerio (zebrafish has a remarkable ability to generate newborn neurons in its brain at adult stages of its lifespan-a process called adult neurogenesis. This ability relies on proliferating ventricular progenitors and is in striking contrast to mammalian brains that have rather restricted capacity for adult neurogenesis. Therefore, investigating the zebrafish brain can help not only to elucidate the molecular mechanisms of widespread adult neurogenesis in a vertebrate species, but also to design therapies in humans with what we learn from this teleost. Yet, understanding the cellular behavior and molecular programs underlying different biological processes in the adult zebrafish brain requires techniques that allow manipulation of gene function. As a complementary method to the currently used misexpression techniques in zebrafish, such as transgenic approaches or electroporation-based delivery of DNA, we devised a cerebroventricular microinjection (CVMI-assisted knockdown protocol that relies on vivo morpholino oligonucleotides, which do not require electroporation for cellular uptake. This rapid method allows uniform and efficient knockdown of genes in the ventricular cells of the zebrafish brain, which contain the neurogenic progenitors. We also provide data on the use of CVMI for growth factor administration to the brain--in our case FGF8, which modulates the proliferation rate of the ventricular cells. In this paper, we describe the CVMI method and discuss its potential uses in zebrafish.

  6. Neocuproine Ablates Melanocytes in Adult Zebrafish

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    O'Reilly-Pol, Thomas; Johnson, Stephen L.

    2008-01-01

    The simplest regeneration experiments involve the ablation of a single cell type. While methods exist to ablate the melanocytes of the larval zebrafish,1,2 no convenient method exists to ablate melanocytes in adult zebrafish. Here, we show that the copper chelator neocuproine (NCP) causes fragmentation and disappearance of melanin in adult zebrafish melanocytes. Adult melanocytes expressing eGFP under the control of a melanocyte-specific promoter also lose eGFP fluorescence in the presence of...

  7. ZFOR2, a new opioid receptor-like gene from the teleost zebrafish (Danio rerio).

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    Barrallo, A; González-Sarmiento, R; Alvar, F; Rodríguez, R E

    2000-12-08

    A new opioid receptor-like (ZFOR2) has been cloned and characterized in an anamniote vertebrate, the teleost zebrafish (Danio rerio). ZFOR2 encodes a 384-amino-acid protein with seven potential transmembrane domains, and its predicted amino acid sequence presents an overall 74% degree of identity to mammalian mu opioid receptors. Its inclusion in a dendrogram generated from the alignment of the opioid receptor's protein sequences, confirms its classification as a mu opioid receptor. Divergences in sequence are greater in the regions corresponding to extracellular loops, suggesting possible differences in ligand selectivity with respect to the classical mu opioid receptors. The genomic structure of ZFOR2 is also highly conserved throughout the phylogenetic scale, supporting the origin of opioid receptors early in evolution. Nevertheless, ZFOR2 lacks the fourth exon found in human and rodent mu opioid receptors, that is known to be involved in desensibilization and internalization processes.

  8. Adult zebrafish model for pneumococcal pathogenesis.

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    Saralahti, Anni; Piippo, Hannaleena; Parikka, Mataleena; Henriques-Normark, Birgitta; Rämet, Mika; Rounioja, Samuli

    2014-02-01

    Streptococcus pneumoniae (pneumococcus) is a leading cause of community acquired pneumonia, septicemia, and meningitis. Due to incomplete understanding of the host and bacterial factors contributing to these diseases optimal treatment and prevention methods are lacking. In the present study we examined whether the adult zebrafish (Danio rerio) can be used to investigate the pathophysiology of pneumococcal diseases. Here we show that both intraperitoneal and intramuscular injections of the pneumococcal strain TIGR4 cause a fulminant, dose-dependent infection in adult zebrafish, while isogenic mutant bacteria lacking the polysaccharide capsule, autolysin, or pneumolysin are attenuated in the model. Infection through the intraperitoneal route is characterized by rapid expansion of pneumococci in the bloodstream, followed by penetration of the blood-brain barrier and progression to meningitis. Using Rag1 mutant zebrafish, which are devoid of somatic recombination and thus lack adaptive immune responses, we show that clearance of pneumococci in adult zebrafish depends mainly on innate immune responses. In conclusion, this study provides evidence that the adult zebrafish can be used as a model for a pneumococcal infection, and that it can be used to study both host and bacterial factors involved in the pathogenesis. However, our results do not support the use of the zebrafish in studies on the role of adaptive immunity in pneumococcal disease or in the development of new pneumococcal vaccines.

  9. Dissection of the adult zebrafish kidney.

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    Gerlach, Gary F; Schrader, Lauran N; Wingert, Rebecca A

    2011-08-29

    Researchers working in the burgeoning field of adult stem cell biology seek to understand the signals that regulate the behavior and function of stem cells during normal homeostasis and disease states. The understanding of adult stem cells has broad reaching implications for the future of regenerative medicine. For example, better knowledge about adult stem cell biology can facilitate the design of therapeutic strategies in which organs are triggered to heal themselves or even the creation of methods for growing organs in vitro that can be transplanted into humans. The zebrafish has become a powerful animal model for the study of vertebrate cell biology. There has been extensive documentation and analysis of embryonic development in the zebrafish. Only recently have scientists sought to document adult anatomy and surgical dissection techniques, as there has been a progressive movement within the zebrafish community to broaden the applications of this research organism to adult studies. For example, there are expanding interests in using zebrafish to investigate the biology of adult stem cell populations and make sophisticated adult models of diseases such as cancer. Historically, isolation of the zebrafish adult kidney has been instrumental for studying hematopoiesis, as the kidney is the anatomical location of blood cell production in fish. The kidney is composed of nephron functional units found in arborized arrangements, surrounded by hematopoietic tissue that is dispersed throughout the intervening spaces. The hematopoietic component consists of hematopoietic stem cells (HSCs) and their progeny that inhabit the kidney until they terminally differentiate. In addition, it is now appreciated that a group of renal stem/progenitor cells (RPCs) also inhabit the zebrafish kidney organ and enable both kidney regeneration and growth, as observed in other fish species. In light of this new discovery, the zebrafish kidney is one organ that houses the location of two

  10. Microanatomy of adult zebrafish extraocular muscles.

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    Daniel S Kasprick

    Full Text Available Binocular vision requires intricate control of eye movement to align overlapping visual fields for fusion in the visual cortex, and each eye is controlled by 6 extraocular muscles (EOMs. Disorders of EOMs are an important cause of symptomatic vision loss. Importantly, EOMs represent specialized skeletal muscles with distinct gene expression profile and susceptibility to neuromuscular disorders. We aim to investigate and describe the anatomy of adult zebrafish extraocular muscles (EOMs to enable comparison with human EOM anatomy and facilitate the use of zebrafish as a model for EOM research. Using differential interference contrast (DIC, epifluorescence microscopy, and precise sectioning techniques, we evaluate the anatomy of zebrafish EOM origin, muscle course, and insertion on the eye. Immunofluorescence is used to identify components of tendons, basement membrane and neuromuscular junctions (NMJs, and to analyze myofiber characteristics. We find that adult zebrafish EOM insertions on the globe parallel the organization of human EOMs, including the close proximity of specific EOM insertions to one another. However, analysis of EOM origins reveals important differences between human and zebrafish, such as the common rostral origin of both oblique muscles and the caudal origin of the lateral rectus muscles. Thrombospondin 4 marks the EOM tendons in regions that are highly innervated, and laminin marks the basement membrane, enabling evaluation of myofiber size and distribution. The NMJs appear to include both en plaque and en grappe synapses, while NMJ density is much higher in EOMs than in somatic muscles. In conclusion, zebrafish and human EOM anatomy are generally homologous, supporting the use of zebrafish for studying EOM biology. However, anatomic differences exist, revealing divergent evolutionary pressures.

  11. Hypoxia-induced retinopathy model in adult zebrafish

    DEFF Research Database (Denmark)

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

    2010-01-01

    . In this article, we describe protocols that create hypoxia-induced retinopathy in adult zebrafish. Adult fli1: EGFP zebrafish are placed in hypoxic water for 3-10 d and retinal neovascularization is analyzed using confocal microscopy. It usually takes 11 d to obtain conclusive results using the hypoxia...

  12. How to make a teleost adenohypophysis: molecular pathways of pituitary development in zebrafish.

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    Pogoda, Hans-Martin; Hammerschmidt, Matthias

    2009-11-27

    The anterior pituitary gland, or adenohypophysis (AH), represents the key component of the vertebrate hypothalamo-hypophyseal axis, where it functions at the interphase of the nervous and endocrine system to regulate basic body functions like growth, metabolism and reproduction. For developmental biologists, the adenohypophysis serves as an excellent model system for the studies of organogenesis and differential cell fate specification. Previous research, mainly done in mouse, identified numerous extrinsic signaling cues and intrinsic transcription factors that orchestrate the gland's developmental progression. In the past years, the zebrafish has emerged as a powerful tool to elucidate the genetic networks controlling vertebrate development, behavior and disease. Based on mutants isolated in forward genetic screens and on gene knock-downs using morpholino oligonucleotide (oligo) antisense technology, our current understanding of the molecular machinery driving adenohypophyseal ontogeny could be considerably improved. In addition, comparative analyses have shed further light onto the evolution of this rather recently invented organ. The goal of this review is to summarize current knowledge of the genetic and molecular control of zebrafish pituitary development, with special focus on most recent findings, including some thus far unpublished data from our own laboratory on the transcription factor Six1. In addition, zebrafish data will be discussed in comparison with current understanding of adenohypophysis development in mouse.

  13. Efficient expression of transgenes in adult zebrafish by electroporation

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    Rao S Hari

    2005-10-01

    Full Text Available Abstract Background Expression of transgenes in muscle by injection of naked DNA is widely practiced. Application of electrical pulses at the site of injection was demonstrated to improve transgene expression in muscle tissue. Zebrafish is a precious model to investigate developmental biology in vertebrates. In this study we investigated the effect of electroporation on expression of transgenes in 3–6 month old adult zebrafish. Results Electroporation parameters such as number of pulses, voltage and amount of plasmid DNA were optimized and it was found that 6 pulses of 40 V·cm-1 at 15 μg of plasmid DNA per fish increased the luciferase expression 10-fold compared to controls. Similar enhancement in transgene expression was also observed in Indian carp (Labeo rohita. To establish the utility of adult zebrafish as a system for transient transfections, the strength of the promoters was compared in A2 cells and adult zebrafish after electroporation. The relative strengths of the promoters were found to be similar in cell lines and in adult zebrafish. GFP fluorescence in tissues after electroporation was also studied by fluorescence microscopy. Conclusion Electroporation after DNA injection enhances gene expression 10-fold in adult zebrafish. Electroporation parameters for optimum transfection of adult zebrafish with tweezer type electrode were presented. Enhanced reporter gene expression upon electroporation allowed comparison of strengths of the promoters in vivo in zebrafish.

  14. Positive selection pressure within teleost Toll-like receptors tlr21 and tlr22 subfamilies and their response to temperature stress and microbial components in zebrafish.

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    Sundaram, Arvind Y M; Consuegra, Sonia; Kiron, Viswanath; Fernandes, Jorge M O

    2012-09-01

    Toll-like receptors (TLRs) play a crucial role in host defence, since they trigger immune response following recognition of pathogen-associated molecular patterns (PAMPs) in potential infectious agents. TLRs have been found in numerous organisms, including mammals, birds and teleosts. Some TLR members are commonly retained across all species, whilst others were lost, gained or diverged independently during evolution. Our knowledge about the evolution and specific functions of tlr21, tlr22 and tlr23 in teleosts are still scarce. Phylogenetic analysis of 18 tlr13, tlr21, tlr22 and tlr23 genes from 9 different fish species divided them in two groups. All tlr21 genes were under the first clade, while the second comprised tlr22, tlr23 and tlr13 from Atlantic salmon. Evidence of positive selection was detected at three sites within the leucine-rich repeat regions of Tlr22, which may influence PAMP recognition. Immunostimulation experiments revealed that expression of zebrafish tlr22 is modulated by several unrelated PAMPs. Up to a 3-fold increase in tlr21 and tlr22 expression was detected in larvae exposed to immunostimulants such as lipopolysaccharide, peptidoglycan or poly I:C. We found that zebrafish tlrs are expressed mainly in immune-related organs, such as spleen and kidney as well as in testis and temperature stress did not have an effect on the expression of tlr21 and tlr22 in the early stages of development in zebrafish larvae. Our data indicates that these teleost tlrs may play a role in innate host defence. In particular, tlr22 is evolving under positive selection, which indicates functional diversification and adaptation of the response to different PAMPs.

  15. Molecular characterization of retinal stem cells and their niches in adult zebrafish

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    Barthel Linda K

    2006-07-01

    Full Text Available Abstract Background The persistence in adult teleost fish of retinal stem cells that exhibit all of the features of true 'adult stem cells' – self-renewal, multipotency, and the capacity to respond to injury by mitotic activation with the ability to regenerate differentiated tissues – has been known for several decades. However, the specialized cellular and molecular characteristics of these adult retinal stem cells and the microenvironmental niches that support their maintenance in the differentiated retina and regulate their activity during growth and regeneration have not yet been elucidated. Results Our data show that the zebrafish retina has two kinds of specialized niches that sustain retinal stem cells: 1 a neuroepithelial germinal zone at the interface between neural retina and ciliary epithelium, called the ciliary marginal zone (CMZ, a continuous annulus around the retinal circumference, and 2 the microenvironment around some Müller glia in the differentiated retina. In the uninjured retina, scattered Müller glia (more frequently those in peripheral retina are associated with clusters of proliferating retinal progenitors that are restricted to the rod photoreceptor lineage, but following injury, the Müller-associated retinal progenitors can function as multipotent retinal stem cells to regenerate other types of retinal neurons. The CMZ has several features in common with the neurogenic niches in the adult mammalian brain, including access to the apical epithelial surface and a close association with blood vessels. Müller glia in the teleost retina have a complex response to local injury that includes some features of reactive gliosis (up-regulation of glial fibrillary acidic protein, GFAP, and re-entry into the cell cycle together with dedifferentiation and re-acquisition of phenotypic and molecular characteristics of multipotent retinal progenitors in the CMZ (diffuse distribution of N-cadherin, activation of Notch

  16. 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.

  17. Whole-body and multispectral photoacoustic imaging of adult zebrafish

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    Huang, Na; Xi, Lei

    2016-10-01

    Zebrafish is a top vertebrate model to study developmental biology and genetics, and it is becoming increasingly popular for studying human diseases due to its high genome similarity to that of humans and the optical transparency in embryonic stages. However, it becomes difficult for pure optical imaging techniques to volumetric visualize the internal organs and structures of wild-type zebrafish in juvenile and adult stages with excellent resolution and penetration depth. Even with the establishment of mutant lines which remain transparent over the life cycle, it is still a challenge for pure optical imaging modalities to image the whole body of adult zebrafish with micro-scale resolution. However, the method called photoacoustic imaging that combines all the advantages of the optical imaging and ultrasonic imaging provides a new way to image the whole body of the zebrafish. In this work, we developed a non-invasive photoacoustic imaging system with optimized near-infrared illumination and cylindrical scanning to image the zebrafish. The lateral and axial resolution yield to 80 μm and 600 μm, respectively. Multispectral strategy with wavelengths from 690 nm to 930 nm was employed to image various organs inside the zebrafish. From the reconstructed images, most major organs and structures inside the body can be precisely imaged. Quantitative and statistical analysis of absorption for organs under illumination with different wavelengths were carried out.

  18. Spatial distribution and cellular composition of adult brain proliferative zones in the teleost, Gymnotus omarorum

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    Valentina eOlivera-Pasilio

    2014-09-01

    Full Text Available Proliferation of stem/progenitor cells during development provides for the generation of mature cell types in the CNS. While adult brain proliferation is highly restricted in the mammals, it is widespread in teleosts. The extent of adult neural proliferation in the weakly electric fish, Gymnotus omarorum has not yet been described. To address this, we used double thymidine analog pulse-chase labeling of proliferating cells to identify brain proliferation zones, characterize their cellular composition, and analyze the fate of newborn cells in adult G. omarorum. Short thymidine analog chase periods revealed the ubiquitous distribution of adult brain proliferation, similar to other teleosts, particularly Apteronotus leptorhynchus. Proliferating cells were abundant at the ventricular-subventricular lining of the ventricular-cisternal system, adjacent to the telencephalic subpallium, the diencephalic preoptic region and hypothalamus, and the mesencephalic tectum opticum and torus semicircularis. Extraventricular proliferation zones, located distant from the ventricular-cisternal system surface, were found in all divisions of the rombencephalic cerebellum. We also report a new adult proliferation zone at the caudal-lateral border of the electrosensory lateral line lobe. All proliferation zones showed a heterogeneous cellular composition. The use of short (24hs and long (30d chase periods revealed abundant fast cycling cells (potentially intermediate amplifiers, sparse slow cycling (potentially stem cells, cells that appear to have entered a quiescent state, and cells that might correspond to migrating newborn neural cells. Their abundance and migration distance differed among proliferation zones: greater numbers and longer range and/or pace of migrating cells were associated with subpallial and cerebellar proliferation zones.

  19. Pharmacological Modulation of Hemodynamics in Adult Zebrafish In Vivo.

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    Daniel Brönnimann

    Full Text Available Hemodynamic parameters in zebrafish receive increasing attention because of their important role in cardiovascular processes such as atherosclerosis, hematopoiesis, sprouting and intussusceptive angiogenesis. To study underlying mechanisms, the precise modulation of parameters like blood flow velocity or shear stress is centrally important. Questions related to blood flow have been addressed in the past in either embryonic or ex vivo-zebrafish models but little information is available for adult animals. Here we describe a pharmacological approach to modulate cardiac and hemodynamic parameters in adult zebrafish in vivo.Adult zebrafish were paralyzed and orally perfused with salt water. The drugs isoprenaline and sodium nitroprusside were directly applied with the perfusate, thus closely resembling the preferred method for drug delivery in zebrafish, namely within the water. Drug effects on the heart and on blood flow in the submental vein were studied using electrocardiograms, in vivo-microscopy and mathematical flow simulations.Under control conditions, heart rate, blood flow velocity and shear stress varied less than ± 5%. Maximal chronotropic effects of isoprenaline were achieved at a concentration of 50 μmol/L, where it increased the heart rate by 22.6 ± 1.3% (n = 4; p < 0.0001. Blood flow velocity and shear stress in the submental vein were not significantly increased. Sodium nitroprusside at 1 mmol/L did not alter the heart rate but increased blood flow velocity by 110.46 ± 19.64% (p = 0.01 and shear stress by 117.96 ± 23.65% (n = 9; p = 0.03.In this study, we demonstrate that cardiac and hemodynamic parameters in adult zebrafish can be efficiently modulated by isoprenaline and sodium nitroprusside. Together with the suitability of the zebrafish for in vivo-microscopy and genetic modifications, the methodology described permits studying biological processes that are dependent on hemodynamic alterations.

  20. A Surgery Protocol for Adult Zebrafish Spinal Cord Injury

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    Ping Fang; Jin-Fei Lin; Hong-Chao Pan; Yan-Qin Shen; Melitta Schachner

    2012-01-01

    Adult zebrafish has a remarkable capability to recover from spinal cord injury,providing an excellent model for studying neuroregeneration.Here we list equipment and reagents,and give a detailed protocol for complete transection of the adult zebrafish spinal cord.In this protocol,potential problems and their solutions are described so that the zebrafish spinal cord injury model can be more easily and reproducibly performed.In addition,two assessments are introduced to monitor the success of the surgery and functional recovery:one test to assess free swimming capability and the other test to assess extent of neuroregeneration by in vivo anterograde axonal tracing.In the swimming behavior test,successful complete spinal cord transection is monitored by the inability of zebrafish to swim freely for 1 week after spinal cord injury,followed by the gradual reacquisition of full locomotor ability within 6 weeks after injury.As a morphometric correlate,anterograde axonal tracing allows the investigator to monitor the ability of regenerated axons to cross the lesion site and increasingly extend into the gray and white matter with time after injury,confirming functional recovery.This zebrafish model provides a paradigm for recovery from spinal cord injury,enabling the identification of pathways and components of neuroregeneration.

  1. Adrenocortical and adrenomedullary homologs in eight species of adult and developing teleosts: morphology, histology, and immunohistochemistry.

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    Grassi Milano, E; Basari, F; Chimenti, C

    1997-12-01

    Morphology, histology, and immunohistochemistry of the adrenocortical and adrenomedullary homologs (adrenal glands) of the following developing and adult teleosts were examined: Salmoniformes-Oncorhynchus mykiss (rainbow trout), Salmo trutta fario (brown trout), Coregonus lavaretus (white fish); Cyprinodontiformes-Gambusia affinis (mosquito fish). Perciformes-Dicentrarchus labrax (sea bass), Sparus aurata (sea bream), Diplodus sargus (white bream), Oblada melanura (saddled bream). The anatomical relationships of the gland with the renal system and venous vessels were also noted. In adults of all species steroidogenic and catecholaminergic chromaffin cells were found in the head kidney, which is pronephric in origin and subsequently transformed into a hematopoietic lymphatic organ. In Perciformes, chromaffin cells are distributed around the anterior and posterior cardinal veins and ducts of Cuvier; in Salmoniformes, around the posterior cardinal veins and in the hematopoietic tissue; and in G. affinis, around the ducts of Cuvier and posterior cardinal veins, while a few are visible also around the sinus venosus. In Perciformes and Salmoniformes, numerous chromaffin cells are also present in the posterior kidney, derived from the opisthonephros, in contact with the caudal vein. Steroidogenic cells are always confined to the head kidney. During development chromaffin and steroidogenic cells appear early after hatching in the pronephric kidney, at the level of the ducts of Cuvier and of the cephalic part of the posterior cardinal veins. Later, chromaffin cells in Perciformes reach the anterior cardinal veins, and subsequently, in both Perciformes and Salmoniformes, they reach the developing posterior kidney. Their localization along the posterior kidney is still in progress about 4 months after hatching and is completed about a year after hatching. These findings support the concept that the structure of the adrenal gland in teleosts is intermediate between that of the

  2. In vivo electroporation of morpholinos into the regenerating adult zebrafish tail fin.

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    Hyde, David R; Godwin, Alan R; Thummel, Ryan

    2012-03-29

    Certain species of urodeles and teleost fish can regenerate their tissues. Zebrafish have become a widely used model to study the spontaneous regeneration of adult tissues, such as the heart, retina, spinal cord, optic nerve, sensory hair cells, and fins. The zebrafish fin is a relatively simple appendage that is easily manipulated to study multiple stages in epimorphic regeneration. Classically, fin regeneration was characterized by three distinct stages: wound healing, blastema formation, and fin outgrowth. After amputating part of the fin, the surrounding epithelium proliferates and migrates over the wound. At 33 °C, this process occurs within six hours post-amputation (hpa, Figure 1B). Next, underlying cells from different lineages (ex. bone, blood, glia, fibroblast) re-enter the cell cycle to form a proliferative blastema, while the overlying epidermis continues to proliferate (Figure 1D). Outgrowth occurs as cells proximal to the blastema re-differentiate into their respective lineages to form new tissue (Figure 1E). Depending on the level of the amputation, full regeneration is completed in a week to a month. The expression of a large number of gene families, including wnt, hox, fgf, msx, retinoic acid, shh, notch, bmp, and activin-betaA genes, is up-regulated during specific stages of fin regeneration. However, the roles of these genes and their encoded proteins during regeneration have been difficult to assess, unless a specific inhibitor for the protein exists, a temperature-sensitive mutant exists or a transgenic animal (either overexpressing the wild-type protein or a dominant-negative protein) was generated. We developed a reverse genetic technique to quickly and easily test the function of any gene during fin regeneration. Morpholino oligonucleotides are widely used to study loss of specific proteins during zebrafish, Xenopus, chick, and mouse development. Morpholinos basepair with a complementary RNA sequence to either block pre-mRNA splicing or m

  3. Modeling GATAD1-Associated Dilated Cardiomyopathy in Adult Zebrafish

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    Jingchun Yang

    2016-01-01

    Full Text Available Animal models have played a critical role in validating human dilated cardiomyopathy (DCM genes, particularly those that implicate novel mechanisms for heart failure. However, the disease phenotype may be delayed due to age-dependent penetrance. For this reason, we generated an adult zebrafish model, which is a simpler vertebrate model with higher throughput than rodents. Specifically, we studied the zebrafish homologue of GATAD1, a recently identified gene for adult-onset autosomal recessive DCM. We showed cardiac expression of gatad1 transcripts, by whole mount in situ hybridization in zebrafish embryos, and demonstrated nuclear and sarcomeric I-band subcellular localization of Gatad1 protein in cardiomyocytes, by injecting a Tol2 plasmid encoding fluorescently-tagged Gatad1. We next generated gatad1 knock-out fish lines by TALEN technology and a transgenic fish line that expresses the human DCM GATAD1-S102P mutation in cardiomyocytes. Under stress conditions, longitudinal studies uncovered heart failure (HF-like phenotypes in stable KO mutants and a tendency toward HF phenotypes in transgenic lines. Based on these efforts of studying a gene-based inherited cardiomyopathy model, we discuss the strengths and bottlenecks of adult zebrafish as a new vertebrate model for assessing candidate cardiomyopathy genes.

  4. A genomic view of the NOD-like receptor family in teleost fish: Identification of a novel NLR subfamily in zebrafish

    Science.gov (United States)

    Laing, K.J.; Purcell, M.K.; Winton, J.R.; Hansen, J.D.

    2008-01-01

    Background. A large multigene family of NOD-like receptor (NLR) molecules have been described in mammals and implicated in immunity and apoptosis. Little information, however, exists concerning this gene family in non-mammalian taxa. This current study, therefore, provides an in-depth investigation of this gene family in lower vertebrates including extensive phylogenetic comparison of zebrafish NLRs with orthologs in tetrapods, and analysis of their tissue-specific expression. Results. Three distinct NLR subfamilies were identified by mining genome databases of various non-mammalian vertebrates; the first subfamily (NLR-A) resembles mammalian NODs, the second (NLR-B) resembles mammalian NALPs, while the third (NLR-C) appears to be unique to teleost fish. In zebrafish, NLR-A and NLR-B subfamilies contain five and six genes respectively. The third subfamily is large, containing several hundred NLR-C genes, many of which are predicted to encode a C-terminal B30.2 domain. This subfamily most likely evolved from a NOD3-like molecule. Gene predictions for zebrafish NLRs were verified using sequence derived from ESTs or direct sequencing of cDNA. Reverse-transcriptase (RT)-PCR analysis confirmed expression of representative genes from each subfamily in selected tissues. Conclusion. Our findings confirm the presence of multiple NLR gene orthologs, which form a large multigene family in teleostei. Although the functional significance of the three major NLR subfamilies is unclear, we speculate that conservation and abundance of NLR molecules in all teleostei genomes, reflects an essential role in cellular control, apoptosis or immunity throughout bony fish. ?? 2008 Laing et al; licensee BioMed Central Ltd.

  5. A genomic view of the NOD-like receptor family in teleost fish: identification of a novel NLR subfamily in zebrafish

    Directory of Open Access Journals (Sweden)

    Winton James R

    2008-02-01

    Full Text Available Abstract Background A large multigene family of NOD-like receptor (NLR molecules have been described in mammals and implicated in immunity and apoptosis. Little information, however, exists concerning this gene family in non-mammalian taxa. This current study, therefore, provides an in-depth investigation of this gene family in lower vertebrates including extensive phylogenetic comparison of zebrafish NLRs with orthologs in tetrapods, and analysis of their tissue-specific expression. Results Three distinct NLR subfamilies were identified by mining genome databases of various non-mammalian vertebrates; the first subfamily (NLR-A resembles mammalian NODs, the second (NLR-B resembles mammalian NALPs, while the third (NLR-C appears to be unique to teleost fish. In zebrafish, NLR-A and NLR-B subfamilies contain five and six genes respectively. The third subfamily is large, containing several hundred NLR-C genes, many of which are predicted to encode a C-terminal B30.2 domain. This subfamily most likely evolved from a NOD3-like molecule. Gene predictions for zebrafish NLRs were verified using sequence derived from ESTs or direct sequencing of cDNA. Reverse-transcriptase (RT-PCR analysis confirmed expression of representative genes from each subfamily in selected tissues. Conclusion Our findings confirm the presence of multiple NLR gene orthologs, which form a large multigene family in teleostei. Although the functional significance of the three major NLR subfamilies is unclear, we speculate that conservation and abundance of NLR molecules in all teleostei genomes, reflects an essential role in cellular control, apoptosis or immunity throughout bony fish.

  6. Distribution of glial cell line-derived neurotrophic factor receptor alpha-1 in the brain of adult zebrafish.

    Science.gov (United States)

    Lucini, Carla; Carla, Lucini; Facello, Bruna; Bruna, Facello; Maruccio, Lucianna; Lucianna, Maruccio; Langellotto, Fernanda; Fernanda, Langellotto; Sordino, Paolo; Paolo, Sordino; Castaldo, Luciana; Luciana, Castaldo

    2010-08-01

    Glial cell line-derived neurotrophic factor (GDNF) is a potent trophic factor for several types of neurons in the central and peripheral nervous systems. The biological activity of GDNF is mediated by a multicomponent receptor complex that includes a common transmembrane signaling component (the rearranged during transfection (RET) proto-oncogene product, a tyrosine kinase receptor) as well as a GDNF family receptor alpha (GFRalpha) subunit, a high-affinity glycosyl phosphatidylinositol (GPI)-linked binding element. Among the four known GFRalpha subunits, GFRalpha1 preferentially binds to GDNF. In zebrafish (Danio rerio) embryos, the expression of the GFRalpha1a and GFRalpha1b genes has been shown in primary motor neurons, the kidney, and the enteric nervous system. To examine the activity of GFRalpha in the adult brain of a lower vertebrate, we have investigated the localization of GFRalpha1a and GFRalpha1b mRNA and the GFRalpha1 protein in zebrafish. GFRalpha1a and GFRalpha1b transcripts were observed in brain extracts by reverse transcription-polymerase chain reaction. Whole-mount in-situ hybridization experiments revealed a wide distribution of GFRalpha1a and GFRalpha1b mRNAs in various regions of the adult zebrafish brain. These included the olfactory bulbs, dorsal and ventral telencephalic area (telencephalon), preoptic area, dorsal and ventral thalamus, posterior tuberculum and hypothalamus (diencephalon), optic tectum (mesencephalon), cerebellum, and medulla oblongata (rhombencephalon). Finally, expression patterns of the GFRalpha1 protein, detected immunohistochemically, correlated well with the mRNA expression and provided further insights into translational activity at the neuroanatomical level. In conclusion, the current study demonstrated that the presence of GFRalpha1 persists beyond the embryonic development of the zebrafish brain and, together with the GDNF ligand, is probably implicated in the brain physiology of an adult teleost fish.

  7. TRPV4 in the sensory organs of adult zebrafish.

    Science.gov (United States)

    Amato, V; Viña, E; Calavia, M G; Guerrera, M C; Laurà, R; Navarro, M; De Carlos, F; Cobo, J; Germanà, A; Vega, J A

    2012-01-01

    TRPV4 is a nonselective cation channel that belongs to the vanilloid (V) subfamily of transient receptor potential (TRP) ion channels. While TRP channels have been found to be involved in sensing temperature, light, pressure, and chemical stimuli, TPRV4 is believed to be primarily a mechanosensor although it can also respond to warm temperatures, acidic pH, and several chemical compounds. In zebrafish, the expression of trpv4 has been studied during embryonic development, whereas its pattern of TPRV4 expression during the adult life has not been thoroughly analyzed. In this study, the occurrence of TRPV4 was addressed in the zebrafish sensory organs at the mRNA (RT-PCR) and protein (Westernblot) levels. Once the occurrence of TRPV4 was demonstrated, the TRPV4 positive cells were identified by using immunohistochemistry. TPRV4 was detected in mantle and sensory cells of neuromasts, in a subpopulation of hair sensory cells in the macula and in the cristae ampullaris of the inner ear, in sensory cells in the taste buds, in crypt neurons and ciliated sensory neurons of the olfactory epithelium, and in cells of the retina. These results demonstrate the presence of TRPV4 in all sensory organs of adult zebrafish and are consistent with the multiple physiological functions suspected for TRPV4 in mammals (mechanosensation, hearing, and temperature sensing), but furthermore suggest potential roles in olfaction and vision in zebrafish.

  8. Whole-body multispectral photoacoustic imaging of adult zebrafish

    Science.gov (United States)

    Huang, Na; Guo, Heng; Qi, Weizhi; Zhang, Zhiwei; Rong, Jian; Yuan, Zhen; Ge, Wei; Jiang, Huabei; Xi, Lei

    2016-01-01

    The zebrafish, an ideal vertebrate for studying developmental biology and genetics, is increasingly being used to understand human diseases, due to its high similarity to the human genome and its optical transparency during embryonic stages. Once the zebrafish has fully developed, especially wild-type breeds, conventional optical imaging techniques have difficulty in imaging the internal organs and structures with sufficient resolution and penetration depth. Even with established mutant lines that remain transparent throughout their life cycle, it is still challenging for purely optical imaging modalities to visualize the organs of juvenile and adult zebrafish at a micro-scale spatial resolution. In this work, we developed a non-invasive three-dimensional photoacoustic imaging platform with an optimized illumination pattern and a cylindrical-scanning-based data collection system to image entire zebrafish with micro-scale resolutions of 80 μm and 600 μm in the lateral and axial directions, respectively. In addition, we employed a multispectral strategy that utilized excitation wavelengths from 690 nm to 930 nm to statistically quantify the relative optical absorption spectrum of major organs. PMID:27699119

  9. Fgf regulates dedifferentiation during skeletal muscle regeneration in adult zebrafish.

    Science.gov (United States)

    Saera-Vila, Alfonso; Kish, Phillip E; Kahana, Alon

    2016-09-01

    Fibroblast growth factors (Fgfs) regulate critical biological processes such as embryonic development, tissue homeostasis, wound healing, and tissue regeneration. In zebrafish, Fgf signaling plays an important role in the regeneration of the spinal cord, liver, heart, fin, and photoreceptors, although its exact mechanism of action is not fully understood. Utilizing an adult zebrafish extraocular muscle (EOM) regeneration model, we demonstrate that blocking Fgf receptor function using either a chemical inhibitor (SU5402) or a dominant-negative transgenic construct (dnFGFR1a:EGFP) impairs muscle regeneration. Adult zebrafish EOMs regenerate through a myocyte dedifferentiation process, which involves a muscle-to-mesenchyme transition and cell cycle reentry by differentiated myocytes. Blocking Fgf signaling reduced cell proliferation and active caspase 3 levels in the regenerating muscle with no detectable levels of apoptosis, supporting the hypothesis that Fgf signaling is involved in the early steps of dedifferentiation. Fgf signaling in regenerating myocytes involves the MAPK/ERK pathway: inhibition of MEK activity with U0126 mimicked the phenotype of the Fgf receptor inhibition on both muscle regeneration and cell proliferation, and activated ERK (p-ERK) was detected in injured muscles by immunofluorescence and western blot. Interestingly, following injury, ERK2 expression is specifically induced and activated by phosphorylation, suggesting a key role in muscle regeneration. We conclude that the critical early steps of myocyte dedifferentiation in EOM regeneration are dependent on Fgf signaling.

  10. Impaired constitutive and regenerative neurogenesis in adult hyperglycemic zebrafish.

    Science.gov (United States)

    Dorsemans, Anne-Claire; Soulé, Stéphanie; Weger, Meltem; Bourdon, Emmanuel; Lefebvre d'Hellencourt, Christian; Meilhac, Olivier; Diotel, Nicolas

    2017-02-15

    A growing body of evidence supports hyperglycemia as a putative contributor to several brain dysfunctions observed in diabetes patients, such as impaired memory capacity, neural plasticity, and neurogenic processes. Thanks to the persistence of radial glial cells acting as neural stem cells, the brain of the adult zebrafish constitutes a relevant model to investigate constitutive and injury-induced neurogenesis in adult vertebrates. However, there is limited understanding of the impact of hyperglycemia on brain dysfunction in the zebrafish model. This work aimed at exploring the impact of acute and chronic hyperglycemia on brain homeostasis and neurogenesis. Acute hyperglycemia was shown to promote gene expression of proinflammatory cytokines (il1β, il6, il8, and tnfα) in the brain and chronic hyperglycemia to impair expression of genes involved in the establishment of the blood-brain barrier (claudin 5a, zona occludens 1a and b). Chronic hyperglycemia also decreased brain cell proliferation in most neurogenic niches throughout the forebrain and the midbrain. By using a stab wound telencephalic injury model, the impact of hyperglycemia on brain repair mechanisms was investigated. Whereas the initial step of parenchymal cell proliferation was not affected by acute hyperglycemia, later proliferation of neural progenitors was significantly decreased by chronic hyperglycemia in the injured brain of fish. Taken together, these data offer new evidence highlighting the evolutionary conserved adverse effects of hyperglycemia on neurogenesis and brain healing in zebrafish. In addition, our study reinforces the utility of zebrafish as a robust model for studying the effects of metabolic disorders on the central nervous system. J. Comp. Neurol. 525:442-458, 2017. © 2016 Wiley Periodicals, Inc.

  11. In vivo high field magnetic resonance imaging and spectroscopy of adult zebrafish

    NARCIS (Netherlands)

    Kabli, Samira

    2009-01-01

    This thesis contains the results of imaging of adult zebrafish by using different MR approaches. We present the first high resolution mMR images of adult zebrafish. To achieve high spatial resolution we used a magnetic field of 9.4T, in combination with strong magnetic field gradients (1000 mT/m) an

  12. Effects of piracetam on behavior and memory in adult zebrafish.

    Science.gov (United States)

    Grossman, Leah; Stewart, Adam; Gaikwad, Siddharth; Utterback, Eli; Wu, Nadine; Dileo, John; Frank, Kevin; Hart, Peter; Howard, Harry; Kalueff, Allan V

    2011-04-25

    Piracetam, a derivative of γ-aminobutyric acid, exerts memory-enhancing and mild anxiolytic effects in human and rodent studies. To examine the drug's behavioral profile further, we assessed its effects on behavioral and endocrine (cortisol) responses of adult zebrafish (Danio rerio)--a novel model species rapidly gaining popularity in neurobehavioral research. Overall, acute piracetam did not affect zebrafish novel tank and light-dark box behavior at mild doses (25-400mg/L), but produced nonspecific behavioral inhibition at 700mg/L. No effects on cortisol levels or inter-/intra-session habituation in the novel tank test were observed for acute or chronic mild non-sedative dose of 200mg/L. In contrast, fish exposed to chronic piracetam at this dose performed significantly better in the cued learning plus-maze test. This observation parallels clinical and rodent literature on the behavioral profile of piracetam, supporting the utility of zebrafish paradigms for testing nootropic agents.

  13. Early dioxin exposure causes toxic effects in adult zebrafish.

    Science.gov (United States)

    Baker, Tracie R; Peterson, Richard E; Heideman, Warren

    2013-09-01

    The acute effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure have been well documented in many vertebrate species. However, less is known about the consequences in adulthood from sublethal exposure during development. To address this, we exposed zebrafish to sublethal levels of TCDD (1h; 50 pg/ml), either in early embryogenesis (day 0) or during sexual determination (3 and 7 weeks), and assessed the effects later in adulthood. We found that exposure during embryogenesis produced few effects on the adults themselves but did affect the offspring of these fish: Malformations and increased mortality were observed in the subsequent generation. Zebrafish exposed during sexual development showed defects in the cranial and axial skeleton as adults. This was most clearly manifested as scoliosis caused by malformation of individual vertebrae. These fish also showed defects in reproduction, producing fewer eggs with lower fertilization success. Both males and females were affected, with males contributing to the decrease in egg release from the females and exposed females contributing to fertilization failure. TCDD exposure at 3 and 7 weeks produced feminization of the population. Surprisingly, part of this was due to the appearance of fish with clearly female bodies, yet carrying testes in place of ovaries. Our results show that exposures that produce little if any impact during development can cause severe consequences during adulthood and present a model for studying this process.

  14. Satellite-like cells contribute to pax7-dependent skeletal muscle repair in adult zebrafish.

    Science.gov (United States)

    Berberoglu, Michael A; Gallagher, Thomas L; Morrow, Zachary T; Talbot, Jared C; Hromowyk, Kimberly J; Tenente, Inês M; Langenau, David M; Amacher, Sharon L

    2017-04-15

    Satellite cells, also known as muscle stem cells, are responsible for skeletal muscle growth and repair in mammals. Pax7 and Pax3 transcription factors are established satellite cell markers required for muscle development and regeneration, and there is great interest in identifying additional factors that regulate satellite cell proliferation, differentiation, and/or skeletal muscle regeneration. Due to the powerful regenerative capacity of many zebrafish tissues, even in adults, we are exploring the regenerative potential of adult zebrafish skeletal muscle. Here, we show that adult zebrafish skeletal muscle contains cells similar to mammalian satellite cells. Adult zebrafish satellite-like cells have dense heterochromatin, express Pax7 and Pax3, proliferate in response to injury, and show peak myogenic responses 4-5 days post-injury (dpi). Furthermore, using a pax7a-driven GFP reporter, we present evidence implicating satellite-like cells as a possible source of new muscle. In lieu of central nucleation, which distinguishes regenerating myofibers in mammals, we describe several characteristics that robustly identify newly-forming myofibers from surrounding fibers in injured adult zebrafish muscle. These characteristics include partially overlapping expression in satellite-like cells and regenerating myofibers of two RNA-binding proteins Rbfox2 and Rbfoxl1, known to regulate embryonic muscle development and function. Finally, by analyzing pax7a; pax7b double mutant zebrafish, we show that Pax7 is required for adult skeletal muscle repair, as it is in the mouse.

  15. New tools for the identification of developmentally regulated enhancer regions in embryonic and adult zebrafish.

    Science.gov (United States)

    Levesque, Mitchell P; Krauss, Jana; Koehler, Carla; Boden, Cindy; Harris, Matthew P

    2013-03-01

    We have conducted a screen to identify developmentally regulated enhancers that drive tissue-specific Gal4 expression in zebrafish. We obtained 63 stable transgenic lines with expression patterns in embryonic or adult zebrafish. The use of a newly identified minimal promoter from the medaka edar locus resulted in a relatively unbiased set of expression patterns representing many tissue types derived from all germ layers. Subsequent detailed characterization of selected lines showed strong and reproducible Gal4-driven GFP expression in diverse tissues, including neurons from the central and peripheral nervous systems, pigment cells, erythrocytes, and peridermal cells. By screening adults for GFP expression, we also isolated lines expressed in tissues of the adult zebrafish, including scales, fin rays, and joints. The new and efficient minimal promoter and large number of transactivating driver-lines we identified will provide the zebrafish community with a useful resource for further enhancer trap screening, as well as precise investigation of tissue-specific processes in vivo.

  16. Advanced echocardiography in adult zebrafish reveals delayed recovery of heart function after myocardial cryoinjury.

    Directory of Open Access Journals (Sweden)

    Selina J Hein

    Full Text Available Translucent zebrafish larvae represent an established model to analyze genetics of cardiac development and human cardiac disease. More recently adult zebrafish are utilized to evaluate mechanisms of cardiac regeneration and by benefiting from recent genome editing technologies, including TALEN and CRISPR, adult zebrafish are emerging as a valuable in vivo model to evaluate novel disease genes and specifically validate disease causing mutations and their underlying pathomechanisms. However, methods to sensitively and non-invasively assess cardiac morphology and performance in adult zebrafish are still limited. We here present a standardized examination protocol to broadly assess cardiac performance in adult zebrafish by advancing conventional echocardiography with modern speckle-tracking analyses. This allows accurate detection of changes in cardiac performance and further enables highly sensitive assessment of regional myocardial motion and deformation in high spatio-temporal resolution. Combining conventional echocardiography measurements with radial and longitudinal velocity, displacement, strain, strain rate and myocardial wall delay rates after myocardial cryoinjury permitted to non-invasively determine injury dimensions and to longitudinally follow functional recovery during cardiac regeneration. We show that functional recovery of cryoinjured hearts occurs in three distinct phases. Importantly, the regeneration process after cryoinjury extends far beyond the proposed 45 days described for ventricular resection with reconstitution of myocardial performance up to 180 days post-injury (dpi. The imaging modalities evaluated here allow sensitive cardiac phenotyping and contribute to further establish adult zebrafish as valuable cardiac disease model beyond the larval developmental stage.

  17. Husbandry stress exacerbates mycobacterial infections in adult zebrafish, Danio rerio (Hamilton)

    Science.gov (United States)

    Ramsay, J.M.; Watral, V.; Schreck, C.B.; Kent, M.L.

    2009-01-01

    Mycobacteria are significant pathogens of laboratory zebrafish, Danio rerio (Hamilton). Stress is often implicated in clinical disease and morbidity associated with mycobacterial infections but has yet to be examined with zebrafish. The aim of this study was to examine the effects of husbandry stressors on zebrafish infected with mycobacteria. Adult zebrafish were exposed to Mycobacterium marinum or Mycobacterium chelonae, two species that have been associated with disease in zebrafish. Infected fish and controls were then subjected to chronic crowding and handling stressors and examined over an 8-week period. Whole-body cortisol was significantly elevated in stressed fish compared to non-stressed fish. Fish infected with M. marinum ATCC 927 and subjected to husbandry stressors had 14% cumulative mortality while no mortality occurred among infected fish not subjected to husbandry stressors. Stressed fish, infected with M. chelonae H1E2 from zebrafish, were 15-fold more likely to be infected than non-stressed fish at week 8 post-injection. Sub-acute, diffuse infections were more common among stressed fish infected with M. marinum or M. chelonae than non-stressed fish. This is the first study to demonstrate an effect of stress and elevated cortisol on the morbidity, prevalence, clinical disease and histological presentation associated with mycobacterial infections in zebrafish. Minimizing husbandry stress may be effective at reducing the severity of outbreaks of clinical mycobacteriosis in zebrafish facilities. ?? 2009 Blackwell Publishing Ltd.

  18. Sustained action of developmental ethanol exposure on the cortisol response to stress in zebrafish larvae and adults.

    Directory of Open Access Journals (Sweden)

    Matteo Baiamonte

    Full Text Available Ethanol exposure during pregnancy is one of the leading causes of preventable birth defects, leading to a range of symptoms collectively known as fetal alcohol spectrum disorder. More moderate levels of prenatal ethanol exposure lead to a range of behavioural deficits including aggression, poor social interaction, poor cognitive performance and increased likelihood of addiction in later life. Current theories suggest that adaptation in the hypothalamo-pituitary-adrenal (HPA axis and neuroendocrine systems contributes to mood alterations underlying behavioural deficits and vulnerability to addiction. In using zebrafish (Danio rerio, the aim is to determine whether developmental ethanol exposure provokes changes in the hypothalamo-pituitary-interrenal (HPI axis (the teleost equivalent of the HPA, as it does in mammalian models, therefore opening the possibilities of using zebrafish to elucidate the mechanisms involved, and to test novel therapeutics to alleviate deleterious symptoms.The results showed that developmental exposure to ambient ethanol, 20mM-50mM 1-9 days post fertilisation, had immediate effects on the HPI, markedly reducing the cortisol response to air exposure stress, as measured by whole body cortisol content. This effect was sustained in adults 6 months later. Morphology, growth and locomotor activity of the animals were unaffected, suggesting a specific action of ethanol on the HPI. In this respect the data are consistent with mammalian results, although they contrast with the higher corticosteroid stress response reported in rats after developmental ethanol exposure. The mechanisms that underlie the specific sensitivity of the HPI to ethanol require elucidation.

  19. Normal anatomy and histology of the adult zebrafish.

    Science.gov (United States)

    Menke, Aswin L; Spitsbergen, Jan M; Wolterbeek, Andre P M; Woutersen, Ruud A

    2011-08-01

    The zebrafish has been shown to be an excellent vertebrate model for studying the roles of specific genes and signaling pathways. The sequencing of its genome and the relative ease with which gene modifications can be performed have led to the creation of numerous human disease models that can be used for testing the potential and the toxicity of new pharmaceutical compounds. Many pharmaceutical companies already use the zebrafish for prescreening purposes. So far, the focus has been on ecotoxicity and the effects on embryonic development, but there is a trend to expand the use of the zebrafish with acute, subchronic, and chronic toxicity studies that are currently still carried out with the more conventional test animals such as rodents. However, before we can fully realize the potential of the zebrafish as an animal model for understanding human development, disease, and toxicology, we must first greatly advance our knowledge of normal zebrafish physiology, anatomy, and histology. To further this knowledge, we describe, in the present article, location and histology of the major zebrafish organ systems with a brief description of their function.

  20. Rhodopsin expression in the zebrafish pineal gland from larval to adult stage.

    Science.gov (United States)

    Magnoli, Domenico; Zichichi, Rosalia; Laurà, Rosaria; Guerrera, Maria Cristina; Campo, Salvatore; de Carlos, Felix; Suárez, Alberto Álvarez; Abbate, Francesco; Ciriaco, Emilia; Vega, Jose Antonio; Germanà, Antonino

    2012-03-09

    The zebrafish pineal gland plays an important role in different physiological functions including the regulation of the circadian clock. In the fish pineal gland the pinealocytes are made up of different segments: outer segment, inner segment and basal pole. Particularly, in the outer segment the rhodopsin participates in the external environment light reception that represents the first biochemical step in the melatonin production. It is well known that the rhodopsin in the adult zebrafish is well expressed in the pineal gland but both the expression and the cellular localization of this protein during development remain still unclear. In this study using qRT-PCR, sequencing and immunohistochemistry the expression as well as the protein localization of the rhodopsin in the zebrafish from larval (10 dpf) to adult stage (90 dpf) were demonstrated. The rhodopsin mRNA expression presents a peak of expression at 10 dpf, a further reduction to 50 dpf before increasing again in the adult stage. Moreover, the cellular localization of the rhodopsin-like protein was always localized in the pinealocyte at all ages examined. Our results demonstrated the involvement of the rhodopsin in the zebrafish pineal gland physiology particularly in the light capture during the zebrafish lifespan.

  1. Organization of the histaminergic system in adult zebrafish (Danio rerio) brain: neuron number, location, and cotransmitters.

    Science.gov (United States)

    Sundvik, Maria; Panula, Pertti

    2012-12-01

    Histamine is an essential factor in the ascending arousal system (AAS) during motivated behaviors. Histamine and hypocretin/orexin (hcrt) are proposed to be responsible for different aspects of arousal and wakefulness, histamine mainly for cognitive and motivated behaviors. In this study we visualized the entire histaminergic neuron population in adult male and female zebrafish brain and quantified the histaminergic neuron numbers. There were 40-45 histaminergic neurons in both male and female zebrafish brain. Further, we identified cotransmitters of histaminergic neurons in the ventrocaudal hypothalamus, i.e., around the posterior recess (PR) in adult zebrafish. Galanin, γ-aminobutyric acid (GABA), and thyrotropin-releasing hormone (TRH) were colocalized with histamine in some but not all neurons, a result that was verified by intracerebroventricular injections of colchicine into adult zebrafish. Fibers immunoreactive (ir) for galanin, GABA, TRH, or methionine-enkephalin (mENK) were dense in the ventrocaudal hypothalamus around the histaminergic neurons. In histamine-ir fibers TRH and galanin immunoreactivities were also detected in the ventral telencephalon. All these neurotransmitters are involved in maintaining the equilibrium of the sleep-wake state. Our results are in accordance with results from rats, further supporting the use of zebrafish as a tool to study molecular mechanisms underlying complex behaviors.

  2. Reducing the noise in behavioral assays: sex and age in adult zebrafish locomotion.

    Science.gov (United States)

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

    2012-12-01

    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 moved to report the age and sex of all animals used in their studies. Zebrafish work does not yet carry the same mandate. In this study, we evaluated sex and age differences in locomotion behavior. We found that age was a significant factor in locomotion, as was sex within a given age group. In short, as zebrafish age, they appear to show less base level locomotion. With regard to sex, younger (10 months) zebrafish showed more locomotion in males, while older zebrafish (22 months) showed more movement in females. These findings have led us to suggest that those using the zebrafish for behavioral studies control for age and sex within their experimental design and report these descriptors in their methods.

  3. The photoreceptive cells of the pineal gland in adult zebrafish (Danio rerio).

    Science.gov (United States)

    Laurà, Rosaria; Magnoli, Domenico; Zichichi, Rosalia; Guerrera, Maria Cristina; De Carlos, Felix; Suárez, Alberto Álvarez; Abbate, Francesco; Ciriaco, Emilia; Vega, Jose Antonio; Germanà, Antonino

    2012-03-01

    The zebrafish pineal gland plays a fundamental role in the regulation of the circadian rhythm through the melatonin secretion. The pinealocytes, also called photoreceptive cells, are considered the morphofunctional unit of pineal gland. In literature, the anatomical features, the cellular characteristics, and the pinealocytes morphology of zebrafish pineal gland have not been previously described in detail. Therefore, this study was undertaken to analyze the structure and ultrastructure, as well as the immunohistochemical profile of the zebrafish pineal gland with particular reference to the pinealocytes. Here, we demonstrated, using RT-PCR, immunohistochemistry and transmission electron microscopy, the expression of the mRNA for rhodopsin in the pineal gland of zebrafish, as well as its cellular localization exclusively in the pinealocytes of adult zebrafish. Moreover, the ultrastructural observations demonstrated that the pinealocytes were constituted by an outer segment with numerous lamellar membranes, an inner segment with many mitochondria, and a basal pole with the synapses. Our results taken together demonstrated a central role of zebrafish pinealocytes in the control of pineal gland functions.

  4. Identifying proteins in zebrafish embryos using spectral libraries generated from dissected adult organs and tissues.

    Science.gov (United States)

    van der Plas-Duivesteijn, Suzanne J; Mohammed, Yassene; Dalebout, Hans; Meijer, Annemarie; Botermans, Anouk; Hoogendijk, Jordy L; Henneman, Alex A; Deelder, André M; Spaink, Herman P; Palmblad, Magnus

    2014-03-07

    Spectral libraries provide a sensitive and accurate method for identifying peptides from tandem mass spectra, complementary to searching genome-derived databases or sequencing de novo. Their application requires comprehensive libraries including peptides from low-abundant proteins. Here we describe a method for constructing such libraries using biological differentiation to "fractionate" the proteome by harvesting adult organs and tissues and build comprehensive libraries for identifying proteins in zebrafish (Danio rerio) embryos and larvae (an important and widely used model system). Hierarchical clustering using direct comparison of spectra was used to prioritize organ selection. The resulting and publicly available library covers 14,164 proteins, significantly improved the number of peptide-spectrum matches in zebrafish developmental stages, and can be used on data from different instruments and laboratories. The library contains information on tissue and organ expression of these proteins and is also applicable for adult experiments. The approach itself is not limited to zebrafish but would work for any model system.

  5. Retinoic acid signaling controls the formation, proliferation and survival of the blastema during adult zebrafish fin regeneration.

    Science.gov (United States)

    Blum, Nicola; Begemann, Gerrit

    2012-01-01

    Adult teleosts rebuild amputated fins through a proliferation-dependent process called epimorphic regeneration, in which a blastema of cycling progenitor cells replaces the lost fin tissue. The genetic networks that control formation of blastema cells from formerly quiescent stump tissue and subsequent blastema function are still poorly understood. Here, we investigated the cellular and molecular consequences of genetically interfering with retinoic acid (RA) signaling for the formation of the zebrafish blastema. We show that RA signaling is upregulated within the first few hours after fin amputation in the stump mesenchyme, where it controls Fgf, Wnt/β-catenin and Igf signaling. Genetic inhibition of the RA pathway at this stage blocks blastema formation by inhibiting cell cycle entry of stump cells and impairs the formation of the basal epidermal layer, a signaling center in the wound epidermis. In the established blastema, RA signaling remains active to ensure the survival of the highly proliferative blastemal population by controlling expression of the anti-apoptotic factor bcl2. In addition, RA signaling maintains blastema proliferation through the activation of growth-stimulatory signals mediated by Fgf and Wnt/β-catenin signaling, as well as by reducing signaling through the growth-inhibitory non-canonical Wnt pathway. The endogenous roles of RA in adult vertebrate appendage regeneration are uncovered here for the first time. They provide a mechanistic framework to understand previous observations in salamanders that link endogenous sources of RA to the regeneration process itself and support the hypothesis that the RA signaling pathway is an essential component of vertebrate tissue regeneration.

  6. Subdivisions of the adult zebrafish pallium based on molecular marker analysis

    Science.gov (United States)

    Ganz, Julia; Kroehne, Volker; Freudenreich, Dorian; Machate, Anja; Geffarth, Michaela; Braasch, Ingo; Kaslin, Jan; Brand, Michael

    2015-01-01

    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 system as a model

  7. Nonhatching Decapsulated Artemia Cysts As a Replacement to Artemia Nauplii in Juvenile and Adult Zebrafish Culture.

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    Tye, Marc; Rider, Dana; Duffy, Elizabeth A; Seubert, Adam; Lothert, Brogen; Schimmenti, Lisa A

    2015-12-01

    Feeding Artemia nauplii as the main nutrition source for zebrafish is a common practice for many research facilities. Culturing live feed can be time-consuming and requires additional equipment to be purchased, maintained, and cleaned. Nonhatching decapsulated Artemia cysts (decaps) are a commercially available product that can be fed directly to fish. Several other ornamental fish species have been successfully cultured using decaps. Replacing Artemia nauplii with decaps could reduce the overall time and costs associated with the operation of a zebrafish facility. The objective of this study was to determine if decaps could be a suitable replacement to Artemia nauplii in juvenile and adult zebrafish culture. Wild-type zebrafish were fed one of three dietary treatments: decaps only, nauplii only, or a standard consisting of nauplii plus a commercially prepared pellet food. Survival, growth (length and weight), and embryo production were analyzed between the treatments. Fish receiving the decap diet demonstrated a significantly higher growth and embryo production when compared to the fish receiving the nauplii-only diet. When comparing the decap fish to the standard fish, no significant difference was found in mean survival, mean weight at 90 days postfertilization, or mean embryo production. It was determined that nonhatching decapsulated Artemia cysts can be used as a suitable replacement to Artemia nauplii in juvenile and adult zebrafish culture.

  8. Oxazolone-Induced Intestinal Inflammation in Adult Zebrafish

    NARCIS (Netherlands)

    Brugman, Sylvia; Nieuwenhuis, EES

    2017-01-01

    Zebrafish are an excellent model for the study of intestinal immunity. The availability of several transgenic reporter fish for different innate and adaptive immune cells and the high homology in terms of gut function and morphology enables in depth analysis of the process of intestinal inflammation

  9. Oxazolone-induced intestinal inflammation in adult zebrafish

    NARCIS (Netherlands)

    Brugman, Sylvia; Nieuwenhuis, Edward E.S.

    2017-01-01

    Zebrafish are an excellent model for the study of intestinal immunity. The availability of several transgenic reporter fish for different innate and adaptive immune cells and the high homology in terms of gut function and morphology enables in depth analysis of the process of intestinal inflammat

  10. Normal anatomy and histology of the adult zebrafish

    NARCIS (Netherlands)

    Menke, H.; Spitsbergen, J.M.; Wolterbeek, A.P.; Woutersen, R.A.

    2011-01-01

    The zebrafish has been shown to be an excellent vertebrate model for studying the roles of specific genes and signaling pathways. The sequencing of its genome and the relative ease with which gene modifications can be performed have led to the creation of numerous human disease models that can be us

  11. Cell fate determination in zebrafish embryonic and adult muscle development

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    Tee, J.M.

    2010-01-01

    We are interested in how the genetic basis of muscle precursor cells determines the outcome of the muscle cell fate, and thus leading to disruption in muscle formation and maintenance. We utilized the zebrafish carrying mutations in both Axin1 and Apc1, resulting in overactivation of the Wnt/beta-ca

  12. Assessment of locomotion behavior in adult Zebrafish after acute exposure to different pharmacological reference compounds

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    Pankaj Gupta

    2014-01-01

    Full Text Available Objectives: The objective of the present study was to assess locomotor behavior of adult zebrafish after acute exposure to different pharmacological reference compounds. Materials and Methods: Adult zebrafish of 4-5-months-old were exposed to different concentrations of known reference compounds for 15 min. The test was conducted separately for each drug concentration as well as control. Locomotor activity parameters viz. distance travelled, speed, total mobile time, and total immobile time were recorded for each animal during the exposure period. Results: Out of 11 compounds tested, nine compounds showed decrease in locomotor behavior with significant changes in distance travelled, speed, total mobile time, and total immobile time. Caffeine exhibited biphasic response in locomotion behavior, while scopolamine failed to induce any significant changes. Conclusion: In view of the above findings, these results suggested that exposure of adult zebrafish with different known compounds produce the expected changes in the locomotion behavior; therefore, adult zebrafish can be used an alternative approach for the assessment of new chemical entities for their effect on locomotor behavior.

  13. Standardized echocardiographic assessment of cardiac function in normal adult zebrafish and heart disease models

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    Wang, Louis W.; Huttner, Inken G.; Santiago, Celine F.; Kesteven, Scott H.; Yu, Ze-Yan; Feneley, Michael P.

    2017-01-01

    ABSTRACT The zebrafish (Danio rerio) is an increasingly popular model organism in cardiovascular research. Major insights into cardiac developmental processes have been gained by studies of embryonic zebrafish. However, the utility of zebrafish for modeling adult-onset heart disease has been limited by a lack of robust methods for in vivo evaluation of cardiac function. We established a physiological protocol for underwater zebrafish echocardiography using high frequency ultrasound, and evaluated its reliability in detecting altered cardiac function in two disease models. Serial assessment of cardiac function was performed in wild-type zebrafish aged 3 to 12 months and the effects of anesthetic agents, age, sex and background strain were evaluated. There was a varying extent of bradycardia and ventricular contractile impairment with different anesthetic drugs and doses, with tricaine 0.75 mmol l−1 having a relatively more favorable profile. When compared with males, female fish were larger and had more measurement variability. Although age-related increments in ventricular chamber size were greater in females than males, there were no sex differences when data were normalized to body size. Systolic ventricular function was similar in both sexes at all time points, but differences in diastolic function were evident from 6 months onwards. Wild-type fish of both sexes showed a reliance on atrial contraction for ventricular diastolic filling. Echocardiographic evaluation of adult zebrafish with diphtheria toxin-induced myocarditis or anemia-induced volume overload accurately identified ventricular dilation and altered contraction, with suites of B-mode, ventricular strain, pulsed-wave Doppler and tissue Doppler indices showing concordant changes indicative of myocardial hypocontractility or hypercontractility, respectively. Repeatability, intra-observer and inter-observer correlations for echocardiographic measurements were high. We demonstrate that high

  14. Embryonic alcohol exposure impairs associative learning performance in adult zebrafish.

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    Fernandes, Yohaan; Tran, Steven; Abraham, Emil; Gerlai, Robert

    2014-05-15

    The zebrafish has been proposed for modeling fetal alcohol spectrum disorders (FASD). Previous FASD research with zebrafish employed high concentrations of alcohol and/or long exposure periods. Here, we exposed zebrafish eggs to low doses of alcohol (0, 0.25, 0.50, 0.75 and 1.0% (vol/vol); external bath application of which 1/20th may reach the inside of the egg) at 16-h post-fertilization (hpf) and only for a short duration (2h) in the hope to avoid gross morphological aberrations and to mimic the more frequent FASD exposure levels. Upon reaching adulthood the exposed and control zebrafish were tested for their associative learning performance in a plus-maze. Embryonic alcohol exposure led to no gross anatomical abnormalities and did not increase mortality. Unexposed (control) zebrafish showed excellent acquisition of association between a conditioned visual stimulus (CS) and food reward, demonstrated by their preference for the target zone of the maze that contained the CS during a probe trial in the absence of reward. However, alcohol-exposed fish showed no such preference and performed indistinguishable from random chance. Locomotor activity during training and the probe trial or the amount of food consumed during training did not differ between the embryonic alcohol exposed and unexposed (control) fish, suggesting that the impaired learning performance found was unlikely to be caused by altered motivation or motor function. Our results suggest that even very small amounts of alcohol reaching the embryo for only a short duration of time may have long lasting deleterious effects on cognitive function in vertebrates.

  15. Intestinal upregulation of melanin-concentrating hormone in TNBS-induced enterocolitis in adult zebrafish.

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    Brenda M Geiger

    Full Text Available BACKGROUND: Melanin-concentrating hormone (MCH, an evolutionarily conserved appetite-regulating neuropeptide, has been recently implicated in the pathogenesis of inflammatory bowel disease (IBD. Expression of MCH is upregulated in inflamed intestinal mucosa in humans with colitis and MCH-deficient mice treated with trinitrobenzene-sulfonic acid (TNBS develop an attenuated form of colitis compared to wild type animals. Zebrafish have emerged as a new animal model of IBD, although the majority of the reported studies concern zebrafish larvae. Regulation MCH expression in the adult zebrafish intestine remains unknown. METHODS: In the present study we induced enterocolitis in adult zebrafish by intrarectal administration of TNBS. Follow-up included survival analysis, histological assessment of changes in intestinal architecture, and assessment of intestinal infiltration by myeloperoxidase positive cells and cytokine transcript levels. RESULTS: Treatment with TNBS dose-dependently reduced fish survival. This response required the presence of an intact microbiome, since fish pre-treated with vancomycin developed less severe enterocolitis. At 6 hours post-challenge, we detected a significant influx of myeloperoxidase positive cells in the intestine and upregulation of both proinflammatory and anti-inflammatory cytokines. Most importantly, and in analogy to human IBD and TNBS-induced mouse experimental colitis, we found increased intestinal expression of MCH and its receptor in TNBS-treated zebrafish. CONCLUSIONS: Taken together these findings not only establish a model of chemically-induced experimental enterocolitis in adult zebrafish, but point to effects of MCH in intestinal inflammation that are conserved across species.

  16. Morphological and molecular evidence for functional organization along the rostrocaudal axis of the adult zebrafish intestine

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    Lam Siew

    2010-06-01

    Full Text Available Abstract Background The zebrafish intestine is a simple tapered tube that is folded into three sections. However, whether the intestine is functionally similar along its length remains unknown. Thus, a systematic structural and functional characterization of the zebrafish intestine is desirable for future studies of the digestive tract and the intestinal biology and development. Results To characterize the structure and function of the adult zebrafish intestine, we divided the intestine into seven roughly equal-length segments, S1-S7, and systematically examined the morphology of the mucosal lining, histology of the epithelium, and molecular signatures from transcriptome analysis. Prominent morphological features are circumferentially-oriented villar ridges in segments S1-S6 and the absence of crypts. Molecular characterization of the transcriptome from each segment shows that segments S1-S5 are very similar while S6 and S7 unique. Gene ontology analyses reveal that S1-S5 express genes whose functions involve metabolism of carbohydrates, transport of lipids and energy generation, while the last two segments display relatively limited function. Based on comparative Gene Set Enrichment Analysis, the first five segments share strong similarity with human and mouse small intestine while S6 shows similarity with human cecum and rectum, and S7 with human rectum. The intestinal tract does not display the anatomical, morphological, and molecular signatures of a stomach and thus we conclude that this organ is absent from the zebrafish digestive system. Conclusions Our genome-wide gene expression data indicate that, despite the lack of crypts, the rostral, mid, and caudal portions of the zebrafish intestine have distinct functions analogous to the mammalian small and large intestine, respectively. Organization of ridge structures represents a unique feature of zebrafish intestine, though they produce similar cross sections to mammalian intestines

  17. Major vault protein promotes locomotor recovery and regeneration after spinal cord injury in adult zebrafish.

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    Pan, Hong-Chao; Lin, Jin-Fei; Ma, Li-Ping; Shen, Yan-Qin; Schachner, Melitta

    2013-01-01

    In contrast to mammals, adult zebrafish recover locomotor functions after spinal cord injury (SCI), in part due to axonal regrowth and regeneration permissivity of the central nervous system. Upregulation of major vault protein (MVP) expression after spinal cord injury in the brainstem of the adult zebrafish prompted us to probe for its contribution to recovery after SCI. MVP is a multifunctional protein expressed not only in many types of tumours but also in the nervous system, where its importance for regeneration is, however, unclear. Using an established zebrafish SCI model, we found that MVP mRNA and protein expression levels were increased in ependymal cells in the spinal cord caudal to the lesion site at 6 and 11 days after SCI. Double immunolabelling showed that MVP was co-localised with Islet-1 or tyrosine hydroxylase around the central canal of the spinal cord in sham-injured control fish and injured fish 11 days after surgery. MVP co-localised with the neural stem cell marker nestin in ependymal cells after injury. By using an in vivo morpholino-based knock-down approach, we found that the distance moved by MVP morpholino-treated fish was reduced at 4, 5 and 6 weeks after SCI when compared to fish treated with standard control morpholino. Knock-down of MVP resulted in reduced regrowth of axons from brainstem neurons into the spinal cord caudal to the lesion site. These results indicate that MVP supports locomotor recovery and axonal regrowth after SCI in adult zebrafish.

  18. Zebrafish adult-derived hypothalamic neurospheres generate gonadotropin-releasing hormone (GnRH neurons

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    Christian Cortés-Campos

    2015-09-01

    Full Text Available Gonadotropin-releasing hormone (GnRH is a hypothalamic decapeptide essential for fertility in vertebrates. Human male patients lacking GnRH and treated with hormone therapy can remain fertile after cessation of treatment suggesting that new GnRH neurons can be generated during adult life. We used zebrafish to investigate the neurogenic potential of the adult hypothalamus. Previously we have characterized the development of GnRH cells in the zebrafish linking genetic pathways to the differentiation of neuromodulatory and endocrine GnRH cells in specific regions of the brain. Here, we developed a new method to obtain neural progenitors from the adult hypothalamus in vitro. Using this system, we show that neurospheres derived from the adult hypothalamus can be maintained in culture and subsequently differentiate glia and neurons. Importantly, the adult derived progenitors differentiate into neurons containing GnRH and the number of cells is increased through exposure to either testosterone or GnRH, hormones used in therapeutic treatment in humans. Finally, we show in vivo that a neurogenic niche in the hypothalamus contains GnRH positive neurons. Thus, we demonstrated for the first time that neurospheres can be derived from the hypothalamus of the adult zebrafish and that these neural progenitors are capable of producing GnRH containing neurons.

  19. Zebrafish adult-derived hypothalamic neurospheres generate gonadotropin-releasing hormone (GnRH) neurons

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    Cortés-Campos, Christian; Letelier, Joaquín; Ceriani, Ricardo; Whitlock, Kathleen E.

    2015-01-01

    ABSTRACT Gonadotropin-releasing hormone (GnRH) is a hypothalamic decapeptide essential for fertility in vertebrates. Human male patients lacking GnRH and treated with hormone therapy can remain fertile after cessation of treatment suggesting that new GnRH neurons can be generated during adult life. We used zebrafish to investigate the neurogenic potential of the adult hypothalamus. Previously we have characterized the development of GnRH cells in the zebrafish linking genetic pathways to the differentiation of neuromodulatory and endocrine GnRH cells in specific regions of the brain. Here, we developed a new method to obtain neural progenitors from the adult hypothalamus in vitro. Using this system, we show that neurospheres derived from the adult hypothalamus can be maintained in culture and subsequently differentiate glia and neurons. Importantly, the adult derived progenitors differentiate into neurons containing GnRH and the number of cells is increased through exposure to either testosterone or GnRH, hormones used in therapeutic treatment in humans. Finally, we show in vivo that a neurogenic niche in the hypothalamus contains GnRH positive neurons. Thus, we demonstrated for the first time that neurospheres can be derived from the hypothalamus of the adult zebrafish and that these neural progenitors are capable of producing GnRH containing neurons. PMID:26209533

  20. Treatment of Glucocorticoids Inhibited Early Immune Responses and Impaired Cardiac Repair in Adult Zebrafish.

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    Wei-Chang Huang

    Full Text Available Myocardial injury, such as myocardial infarction (MI, can lead to drastic heart damage. Zebrafish have the extraordinary ability to regenerate their heart after a severe injury. Upon ventricle resection, fibrin clots seal the wound and serve as a matrix for recruiting myeloid-derived phagocytes. Accumulated neutrophils and macrophages not only reduce the risk of infection but also secrete cytokines and growth factors to promote tissue repair. However, the underlying cellular and molecular mechanisms for how immune responses are regulated during the early stages of cardiac repair are still unclear. We investigated the role and programming of early immune responses during zebrafish heart regeneration. We found that zebrafish treated with an anti-inflammatory glucocorticoid had significantly reduced heart regenerative capacities, consistent with findings in other higher vertebrates. Moreover, inhibiting the inflammatory response led to excessive collagen deposition. A microarray approach was used to assess the differential expression profiles between zebrafish hearts with normal or impaired healing. Combining cytokine profiling and immune-staining, our data revealed that impaired heart regeneration could be due to reduced phagocyte recruitment, leading to diminished angiogenesis and cell proliferation post-cardiac injury. Despite their robust regenerative ability, our study revealed that glucocorticoid treatment could effectively hinder cardiac repair in adult zebrafish by interfering with the inflammatory response. Our findings may help to clarify the initiation of cardiac repair, which could be used to develop a therapeutic intervention that may enhance cardiac repair in humans to compensate for the loss of cardiomyocytes after an MI.

  1. The chemokine receptor cxcr5 regulates the regenerative neurogenesis response in the adult zebrafish brain

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    Kizil Caghan

    2012-07-01

    Full Text Available Abstract Background Unlike mammals, zebrafish exhibits extensive neural regeneration after injury in adult stages of its lifetime due to the neurogenic activity of the radial glial cells. However, the genes involved in the regenerative neurogenesis response of the zebrafish brain are largely unknown. Thus, understanding the underlying principles of this regeneration capacity of the zebrafish brain is an interesting research realm that may offer vast clinical ramifications. Results In this paper, we characterized the expression pattern of cxcr5 and analyzed the function of this gene during adult neurogenesis and regeneration of the zebrafish telencephalon. We found that cxcr5 was upregulated transiently in the RGCs and neurons, and the expression in the immune cells such as leukocytes was negligible during both adult neurogenesis and regeneration. We observed that the transgenic misexpression of cxcr5 in the ventricular cells using dominant negative and full-length variants of the gene resulted in altered proliferation and neurogenesis response of the RGCs. When we knocked down cxcr5 using antisense morpholinos and cerebroventricular microinjection, we observed outcomes similar to the overexpression of the dominant negative cxcr5 variant. Conclusions Thus, based on our results, we propose that cxcr5 imposes a proliferative permissiveness to the radial glial cells and is required for differentiation of the RGCs to neurons, highlighting novel roles of cxcr5 in the nervous system of vertebrates. We therefore suggest that cxcr5 is an important cue for ventricular cell proliferation and regenerative neurogenesis in the adult zebrafish telencephalon. Further studies on the role of cxcr5 in mediating neuronal replenishment have the potential to produce clinical ramifications in efforts for regenerative therapeutic applications for human neurological disorders or acute injuries.

  2. F-spondin/spon1b expression patterns in developing and adult zebrafish.

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    Veronica Akle

    Full Text Available F-spondin, an extracellular matrix protein, is an important player in embryonic morphogenesis and CNS development, but its presence and role later in life remains largely unknown. We generated a transgenic zebrafish in which GFP is expressed under the control of the F-spondin (spon1b promoter, and used it in combination with complementary techniques to undertake a detailed characterization of the expression patterns of F-spondin in developing and adult brain and periphery. We found that F-spondin is often associated with structures forming long neuronal tracts, including retinal ganglion cells, the olfactory bulb, the habenula, and the nucleus of the medial longitudinal fasciculus (nMLF. F-spondin expression coincides with zones of adult neurogenesis and is abundant in CSF-contacting secretory neurons, especially those in the hypothalamus. Use of this new transgenic model also revealed F-spondin expression patterns in the peripheral CNS, notably in enteric neurons, and in peripheral tissues involved in active patterning or proliferation in adults, including the endoskeleton of zebrafish fins and the continuously regenerating pharyngeal teeth. Moreover, patterning of the regenerating caudal fin following fin amputation in adult zebrafish was associated with F-spondin expression in the blastema, a proliferative region critical for tissue reconstitution. Together, these findings suggest major roles for F-spondin in the CNS and periphery of the developing and adult vertebrate.

  3. Photoacoustic Tomography Imaging of the Adult Zebrafish by Using Unfocused and Focused High-Frequency Ultrasound Transducers

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    Yubin Liu

    2016-11-01

    Full Text Available The zebrafish model provides an essential platform for the study of human diseases or disorders due to the possession of about 87% homologous genes with human. However, it is still very challenging to noninvasively visualize the structure and function of adult zebrafish based on available optical imaging techniques. In this study, photoacoustic tomography (PAT was utilized for high-resolution imaging of adult zebrafish by using focused and unfocused high-frequency (10 MHz ultrasound transducers. We examined and compared the imaging results from the two categories of transducers with in vivo experimental tests, in which we discovered that the unfocused transducer is able to identify the inner organs of adult zebrafish with higher contrast but limited regional resolution, whereas the findings from the focused transducer were with high resolution but limited regional contrast for the recovered inner organs.

  4. Time-Gated Optical Projection Tomography Allows Visualization of Adult Zebrafish Internal Structures

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    Foglia, Efrem Alessandro; Pistocchi, Anna; D'Andrea, Cosimo; Valentini, Gianluca; Cubeddu, Rinaldo; De Silvestri, Sandro; Cerullo, Giulio; Cotelli, Franco

    2012-01-01

    Optical imaging through biological samples is compromised by tissue scattering and currently various approaches aim to overcome this limitation. In this paper we demonstrate that an all optical technique, based on non-linear upconversion of infrared ultrashort laser pulses and on multiple view acquisition, allows the reduction of scattering effects in tomographic imaging. This technique, namely Time-Gated Optical Projection Tomography (TGOPT), is used to reconstruct three dimensionally the internal structure of adult zebrafish without staining or clearing agents. This method extends the use of Optical Projection Tomography to optically diffusive samples yielding reconstructions with reduced artifacts, increased contrast and improved resolution with respect to those obtained with non-gated techniques. The paper shows that TGOPT is particularly suited for imaging the skeletal system and nervous structures of adult zebrafish. PMID:23185643

  5. zebraflash transgenic lines for in vivo bioluminescence imaging of stem cells and regeneration in adult zebrafish.

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    Chen, Chen-Hui; Durand, Ellen; Wang, Jinhu; Zon, Leonard I; Poss, Kenneth D

    2013-12-01

    The zebrafish has become a standard model system for stem cell and tissue regeneration research, based on powerful genetics, high tissue regenerative capacity and low maintenance costs. Yet, these studies can be challenged by current limitations of tissue visualization techniques in adult animals. Here we describe new imaging methodology and present several ubiquitous and tissue-specific luciferase-based transgenic lines, which we have termed zebraflash, that facilitate the assessment of regeneration and engraftment in freely moving adult zebrafish. We show that luciferase-based live imaging reliably estimates muscle quantity in an internal organ, the heart, and can longitudinally follow cardiac regeneration in individual animals after major injury. Furthermore, luciferase-based detection enables visualization and quantification of engraftment in live recipients of transplanted hematopoietic stem cell progeny, with advantages in sensitivity and gross spatial resolution over fluorescence detection. Our findings present a versatile resource for monitoring and dissecting vertebrate stem cell and regeneration biology.

  6. Effect of radiation dose-rate on hematopoietic cell engraftment in adult zebrafish.

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    Tiffany J Glass

    Full Text Available Although exceptionally high radiation dose-rates are currently attaining clinical feasibility, there have been relatively few studies reporting the biological consequences of these dose-rates in hematopoietic cell transplant (HCT. In zebrafish models of HCT, preconditioning before transplant is typically achieved through radiation alone. We report the comparison of outcomes in adult zebrafish irradiated with 20 Gy at either 25 or 800 cGy/min in the context of experimental HCT. In non-transplanted irradiated fish we observed no substantial differences between dose-rate groups as assessed by fish mortality, cell death in the kidney, endogenous hematopoietic reconstitution, or gene expression levels of p53 and ddb2 (damage-specific DNA binding protein 2 in the kidney. However, following HCT, recipients conditioned with the higher dose rate showed significantly improved donor-derived engraftment at 9 days post transplant (p ≤ 0.0001, and improved engraftment persisted at 31 days post transplant. Analysis for sdf-1a expression, as well as transplant of hematopoietic cells from cxcr4b -/- zebrafish, (odysseus, cumulatively suggest that the sdf-1a/cxcr4b axis is not required of donor-derived cells for the observed dose-rate effect on engraftment. Overall, the adult zebrafish model of HCT indicates that exceptionally high radiation dose-rates can impact HCT outcome, and offers a new system for radiobiological and mechanistic interrogation of this phenomenon. Key words: Radiation dose rate, Total Marrow Irradiation (TMI, Total body irradiation (TBI, SDF-1, Zebrafish, hematopoietic cell transplant.

  7. Teleost Chemokines and Their Receptors

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    Steve Bird

    2015-11-01

    Full Text Available Chemokines are a superfamily of cytokines that appeared about 650 million years ago, at the emergence of vertebrates, and are responsible for regulating cell migration under both inflammatory and physiological conditions. The first teleost chemokine gene was reported in rainbow trout in 1998. Since then, numerous chemokine genes have been identified in diverse fish species evidencing the great differences that exist among fish and mammalian chemokines, and within the different fish species, as a consequence of extensive intrachromosomal gene duplications and different infectious experiences. Subsequently, it has only been possible to establish clear homologies with mammalian chemokines in the case of some chemokines with well-conserved homeostatic roles, whereas the functionality of other chemokine genes will have to be independently addressed in each species. Despite this, functional studies have only been undertaken for a few of these chemokine genes. In this review, we describe the current state of knowledge of chemokine biology in teleost fish. We have mainly focused on those species for which more research efforts have been made in this subject, specially zebrafish (Danio rerio, rainbow trout (Oncorhynchus mykiss and catfish (Ictalurus punctatus, outlining which genes have been identified thus far, highlighting the most important aspects of their expression regulation and addressing any known aspects of their biological role in immunity. Finally, we summarise what is known about the chemokine receptors in teleosts and provide some analysis using recently available data to help characterise them more clearly.

  8. The Behavioral Effects of Single Housing and Environmental Enrichment on Adult Zebrafish (Danio rerio).

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    Collymore, Chereen; Tolwani, Ravi J; Rasmussen, Skye

    2015-05-01

    Environmental enrichment provides laboratory-housed species the opportunity to express natural behavior and exert control over their home environment, thereby minimizing stress. We sought to determine whether providing an artificial plant in the holding tank as enrichment influenced anxiety-like behaviors and place-preference choice in adult zebrafish. Fish were housed singly or in social groups of 5 for 3 wk in 1 of 4 experimental housing environments: single-housed enriched (n = 30), single-housed barren (n = 30), group-housed enriched (n = 30), and group-housed barren (n = 30). On week 4, individual fish were selected randomly from each of the experimental housing environments and tested by using novel-tank, light-dark, and place-preference tests. Housing fish singly in a barren environment increased anxiety-like behaviors in the novel-tank and light-dark behavioral tests. Single-housed zebrafish in barren tanks as well as zebrafish group-housed with conspecifics, both with and without plant enrichment, spent more time associating with conspecifics than with the artificial plant enrichment device during the place-preference test. Single-housed fish maintained in enriched tanks displayed no preference between a compartment with conspecifics or an artificial plant. Our results suggest the addition of an artificial plant as enrichment may benefit single-housed zebrafish when social housing is not possible.

  9. Identification and characterization of the pumilio-2 expressed in zebrafish embryos and adult tissues.

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    Wang, Huan Nan; Xu, Yan; Tao, Ling Jie; Zhou, Jian; Qiu, Meng Xi; Teng, Yu Hang; Deng, Feng Jiao

    2012-03-01

    Pumilio proteins regulate the translation of specific proteins required for germ cell development and morphogenesis. In the present study, we have identified the pumilio-2 in zebrafish and analyze its expression in adult tissues and early embryos. Pumilio-2 codes for the full-length Pumilio-2 protein and contains a PUF-domain. When compared to the mammalian and avian Pumilio-2 proteins, zebrafish Pumilio-2 protein was found to contain an additional sequence of 24 amino acid residues within the PUF-domain. Zebrafish pumilio-2 mRNA is expressed in the ovary, testis, liver, kidney and brain but is absent in the heart and muscle as detected by RT-PCR. The results of in situ hybridization indicate that transcripts of pumilio-2 are distributed in all blastomeres from the 1-cell stage to the sphere stage and accumulate in the head and tail during the 60%-epiboly and 3-somite stages. Transcripts were also detected in the brain and neural tube of the 24 h post-fertilization (hpf) embryos. Western blot analyses indicate that the Pumilio-2 protein is strongly expressed in the ovary, testis and brain but not in other tissues. These data suggest that pumilio-2 plays an important role in the development of the zebrafish germ cells and nervous system.

  10. Insights into the evolutionary history of the vertebrate zic3 locus from a teleost-specific zic6 gene in the zebrafish, Danio rerio.

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    Keller, Michael J; Chitnis, Ajay B

    2007-07-01

    The Zic gene family of zinc-finger transcription factors includes five orthologues, zic1-5, that are common to the Euteleostian vertebrates (fish, frogs, birds, and mammals). The Zic genes have been implicated as regulators of a number of critical developmental processes, including neurulation, neuronal differentiation, neural crest specification, the establishment of left-right asymmetry, and regulation of cell proliferation. The different Zic genes encode proteins that are expressed in broadly overlapping spatial domains, have conserved DNA-binding domains that recognize a common motif, are capable of physical interactions, and can co-regulate one another's transcription. Thus, the transcriptional regulation of individual proteins and their effects on downstream targets must be assessed within the context of co-expression with other family members. We describe a novel gene, zic6, that is specific to the teleost fishes and lacks the lateral and rostral expression domains typical of the other Zic family members. We present evidence that zic6 is an ancestral locus arising by chromosomal duplication early in the Euteleostomi that was subsequently lost in the terrestrial vertebrates.

  11. Long-term (30 days toxicity of NiO nanoparticles for adult zebrafish Danio rerio

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    Kovrižnych Jevgenij A.

    2014-03-01

    Full Text Available Nickel oxide in the form of nanoparticles (NiO NPs is extensively used in different industrial branches. In a test on adult zebrafish, the acute toxicity of NiO NPs was shown to be low, however longlasting contact with this compound can lead to its accumulation in the tissues and to increased toxicity. In this work we determined the 30-day toxicity of NiO NPs using a static test for zebrafish Danio rerio. We found the 30-day LC50 value to be 45.0 mg/L, LC100 (minimum concentration causing 100% mortality was 100.0 mg/L, and LC0 (maximum concentration causing no mortality was 6.25 mg/L for adult individuals of zebrafish. Considering a broad use of Ni in the industry, NiO NPs chronic toxicity may have a negative impact on the population of aquatic organisms and on food web dynamics in aquatic systems.

  12. Characterization of Proliferating Neural Progenitors after Spinal Cord Injury in Adult Zebrafish.

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    Subhra Prakash Hui

    Full Text Available Zebrafish can repair their injured brain and spinal cord after injury unlike adult mammalian central nervous system. Any injury to zebrafish spinal cord would lead to increased proliferation and neurogenesis. There are presences of proliferating progenitors from which both neuronal and glial loss can be reversed by appropriately generating new neurons and glia. We have demonstrated the presence of multiple progenitors, which are different types of proliferating populations like Sox2+ neural progenitor, A2B5+ astrocyte/ glial progenitor, NG2+ oligodendrocyte progenitor, radial glia and Schwann cell like progenitor. We analyzed the expression levels of two common markers of dedifferentiation like msx-b and vimentin during regeneration along with some of the pluripotency associated factors to explore the possible role of these two processes. Among the several key factors related to pluripotency, pou5f1 and sox2 are upregulated during regeneration and associated with activation of neural progenitor cells. Uncovering the molecular mechanism for endogenous regeneration of adult zebrafish spinal cord would give us more clues on important targets for future therapeutic approach in mammalian spinal cord repair and regeneration.

  13. 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

    was not significantly affected, but yolk-sac oedema tended to increase in frequency in exposed groups with time. Our results show that dietary exposure to TBP, at concentrations found in marine organisms that are part of the natural diet of wild fish, can interfere with reproduction in zebrafish. We also observed low......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...

  14. Mycobacterium marinum causes a latent infection that can be reactivated by gamma irradiation in adult zebrafish.

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    Mataleena Parikka

    2012-09-01

    Full Text Available The mechanisms leading to latency and reactivation of human tuberculosis are still unclear, mainly due to the lack of standardized animal models for latent mycobacterial infection. In this longitudinal study of the progression of a mycobacterial disease in adult zebrafish, we show that an experimental intraperitoneal infection with a low dose (≈ 35 bacteria of Mycobacterium marinum, results in the development of a latent disease in most individuals. The infection is characterized by limited mortality (25%, stable bacterial loads 4 weeks following infection and constant numbers of highly organized granulomas in few target organs. The majority of bacteria are dormant during a latent mycobacterial infection in zebrafish, and can be activated by resuscitation promoting factor ex vivo. In 5-10% of tuberculosis cases in humans, the disease is reactivated usually as a consequence of immune suppression. In our model, we are able to show that reactivation can be efficiently induced in infected zebrafish by γ-irradiation that transiently depletes granulo/monocyte and lymphocyte pools, as determined by flow cytometry. This immunosuppression causes reactivation of the dormant mycobacterial population and a rapid outgrowth of bacteria, leading to 88% mortality in four weeks. In this study, the adult zebrafish presents itself as a unique non-mammalian vertebrate model for studying the development of latency, regulation of mycobacterial dormancy, as well as reactivation of latent or subclinical tuberculosis. The possibilities for screening for host and pathogen factors affecting the disease progression, and identifying novel therapeutic agents and vaccine targets make this established model especially attractive.

  15. Endosulfan affects health variables in adult zebrafish (Danio rerio) and induces alterations in larvae development

    DEFF Research Database (Denmark)

    Velasco-Santamaria, Y. M.; Handy, R. D.; Sloman, K. A.

    2011-01-01

    Adult zebrafish (Danio rerio) were exposed to 0 (control), 0.16 or 0.48 mu g/L of the insecticide, endosulfan, for 28 days. Haematology, whole body ions, thiobarbituric acid reactive substances (TBARS), Na(+)K(+)-ATPase, organ histology and reproduction were assessed in adults. The resulting...... offspring were examined for latent effects on development (heart rate and morphometrics). On day 14, adult fish exposed to 0.16 mu g/L endosulfan showed significantly lower red blood cell counts than those exposed to 0.48 mu g/L endosulfan; adult fish exposed to 0.16 ug/L also showed elevated TBARS compared...... to controls. Both concentrations of endosulfan caused a 4.0 fold increase in Na(+)K(+)-ATPase activity compared to controls (ANOVA, p fish exposed to endosulfan had fewer, enlarged hepatocytes, with cell diameters greater than the controls (ANOVA, p

  16. Regulation of gene expression mediating indeterminate muscle growth in teleosts.

    Science.gov (United States)

    Ahammad, A K Shakur; Asaduzzaman, Md; Asakawa, Shuichi; Watabe, Shugo; Kinoshita, Shigeharu

    2015-08-01

    Teleosts are unique among vertebrates due to their indeterminate muscle growth, i.e., continued production of neonatal muscle fibers until death. However, the molecular mechanism(s) underlying this property is unknown. Here, we focused on the torafugu (Takifugu rubripes) myosin heavy chain gene, MYHM2528-1, which is specifically expressed in neonatal muscle fibers produced by indeterminate muscle growth. We examined the flanking region of MYHM2528-1 through an in vivo reporter assay using zebrafish (Danio rerio) and identified a 2100 bp 5'-flanking sequence that contained sufficient promoter activity to allow specific gene expression. The effects of enhanced promoter activity were observed at the outer region of the fast muscle and the dorsal edge of slow muscle in zebrafish larvae. At the juvenile stage, the promoter was specifically activated in small diameter muscle fibers scattered throughout fast muscle and in slow muscle near the septum separating slow and fast muscles. This spatio-temporal promoter activity overlapped with known myogenic zones involved in teleost indeterminate muscle growth. A deletion mutant analysis revealed that the -2100 to -600 bp 5'flanking sequence of MYHM2528-1 is essential for promoter activity. This region contains putative binding sites for several representative myogenesis-related transcription factors and nuclear factor of activated T-cell (NFAT), a transcription activator involved in regeneration of mammalian adult skeletal muscle. A significant reduction in the promoter activity of the MYHM2528-1 deletion constructs was observed in accordance with a reduction in the number of these binding sites, suggesting the involvement of specific transcription factors in indeterminate muscle growth.

  17. Methylmercury Induced Neurotoxicity and the Influence of Selenium in the Brains of Adult Zebrafish (Danio rerio

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    Josef Daniel Rasinger

    2017-03-01

    Full Text Available The neurotoxicity of methylmercury (MeHg is well characterised, and the ameliorating effects of selenium have been described. However, little is known about the molecular mechanisms behind this contaminant-nutrient interaction. We investigated the influence of selenium (as selenomethionine, SeMet and MeHg on mercury accumulation and protein expression in the brain of adult zebrafish (Danio rerio. Fish were fed diets containing elevated levels of MeHg and/or SeMet in a 2 × 2 full factorial design for eight weeks. Mercury concentrations were highest in the brain tissue of MeHg-exposed fish compared to the controls, whereas lower levels of mercury were found in the brain of zebrafish fed both MeHg and SeMet compared with the fish fed MeHg alone. The expression levels of proteins associated with gap junction signalling, oxidative phosphorylation, and mitochondrial dysfunction were significantly (p < 0.05 altered in the brain of zebrafish after exposure to MeHg and SeMet alone or in combination. Analysis of upstream regulators indicated that these changes were linked to the mammalian target of rapamycin (mTOR pathways, which were activated by MeHg and inhibited by SeMet, possibly through a reactive oxygen species mediated differential activation of RICTOR, the rapamycin-insensitive binding partner of mTOR.

  18. Regulation of Müller glial dependent neuronal regeneration in the damaged adult zebrafish retina.

    Science.gov (United States)

    Gorsuch, Ryne A; Hyde, David R

    2014-06-01

    This article examines our current knowledge underlying the mechanisms involved in neuronal regeneration in the adult zebrafish retina. Zebrafish, which has the capacity to regenerate a wide variety of tissues and organs (including the fins, kidney, heart, brain, and spinal cord), has become the premier model system to study retinal regeneration due to the robustness and speed of the response and the variety of genetic tools that can be applied to study this question. It is now well documented that retinal damage induces the resident Müller glia to dedifferentiate and reenter the cell cycle to produce neuronal progenitor cells that continue to proliferate, migrate to the damaged retinal layer and differentiate into the missing neuronal cell types. Increasing our understanding of how these cellular events are regulated and occur in response to neuronal damage may provide critical information that can be applied to stimulating a regeneration response in the mammalian retina. In this review, we will focus on the genes/proteins that regulate zebrafish retinal regeneration and will attempt to critically evaluate how these factors may interact to correctly orchestrate the definitive cellular events that occur during regeneration.

  19. Zebrafish Development: High-throughput Test Systems to Assess Developmental Toxicity

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    Abstract Because of its developmental concordance, ease of handling and rapid development, the small teleost, zebrafish (Danio rerio), is frequently promoted as a vertebrate model for medium-throughput developmental screens. This present chapter discusses zebrafish as an altern...

  20. Aminoglycoside-induced hair cell death of inner ear organs causes functional deficits in adult zebrafish (Danio rerio.

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    Phillip M Uribe

    Full Text Available Aminoglycoside antibiotics, like gentamicin, kill inner ear sensory hair cells in a variety of species including chickens, mice, and humans. The zebrafish (Danio rerio has been used to study hair cell cytotoxicity in the lateral line organs of larval and adult animals. Little is known about whether aminoglycosides kill the hair cells within the inner ear of adult zebrafish. We report here the ototoxic effects of gentamicin on hair cells in the saccule, the putative hearing organ, and utricle of zebrafish. First, adult zebrafish received a single 30 mg/kg intraperitoneal injection of fluorescently-tagged gentamicin (GTTR to determine the distribution of gentamicin within inner ear sensory epithelia. After 4 hours, GTTR was observed in hair cells throughout the saccular and utriclar sensory epithelia. To assess the ototoxic effects of gentamicin, adult zebrafish received a single 250 mg/kg intraperitoneal injection of gentamicin and, 24 hours later, auditory evoked potential recordings (AEPs revealed significant shifts in auditory thresholds compared to untreated controls. Zebrafish were then euthanized, the inner ear fixed, and labeled for apoptotic cells (TUNEL reaction, and the stereociliary bundles of hair cells labeled with fluorescently-tagged phalloidin. Whole mounts of the saccule and utricle were imaged and cells counted. There were significantly more TUNEL-labeled cells found in both organs 4 hours after gentamicin injection compared to vehicle-injected controls. As expected, significantly fewer hair cell bundles were found along the rostral-caudal axis of the saccule and in the extrastriolar and striolar regions of the utricle in gentamicin-treated animals compared to untreated controls. Therefore, as in other species, gentamicin causes significant inner ear sensory hair cell death and auditory dysfunction in zebrafish.

  1. Strong static magnetic fields elicit swimming behaviors consistent with direct vestibular stimulation in adult zebrafish.

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    Bryan K Ward

    Full Text Available Zebrafish (Danio rerio offer advantages as model animals for studies of inner ear development, genetics and ototoxicity. However, traditional assessment of vestibular function in this species using the vestibulo-ocular reflex requires agar-immobilization of individual fish and specialized video, which are difficult and labor-intensive. We report that using a static magnetic field to directly stimulate the zebrafish labyrinth results in an efficient, quantitative behavioral assay in free-swimming fish. We recently observed that humans have sustained nystagmus in high strength magnetic fields, and we attributed this observation to magnetohydrodynamic forces acting on the labyrinths. Here, fish were individually introduced into the center of a vertical 11.7T magnetic field bore for 2-minute intervals, and their movements were tracked. To assess for heading preference relative to a magnetic field, fish were also placed in a horizontally oriented 4.7T magnet in infrared (IR light. A sub-population was tested again in the magnet after gentamicin bath to ablate lateral line hair cell function. Free-swimming adult zebrafish exhibited markedly altered swimming behavior while in strong static magnetic fields, independent of vision or lateral line function. Two-thirds of fish showed increased swimming velocity or consistent looping/rolling behavior throughout exposure to a strong, vertically oriented magnetic field. Fish also demonstrated altered swimming behavior in a strong horizontally oriented field, demonstrating in most cases preferred swimming direction with respect to the field. These findings could be adapted for 'high-throughput' investigations of the effects of environmental manipulations as well as for changes that occur during development on vestibular function in zebrafish.

  2. Zebrafish whole-adult-organism chemogenomics for large-scale predictive and discovery chemical biology.

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    Siew Hong Lam

    2008-07-01

    Full Text Available The ability to perform large-scale, expression-based chemogenomics on whole adult organisms, as in invertebrate models (worm and fly, is highly desirable for a vertebrate model but its feasibility and potential has not been demonstrated. We performed expression-based chemogenomics on the whole adult organism of a vertebrate model, the zebrafish, and demonstrated its potential for large-scale predictive and discovery chemical biology. Focusing on two classes of compounds with wide implications to human health, polycyclic (halogenated aromatic hydrocarbons [P(HAHs] and estrogenic compounds (ECs, we generated robust prediction models that can discriminate compounds of the same class from those of different classes in two large independent experiments. The robust expression signatures led to the identification of biomarkers for potent aryl hydrocarbon receptor (AHR and estrogen receptor (ER agonists, respectively, and were validated in multiple targeted tissues. Knowledge-based data mining of human homologs of zebrafish genes revealed highly conserved chemical-induced biological responses/effects, health risks, and novel biological insights associated with AHR and ER that could be inferred to humans. Thus, our study presents an effective, high-throughput strategy of capturing molecular snapshots of chemical-induced biological states of a whole adult vertebrate that provides information on biomarkers of effects, deregulated signaling pathways, and possible affected biological functions, perturbed physiological systems, and increased health risks. These findings place zebrafish in a strategic position to bridge the wide gap between cell-based and rodent models in chemogenomics research and applications, especially in preclinical drug discovery and toxicology.

  3. Culture of Adult Transgenic Zebrafish Retinal Explants for Live-cell Imaging by Multiphoton Microscopy.

    Science.gov (United States)

    Lahne, Manuela; Gorsuch, Ryne A; Nelson, Craig M; Hyde, David R

    2017-02-24

    An endogenous regeneration program is initiated by Müller glia in the adult zebrafish (Danio rerio) retina following neuronal damage and death. The Müller glia re-enter the cell cycle and produce neuronal progenitor cells that undergo subsequent rounds of cell divisions and differentiate into the lost neuronal cell types. Both Müller glia and neuronal progenitor cell nuclei replicate their DNA and undergo mitosis in distinct locations of the retina, i.e. they migrate between the basal Inner Nuclear Layer (INL) and the Outer Nuclear Layer (ONL), respectively, in a process described as Interkinetic Nuclear Migration (INM). INM has predominantly been studied in the developing retina. To examine the dynamics of INM in the adult regenerating zebrafish retina in detail, live-cell imaging of fluorescently-labeled Müller glia/neuronal progenitor cells is required. Here, we provide the conditions to isolate and culture dorsal retinas from Tg[gfap:nGFP](mi2004) zebrafish that were exposed to constant intense light for 35 h. We also show that these retinal cultures are viable to perform live-cell imaging experiments, continuously acquiring z-stack images throughout the thickness of the retinal explant for up to 8 h using multiphoton microscopy to monitor the migratory behavior of gfap:nGFP-positive cells. In addition, we describe the details to perform post-imaging analysis to determine the velocity of apical and basal INM. To summarize, we established conditions to study the dynamics of INM in an adult model of neuronal regeneration. This will advance our understanding of this crucial cellular process and allow us to determine the mechanisms that control INM.

  4. Relaxin gene family in teleosts: phylogeny, syntenic mapping, selective constraint, andexpression analysis

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    Glen Peter

    2009-12-01

    Full Text Available Abstract Background In recent years, the relaxin family of signaling molecules has been shown to play diverse roles in mammalian physiology, but little is known about its diversity or physiology in teleosts, an infraclass of the bony fishes comprising ~ 50% of all extant vertebrates. In this paper, 32 relaxin family sequences were obtained by searching genomic and cDNA databases from eight teleost species; phylogenetic, molecular evolutionary, and syntenic data analyses were conducted to understand the relationship and differential patterns of evolution of relaxin family genes in teleosts compared with mammals. Additionally, real-time quantitative PCR was used to confirm and assess the tissues of expression of five relaxin family genes in Danio rerio and in situ hybridization used to assess the site-specific expression of the insulin 3-like gene in D. rerio testis. Results Up to six relaxin family genes were identified in each teleost species. Comparative syntenic mapping revealed that fish possess two paralogous copies of human RLN3, which we call rln3a and rln3b, an orthologue of human RLN2, rln, two paralogous copies of human INSL5, insl5a and insl5b, and an orthologue of human INSL3, insl3. Molecular evolutionary analyses indicated that: rln3a, rln3b and rln are under strong evolutionary constraint, that insl3 has been subject to moderate rates of sequence evolution with two amino acids in insl3/INSL3 showing evidence of positively selection, and that insl5b exhibits a higher rate of sequence evolution than its paralogue insl5a suggesting that it may have been neo-functionalized after the teleost whole genome duplication. Quantitative PCR analyses in D. rerio indicated that rln3a and rln3b are expressed in brain, insl3 is highly expressed in gonads, and that there was low expression of both insl5 genes in adult zebrafish. Finally, in situ hybridization of insl3 in D. rerio testes showed highly specific hybridization to interstitial Leydig

  5. Notch receptor expression in neurogenic regions of the adult zebrafish brain.

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    Vanessa de Oliveira-Carlos

    Full Text Available The adult zebrash brain has a remarkable constitutive neurogenic capacity. The regulation and maintenance of its adult neurogenic niches are poorly understood. In mammals, Notch signaling is involved in stem cell maintenance both in embryonic and adult CNS. To better understand how Notch signaling is involved in stem cell maintenance during adult neurogenesis in zebrafish we analysed Notch receptor expression in five neurogenic zones of the adult zebrafish brain. Combining proliferation and glial markers we identified several subsets of Notch receptor expressing cells. We found that 90 [Formula: see text] of proliferating radial glia express notch1a, notch1b and notch3. In contrast, the proliferating non-glial populations of the dorsal telencephalon and hypothalamus rarely express notch3 and about half express notch1a/1b. In the non-proliferating radial glia notch3 is the predominant receptor throughout the brain. In the ventral telencephalon and in the mitotic area of the optic tectum, where cells have neuroepithelial properties, notch1a/1b/3 are expressed in most proliferating cells. However, in the cerebellar niche, although progenitors also have neuroepithelial properties, only notch1a/1b are expressed in a high number of PCNA [Formula: see text] cells. In this region notch3 expression is mostly in Bergmann glia and at low levels in few PCNA [Formula: see text] cells. Additionally, we found that in the proliferation zone of the ventral telencephalon, Notch receptors display an apical high to basal low gradient of expression. Notch receptors are also expressed in subpopulations of oligodendrocytes, neurons and endothelial cells. We suggest that the partial regional heterogeneity observed for Notch expression in progenitor cells might be related to the cellular diversity present in each of these neurogenic niches.

  6. Neurochemical coding of enteric neurons in adult and embryonic zebrafish (Danio rerio).

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    Uyttebroek, Leen; Shepherd, Iain T; Harrisson, Fernand; Hubens, Guy; Blust, Ronny; Timmermans, Jean-Pierre; Van Nassauw, Luc

    2010-11-01

    Although the morphology and development of the zebrafish enteric nervous system have been extensively studied, the precise neurochemical coding of enteric neurons and their proportional enteric distribution are currently not known. By using immunohistochemistry, we determined the proportional expression and coexpression of neurochemical markers in the embryonic and adult zebrafish intestine. Tyrosine hydroxylase (TH), vasoactive intestinal peptide (VIP), and pituitary adenylate cyclase-activating peptide (PACAP) were observed only in nerve fibers, whereas other markers were also detected in neuronal cell bodies. Calretinin and calbindin had similar distributions. In embryos, all markers, except for choline acetyltransferase (ChAT) and TH, were present from 72 hours postfertilization. Nitrergic neurons, evenly distributed and remaining constant in time, constituted the major neuronal subpopulation. The neuronal proportions of the other markers increased during development and were characterized by regional differences. In the adult, all markers examined were expressed in the enteric nervous system. A large percentage of enteric neurons displayed calbindin and calretinin, and serotonin was the only marker showing significant distribution differences in the three intestinal regions. Colocalization studies showed that serotonin was not coexpressed with any of the other markers. At least five neuronal subpopulations were determined: a serotonergic, a nitrergic noncholinergic, two cholinergic nonnitrergic subpopulations along with one subpopulation expressing both ChAT and neuronal nitric oxide synthase. Analysis of nerve fibers revealed that nitrergic neurons coexpress VIP and PACAP, and that nitrergic neurons innervate the tunica muscularis, whereas serotonergic and cholinergic nonnitrergic neurons innervate the lamina propria and the tunica muscularis.

  7. 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

  8. Neurodevelopment. Live imaging of adult neural stem cell behavior in the intact and injured zebrafish brain.

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    Barbosa, Joana S; Sanchez-Gonzalez, Rosario; Di Giaimo, Rossella; Baumgart, Emily Violette; Theis, Fabian J; Götz, Magdalena; Ninkovic, Jovica

    2015-05-15

    Adult neural stem cells are the source for restoring injured brain tissue. We used repetitive imaging to follow single stem cells in the intact and injured adult zebrafish telencephalon in vivo and found that neurons are generated by both direct conversions of stem cells into postmitotic neurons and via intermediate progenitors amplifying the neuronal output. We observed an imbalance of direct conversion consuming the stem cells and asymmetric and symmetric self-renewing divisions, leading to depletion of stem cells over time. After brain injury, neuronal progenitors are recruited to the injury site. These progenitors are generated by symmetric divisions that deplete the pool of stem cells, a mode of neurogenesis absent in the intact telencephalon. Our analysis revealed changes in the behavior of stem cells underlying generation of additional neurons during regeneration.

  9. Alternate Immersion in an External Glucose Solution Differentially Affects Blood Sugar Values in Older Versus Younger Zebrafish Adults.

    Science.gov (United States)

    Connaughton, Victoria P; Baker, Cassandra; Fonde, Lauren; Gerardi, Emily; Slack, Carly

    2016-04-01

    Recently, zebrafish have been used to examine hyperglycemia-induced complications (retinopathy and neuropathy), as would occur in individuals with diabetes. Current models to induce hyperglycemia in zebrafish include glucose immersion and streptozotocin injections. Both are effective, although neither is reported to elevate blood sugar values for more than 1 month. In this article, we report differences in hyperglycemia induction and maintenance in young (4-11 months) versus old (1-3 years) zebrafish adults. In particular, older fish immersed in an alternating constant external glucose solution (2%) for 2 months displayed elevated blood glucose levels for the entire experimental duration. In contrast, younger adults displayed only transient hyperglycemia, suggesting the fish were acclimating to the glucose exposure protocol. However, modifying the immersion protocol to include a stepwise increasing glucose concentration (from 1% → 2%→3%) resulted in maintained hyperglycemia in younger zebrafish adults for up to 2 months. Glucose-exposed younger fish collected after 8 weeks of exposure also displayed a significant decrease in wet weight. Taken together, these data suggest different susceptibilities to hyperglycemia in older and younger fish and that stepwise increasing glucose concentrations of 1% are required for maintenance of hyperglycemia in younger adults, with higher concentrations of glucose resulting in greater increases in blood sugar values.

  10. Inter-Specific Differences in Numerical Abilities Among Teleost Fish

    Science.gov (United States)

    Agrillo, Christian; Miletto Petrazzini, Maria Elena; Tagliapietra, Christian; Bisazza, Angelo

    2012-01-01

    Adults, infants and non-human primates are thought to possess similar non-verbal numerical systems, but there is considerable debate regarding whether all vertebrates share the same numerical abilities. Despite an abundance of studies, cross-species comparison remains difficult because the methodology employed and the context of species examination vary considerably across studies. To fill this gap, we used the same procedure, stimuli, and numerical contrasts to compare quantity abilities of five teleost fish: redtail splitfin, guppies, zebrafish, Siamese fighting fish, and angelfish. Subjects were trained to discriminate between two sets of geometrical figures using a food reward. Fish initially were trained on an easy numerical ratio (5 vs. 10 and 6 vs. 12). Once they reached the learning criterion, they were subjected to non-reinforced probe trials in which the set size was constant but numerical ratios varied (8 vs. 12 and 9 vs. 12). They also were subjected to probe trials in which the ratio was constant, but the total set size was increased (25 vs. 50) or decreased (2 vs. 4). Overall, fish generalized to numerosities with a 0.67 ratio, but failed with a 0.75 ratio; they generalized to a smaller set size, but not to a larger one. Only minor differences were observed among the five species. However, in one species, zebrafish, the proportion of individuals reaching the learning criterion was much smaller than in the others. In a control experiment, zebrafish showed a similar lower performance in shape discrimination, suggesting that the observed difference resulted from the zebrafish’s difficulty in learning this procedure rather than from a cross-species variation in the numerical domain. PMID:23162517

  11. Inter-specific differences in numerical abilities among teleost fish

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    Christian eAgrillo

    2012-11-01

    Full Text Available Adults, infants and non-human primates are thought to possess similar non-verbal numerical systems, but there is considerable debate regarding whether all vertebrates share the same numerical abilities. Despite an abundance of studies, cross-species comparison remains difficult because the methodology employed and the context of species examination vary considerably across studies. To fill this gap, we used the same procedure, stimuli and numerical contrasts to compare quantity abilities of five teleost fish: redtail splitfin, guppies, zebrafish, Siamese fighting fish, and angelfish. Subjects were trained to discriminate between two sets of geometrical figures using a food reward. Fish initially were trained on an easy numerical ratio (5 vs. 10 and 6 vs. 12. Once they reached the learning criterion, they were subjected to non-reinforced probe trials in which the set size was constant but numerical ratios varied (8 vs. 12 and 9 vs. 12. They also were subjected to probe trials in which the ratio was constant, but the total set size was increased (25 vs. 50 or decreased (2 vs. 4. Overall, fish generalized to numerosities with a 0.67 ratio, but failed with a 0.75 ratio; they generalized to a smaller set size, but not to a larger one. Only minor differences were observed among the five species. However, in one species, zebrafish, the proportion of individuals reaching the learning criterion was much smaller than in the others. In a control experiment, zebrafish showed a similar lower performance in shape discrimination, suggesting that the observed difference resulted from the zebrafish’s difficulty in learning this procedure rather than from a cross-species variation in the numerical domain.

  12. Triclosan Exposure Is Associated with Rapid Restructuring of the Microbiome in Adult Zebrafish.

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    Christopher A Gaulke

    Full Text Available Growing evidence indicates that disrupting the microbial community that comprises the intestinal tract, known as the gut microbiome, can contribute to the development or severity of disease. As a result, it is important to discern the agents responsible for microbiome disruption. While animals are frequently exposed to a diverse array of environmental chemicals, little is known about their effects on gut microbiome stability and structure. Here, we demonstrate how zebrafish can be used to glean insight into the effects of environmental chemical exposure on the structure and ecological dynamics of the gut microbiome. Specifically, we exposed forty-five adult zebrafish to triclosan-laden food for four or seven days or a control diet, and analyzed their microbial communities using 16S rRNA amplicon sequencing. Triclosan exposure was associated with rapid shifts in microbiome structure and diversity. We find evidence that several operational taxonomic units (OTUs associated with the family Enterobacteriaceae appear to be susceptible to triclosan exposure, while OTUs associated with the genus Pseudomonas appeared to be more resilient and resistant to exposure. We also found that triclosan exposure is associated with topological alterations to microbial interaction networks and results in an overall increase in the number of negative interactions per microbe in these networks. Together these data indicate that triclosan exposure results in altered composition and ecological dynamics of microbial communities in the gut. Our work demonstrates that because zebrafish afford rapid and inexpensive interrogation of a large number of individuals, it is a useful experimental system for the discovery of the gut microbiome's interaction with environmental chemicals.

  13. Comparative Analysis of the Testis and Ovary Transcriptomes in Zebrafish by Combining Experimental and Computational Tools

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    Laszlo Orban

    2006-04-01

    Full Text Available Studies on the zebrafish model have contributed to our understanding of several important developmental processes, especially those that can be easily studied in the embryo. However, our knowledge on late events such as gonad differentiation in the zebrafish is still limited. Here we provide an analysis on the gene sets expressed in the adult zebrafish testis and ovary in an attempt to identify genes with potential role in (zebrafish gonad development and function. We produced 10 533 expressed sequence tags (ESTs from zebrafish testis or ovary and downloaded an additional 23 642 gonad-derived sequences from the zebrafish EST database. We clustered these sequences together with over 13 000 kidney-derived zebrafish ESTs to study partial transcriptomes for these three organs. We searched for genes with gonad-specific expression by screening macroarrays containing at least 2600 unique cDNA inserts with testis-, ovary- and kidney-derived cDNA probes. Clones hybridizing to only one of the two gonad probes were selected, and subsequently screened with computational tools to identify 72 genes with potentially testis-specific and 97 genes with potentially ovary-specific expression, respectively. PCR-amplification confirmed gonad-specificity for 21 of the 45 clones tested (all without known function. Our study, which involves over 47 000 EST sequences and specialized cDNA arrays, is the first analysis of adult organ transcriptomes of zebrafish at such a scale. The study of genes expressed in adult zebrafish testis and ovary will provide useful information on regulation of gene expression in teleost gonads and might also contribute to our understanding of the development and differentiation of reproductive organs in vertebrates.

  14. Aquaporin 4 in the sensory organs of adult zebrafish (Danio rerio).

    Science.gov (United States)

    Zichichi, Rosalia; Magnoli, Domenico; Montalbano, Giuseppe; Laurà, Rosaria; Vega, José A; Ciriaco, Emilia; Germanà, Antonino

    2011-04-12

    The aquaporins (AQPs) are a family (AQP-AQP10) of transmembrane channel proteins that mediate the transport of water, ions, gases, and small molecules across the cell membrane, thus regulating cell homeostasis. AQP4 has the highest water permeability and it is involved in hearing and vision in mammals. Here, we used immunohistochemistry to map the presence of AQP4 in the sensory organs of adult zebrafish. The antibody used detected by Western blot proteins of 34 kDa (equivalent to that of mammalian AQP4) and maps in the sensory cells of taste buds, the hair sensory cells of the neuromast and of the maculae, and cristae ampullaris of the inner ear. Moreover, the retinal photoreceptors display AQP4 immunoreactivity. The non-sensory cells in these organs were AQP4 negative. These results suggest that the AQP4 could play a role in the regulation of water balance and ion transport in the sensory cells of zebrafish, bringing new data for the utilizing of this experimental model in the biology of sensory system.

  15. Exposure to Zinc Sulfate Results in Differential Effects on Olfactory Sensory Neuron Subtypes in Adult Zebrafish

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    James T. Hentig

    2016-08-01

    Full Text Available Zinc sulfate is a known olfactory toxicant, although its specific effects on the olfactory epithelium of zebrafish are unknown. Olfactory organs of adult zebrafish were exposed to zinc sulfate and, after 2, 3, 5, 7, 10 or 14 days, fish were processed for histological, immunohistochemical, ultrastructural, and behavioral analyses. Severe morphological disruption of the olfactory organ was observed two days following zinc sulfate exposure, including fusion of lamellae, epithelial inflammation, and significant loss of anti-calretinin labeling. Scanning electron microscopy revealed the apical surface of the sensory region was absent of ciliated structures, but microvilli were still present. Behavioral analysis showed significant loss of the ability to perceive bile salts and some fish also had no response to amino acids. Over the next several days, olfactory organ morphology, epithelial structure, and anti-calretinin labeling returned to control-like conditions, although the ability to perceive bile salts remained lost until day 14. Thus, exposure to zinc sulfate results in rapid degeneration of the olfactory organ, followed by restoration of morphology and function within two weeks. Zinc sulfate appears to have a greater effect on ciliated olfactory sensory neurons than on microvillous olfactory sensory neurons, suggesting differential effects on sensory neuron subtypes.

  16. Effects of butachlor on reproduction and hormone levels in adult zebrafish (Danio rerio).

    Science.gov (United States)

    Chang, Juhua; Liu, Shaoying; Zhou, Shengli; Wang, Minghua; Zhu, Guonian

    2013-01-01

    Butachlor, a chloracetamide herbicide, is widely used in China. In the present study, paired adult male and female zebrafish (Danio rerio) were exposed to various concentrations of butachlor (0, 25, 50 and 100 μg/L) for 30 days, and the effects on reproduction and endocrine disruption were evaluated using fecundity, condition factor (CF), gonadosomatic index (GSI), liver somatic index (LSI), plasma vitellogenin (VTG), sex steroids and thyroid hormone levels as endpoints. Our results showed that the mean fecundity rates were significantly decreased at 50 and 100 μg/L butachlor during the 30-day exposure period. At the end of the exposure period, no significant changes were observed in CF and LSI in both females and males, while GSI was significantly reduced in males at 50 and 100 μg/L butachlor. At 100 μg/L butachlor, plasma testosterone (T) and 17β-estradiol (E2) levels were significantly decreased in females, while plasma VTG level was significantly increased in males. Plasma thyroxine (T4) and triiodothyronine (T3) levels were significantly increased at 50 and 100 μg/L butachlor in males, and at 100 μg/L in females. This work demonstrated that butachlor adversely affected the normal reproductive success of zebrafish, and disrupted the thyroid and sex steroid endocrine systems, which provides the basis for the estimated ecological risk during butachlor exposure.

  17. Long-term exposure to paraquat alters behavioral parameters and dopamine levels in adult zebrafish (Danio rerio).

    Science.gov (United States)

    Bortolotto, Josiane W; Cognato, Giana P; Christoff, Raissa R; Roesler, Laura N; Leite, Carlos E; Kist, Luiza W; Bogo, Mauricio R; Vianna, Monica R; Bonan, Carla D

    2014-04-01

    Chronic exposure to paraquat (Pq), a toxic herbicide, can result in Parkinsonian symptoms. This study evaluated the effect of the systemic administration of Pq on locomotion, learning and memory, social interaction, tyrosine hydroxylase (TH) expression, dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) levels, and dopamine transporter (DAT) gene expression in zebrafish. Adult zebrafish received an i.p. injection of either 10 mg/kg (Pq10) or 20 mg/kg (Pq20) of Pq every 3 days for a total of six injections. Locomotion and distance traveled decreased at 24 h after each injection in both treatment doses. In addition, both Pq10- and Pq20-treated animals exhibited differential effects on the absolute turn angle. Nonmotor behaviors were also evaluated, and no changes were observed in anxiety-related behaviors or social interactions in Pq-treated zebrafish. However, Pq-treated animals demonstrated impaired acquisition and consolidation of spatial memory in the Y-maze task. Interestingly, dopamine levels increased while DOPAC levels decreased in the zebrafish brain after both treatments. However, DAT expression decreased in the Pq10-treated group, and there was no change in the Pq20-treated group. The amount of TH protein showed no significant difference in the treated group. Our study establishes a new model to study Parkinson-associated symptoms in zebrafish that have been chronically treated with Pq.

  18. Zebrafish: a novel research tool for cardiac (patho)electrophysiology and ion channel disorders.

    Science.gov (United States)

    Verkerk, Arie O; Remme, Carol Ann

    2012-01-01

    The zebrafish is a cold-blooded tropical freshwater teleost with two-chamber heart morphology. A major advantage of the zebrafish for heart studies is that the embryo is transparent, allowing for easy assessment of heart development, heart rate analysis and phenotypic characterization. Moreover, rapid and effective gene-specific knockdown can be achieved using morpholino oligonucleotides. Lastly, zebrafish are small in size, are easy to maintain and house, grow fast, and have large offspring size, making them a cost-efficient research model. Zebrafish embryonic and adult heart rates as well as action potential (AP) shape and duration and electrocardiogram morphology closely resemble those of humans. However, whether the zebrafish is truly an attractive alternative model for human cardiac electrophysiology depends on the presence and gating properties of the various ion channels in the zebrafish heart, but studies into the latter are as yet limited. The rapid component of the delayed rectifier K(+) current (I(Kr)) remains the best characterized and validated ion current in zebrafish myocytes, and zebrafish may represent a valuable model to investigate human I(Kr) channel-related disease, including long QT syndrome. Arguments against the use of zebrafish as model for human cardiac (patho)electrophysiology include its cold-bloodedness and two-chamber heart morphology, absence of t-tubuli, sarcoplamatic reticulum function, and a different profile of various depolarizing and repolarizing ion channels, including a limited Na(+) current density. Based on the currently available literature, we propose that zebrafish may constitute a relevant research model for investigating ion channel disorders associated with abnormal repolarization, but may be less suitable for studying depolarization disorders or Ca(2+)-modulated arrhythmias.

  19. Zebrafish: a novel research tool for cardiac (pathoelectrophysiology and ion channel disorders

    Directory of Open Access Journals (Sweden)

    Arie O Verkerk

    2012-07-01

    Full Text Available The zebrafish is a cold-blooded tropical freshwater teleost with a two-chamber heart morphology, typical for non-mammalian vertebrates. A major advantage of the zebrafish for heart studies is that the embryo is transparent, allowing for easy assessment of heart development, heart rate analysis and phenotypic characterization. Moreover, rapid and effective gene-specific knockdown can be achieved using morpholino oligonucleotides. Lastly, zebrafish are small in size, are easy to maintain and house, grow fast, and have large offspring size, making them a cost-efficient research model. Zebrafish embryonic and adult heart rates as well as action potential shape and duration and electrocardiogram morphology closely resemble those of humans. However, whether the zebrafish is truly an attractive alternative model for human cardiac electrophysiology depends on the presence and gating properties of the various ion channels in the zebrafish heart, but studies into the latter are as yet limited. The rapid component of the delayed rectifier K+ current (IKr remains the best characterized and validated ion current in zebrafish myocytes, and zebrafish may represent a valuable model to investigate human IKr channel related disease, including long QT syndrome. Arguments against the use of zebrafish as model for human cardiac (pathoelectrophysiology include its cold-bloodedness and two-chamber heart morphology, absence of t-tubuli, sarcoplamatic reticulum function, and a different profile of various depolarizing and repolarizing ion channels, including a limited Na+ current density. Based on the currently available literature, we propose that zebrafish may constitute a relevant research model for investigating ion channel disorders associated with abnormal repolarization, but may be less suitable for studying depolarization disorders or Ca2+-modulated arrhythmias.

  20. Development and specification of cerebellar stem and progenitor cells in zebrafish: from embryo to adult

    OpenAIRE

    Kaslin, Jan; Kroehne, Volker; Benato, Francesca; Argenton, Francesco; Brand, Michael

    2013-01-01

    Background Teleost fish display widespread post-embryonic neurogenesis originating from many different proliferative niches that are distributed along the brain axis. During the development of the central nervous system (CNS) different cell types are produced in a strict temporal order from increasingly committed progenitors. However, it is not known whether diverse neural stem and progenitor cell types with restricted potential or stem cells with broad potential are maintained in the teleost...

  1. Reduced swim performance and aerobic capacity in adult zebrafish exposed to waterborne selenite.

    Science.gov (United States)

    Massé, Anita J; Thomas, Jith K; Janz, David M

    2013-04-01

    Although dietary exposure of adult fish to organoselenium in contaminated aquatic ecosystems has been reported to bioaccumulate and cause larval deformities in offspring, subtle physiological effects produced through low level waterborne selenium exposure in fish such as swim performance and aerobic capacity have not been investigated. To evaluate potential effects of selenite on these responses, adult zebrafish (Danio rerio) were exposed to nominal aqueous concentrations of 0, 10 or 100 μg/L sodium selenite for 14 days. Upon completion of the exposure period, fish underwent two successive swim trials in a swim tunnel respirometer to determine critical swim speed (Ucrit), oxygen consumption (MO2), standard and active metabolic rates, aerobic scope (AS) and cost of transport (COT) followed by analysis of whole body triglyceride and glycogen concentrations. Selenite exposure had a significant negative effect on Ucrit and aerobic capacity. Active metabolic rates and AS significantly decreased in both selenite exposure groups after the second swim trial. No significant effect was observed in MO2, standard metabolic rate, COT, triglyceride and glycogen levels, or condition factor between groups. These results suggest that aqueous selenite exposure at environmentally relevant concentrations produces adverse effects on aerobic capacity that can diminish endurance and maximum swim speeds, which may lower fish survivability.

  2. Adult zebrafish intestine resection: a novel model of short bowel syndrome, adaptation, and intestinal stem cell regeneration.

    Science.gov (United States)

    Schall, K A; Holoyda, K A; Grant, C N; Levin, D E; Torres, E R; Maxwell, A; Pollack, H A; Moats, R A; Frey, M R; Darehzereshki, A; Al Alam, D; Lien, C; Grikscheit, T C

    2015-08-01

    Loss of significant intestinal length from congenital anomaly or disease may lead to short bowel syndrome (SBS); intestinal failure may be partially offset by a gain in epithelial surface area, termed adaptation. Current in vivo models of SBS are costly and technically challenging. Operative times and survival rates have slowed extension to transgenic models. We created a new reproducible in vivo model of SBS in zebrafish, a tractable vertebrate model, to facilitate investigation of the mechanisms of intestinal adaptation. Proximal intestinal diversion at segment 1 (S1, equivalent to jejunum) was performed in adult male zebrafish. SBS fish emptied distal intestinal contents via stoma as in the human disease. After 2 wk, S1 was dilated compared with controls and villus ridges had increased complexity, contributing to greater villus epithelial perimeter. The number of intervillus pockets, the intestinal stem cell zone of the zebrafish increased and contained a higher number of bromodeoxyuridine (BrdU)-labeled cells after 2 wk of SBS. Egf receptor and a subset of its ligands, also drivers of adaptation, were upregulated in SBS fish. Igf has been reported as a driver of intestinal adaptation in other animal models, and SBS fish exposed to a pharmacological inhibitor of the Igf receptor failed to demonstrate signs of intestinal adaptation, such as increased inner epithelial perimeter and BrdU incorporation. We describe a technically feasible model of human SBS in the zebrafish, a faster and less expensive tool to investigate intestinal stem cell plasticity as well as the mechanisms that drive intestinal adaptation.

  3. Enantio-alteration of gene transcription associated with bioconcentration in adult zebrafish (Danio rerio) exposed to chiral PCB149

    Science.gov (United States)

    Chai, Tingting; Cui, Feng; Mu, Pengqian; Yang, Yang; Xu, Nana; Yin, Zhiqiang; Jia, Qi; Yang, Shuming; Qiu, Jing; Wang, Chengju

    2016-01-01

    Enantioselective enrichment of chiral PCB149 (2,2’,3,4’,5’,6-hexachlorobiphenyl) was analysed in adult zebrafish (Danio rerio) exposed to the racemate, (‑)-PCB149, and (+)-PCB149. Greater enrichment of (‑)-PCB149 compared to (+) PCB149 was observed following 0.5 ng/L exposure; however, as the exposure time and concentration increased, racemic enrichment was observed in adult fish exposed to the racemate. No biotransformation between the two isomers was observed in fish exposed to single enantiomers. When zebrafish were exposed to different forms of chiral PCB149, enantioselective expression of genes associated with polychlorinated biphenyls (PCBs) was observed in brain and liver tissues and enantioselective correlations between bioconcentration and target gene expression levels were observed in brain and liver tissues. The strong positive correlations between expression levels of target genes (alox5a and alox12) and PCB149 bioconcentration suggest that prolonged exposure to the racemate of chiral PCB149 may result in inflammation-associated diseases. Prolonged exposure to (‑)-PCB149 may also affect metabolic pathways such as dehydrogenation and methylation in the brain tissues of adult zebrafish. Hepatic expression levels of genes related to the antioxidant system were significantly negatively correlated with bioconcentration following exposure to (+)-PCB149.

  4. Feed and feeding regime affect growth rate and gonadosomatic index of adult zebrafish (Danio rerio).

    Science.gov (United States)

    Gonzales, John M; Law, Sheran Hiu Wan

    2013-12-01

    A 5-week study was conducted to evaluate commercially available Artemia, Ziegler zebrafish diet, and Calamac diet fed in five different feeding regimes on the growth and reproductive development of 7-month-old zebrafish. Zebrafish were fed to satiation three times daily during the normal work week and twice daily during the weekend and holidays. Zebrafish in dietary groups CCC (Calamac three times daily) and CCA (Calamac twice daily, Artemia once daily) had a significantly (pzebrafish in dietary group 5 had significantly larger gonadosomatic index (GSI) values than all other groups, while female zebrafish in dietary group CCC had significantly larger GSI values than all other groups. No differences in the fatty acid content of female gonads were detected. Zebrafish fed solely Artemia had the greatest weight loss and lowest GSI values. Preliminary evidence of protein sparing in zebrafish is reported. Collectively, this study sheds more light into the effects of the use of commercially available feeds and feeding regime on the rearing of zebrafish.

  5. Evolutionary patterns and selective pressures of odorant/pheromone receptor gene families in teleost fishes.

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    Yasuyuki Hashiguchi

    Full Text Available BACKGROUND: Teleost fishes do not have a vomeronasal organ (VNO, and their vomeronasal receptors (V1Rs, V2Rs are expressed in the main olfactory epithelium (MOE, as are odorant receptors (ORs and trace amine-associated receptors (TAARs. In this study, to obtain insights into the functional distinction among the four chemosensory receptor families in teleost fishes, their evolutionary patterns were examined in zebrafish, medaka, stickleback, fugu, and spotted green pufferfish. METHODOLOGY/PRINCIPAL FINDINGS: Phylogenetic analysis revealed that many lineage-specific gene gains and losses occurred in the teleost fish TAARs, whereas only a few gene gains and losses have taken place in the teleost fish vomeronasal receptors. In addition, synonymous and nonsynonymous nucleotide substitution rate ratios (K(A/K(S in TAARs tended to be higher than those in ORs and V2Rs. CONCLUSIONS/SIGNIFICANCE: Frequent gene gains/losses and high K(A/K(S in teleost TAARs suggest that receptors in this family are used for detecting some species-specific chemicals such as pheromones. Conversely, conserved repertoires of V1R and V2R families in teleost fishes may imply that receptors in these families perceive common odorants for teleosts, such as amino acids. Teleost ORs showed intermediate evolutionary pattern between TAARs and vomeronasal receptors. Many teleost ORs seem to be used for common odorants, but some ORs may have evolved to recognize lineage-specific odors.

  6. Neurotoxicity of neem commercial formulation (Azadirachta indica A. Juss) in adult zebrafish (Danio rerio).

    Science.gov (United States)

    Bernardi, M M; Dias, S G; Barbosa, V E

    2013-11-01

    The neurotoxic effects of a commercial formulation of Azadirachta indica A. Juss, also called neem or nim, in adult zebrafish were determined using behavioral models. General activity, anxiety-like effects, and learning and memory in a passive avoidance task were assessed after exposure to 20 or 40 μl/L neem. The results showed that 20 μl/L neem reduced the number of runs. Both neem concentrations increased the number of climbs to the water surface, and 40 μl/L increased the number of tremors. In the anxiety test, the 20 μl/L dose increased the number of entries in the light side compared with controls, but the latency to enter the dark side and the freezing behavior in this side did not changed. In relation to controls, the 40 μl/L neem reduced the latency to enter in the light side, did not change the number of entries in this side and increased freezing behavior in the light side. In the passive avoidance test, pre-training and pre-test neem exposure to 40 μl/L decreased the response to the learning task. Thus, no impairment was observed in this behavioral test. We conclude that neem reduced general activity and increased anxiety-like behavior but did not affect learning and memory.

  7. Transcription of the SCL/TAL1 interrupting Locus (Stil) is required for cell proliferation in adult Zebrafish Retinas.

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    Sun, Lei; Li, Ping; Carr, Aprell L; Gorsuch, Ryne; Yarka, Clare; Li, Jingling; Bartlett, Michael; Pfister, Delaney; Hyde, David R; Li, Lei

    2014-03-07

    The human oncogene SCL/TAL1 interrupting locus (Stil) is highly conserved in vertebrate species. Previously, we identified a homolog of the Stil gene in zebrafish mutant (night blindness b, nbb), which showed neural defects in the retina (e.g. dopaminergic cell degeneration and/or lack of regeneration). In this research, we examined the roles of Stil in cell proliferation after degeneration in adult zebrafish retinas. We demonstrated that knockdown of Stil gene expression or inhibition of Sonic hedgehog (Shh) signaling transduction decreases the rate of cell proliferation. In contrast, activation of Shh signal transduction promotes cell proliferation. In nbb(+/-) retinas, inhibition of SUFU (a repressor in the Shh pathway) rescues the defects in cell proliferation due to down-regulation of Stil gene expression. The latter data suggest that Stil play a role in cell proliferation through the Shh signal transduction pathway.

  8. Transcription of the SCL/TAL1 Interrupting Locus (Stil) Is Required for Cell Proliferation in Adult Zebrafish Retinas*

    Science.gov (United States)

    Sun, Lei; Li, Ping; Carr, Aprell L.; Gorsuch, Ryne; Yarka, Clare; Li, Jingling; Bartlett, Michael; Pfister, Delaney; Hyde, David R.; Li, Lei

    2014-01-01

    The human oncogene SCL/TAL1 interrupting locus (Stil) is highly conserved in vertebrate species. Previously, we identified a homolog of the Stil gene in zebrafish mutant (night blindness b, nbb), which showed neural defects in the retina (e.g. dopaminergic cell degeneration and/or lack of regeneration). In this research, we examined the roles of Stil in cell proliferation after degeneration in adult zebrafish retinas. We demonstrated that knockdown of Stil gene expression or inhibition of Sonic hedgehog (Shh) signaling transduction decreases the rate of cell proliferation. In contrast, activation of Shh signal transduction promotes cell proliferation. In nbb+/− retinas, inhibition of SUFU (a repressor in the Shh pathway) rescues the defects in cell proliferation due to down-regulation of Stil gene expression. The latter data suggest that Stil play a role in cell proliferation through the Shh signal transduction pathway. PMID:24469449

  9. Dynamics of miRNA transcriptome during gonadal development of zebrafish

    Science.gov (United States)

    Presslauer, Christopher; Tilahun Bizuayehu, Teshome; Kopp, Martina; Fernandes, Jorge M. O.; Babiak, Igor

    2017-01-01

    Studies in non-teleost vertebrates have found microRNAs (miRNAs) to be essential for proper gonadal development. However, comparatively little is known about their role during gonadal development in teleost fishes. So far in zebrafish, a model teleost, transcript profiling throughout gonadal development has not been established because of a tiny size of an organ in juvenile stages and its poor distinguishability from surrounding tissues. We performed small RNA sequencing on isolated gonads of See-Thru-Gonad line, from the undifferentiated state at 3 weeks post fertilization (wpf) to fully mature adults at 24 wpf. We identified 520 gonadal mature miRNAs; 111 of them had significant changes in abundance over time, while 50 miRNAs were either testis- or ovary-enriched significantly in at least one developmental stage. We characterized patterns of miRNA abundance over time including isomiR variants. We identified putative germline versus gonadal somatic miRNAs through differential small RNA sequencing of isolated gametes versus the whole gonads. This report is the most comprehensive analysis of the miRNA repertoire in zebrafish gonads during the sexual development to date and provides an important database from which functional studies can be performed. PMID:28262836

  10. Swimming-induced exercise promotes hypertrophy and vascularization of fast skeletal muscle fibres and activation of myogenic and angiogenic transcriptional programs in adult zebrafish

    NARCIS (Netherlands)

    Palstra, A.P.; Rovira, M.; Rizo-Roca, D.; Torrella, J.R.; Spaink, H.P.; Planas, J.V.

    2014-01-01

    Background The adult skeletal muscle is a plastic tissue with a remarkable ability to adapt to different levels of activity by altering its excitability, its contractile and metabolic phenotype and its mass. We previously reported on the potential of adult zebrafish as a tractable experimental model

  11. Midkine-a protein localization in the developing and adult retina of the zebrafish and its function during photoreceptor regeneration.

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    Esther Gramage

    Full Text Available Midkine is a heparin binding growth factor with important functions in neuronal development and survival, but little is known about its function in the retina. Previous studies show that in the developing zebrafish, Midkine-a (Mdka regulates cell cycle kinetics in retinal progenitors, and following injury to the adult zebrafish retina, mdka is strongly upregulated in Müller glia and the injury-induced photoreceptor progenitors. Here we provide the first data describing Mdka protein localization during different stages of retinal development and during the regeneration of photoreceptors in adults. We also experimentally test the role of Mdka during photoreceptor regeneration. The immuno-localization of Mdka reflects the complex spatiotemporal pattern of gene expression and also reveals the apparent secretion and extracellular trafficking of this protein. During embryonic retinal development the Mdka antibodies label all mitotically active cells, but at the onset of neuronal differentiation, immunostaining is also localized to the nascent inner plexiform layer. Starting at five days post fertilization through the juvenile stage, Mdka immunostaining labels the cytoplasm of horizontal cells and the overlying somata of rod photoreceptors. Double immunolabeling shows that in adult horizontal cells, Mdka co-localizes with markers of the Golgi complex. Together, these data are interpreted to show that Mdka is synthesized in horizontal cells and secreted into the outer nuclear layer. In adults, Mdka is also present in the end feet of Müller glia. Similar to mdka gene expression, Mdka in horizontal cells is regulated by circadian rhythms. After the light-induced death of photoreceptors, Mdka immuonolabeling is localized to Müller glia, the intrinsic stem cells of the zebrafish retina, and proliferating photoreceptor progenitors. Knockdown of Mdka during photoreceptor regeneration results in less proliferation and diminished regeneration of rod

  12. Cypermethrin has the potential to induce hepatic oxidative stress, DNA damage and apoptosis in adult zebrafish (Danio rerio).

    Science.gov (United States)

    Jin, Yuanxiang; Zheng, Shanshan; Pu, Yue; Shu, Linjun; Sun, Liwei; Liu, Weiping; Fu, Zhengwei

    2011-01-01

    Cypermethrin (CYP), a widely used Type II pyrethroid pesticide, is one of the most common contaminants in the freshwater aquatic system. We studied the effects of CYP exposure on the induction of hepatic oxidative stress, DNA damage and the alteration of gene expression related to apoptosis in adult zebrafish. Hepatic mRNA levels for the genes encoding antioxidant proteins, such as Cu/Zn-Sod, Mn-Sod, Cat, and Gpx, were significantly upregulated when zebrafish were exposed to various concentrations of CYP for 4 or 8 days. In addition, the main genes related to fatty acid β-oxidation and the mitochondrial genes related to respiration and ATP synthesis were also significantly upregulated after exposure to high concentrations (1 and 3 μg L(-1)) of CYP for 4 or 8 days. Moreover, in a comet assay of zebrafish hepatocytes, tail DNA, tail length, tail moment and Olive tail moment increased in a concentration-dependent manner. The significant induction (pzebrafish. This information will be helpful in fully understanding the mechanism of aquatic toxicology induced by CYP in fish.

  13. Inhibition of phosphorylated tyrosine hydroxylase attenuates ethanol-induced hyperactivity in adult zebrafish (Danio rerio).

    Science.gov (United States)

    Nowicki, Magda; Tran, Steven; Chatterjee, Diptendu; Gerlai, Robert

    2015-11-01

    Zebrafish have been successfully employed in the study of the behavioural and biological effects of ethanol. Like in mammals, low to moderate doses of ethanol induce motor hyperactivity in zebrafish, an effect that has been attributed to the activation of the dopaminergic system. Acute ethanol exposure increases dopamine (DA) in the zebrafish brain, and it has been suggested that tyrosine hydroxylase, the rate-limiting enzyme of DA synthesis, may be activated in response to ethanol via phosphorylation. The current study employed tetrahydropapaveroline (THP), a selective inhibitor of phosphorylated tyrosine hydroxylase, for the first time, in zebrafish. We treated zebrafish with a THP dose that did not alter baseline motor responses to examine whether it can attenuate or abolish the effects of acute exposure to alcohol (ethanol) on motor activity, on levels of DA, and on levels of dopamine's metabolite 3,4-dihydroxyphenylacetic acid (DOPAC). We found that 60-minute exposure to 1% alcohol induced motor hyperactivity and an increase in brain DA. Both of these effects were attenuated by pre-treatment with THP. However, no differences in DOPAC levels were found among the treatment groups. These findings suggest that tyrosine hydroxylase is activated via phosphorylation to increase DA synthesis during alcohol exposure in zebrafish, and this partially mediates alcohol's locomotor stimulant effects. Future studies will investigate other potential candidates in the molecular pathway to further decipher the neurobiological mechanism that underlies the stimulatory properties of this popular psychoactive drug.

  14. Fast gene transfer into the adult zebrafish brain by herpes simplex virus 1 (HSV-1 and electroporation: methods and optogenetic applications

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    Ming eZou

    2014-05-01

    Full Text Available The zebrafish has various advantages as a model organism to analyze the structure and function of neural circuits but efficient viruses or other tools for fast gene transfer are lacking. We show that transgenes can be introduced directly into the adult zebrafish brain by herpes simplex type I viruses (HSV-1 or electroporation. We developed a new procedure to target electroporation to defined brain areas and identified promoters that produced strong long-term expression. The fast workflow of electroporation was exploited to express multiple channelrhodopsin-2 variants and genetically encoded calcium indicators in telencephalic neurons for measurements of neuronal activity and synaptic connectivity. The results demonstrate that HSV-1 and targeted electroporation are efficient tools for gene delivery into the zebrafish brain, similar to adeno-associated viruses and lentiviruses in other species. These methods fill an important gap in the spectrum of molecular tools for zebrafish and are likely to have a wide range of applications.

  15. Monitoring of single-cell responses in the optic tectum of adult zebrafish with dextran-coupled calcium dyes delivered via local electroporation.

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    Vanessa Kassing

    Full Text Available The zebrafish (Danio rerio has become one of the major animal models for in vivo examination of sensory and neuronal computation. Similar to Xenopus tadpoles neural activity in the optic tectum, the major region controlling visually guided behavior, can be examined in zebrafish larvae by optical imaging. Prerequisites of these approaches are usually the transparency of larvae up to a certain age and the use of two-photon microscopy. This principle of fluorescence excitation was necessary to suppress crosstalk between signals from individual neurons, which is a critical issue when using membrane-permeant dyes. This makes the equipment to study neuronal processing costly and limits the approach to the study of larvae. Thus there is lack of knowledge about the properties of neurons in the optic tectum of adult animals. We established a procedure to circumvent these problems, enabling in vivo calcium imaging in the optic tectum of adult zebrafish. Following local application of dextran-coupled dyes single-neuron activity of adult zebrafish can be monitored with conventional widefield microscopy, because dye labeling remains restricted to tens of neurons or less. Among the neurons characterized with our technique we found neurons that were selective for a certain pattern orientation as well as neurons that responded in a direction-selective way to visual motion. These findings are consistent with previous studies and indicate that the functional integrity of neuronal circuits in the optic tectum of adult zebrafish is preserved with our staining technique. Overall, our protocol for in vivo calcium imaging provides a useful approach to monitor visual responses of individual neurons in the optic tectum of adult zebrafish even when only widefield microscopy is available. This approach will help to obtain valuable insight into the principles of visual computation in adult vertebrates and thus complement previous work on developing visual circuits.

  16. Her4-positive population in the tectum opticum is proliferating neural precursors in the adult zebrafish brain.

    Science.gov (United States)

    Jung, Seung-Hyun; Kim, Hyung-Seok; Ryu, Jae-Ho; Gwak, Jung-Woo; Bae, Young-Ki; Kim, Cheol-Hee; Yeo, Sang-Yeob

    2012-06-01

    Previous studies have shown that Notch signaling not only regulates the number of early differentiating neurons, but also maintains proliferating neural precursors in the neural tube. Although it is well known that Notch signaling is closely related to the differentiation of adult neural stem cells, none of transgenic zebrafish provides a tool to figure out the relationship between Notch signaling and the differentiation of neural precursors. The goal of this study was to characterize Her4-positive cells by comparing the expression of a fluorescent Her4 reporter in Tg[her4-dRFP] animals with a GFAP reporter in Tg[gfap-GFP] adult zebrafish. BrdU incorporation indicated that dRFP-positive cells were proliferating and a double labeling assay revealed that a significant fraction of the Her4-dRFP positive population was also GFAP-GFP positive. Our observations suggest that a reporter line with Notch-dependent gene expression can provide a tool to examine proliferating neural precursors and/or neuronal/glial precursors in the development of the adult nervous system to examine the model in which Notch signaling maintains proliferating neural precursors in the neural tube.

  17. CLARITY and PACT-based imaging of adult zebrafish and mouse for whole-animal analysis of infections

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    Mark R. Cronan

    2015-12-01

    Full Text Available Visualization of infection and the associated host response has been challenging in adult vertebrates. Owing to their transparency, zebrafish larvae have been used to directly observe infection in vivo; however, such larvae have not yet developed a functional adaptive immune system. Cells involved in adaptive immunity mature later and have therefore been difficult to access optically in intact animals. Thus, the study of many aspects of vertebrate infection requires dissection of adult organs or ex vivo isolation of immune cells. Recently, CLARITY and PACT (passive clarity technique methodologies have enabled clearing and direct visualization of dissected organs. Here, we show that these techniques can be applied to image host-pathogen interactions directly in whole animals. CLARITY and PACT-based clearing of whole adult zebrafish and Mycobacterium tuberculosis-infected mouse lungs enables imaging of mycobacterial granulomas deep within tissue to a depth of more than 1 mm. Using established transgenic lines, we were able to image normal and pathogenic structures and their surrounding host context at high resolution. We identified the three-dimensional organization of granuloma-associated angiogenesis, an important feature of mycobacterial infection, and characterized the induction of the cytokine tumor necrosis factor (TNF within the granuloma using an established fluorescent reporter line. We observed heterogeneity in TNF induction within granuloma macrophages, consistent with an evolving view of the tuberculous granuloma as a non-uniform, heterogeneous structure. Broad application of this technique will enable new understanding of host-pathogen interactions in situ.

  18. Development of Alginate Microspheres Containing Chuanxiong for Oral Administration to Adult Zebrafish

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    Li-Jen Lin

    2016-01-01

    Full Text Available Oral administration of Traditional Chinese Medicine (TCM by patients is the common way to treat health problems. Zebrafish emerges as an excellent animal model for the pharmacology investigation. However, the oral delivery system of TCM in zebrafish has not been established so far. This issue was addressed by development of alginate microparticles for oral delivery of chuanxiong, a TCM that displays antifibrotic and antiproliferative effects on hepatocytes. The delivery microparticles were prepared from gelification of alginate containing various levels of chuanxiong. The chuanxiong-encapsulated alginate microparticles were characterized for their solubility, structure, encapsulation efficiency, the cargo release profile, and digestion in gastrointestinal tract of zebrafish. Encapsulation of chuanxiong resulted in more compact structure and the smaller size of microparticles. The release rate of chuanxiong increased for alginate microparticles carrying more chuanxiong in simulated intestinal fluid. This remarkable feature ensures the controlled release of encapsulated cargos in the gastrointestinal tract of zebrafish. Moreover, chuanxiong-loaded alginate microparticles were moved to the end of gastrointestinal tract after oral administration for 6 hr and excreted from the body after 16 hr. Therefore, our developed method for oral administration of TCM in zebrafish is useful for easy and rapid evaluation of the drug effect on disease.

  19. A new zebrafish bone crush injury model

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    Sara Sousa

    2012-07-01

    While mammals have a limited capacity to repair bone fractures, zebrafish can completely regenerate amputated bony fin rays. Fin regeneration in teleosts has been studied after partial amputation of the caudal fin, which is not ideal to model human bone fractures because it involves substantial tissue removal, rather than local tissue injury. In this work, we have established a bone crush injury model in zebrafish adult caudal fin, which consists of the precise crush of bony rays with no tissue amputation. Comparing these two injury models, we show that the initial stages of injury response are the same regarding the activation of wound healing molecular markers. However, in the crush assay the expression of the blastema marker msxb appears later than during regeneration after amputation. Following the same trend, bone cells deposition and expression of genes involved in skeletogenesis are also delayed. We further show that bone and blood vessel patterning is also affected. Moreover, analysis of osteopontin and Tenascin-C reveals that they are expressed at later stages in crushed tissue, suggesting that in this case bone repair is prolonged for longer than in the case of regeneration after amputation. Due to the nature of the trauma inflicted, the crush injury model seems more similar to fracture bone repair in mammals than bony ray amputation. Therefore, the new model that we present here may help to identify the key processes that regulate bone fracture and contribute to improve bone repair in humans.

  20. Molecular psychiatry of zebrafish.

    Science.gov (United States)

    Stewart, A M; Ullmann, J F P; Norton, W H J; Parker, M O; Brennan, C H; Gerlai, R; Kalueff, A V

    2015-02-01

    Due to their well-characterized neural development and high genetic homology to mammals, zebrafish (Danio rerio) have emerged as a powerful model organism in the field of biological psychiatry. Here, we discuss the molecular psychiatry of zebrafish, and its implications for translational neuroscience research and modeling central nervous system (CNS) disorders. In particular, we outline recent genetic and technological developments allowing for in vivo examinations, high-throughput screening and whole-brain analyses in larval and adult zebrafish. We also summarize the application of these molecular techniques to the understanding of neuropsychiatric disease, outlining the potential of zebrafish for modeling complex brain disorders, including attention-deficit/hyperactivity disorder (ADHD), aggression, post-traumatic stress and substance abuse. Critically evaluating the advantages and limitations of larval and adult fish tests, we suggest that zebrafish models become a rapidly emerging new field in modern molecular psychiatry research.

  1. Identification and functional evidence of GABAergic neurons in parts of the brain of adult zebrafish (Danio rerio).

    Science.gov (United States)

    Kim, Yong-Jung; Nam, Ryoung-Hee; Yoo, Young Mi; Lee, Chang-Joong

    2004-01-23

    The distribution of GABA-containing neurons was studied in the brain of the adult zebrafish by Nissl staining and immunohistochemistry. GABA immunoreactivity (GABA-IR) was demonstrated in parts of the brain such as olfactory bulb (OB), telencephalon, tectum stratum, and in the hypothalamus. GABA-IR appeared in the area where Nissl-stained cell bodies were abundant. The internal cellular layer of the OB was most densely stained by Nissl staining, and also showed a high level of GABA-IR. The telencephalon and the hypothalamus revealed a similar pattern to the OB in terms of Nissl staining and GABA-IR. However, the distribution and shape of stained cells of the tectum stratum were distinct from those in other regions: Nissl-stained neurons were ubiquitously present throughout all cellular layers including the stratum griseum centrale, the stratum album centrale (SAC), and the stratum periventriculare (SP). However, GABA-IR was weakly expressed in a limited number of neurons only in the SAC and SP. Whether GABA serves as an inhibitory neurotransmitter was also tested in the isolated telencephalon preparation by using extracellular field potential recordings. The synaptic activity recorded in the posterior dorsal telencephalon in response to the electrical stimulation of the anterior dorsal telencephalon was increased in the presence of the GABAA receptor antagonist, BMI, suggesting an inhibitory role for GABA-immunoreactive neurons in the adult brain of the zebrafish.

  2. Subdivisions of the adult zebrafish pallium based on molecular marker analysis [v1; ref status: indexed, http://f1000r.es/4m2

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    Julia Ganz

    2014-12-01

    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

  3. The odorant receptor repertoire of teleost fish

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    Alioto Tyler S

    2005-12-01

    Full Text Available Abstract Background Vertebrate odorant receptors comprise three types of G protein-coupled receptors: the OR, V1R and V2R receptors. The OR superfamily contains over 1,000 genes in some mammalian species, representing the largest gene superfamily in the mammalian genome. Results To facilitate an informed analysis of OR gene phylogeny, we identified the complete set of 143 OR genes in the zebrafish genome, as well as the OR repertoires in two pufferfish species, fugu (44 genes and tetraodon (42 genes. Although the genomes analyzed here contain fewer genes than in mammalian species, the teleost OR genes can be grouped into a larger number of major clades, representing greater overall OR diversity in the fish. Conclusion Based on the phylogeny of fish and mammalian repertoires, we propose a model for OR gene evolution in which different ancestral OR genes or gene families were selectively lost or expanded in different vertebrate lineages. In addition, our calculations of the ratios of non-synonymous to synonymous codon substitutions among more recently expanding OR subgroups in zebrafish implicate residues that may be involved in odorant binding.

  4. In vivo spectroscopic photoacoustic tomography imaging of a far red fluorescent protein expressed in the exocrine pancreas of adult zebrafish

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    Liu, Mengyang; Schmitner, Nicole; Sandrian, Michelle G.; Zabihian, Behrooz; Hermann, Boris; Salvenmoser, Willi; Meyer, Dirk; Drexler, Wolfgang

    2014-03-01

    Fluorescent proteins brought a revolution in life sciences and biological research in that they make a powerful tool for researchers to study not only the structural and morphological information, but also dynamic and functional information in living cells and organisms. While green fluorescent proteins (GFP) have become a common labeling tool, red-shifted or even near infrared fluorescent proteins are becoming the research focus due to the fact that longer excitation wavelengths are more suitable for deep tissue imaging. In this study, E2-Crimson, a far red fluorescent protein whose excitation wavelength is 611 nm, was genetically expressed in the exocrine pancreas of adult zebrafish. Using spectroscopic all optical detection photoacoustic tomography, we mapped the distribution of E2-Crimson in 3D after imaging the transgenic zebrafish in vivo using two different wavelengths. With complementary morphological information provided by imaging the same fish using a spectral domain optical coherence tomography system, the E2-Crimson distribution acquired from spectroscopic photoacoustic tomography was confirmed in 2D by epifluorescence microscopy and in 3D by histology. To the authors' knowledge, this is the first time a far red fluorescent protein is imaged in vivo by spectroscopic photoacoustic tomography. Due to the regeneration feature of zebrafish pancreas, this work preludes the longitudinal studies of animal models of diseases such as pancreatitis by spectroscopic photoacoustic tomography. Since the effective penetration depth of photoacoustic tomography is beyond the transport mean free path length, other E2-Crimson labeled inner organs will also be able to be studied dynamically using spectroscopic photoacoustic tomography.

  5. Melanophore migration and survival during zebrafish adult pigment stripe development require the immunoglobulin superfamily adhesion molecule Igsf11.

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    Dae Seok Eom

    Full Text Available The zebrafish adult pigment pattern has emerged as a useful model for understanding the development and evolution of adult form as well as pattern-forming mechanisms more generally. In this species, a series of horizontal melanophore stripes arises during the larval-to-adult transformation, but the genetic and cellular bases for stripe formation remain largely unknown. Here, we show that the seurat mutant phenotype, consisting of an irregular spotted pattern, arises from lesions in the gene encoding Immunoglobulin superfamily member 11 (Igsf11. We find that Igsf11 is expressed by melanophores and their precursors, and we demonstrate by cell transplantation and genetic rescue that igsf11 functions autonomously to this lineage in promoting adult stripe development. Further analyses of cell behaviors in vitro, in vivo, and in explant cultures ex vivo demonstrate that Igsf11 mediates adhesive interactions and that mutants for igsf11 exhibit defects in both the migration and survival of melanophores and their precursors. These findings identify the first in vivo requirements for igsf11 as well as the first instance of an immunoglobulin superfamily member functioning in pigment cell development and patterning. Our results provide new insights into adult pigment pattern morphogenesis and how cellular interactions mediate pattern formation.

  6. Distribution of cannabinoid receptor 1 in the CNS of zebrafish.

    Science.gov (United States)

    Lam, C S; Rastegar, S; Strähle, U

    2006-01-01

    The cannabinoid receptor 1 (Cb1) mediates the psychoactive effect of marijuana. In mammals, there is abundant evidence advocating the importance of cannabinoid signaling; activation of Cb1 exerts diverse functions, chiefly by its ability to modulate neurotransmission. Thus, much attention has been devoted to understand its role in health and disease and to evaluate its therapeutic potential. Here, we have cloned zebrafish cb1 and investigated its expression in developing and adult zebrafish brain. Sequence analysis showed that there is a high degree of conservation, especially in residues demonstrated to be critical for function in mammals. In situ hybridization revealed that zebrafish cb1 appears first in the preoptic area at 24 hours post-fertilization. Subsequently, transcripts are detected in the dorsal telencephalon, hypothalamus, pretectum and torus longitudinalis. A similar pattern of expression is recapitulated in the adult brain. While cb1 is intensively stained in the medial zone of the dorsal telencephalon, expression elsewhere is weak by comparison. In particular, localization of cb1 in the telencephalic periventricular matrix is suggestive of the involvement of Cb1 in neurogenesis, bearing strong resemblance in terms of expression and function to the proliferative mammalian hippocampal formation. In addition, a gradient-like expression of cb1 is detected in the torus longitudinalis, a teleost specific neural tissue. In relation to dopaminergic neurons in the diencephalic posterior tuberculum (considered to be the teleostean homologue of the mammalian midbrain dopaminergic system), both cb1 and tyrosine hydroxylase-expressing cells occupy non-overlapping domains. However there is evidence that they are co-localized in the caudal zone of the hypothalamus, implying a direct modulation of dopamine release in this particular region. Collectively, our data indicate the propensity of zebrafish cb1 to participate in multiple neurological processes.

  7. Effect of triazophos, fipronil and their mixture on miRNA expression in adult zebrafish.

    Science.gov (United States)

    Wang, Xingxing; Zhou, Shengli; Ding, Xianfeng; Zhu, Guonian; Guo, Jiangfeng

    2010-10-01

    MicroRNA (miRNA) plays a crucial role in gene expression regulation. However, no data are available on change of miRNA expression of zebrafish (Danio rerio) after treatment with pesticides. We evaluated the effect of fipronil (5-amino-1-[2, 6-dichloro-4-(trifluoromethyl) phenyl]-4-[(trifluoromethyl) sulfinyl]-1H-pyrazole-3-carbonitrile) and triazophos (3-(O, O-diethyl)-1-phenyl thiophosphoryl-1, 2, 4-triazol) and their mixture on miRNA expression in zebrafish.MiRNA expression profiles in zebrafish were altered after treatment with these chemicals. An association between these chemicals and the expression of 21 miRNAs was found 96 h after treatment. Among them, 14 miRNAs were differentially expressed due to the treatments with fipronil, triazophos and their mixture; 5 miRNAs showed altered expression level after treatment with formulations of these chemicals; miR-29b and miR-738 were differentially expressed after treatment with adjuvants. MiRNAs might present a novel toxicological response that could be used as a toxicological biomarker and have a different direction for future investigations of their association with miRNAs involved in chemical related diseases.

  8. Zebrafish as a model for investigating developmental lead (Pb) neurotoxicity as a risk factor in adult neurodegenerative disease: a mini-review.

    Science.gov (United States)

    Lee, Jinyoung; Freeman, Jennifer L

    2014-07-01

    Lead (Pb) exposure has long been recognized to cause neurological alterations in both adults and children. While most of the studies in adults are related to higher dose exposure, epidemiological studies indicate cognitive decline and neurobehavioral alterations in children associated with lower dose environmental Pb exposure (a blood Pb level of 10μg/dL and below). Recent animal studies also now report that an early-life Pb exposure results in pathological hallmarks of Alzheimer's disease later in life. While previous studies evaluating higher Pb exposures in adult animal models and higher occupational Pb exposures in humans have suggested a link between higher dose Pb exposure during adulthood and neurodegenerative disease, these newer studies now indicate a link between an early-life Pb exposure and adult neurodegenerative disease. These studies are supporting the "fetal/developmental origin of adult disease" hypothesis and present a new challenge in our understanding of Pb neurotoxicity. There is a need to expand research in this area and additional model systems are needed. The zebrafish presents as a complementary vertebrate model system with numerous strengths including high genetic homology. Several zebrafish genes orthologous to human genes associated with neurodegenerative diseases including Alzheimer's and Parkinson's diseases are identified and this model is starting to be applied in neurodegenerative disease research. Moreover, the zebrafish is being used in developmental Pb neurotoxicity studies to define genetic mechanisms of toxicity and associated neurobehavioral alterations. While these studies are in their infancy, the genetic and functional conservation of genes associated with neurodegenerative diseases and application in developmental Pb neurotoxicity studies supports the potential for this in vivo model to further investigate the link between developmental Pb exposure and adult neurodegenerative disease pathogenesis. In this review, the

  9. Transcriptomic analyses reveal novel genes with sexually dimorphic expression in the zebrafish gonad and brain.

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    Rajini Sreenivasan

    Full Text Available BACKGROUND: Our knowledge on zebrafish reproduction is very limited. We generated a gonad-derived cDNA microarray from zebrafish and used it to analyze large-scale gene expression profiles in adult gonads and other organs. METHODOLOGY/PRINCIPAL FINDINGS: We have identified 116638 gonad-derived zebrafish expressed sequence tags (ESTs, 21% of which were isolated in our lab. Following in silico normalization, we constructed a gonad-derived microarray comprising 6370 unique, full-length cDNAs from differentiating and adult gonads. Labeled targets from adult gonad, brain, kidney and 'rest-of-body' from both sexes were hybridized onto the microarray. Our analyses revealed 1366, 881 and 656 differentially expressed transcripts (34.7% novel that showed highest expression in ovary, testis and both gonads respectively. Hierarchical clustering showed correlation of the two gonadal transcriptomes and their similarities to those of the brains. In addition, we have identified 276 genes showing sexually dimorphic expression both between the brains and between the gonads. By in situ hybridization, we showed that the gonadal transcripts with the strongest array signal intensities were germline-expressed. We found that five members of the GTP-binding septin gene family, from which only one member (septin 4 has previously been implicated in reproduction in mice, were all strongly expressed in the gonads. CONCLUSIONS/SIGNIFICANCE: We have generated a gonad-derived zebrafish cDNA microarray and demonstrated its usefulness in identifying genes with sexually dimorphic co-expression in both the gonads and the brains. We have also provided the first evidence of large-scale differential gene expression between female and male brains of a teleost. Our microarray would be useful for studying gonad development, differentiation and function not only in zebrafish but also in related teleosts via cross-species hybridizations. Since several genes have been shown to play similar

  10. Effects of chronic dietary selenomethionine exposure on repeat swimming performance, aerobic metabolism and methionine catabolism in adult zebrafish (Danio rerio).

    Science.gov (United States)

    Thomas, Jith K; Wiseman, Steve; Giesy, John P; Janz, David M

    2013-04-15

    In a previous study we reported impaired swimming performance and greater stored energy in adult zebrafish (Danio rerio) after chronic dietary exposure to selenomethionine (SeMet). The goal of the present study was to further investigate effects of chronic exposure to dietary SeMet on repeat swimming performance, oxygen consumption (MO2), metabolic capacities (standard metabolic rate [SMR], active metabolic rate [AMR], factorial aerobic scope [F-AS] and cost of transport [COT]) and gene expression of energy metabolism and methionine catabolism enzymes in adult zebrafish. Fish were fed SeMet at measured concentrations of 1.3, 3.4, 9.8 or 27.5 μg Se/g dry mass (d.m.) for 90 d. At the end of the exposure period, fish from each treatment group were divided into three subgroups: (a) no swim, (b) swim, and (c) repeat swim. Fish from the no swim group were euthanized immediately at 90 d and whole body triglycerides, glycogen and lactate, and gene expression of energy metabolism and methionine catabolism enzymes were determined. Individual fish from the swim group were placed in a swim tunnel respirometer and swimming performance was assessed by determining the critical swimming speed (U(crit)). After both Ucrit and MO2 analyses, fish were euthanized and whole body energy stores and lactate were determined. Similarly, individual fish from the repeat swim group were subjected to two U(crit) tests (U(crit-1) and U(crit-2)) performed with a 60 min recovery period between tests, followed by determination of energy stores and lactate. Impaired swim performance was observed in fish fed SeMet at concentrations greater than 3 μg Se/g in the diet. However, within each dietary Se treatment group, no significant differences between single and repeat U(crits) were observed. Oxygen consumption, SMR and COT were significantly greater, and F-AS was significantly lesser, in fish fed SeMet. Whole body triglycerides were proportional to the concentration of SeMet in the diet. While

  11. Organ-Specific and Size-Dependent Ag Nanoparticle Toxicity in Gills and Intestines of Adult Zebrafish.

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    Osborne, Olivia J; Lin, Sijie; Chang, Chong Hyun; Ji, Zhaoxia; Yu, Xuechen; Wang, Xiang; Lin, Shuo; Xia, Tian; Nel, André E

    2015-10-27

    We studied adult zebrafish to determine whether the size of 20 and 110 nm citrate-coated silver nanoparticles (AgC NPs) differentially impact the gills and intestines, known target organs for Ag toxicity in fish. Following exposure for 4 h, 4 days, or 4 days plus a 7 day depuration period, we obtained different toxicokinetic profiles for different particle sizes, as determined by Ag content of the tissues. Ionic AgNO3 served as a positive control. The gills showed a significantly higher Ag content for the 20 nm particles at 4 h and 4 days than the 110 nm particles, while the values were more similar in the intestines. Both particle types were retained in the intestines even after depuration. These toxicokinetics were accompanied by striking size-dependent differences in the ultrastructural features and histopathology in the target organs in response to the particulates. Ag staining of the gills and intestines confirmed prominent Ag deposition in the basolateral membranes for the 20 nm but not for the 110 nm particles. Furthermore, it was possible to link the site of tissue deposition to disruption of the Na(+)/K(+) ion channel, which is also localized to the basolateral membrane. This was confirmed by a reduction in ATPase activity and immunohistochemical detection of the α subunit of this channel in both target organs, with the 20 nm particles causing significantly higher inhibition and disruption than the larger size particles or AgNO3. These results demonstrate the importance of particle size in determining the hazardous impact of AgNPs in the gills and intestines of adult zebrafish.

  12. Myosin VIIA is a marker for the cone accessory outer segment in zebrafish

    NARCIS (Netherlands)

    Hodel, Corinne; Niklaus, Stephanie; Heidemann, Martina; Klooster, Jan; Kamermans, M.; Biehlmaier, Oliver; Gesemann, Matthias; Neuhauss, Stephan C F

    2014-01-01

    The accessory outer segment, a cytoplasmic structure running alongside the photoreceptor outer segment, has been described in teleost fishes, excluding the model organism zebrafish. So far, the function of the accessory outer segment is unknown. Here, we describe the ultrastructure of the zebrafish

  13. Zebrafish hox clusters and vertebrate genome evolution.

    Science.gov (United States)

    Amores, A; Force, A; Yan, Y L; Joly, L; Amemiya, C; Fritz, A; Ho, R K; Langeland, J; Prince, V; Wang, Y L; Westerfield, M; Ekker, M; Postlethwait, J H

    1998-11-27

    HOX genes specify cell fate in the anterior-posterior axis of animal embryos. Invertebrate chordates have one HOX cluster, but mammals have four, suggesting that cluster duplication facilitated the evolution of vertebrate body plans. This report shows that zebrafish have seven hox clusters. Phylogenetic analysis and genetic mapping suggest a chromosome doubling event, probably by whole genome duplication, after the divergence of ray-finned and lobe-finned fishes but before the teleost radiation. Thus, teleosts, the most species-rich group of vertebrates, appear to have more copies of these developmental regulatory genes than do mammals, despite less complexity in the anterior-posterior axis.

  14. Genome duplication, subfunction partitioning, and lineage divergence: Sox9 in stickleback and zebrafish.

    Science.gov (United States)

    Cresko, William A; Yan, Yi-Lin; Baltrus, David A; Amores, Angel; Singer, Amy; Rodríguez-Marí, Adriana; Postlethwait, John H

    2003-11-01

    Teleosts are the most species-rich group of vertebrates, and a genome duplication (tetraploidization) event in ray-fin fish appears to have preceded this remarkable explosion of biodiversity. What is the relationship of the ray-fin genome duplication to the teleost radiation? Genome duplication may have facilitated lineage divergence by partitioning different ancestral gene subfunctions among co-orthologs of tetrapod genes in different teleost lineages. To test this hypothesis, we investigated gene expression patterns for Sox9 gene duplicates in stickleback and zebrafish, teleosts whose lineages diverged early in Euteleost evolution. Most expression domains appear to have been partitioned between Sox9a and Sox9b before the divergence of stickleback and zebrafish lineages, but some ancestral expression domains were distributed differentially in each lineage. We conclude that some gene subfunctions, as represented by lineage-specific expression domains, may have assorted differently in separate lineages and that these may have contributed to lineage diversification during teleost evolution.

  15. Caffeine protects against memory loss induced by high and non-anxiolytic dose of cannabidiol in adult zebrafish (Danio rerio).

    Science.gov (United States)

    Nazario, Luiza Reali; Antonioli, Régis; Capiotti, Katiucia Marques; Hallak, Jaime Eduardo Cecílio; Zuardi, Antonio Waldo; Crippa, José Alexandre S; Bonan, Carla Denise; da Silva, Rosane Souza

    2015-08-01

    Cannabidiol (CBD) has been investigated in a wide spectrum of clinical approaches due to its psychopharmacological properties. CBD has low affinity for cannabinoid neuroreceptors and agonistic properties to 5-HT receptors. An interaction between cannabinoid and purinergic receptor systems has been proposed. The purpose of this study is to evaluate CBD properties on memory behavioral and locomotor parameters and the effects of pre-treatment of adenosine receptor blockers on CBD impacts on memory using adult zebrafish. CBD (0.1, 0.5, 5, and 10mg/kg) was tested in the avoidance inhibitory paradigm and anxiety task. We analyzed the effect of a long-term caffeine pre-treatment (~20mg/L - four months). Also, acute block of adenosine receptors was performed in co-administration with CBD exposure in the memory assessment. CBD promoted an inverted U-shaped dose-response curve in the anxiety task; in the memory assessment, CBD in the dose of 5mg/Kg promoted the strongest effects without interfering with social and aggressive behavior. Caffeine treatment was able to prevent CBD (5mg/kg) effects on memory when CBD was given after the training session. CBD effects on memory were partially prevented by co-treatment with a specific A2A adenosine receptor antagonist when given prior to or after the training session, while CBD effects after the training session were fully prevented by adenosine A1 receptor antagonist. These results indicated that zebrafish have responses to CBD anxiolytic properties that are comparable to other animal models, and high doses changed memory retention in a way dependent on adenosine.

  16. Strategies of vertebrate neurulation and a re-evaluation of teleost neural tube formation.

    Science.gov (United States)

    Lowery, Laura Anne; Sive, Hazel

    2004-10-01

    The vertebrate neural tube develops by two distinct mechanisms. Anteriorly, in the brain and future trunk (cervicothoracic) region, 'primary neurulation' occurs, where an epithelial sheet rolls or bends into a tube. Posteriorly, in the future lumbar and tail region, the neural tube forms by 'secondary neurulation', where a mesenchymal cell population condenses to form a solid rod that undergoes transformation to an epithelial tube. Teleost neurulation has been described as different from that of other vertebrates. This is principally because the teleost trunk neural tube initially forms a solid rod (the neural keel) that later develops a lumen. This process has also been termed secondary neurulation. However, this description is not accurate since the teleost neural tube derives from an epithelial sheet that folds. This best fits the description of primary neurulation. It has also been suggested that teleost neurulation is primitive, however, both primary and secondary neurulation are found in groups with a more ancient origin than the teleosts. The similarity between neurulation in teleosts and other vertebrates indicates that this group includes viable models (such as the zebrafish) for understanding human neural tube development.

  17. 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

  18. Neurochemical measurements in the zebrafish brain

    OpenAIRE

    Lauren eJones; James eMcCutcheon; Andrew eYoung; William eNorton

    2015-01-01

    The zebrafish is an ideal model organism for behavioural genetics and neuroscience. The high conservation of genes and neurotransmitter pathways between zebrafish and other vertebrates permits the translation of research between species. Zebrafish behaviour can be studied at both larval and adult stages and recent research has begun to establish zebrafish models for human disease. Fast scan cyclic voltammetry (FSCV) is an electrochemical technique that permits the detection of neurotransmitte...

  19. Calcium-activated potassium channel SK1 is widely expressed in the peripheral nervous system and sensory organs of adult zebrafish.

    Science.gov (United States)

    Cabo, R; Zichichi, R; Viña, E; Guerrera, M C; Vázquez, G; García-Suárez, O; Vega, J A; Germanà, A

    2013-10-25

    Sensory cells contain ion channels involved in the organ-specific transduction mechanisms that convert different types of stimuli into electric energy. Here we focus on small-conductance calcium-activated potassium channel 1 (SK1) which plays an important role in all excitable cells acting as feedback regulators in after-hyperpolarization. This study was undertaken to analyze the pattern of expression of SK1 in the zebrafish peripheral nervous system and sensory organs using RT-PRC, Westernblot and immunohistochemistry. Expression of SK1 mRNA was observed at all developmental stages analyzed (from 10 to 100 days post fertilization, dpf), and the antibody used identified a protein with a molecular weight of 70kDa, at 100dpf (regarded to be adult). Cell expressing SK1 in adult animals were neurons of dorsal root and cranial nerve sensory ganglia, sympathetic neurons, sensory cells in neuromasts of the lateral line system and taste buds, crypt olfactory neurons and photoreceptors. Present results report for the first time the expression and the distribution of SK1 in the peripheral nervous system and sensory organs of adult zebrafish, and may contribute to set zebrafish as an interesting experimental model for calcium-activated potassium channels research. Moreover these findings are of potential interest because the potential role of SK as targets for the treatment of neurological diseases and sensory disorders.

  20. The MHC class I genes of zebrafish.

    Science.gov (United States)

    Dirscherl, Hayley; McConnell, Sean C; Yoder, Jeffrey A; de Jong, Jill L O

    2014-09-01

    Major histocompatibility complex (MHC) molecules play a central role in the immune response and in the recognition of non-self. Found in all jawed vertebrate species, including zebrafish and other teleosts, MHC genes are considered the most polymorphic of all genes. In this review we focus on the multi-faceted diversity of zebrafish MHC class I genes, which are classified into three sequence lineages: U, Z, and L. We examine the polygenic, polymorphic, and haplotypic diversity of the zebrafish MHC class I genes, discussing known and postulated functional differences between the different class I lineages. In addition, we provide the first comprehensive nomenclature for the L lineage genes in zebrafish, encompassing at least 15 genes, and characterize their sequence properties. Finally, we discuss how recent findings have shed new light on the remarkably diverse MHC loci of this species.

  1. Duplication of the dystroglycan gene in most branches of teleost fish

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    Giardina Bruno

    2007-05-01

    Full Text Available Abstract Background The dystroglycan (DG complex is a major non-integrin cell adhesion system whose multiple biological roles involve, among others, skeletal muscle stability, embryonic development and synapse maturation. DG is composed of two subunits: α-DG, extracellular and highly glycosylated, and the transmembrane β-DG, linking the cytoskeleton to the surrounding basement membrane in a wide variety of tissues. A single copy of the DG gene (DAG1 has been identified so far in humans and other mammals, encoding for a precursor protein which is post-translationally cleaved to liberate the two DG subunits. Similarly, D. rerio (zebrafish seems to have a single copy of DAG1, whose removal was shown to cause a severe dystrophic phenotype in adult animals, although it is known that during evolution, due to a whole genome duplication (WGD event, many teleost fish acquired multiple copies of several genes (paralogues. Results Data mining of pufferfish (T. nigroviridis and T. rubripes and other teleost fish (O. latipes and G. aculeatus available nucleotide sequences revealed the presence of two functional paralogous DG sequences. RT-PCR analysis proved that both the DG sequences are transcribed in T. nigroviridis. One of the two DG sequences harbours an additional mini-intronic sequence, 137 bp long, interrupting the uncomplicated exon-intron-exon pattern displayed by DAG1 in mammals and D. rerio. A similar scenario emerged also in D. labrax (sea bass, from whose genome we have cloned and sequenced a new DG sequence that also harbours a shorter additional intronic sequence of 116 bp. Western blot analysis confirmed the presence of DG protein products in all the species analysed including two teleost Antarctic species (T. bernacchii and C. hamatus. Conclusion Our evolutionary analysis has shown that the whole-genome duplication event in the Class Actinopterygii (ray-finned fish involved also DAG1. We unravelled new important molecular genetic details

  2. Sequencing and comparative analysis of fugu protocadherin clusters reveal diversity of protocadherin genes among teleosts

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    Rajasegaran Vikneswari

    2007-03-01

    Full Text Available Abstract Background The synaptic cell adhesion molecules, protocadherins, are a vertebrate innovation that accompanied the emergence of the neural tube and the elaborate central nervous system. In mammals, the protocadherins are encoded by three closely-linked clusters (α, β and γ of tandem genes and are hypothesized to provide a molecular code for specifying the remarkably-diverse neural connections in the central nervous system. Like mammals, the coelacanth, a lobe-finned fish, contains a single protocadherin locus, also arranged into α, β and γ clusters. Zebrafish, however, possesses two protocadherin loci that contain more than twice the number of genes as the coelacanth, but arranged only into α and γ clusters. To gain further insight into the evolutionary history of protocadherin clusters, we have sequenced and analyzed protocadherin clusters from the compact genome of the pufferfish, Fugu rubripes. Results Fugu contains two unlinked protocadherin loci, Pcdh1 and Pcdh2, that collectively consist of at least 77 genes. The fugu Pcdh1 locus has been subject to extensive degeneration, resulting in the complete loss of Pcdh1γ cluster. The fugu Pcdh genes have undergone lineage-specific regional gene conversion processes that have resulted in a remarkable regional sequence homogenization among paralogs in the same subcluster. Phylogenetic analyses show that most protocadherin genes are orthologous between fugu and zebrafish either individually or as paralog groups. Based on the inferred phylogenetic relationships of fugu and zebrafish genes, we have reconstructed the evolutionary history of protocadherin clusters in the teleost fish lineage. Conclusion Our results demonstrate the exceptional evolutionary dynamism of protocadherin genes in vertebrates in general, and in teleost fishes in particular. Besides the 'fish-specific' whole genome duplication, the evolution of protocadherin genes in teleost fishes is influenced by lineage

  3. Intraperitoneal Exposure to Nano/Microparticles of Fullerene (C60 Increases Acetylcholinesterase Activity and Lipid Peroxidation in Adult Zebrafish (Danio rerio Brain

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    Gonzalo Ogliari Dal Forno

    2013-01-01

    Full Text Available Even though technologies involving nano/microparticles have great potential, it is crucial to determine possible toxicity of these technological products before extensive use. Fullerenes C60 are nanomaterials with unique physicochemical and biological properties that are important for the development of many technological applications. The aim of this study was to evaluate the consequences of nonphotoexcited fullerene C60 exposure in brain acetylcholinesterase expression and activity, antioxidant responses, and oxidative damage using adult zebrafish as an animal model. None of the doses tested (7.5, 15, and 30 mg/kg altered AChE activity, antioxidant responses, and oxidative damage when zebrafish were exposed to nonphotoexcited C60 nano/microparticles during 6 and 12 hours. However, adult zebrafish exposed to the 30 mg/kg dose for 24 hours have shown enhanced AChE activity and augmented lipid peroxidation (TBARS assays in brain. In addition, the up-regulation of brain AChE activity was neither related to the transcriptional control (RT-qPCR analysis nor to the direct action of nonphotoexcited C60 nano/microparticles on the protein (in vitro results but probably involved a posttranscriptional or posttranslational modulation of this enzymatic activity. Taken together these findings provided further evidence of toxic effects on brain after C60 exposure.

  4. The Zebrafish Neurophenome Database (ZND): a dynamic open-access resource for zebrafish neurophenotypic data.

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    Kyzar, Evan; Zapolsky, Ivan; Green, Jeremy; Gaikwad, Siddharth; Pham, Mimi; Collins, Christopher; Roth, Andrew; Stewart, Adam Michael; St-Pierre, Paul; Hirons, Budd; Kalueff, Allan V

    2012-03-01

    Zebrafish (Danio rerio) are widely used in neuroscience research, where their utility as a model organism is rapidly expanding. Low cost, ease of experimental manipulations, and sufficient behavioral complexity make zebrafish a valuable tool for high-throughput studies in biomedicine. To complement the available repositories for zebrafish genetic information, there is a growing need for the collection of zebrafish neurobehavioral and neurological phenotypes. For this, we are establishing the Zebrafish Neurophenome Database (ZND; www.tulane.edu/∼znpindex/search ) as a new dynamic online open-access data repository for behavioral and related physiological data. ZND, currently focusing on adult zebrafish, combines zebrafish neurophenotypic data with a simple, easily searchable user interface, which allow scientists to view and compare results obtained by other laboratories using various treatments in different testing paradigms. As a developing community effort, ZND is expected to foster innovative research using zebrafish by federating the growing body of zebrafish neurophenotypic data.

  5. Notch regulates blastema proliferation and prevents differentiation during adult zebrafish fin regeneration.

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    Münch, Juliane; González-Rajal, Alvaro; de la Pompa, José Luis

    2013-04-01

    Zebrafish have the capacity to regenerate several organs, including the heart and fins. Fin regeneration is epimorphic, involving the formation at the amputation plane of a mass of undifferentiated, proliferating mesenchymal progenitor-like cells, called blastema. This tissue provides all the cell types that form the fin, so that after damage or amputation the fin pattern and structure are fully restored. How blastema cells remain in this progenitor-like state is poorly understood. Here, we show that the Notch pathway plays an essential role during fin regeneration. Notch signalling is activated during blastema formation and remains active throughout the regeneration process. Chemical inhibition or morpholino-mediated knockdown of Notch signalling impairs fin regeneration via decreased proliferation accompanied by reduced expression of Notch target genes in the blastema. Conversely, overexpression of a constitutively active form of the Notch1 receptor (N1ICD) in the regenerating fin leads to increased proliferation and to the expansion of the blastema cell markers msxe and msxb, as well as increased expression of the proliferation regulator aldh1a2. This blastema expansion prevents regenerative fin outgrowth, as indicated by the reduction in differentiating osteoblasts and the inhibition of bone regeneration. We conclude that Notch signalling maintains blastema cells in a plastic, undifferentiated and proliferative state, an essential requirement for fin regeneration.

  6. Lactobacillus plantarum attenuates anxiety-related behavior and protects against stress-induced dysbiosis in adult zebrafish

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    Davis, Daniel J.; Doerr, Holly M.; Grzelak, Agata K.; Busi, Susheel B.; Jasarevic, Eldin; Ericsson, Aaron C.; Bryda, Elizabeth C.

    2016-01-01

    The consumption of probiotics has become increasingly popular as a means to try to improve health and well-being. Not only are probiotics considered beneficial to digestive health, but increasing evidence suggests direct and indirect interactions between gut microbiota (GM) and the central nervous system (CNS). Here, adult zebrafish were supplemented with Lactobacillus plantarum to determine the effects of probiotic treatment on structural and functional changes of the GM, as well as host neurological and behavioral changes. L. plantarum administration altered the β-diversity of the GM while leaving the major core architecture intact. These minor structural changes were accompanied by significant enrichment of several predicted metabolic pathways. In addition to GM modifications, L. plantarum treatment also significantly reduced anxiety-related behavior and altered GABAergic and serotonergic signaling in the brain. Lastly, L. plantarum supplementation provided protection against stress-induced dysbiosis of the GM. These results underscore the influence commensal microbes have on physiological function in the host, and demonstrate bidirectional communication between the GM and the host. PMID:27641717

  7. Levels of 17beta-estradiol receptors expressed in embryonic and adult zebrafish following in vivo treatment of natural or synthetic ligands.

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    Gayathri Chandrasekar

    Full Text Available The nuclear receptors encompass a group of regulatory proteins involved in a number of physiological processes. The estrogen receptors (ERs, of which one alpha and one beta form exist in mammals function as transcription factors in response to 17beta-estradiol (E2. In zebrafish there are three gene products of estrogen receptors and they are denoted esr1 (ERalpha, esr2a (ERbeta2 and esr2b (ERbeta1. Total RNA of zebrafish early life stages (<3, 6, 12, 24, 48, 72, 96 and 120 hours post fertilization and of adult fish (liver, intestine, eye, heart, brain, ovary, testis, gill, swim bladder and kidney were isolated following in vivo exposures. Using specific primers for each of the three zebrafish ERs the expression levels were quantified using real time PCR methodology. It was shown that in absence of exposure all three estrogen receptors were expressed in adult fish. The levels of expression of two of these three ER genes, the esr1 and esr2a were altered in organs such as liver, intestine, brain and testis in response to ligand (E2, diethylstilbestrol or 4-nonylphenol. During embryogenesis two of the three receptor genes, esr1 and esr2b were expressed, and in presence of ligand the mRNA levels of these two genes increased. The conclusions are i estrogen receptor genes are expressed during early development ii altered expression of esr genes in response to ligand is dependent on the cellular context; iii the estrogenic ligand 4-nonylphenol, a manufactured compound commonly found in sewage of water treatment plants, acts as an agonist of the estrogen receptor during development and has both agonist and antagonist properties in tissues of adult fish. This knowledge of esr gene function in development and in adult life will help to understand mechanisms of interfering mimicking endocrine chemicals in vivo.

  8. Teleost microbiomes: progress towards their characterisation, manipulation and applications in aquaculture and fisheries.

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    Martin eLlewellyn

    2014-06-01

    Full Text Available Indigenous bacterial flora play a critical role in the lives of their vertebrate hosts. In human and mouse models it is increasingly clear that innate and adaptive immunity develop in close consort with the commensal microbiome. Furthermore several aspects of digestion and nutrient metabolism are governed by intestinal microflora. Research on teleosts has responded relatively slowly to the revolution in microbiomics. Nonetheless, progress has been made in biotic and gnotobiotic zebrafish models, defining a core microbiome and describing its role in development. However, microbiome research in other teleost species, especially those important from an aquaculture perspective, has been relatively slow. In this review, we examine progress in teleost microbiome research to date. We discuss teleost microbiomes in health and disease, microbiome ontogeny, prospects for successful microbiome manipulation (especially in an aquaculture setting and attempt to identify important future research themes. We predict an explosion in research in this sector in line with the increasing global demand for fish protein, and the need to find sustainable approaches to improve aquaculture yield. The reduced cost and increasing ease of next generation sequencing technologies provides the technological backing, and the next 10 years will be an exciting time for teleost microbiome research.

  9. Allometric growth of the trunk leads to the rostral shift of the pelvic fin in teleost fishes.

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    Murata, Yumie; Tamura, Mika; Aita, Yusuke; Fujimura, Koji; Murakami, Yasunori; Okabe, Masataka; Okada, Norihiro; Tanaka, Mikiko

    2010-11-01

    The pelvic fin position among teleost fishes has shifted rostrally during evolution, resulting in diversification of both behavior and habitat. We explored the developmental basis for the rostral shift in pelvic fin position in teleost fishes using zebrafish (abdominal pelvic fins) and Nile tilapia (thoracic pelvic fins). Cell fate mapping experiments revealed that changes in the distribution of lateral plate mesodermal cells accompany the trunk-tail protrusion. Presumptive pelvic fin cells are originally located at the body wall adjacent to the anterior limit of hoxc10a expression in the spinal cord, and their position shifts rostrally as the trunk grows. We then showed that the differences in pelvic fin position between zebrafish and Nile tilapia were not due to changes in expression or function of gdf11. We also found that hox-independent motoneurons located above the pelvic fins innervate into the pelvic musculature. Our results suggest that there is a common mechanism among teleosts and tetrapods that controls paired appendage positioning via gdf11, but in teleost fishes the position of prospective pelvic fin cells on the yolk surface shifts as the trunk grows. In addition, teleost motoneurons, which lack lateral motor columns, innervate the pelvic fins in a manner independent of the rostral-caudal patterns of hox expression in the spinal cord.

  10. Persistent effects on adult swim performance and energetics in zebrafish developmentally exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin.

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    Marit, Jordan S; Weber, Lynn P

    2012-01-15

    TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) remains a potent and persistent toxicant in aquatic environments, causing lethal developmental deformities in fish. However, few studies have examined sublethal or persistent effects of developmental TCDD exposure and none have examined its effects on swimming capabilities in sub-adult fish. The objective of the current study was to examine whether effects of TCDD exposure during the critical period of cardiovascular development (2-4 days post fertilization) on swim performance, triglyceride stores and cardiovascular deformities would persist until adulthood in zebrafish. Zebrafish larvae were exposed between 48 and 96 h post fertilization to 1, 0.1, 0.01 ng/L TCDD or DMSO control (0.005%), then raised in clean water for 90 days. Despite having equal survivability, no significant increase in gross deformities and no change in cytochrome P450 1A (CYP1A) activity was observed, while critical swimming speed and dorsal aorta diameter were significantly decreased in TCDD-exposed fish at 90 days. Furthermore, whole body triglycerides were significantly elevated in TCDD-exposed fish both before and after swim testing. Therefore sublethal TCDD exposure during zebrafish development caused a persistent decrease in swim endurance. The cause of this persistent decrease in swim endurance is not known, but may be related to behavioral adaptations limiting swimming capabilities, failure to mobilize triglyceride stores, vascular deformities limiting blood flow to the periphery, or a combination of these factors.

  11. Exposure to perchlorate induces the formation of macrophage aggregates in the trunk kidney of zebrafish and mosquitofish

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    Capps, T.; Mukhi, S.; Rinchard, J.J.; Theodorakis, C.W.; Blazer, V.S.; Patino, R.

    2004-01-01

    Environmental contamination of ground and surface waters by perchlorate, derived from ammonium perchlorate (AP) and other perchlorate salts, is of increasing concern. Exposure to perchlorate can impair the thyroid endocrine system, which is thought to modulate renal and immune function in vertebrates. This study with zebrafish Danio rerio and eastern mosquitofish Gambusia holbrooki examined the histological effects of perchlorate on the trunk kidney, which in teleosts serves excretory and hemopoietic functions and therefore may be a target of perchlorate effects. Adult zebrafish of both sexes were exposed in the laboratory to waterborne, AP-derived perchlorate at measured concentrations of 18 mg/L for 8 weeks. Adult male mosquitofish were exposed to waterborne sodium perchlorate at measured perchlorate concentrations of 1-92 mg/L for 8 weeks. Control fish were kept in untreated water. The region of the body cavity containing the trunk kidney was processed from each fish for histological analysis. Macrophage aggregates (MAs), possible markers of contaminant exposure or immunotoxic effect, were present in the hemopoietic region of the kidney in both species exposed to perchlorate. The estimated percent area of kidney sections occupied by MAs was greater in zebrafish exposed to perchlorate at 18 mg/L (P kidney is affected by exposure to perchlorate. The concentrations of perchlorate at which the effects were noted are relatively high but within the range reported in some contaminated habitats.

  12. 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

  13. A RAD-tag genetic map for the platyfish (Xiphophorus maculatus) reveals mechanisms of karyotype evolution among teleost fish.

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    Amores, Angel; Catchen, Julian; Nanda, Indrajit; Warren, Wesley; Walter, Ron; Schartl, Manfred; Postlethwait, John H

    2014-06-01

    Mammalian genomes can vary substantially in haploid chromosome number even within a small taxon (e.g., 3-40 among deer alone); in contrast, teleost fish genomes are stable (24-25 in 58% of teleosts), but we do not yet understand the mechanisms that account for differences in karyotype stability. Among perciform teleosts, platyfish (Xiphophorus maculatus) and medaka (Oryzias latipes) both have 24 chromosome pairs, but threespine stickleback (Gasterosteus aculeatus) and green pufferfish (Tetraodon nigroviridis) have just 21 pairs. To understand the evolution of teleost genomes, we made a platyfish meiotic map containing 16,114 mapped markers scored on 267 backcross fish. We tiled genomic contigs along the map to create chromosome-length genome assemblies. Genome-wide comparisons of conserved synteny showed that platyfish and medaka karyotypes remained remarkably similar with few interchromosomal translocations but with numerous intrachromosomal rearrangements (transpositions and inversions) since their lineages diverged ∼120 million years ago. Comparative genomics with platyfish shows how reduced chromosome numbers in stickleback and green pufferfish arose by fusion of pairs of ancestral chromosomes after their lineages diverged from platyfish ∼195 million years ago. Zebrafish and human genomes provide outgroups to root observed changes. These studies identify likely genome assembly errors, characterize chromosome fusion events, distinguish lineage-independent chromosome fusions, show that the teleost genome duplication does not appear to have accelerated the rate of translocations, and reveal the stability of syntenies and gene orders in teleost chromosomes over hundreds of millions of years.

  14. Comparative genomic organization and tissue-specific transcription of the duplicated fabp7 and fabp10 genes in teleost fishes.

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    Parmar, Manoj B; Wright, Jonathan M

    2013-11-01

    A whole-genome duplication (WGD) early in the teleost fish lineage makes fish ideal organisms to study the fate of duplicated genes and underlying evolutionary trajectories that have led to the retention of ohnologous gene duplicates in fish genomes. Here, we compare the genomic organization and tissue-specific transcription of the ohnologous fabp7 and fabp10 genes in medaka, three-spined stickleback, and spotted green pufferfish to the well-studied duplicated fabp7 and fabp10 genes of zebrafish. Teleost fabp7 and fabp10 genes contain four exons interrupted by three introns. Polypeptide sequences of Fabp7 and Fabp10 show the highest sequence identity and similarity with their orthologs from vertebrates. Orthology was evident as the ohnologous Fabp7 and Fabp10 polypeptides of teleost fishes each formed distinct clades and clustered together with their orthologs from other vertebrates in a phylogenetic tree. Furthermore, ohnologous teleost fabp7 and fabp10 genes exhibit conserved gene synteny with human FABP7 and chicken FABP10, respectively, which provides compelling evidence that the duplicated fabp7 and fabp10 genes of teleost fishes most likely arose from the well-documented WGD. The tissue-specific distribution of fabp7a, fabp7b, fabp10a, and fabp10b transcripts provides evidence of diverged spatial transcriptional regulation between ohnologous gene duplicates of fabp7 and fabp10 in teleost fishes.

  15. Endosulfan exposure inhibits brain AChE activity and impairs swimming performance in adult zebrafish (Danio rerio).

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    Pereira, Vanessa Maynart; Bortolotto, Josiane Woutheres; Kist, Luiza Wilges; Azevedo, Mariana Barbieri de; Fritsch, Rachel Seemann; Oliveira, Renata da Luz; Pereira, Talita Carneiro Brandão; Bonan, Carla Denise; Vianna, Monica Ryff; Bogo, Maurício Reis

    2012-06-01

    Endosulfan is a broad spectrum organochlorine pesticide that is still widely in use in many developing countries. Following application, endosulfan can get to watercourses through surface runoff from agricultural fields and disturb the non-target aquatic animals including freshwater fish species. Given that the activity of the enzyme acetylcholinesterase (AChE) is one of the most recurrently used biomarkers of exposure to pesticides and there are controversial results concerning the effects of endosulfan exposure and AChE activity in fish, the aim of the present study was to evaluate the effects of endosulfan in brain AChE activity and its gene expression pattern using adult zebrafish (Danio rerio) as an animal model. Moreover, we have analyzed the effects of endosulfan exposure in different parameters of zebrafish swimming activity and in long-term memory formation. After 96 h of exposition, fish in the 2.4 μg endosulfan/L group presented a significant decrease in AChE activity (9.44 ± 1.038 μmol SCh h(-1) mg protein(-1); p=0.0205) when compared to the control group (15.87 ± 1.768 μmol SCh h(-1) mg protein(-1); p=0.0205) which corresponds to approximately 40%. The down-regulation of brain AChE activity is not directly related with the transcriptional control as demonstrated by the RT-qPCR analysis. Our results reinforce AChE activity inhibition as a pathway of endosulfan-induced toxicity in brain of fish species. In addition, exposure to 2.4 μg endosulfan/L during 96 h impaired all exploratory parameters evaluated: decreased line crossings (≈21%, 273.7 ± 28.12 number of line crossings compared to the control group 344.6 ± 21.30, p=0.0483), traveled distance (≈20%, 23.44 ± 2.127 m compared to the control group 29.39 ± 1.585, p=0.0281), mean speed (≈25%, 0.03 ± 0.003 m/s compared to the control group 0.04 ± 0.002, p=0.0275) and body turn angle (≈21%, 69,940 ± 4871 absolute turn angle compared to the control group 88,010 ± 4560, p=0.0114). These

  16. Magnetite-Based Magnetoreceptor Cells in the Olfactory Organ of Rainbow Trout and Zebrafish

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    Kirschvink, J. L.; Cadiou, H.; Dixson, A. D.; Eder, S.; Kobayashi, A.; McNaughton, P. A.; Muhamad, A. N.; Raub, T. D.; Walker, M. M.; Winklhofer, M.; Yuen, B. B.

    2011-12-01

    Many vertebrate and invertebrate animals have a geomagnetic sensory system, but the biophysics and anatomy of how magnetic stimuli are transduced to the nervous system is a challenging problem. Previous work in our laboratories identified single-domain magnetite chains in olfactory epithelium in cells proximal to the ros V nerve, which, in rainbow trout, responds to magnetic fields. Our objectives are to characterize these magnetite-containing cells and determine whether they form part of the mechanism of magnetic field transduction in teleost fishes, as a model for other Vertebrates. Using a combination of reflection mode confocal microscopy and a Prussian Blue technique modified to stain specifically for magnetite, our Auckland group estimated that both juvenile rainbow trout (ca. 7 cm total length) olfactory rosettes have ~200 magnetite-containing cells. The magnetite present in two types of cells within the olfactory epithelium appears to be arranged in intracellular chains. All of our groups (Munich, Auckland, Cambridge and Caltech) have obtained different types of structural evidence that magnetite chains closely associate with the plasma membrane in the cells, even in disaggregated tissues. In addition, our Cambridge group used Ca2+ imaging to demonstrate a clear response by individual magnetite-containing cells to a step change in the intensity of the external magnetic field and a slow change in Ca2+ activity when the external magnetic field was cancelled. In the teleost, zebrafish (Danio rerio), a small (~4 cm adult length in captivity) genetic and developmental biology model organism, our Caltech group detected ferromagnetic material throughout the body, but concentrated in the rostral trunk, using NRM and IRM scans of whole adults. Our analysis suggests greater than one million, 80-100 nm crystals, with Lowrie-Fuller curves strongly consistent with single-domain magnetite in 100-100,000 magnetocytes. Ferromagentic resonance (FMR) spectra show crystals

  17. Development of social behaviour in young zebrafish

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    Elena eDreosti

    2015-08-01

    Full Text Available Adult zebrafish are robustly social animals whereas larvae are not. We designed an assay to determine at what stage of development zebrafish begin to interact with and prefer other fish. One week old zebrafish show no social preference whereas most three week old zebrafish strongly prefer to remain in a compartment where they can view conspecifics. However, for some individuals, the presence of conspecifics drives avoidance instead of attraction. Social preference is dependent on vision and requires viewing fish of a similar age/size. In addition, over the same one to three week period larval zebrafish increasingly tend to coordinate their movements, a simple form of social interaction. Finally, social preference and coupled interactions are differentially modified by an NMDAR antagonist and acute exposure to ethanol, both of which are known to alter social behaviour in adult zebrafish.

  18. Biological Uptake, Distribution, and Depuration of Radio-Labeled Graphene in Adult Zebrafish: Effects of Graphene Size and Natural Organic Matter.

    Science.gov (United States)

    Lu, Kun; Dong, Shipeng; Petersen, Elijah J; Niu, Junfeng; Chang, Xiaofeng; Wang, Peng; Lin, Sijie; Gao, Shixiang; Mao, Liang

    2017-03-28

    The exciting commercial application potential of graphene materials may inevitably lead to their increasing release into the environment where they may pose ecological risks. This study focused on using carbon-14-labeled few-layer graphene (FLG) to determine whether the size of graphene plays a role in its uptake, depuration, and biodistribution in adult zebrafish. After 48 h exposure to larger FLG (L-FLG) at 250 μg/L, the amount of graphene in the organism was close to 48 mg/kg fish dry mass, which was more than 170-fold greater than the body burden of those exposed to the same concentration of smaller FLG (S-FLG). The amount of uptake for both L-FLG and S-FLG increased by a factor of 2.5 and 16, respectively, when natural organic matter (NOM) was added in the exposure suspension. While the L-FLG mainly accumulated in the gut of adult zebrafish, the S-FLG was found in both the gut and liver after exposure with or without NOM. Strikingly, the S-FLG was able to pass through the intestinal wall and enter the intestinal epithelial cells and blood. The presence of NOM increased the quantity of S-FLG in these cells. Exposure to L-FLG or S-FLG also had a significantly different impact on the intestinal microbial community structure.

  19. Demonstration of the Coexistence of Duplicated LH Receptors in Teleosts, and Their Origin in Ancestral Actinopterygians.

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    Gersende Maugars

    Full Text Available Pituitary gonadotropins, FSH and LH, control gonad activity in vertebrates, via binding to their respective receptors, FSHR and LHR, members of GPCR superfamily. Until recently, it was accepted that gnathostomes possess a single FSHR and a single LHR, encoded by fshr and lhcgr genes. We reinvestigated this question, focusing on vertebrate species of key-phylogenetical positions. Genome analyses supported the presence of a single fshr and a single lhcgr in chondrichthyans, and in sarcopterygians including mammals, birds, amphibians and coelacanth. In contrast, we identified a single fshr but two lhgcr in basal teleosts, the eels. We further showed the coexistence of duplicated lhgcr in other actinopterygians, including a non-teleost, the gar, and other teleosts, e.g. Mexican tetra, platyfish, or tilapia. Phylogeny and synteny analyses supported the existence in actinopterygians of two lhgcr paralogs (lhgcr1/ lhgcr2, which do not result from the teleost-specific whole-genome duplication (3R, but likely from a local gene duplication that occurred early in the actinopterygian lineage. Due to gene losses, there was no impact of 3R on the number of gonadotropin receptors in extant teleosts. Additional gene losses during teleost radiation, led to a single lhgcr (lhgcr1 or lhgcr2 in some species, e.g. medaka and zebrafish. Sequence comparison highlighted divergences in the extracellular and intracellular domains of the duplicated lhgcr, suggesting differential properties such as ligand binding and activation mechanisms. Comparison of tissue distribution in the European eel, revealed that fshr and both lhgcr transcripts are expressed in the ovary and testis, but are differentially expressed in non-gonadal tissues such as brain or eye. Differences in structure-activity relationships and tissue expression may have contributed as selective drives in the conservation of the duplicated lhgcr. This study revises the evolutionary scenario and nomenclature of

  20. An update on MyoD evolution in teleosts and a proposed consensus nomenclature to accommodate the tetraploidization of different vertebrate genomes.

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    Daniel J Macqueen

    Full Text Available BACKGROUND: MyoD is a muscle specific transcription factor that is essential for vertebrate myogenesis. In several teleost species, including representatives of the Salmonidae and Acanthopterygii, but not zebrafish, two or more MyoD paralogues are conserved that are thought to have arisen from distinct, possibly lineage-specific duplication events. Additionally, two MyoD paralogues have been characterised in the allotetraploid frog, Xenopus laevis. This has lead to a confusing nomenclature since MyoD paralogues have been named outside of an appropriate phylogenetic framework. METHODS AND PRINCIPAL FINDINGS: Here we initially show that directly depicting the evolutionary relationships of teleost MyoD orthologues and paralogues is hindered by the asymmetric evolutionary rate of Acanthopterygian MyoD2 relative to other MyoD proteins. Thus our aim was to confidently position the event from which teleost paralogues arose in different lineages by a comparative investigation of genes neighbouring myod across the vertebrates. To this end, we show that genes on the single myod-containing chromosome of mammals and birds are retained in both zebrafish and Acanthopterygian teleosts in a striking pattern of double conserved synteny. Further, phylogenetic reconstruction of these neighbouring genes using Bayesian and maximum likelihood methods supported a common origin for teleost paralogues following the split of the Actinopterygii and Sarcopterygii. CONCLUSION: Our results strongly suggest that myod was duplicated during the basal teleost whole genome duplication event, but was subsequently lost in the Ostariophysi (zebrafish and Protacanthopterygii lineages. We propose a sensible consensus nomenclature for vertebrate myod genes that accommodates polyploidization events in teleost and tetrapod lineages and is justified from a phylogenetic perspective.

  1. Structural and functional diversification in the teleost S100 family of calcium-binding proteins

    Directory of Open Access Journals (Sweden)

    Korsching Sigrun I

    2008-02-01

    Full Text Available Abstract Background Among the EF-Hand calcium-binding proteins the subgroup of S100 proteins constitute a large family with numerous and diverse functions in calcium-mediated signaling. The evolutionary origin of this family is still uncertain and most studies have examined mammalian family members. Results We have performed an extensive search in several teleost genomes to establish the s100 gene family in fish. We report that the teleost S100 repertoire comprises fourteen different subfamilies which show remarkable similarity across six divergent teleost species. Individual species feature distinctive subsets of thirteen to fourteen genes that result from local gene duplications and gene losses. Eight of the fourteen S100 subfamilies are unique for teleosts, while six are shared with mammalian species and three of those even with cartilaginous fish. Several S100 family members are found in jawless fish already, but none of them are clear orthologs of cartilaginous or bony fish s100 genes. All teleost s100 genes show the expected structural features and are subject to strong negative selection. Many aspects of the genomic arrangement and location of mammalian s100 genes are retained in the teleost s100 gene family, including a completely conserved intron/exon border between the two EF hands. Zebrafish s100 genes exhibit highly specific and characteristic expression patterns, showing both redundancy and divergence in their cellular expression. In larval tissue expression is often restricted to specific cell types like keratinocytes, hair cells, ionocytes and olfactory receptor neurons as demonstrated by in situ hybridization. Conclusion The origin of the S100 family predates at least the segregation of jawed from jawless fish and some extant family members predate the divergence of bony from cartilaginous fish. Despite a complex pattern of gene gains and losses the total repertoire size is remarkably constant between species. On the expression

  2. Zebrafish embryonic stromal trunk (ZEST) cells support hematopoietic stem and progenitor cell (HSPC) proliferation, survival, and differentiation.

    Science.gov (United States)

    Campbell, Clyde; Su, Tammy; Lau, Ryan P; Shah, Arpit; Laurie, Payton C; Avalos, Brenda; Aggio, Julian; Harris, Elena; Traver, David; Stachura, David L

    2015-12-01

    Forward genetic screens in zebrafish have been used to identify genes essential for the generation of primitive blood and the emergence of hematopoietic stem cells (HSCs), but have not elucidated the genes essential for hematopoietic stem and progenitor cell (HSPC) proliferation and differentiation because of the lack of methodologies to functionally assess these processes. We previously described techniques used to test the developmental potential of HSPCs by culturing them on zebrafish kidney stromal (ZKS) cells, derived from the main site of hematopoiesis in the adult teleost. Here we describe an additional primary stromal cell line we refer to as zebrafish embryonic stromal trunk (ZEST) cells, derived from tissue surrounding the embryonic dorsal aorta, the site of HSC emergence in developing fish. ZEST cells encouraged HSPC differentiation toward the myeloid, lymphoid, and erythroid pathways when assessed by morphologic and quantitative reverse transcription polymerase chain reaction analyses. Additionally, ZEST cells significantly expanded the number of cultured HSPCs in vitro, indicating that these stromal cells are supportive of both HSPC proliferation and multilineage differentiation. Examination of ZEST cells indicates that they express numerous cytokines and Notch ligands and possess endothelial characteristics. Further characterization of ZEST cells should prove to be invaluable in understanding the complex signaling cascades instigated by the embryonic hematopoietic niche required to expand and differentiate HSPCs. Elucidating these processes and identifying possibilities for the modulation of these molecular pathways should allow the in vitro expansion of HSPCs for a multitude of therapeutic uses.

  3. Rumba and Haus3 are essential factors for the maintenance of hematopoietic stem/progenitor cells during zebrafish hematopoiesis.

    Science.gov (United States)

    Du, Linsen; Xu, Jin; Li, Xiuling; Ma, Ning; Liu, Yanmei; Peng, Jinrong; Osato, Motomi; Zhang, Wenqing; Wen, Zilong

    2011-02-01

    The hallmark of vertebrate definitive hematopoiesis is the establishment of the hematopoietic stem/progenitor cell (HSPC) pool during embryogenesis. This process involves a defined ontogenic switching of HSPCs in successive hematopoietic compartments and is evolutionarily conserved from teleost fish to human. In zebrafish, HSPCs originate from the ventral wall of the dorsal aorta (VDA), from which they subsequently mobilize to an intermediate hematopoietic site known as the caudal hematopoietic tissue (CHT) and finally colonize the kidney for adult hematopoiesis. Despite substantial understanding of the ontogeny of HSPCs, the molecular basis governing migration, colonization and maintenance of HSPCs remains to be explored fully. Here, we report the isolation and characterization of two zebrafish mutants, rumba(hkz1) and samba(hkz2), that are defective in generating definitive hematopoiesis. We find that HSPC initiation in the VDA and subsequent homing to the CHT are not affected in these two mutants. However, the further development of HSPCs in the CHT is compromised in both mutants. Positional cloning reveals that Rumba is a novel nuclear C2H2 zinc-finger factor with unknown function and samba encodes an evolutionarily conserved protein that is homologous to human augmin complex subunit 3 (HAUS3). Furthermore, we show that these two factors independently regulate cell cycle progression of HSPCs and are cell autonomously required for HPSC development in the CHT. Our study identifies Rumba and Haus3 as two essential regulators of HSPC maintenance during zebrafish fetal hematopoiesis.

  4. THE TELEOST GUT PERSORBS MICROPARTICULATES

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    Ewen McLean

    2001-06-01

    Full Text Available The ability of the teleost gut to absorb microparticulate material was examined following rectal intubation (3.5 g kg -1 of commercial grade cornstarch (≈21 mm diameter, or potato starch (≈43 mm diameter. Tissue samples were taken from the mid - and hind-gut of control and treated fish 18 h postintubation. Collected samples were processed using standard plastic and staining protocols and resultant photomicrographs examined by computer-assisted image analysis. Cornstarch particles (8-14 mm, were observed to pass from gut lumen to the lamina propria via a paracellular or persorptive route only. No evidence for the like passage of potato starch was found.

  5. Expression of sept3, sept5a and sept5b in the Developing and Adult Nervous System of the Zebrafish (Danio rerio)

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    Helmprobst, Frederik; Lillesaar, Christina; Stigloher, Christian

    2017-01-01

    Septins are a highly conserved family of small GTPases that form cytoskeletal filaments. Their cellular functions, especially in the nervous system, still remain largely enigmatic, but there are accumulating lines of evidence that septins play important roles in neuronal physiology and pathology. In order to further dissect septin function in the nervous system a detailed temporal resolved analysis in the genetically well tractable model vertebrate zebrafish (Danio rerio) is crucially necessary. To close this knowledge gap we here provide a reference dataset describing the expression of selected septins (sept3, sept5a and sept5b) in the zebrafish central nervous system. Strikingly, proliferation zones are devoid of expression of all three septins investigated, suggesting that they have a role in post-mitotic neural cells. Our finding that three septins are mainly expressed in non-proliferative regions was further confirmed by double-stainings with a proliferative marker. Our RNA in situ hybridization (ISH) study, detecting sept3, sept5a and sept5b mRNAs, shows that all three septins are expressed in largely overlapping regions of the developing brain. However, the expression of sept5a is much more confined compared to sept3 and sept5b. In contrast, the expression of all the three analyzed septins is largely similar in the adult brain.

  6. Microcystin-LR acute exposure does not alter in vitro and in vivo ATP, ADP and AMP hydrolysis in adult zebrafish (Danio rerio brain membranes

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    Luiza Wilges Kist

    2012-10-01

    Full Text Available Microcystins (MCs are toxins produced by cyanobacteria during the blooms that could accumulate in aquatic animals and be relocated to higher trophic levels. Adenosine triphosphate (ATP acts as an excitatory neurotransmitter and/or a neuromodulator in the extracellular space playing important roles in physiological and pathological conditions. The aim of this study was, therefore, to evaluate the acute effects of different concentrations of MC-LR on nucleoside triphosphate diphosphohydrolases and 5’-nucleotidade in adult zebrafish (Danio rerio brain membranes. The results have shown no significant changes in ATP, adenosine diphosphate (ADP and adenosine monophosphate (AMP hydrolysis in zebrafish brain membranes. MC-LR in vitro also did not alter ATP, ADP and AMP hydrolysis in the concentrations tested. These findings show that acute exposure to MC-LR did not modulate ectonucleotidase activity in the conditions tested. However, additional studies including chronic exposure should be performed in order to achieve a better understanding about MC-LR toxicity mechanisms in the central nervous system.

  7. Transcriptional impact of organophosphate and metal mixtures on olfaction: copper dominates the chlorpyrifos-induced response in adult zebrafish.

    Science.gov (United States)

    Tilton, Fred A; Tilton, Susan C; Bammler, Theo K; Beyer, Richard P; Stapleton, Patricia L; Scholz, Nathaniel L; Gallagher, Evan P

    2011-04-01

    Chemical exposures in fish have been linked to loss of olfaction leading to an inability to detect predators and prey and decreased survival. However, the mechanisms underlying olfactory neurotoxicity are not well characterized, especially in environmental exposures which involve chemical mixtures. We used zebrafish to characterize olfactory transcriptional responses by two model olfactory inhibitors, the pesticide chlorpyrifos (CPF) and mixtures of CPF with the neurotoxic metal copper (Cu). Microarray analysis was performed on RNA from olfactory tissues of zebrafish exposed to CPF alone or to a mixture of CPF and Cu. Gene expression profiles were analyzed using principal component analysis and hierarchical clustering, whereas gene set analysis was used to identify biological themes in the microarray data. Microarray results were confirmed by real-time PCR on genes serving as potential biomarkers of olfactory injury. In addition, we mined our previously published Cu-induced zebrafish olfactory transcriptional response database (Tilton et al., 2008) for the purposes of discriminating pathways of olfaction impacted by either the individual agents or the CPF-Cu mixture transcriptional signatures. CPF exposure altered the expression of gene pathways associated with cellular morphogenesis and odorant binding, but not olfactory signal transduction, a known olfactory pathway for Cu. The mixture profiles shared genes from the Cu and CPF datasets, whereas some genes were altered only by the mixtures. The transcriptional signature of the mixtures was more similar to that in zebrafish exposed to Cu alone than for CPF. In conclusion, exposure to a mixture containing a common environmental metal and pesticide causes a unique transcriptional signature that is heavily influenced by the metal, even when organophosphate predominates.

  8. Transcriptional impact of organophosphate and metal mixtures on olfaction: Copper dominates the chlorpyrifos-induced response in adult zebrafish

    OpenAIRE

    Tilton, Fred A.; Tilton, Susan C.; Bammler, Theo K.; Beyer, Richard P; Stapleton, Patricia L.; Nathaniel L Scholz; Gallagher, Evan P.

    2011-01-01

    Chemical exposures in fish have been linked to loss of olfaction leading to an inability to detect predators and prey and decreased survival. However, the mechanisms underlying olfactory neurotoxicity are not well characterized, especially in environmental exposures which involve chemical mixtures. We used zebrafish to characterize olfactory transcriptional responses by two model olfactory inhibitors, the pesticide chlorpyrifos (CPF) and mixtures of CPF with the neurotoxic metal copper (Cu). ...

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

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    Eames B Frank

    2012-03-01

    Full Text Available Abstract Background Although the vertebrate skeleton arose in the sea 500 million years ago, our understanding of the molecular fingerprints of chondrocytes and osteoblasts may be biased because it is informed mainly by research on land animals. In fact, the molecular fingerprint of teleost osteoblasts differs in key ways from that of tetrapods, but we do not know the origin of these novel gene functions. They either arose as neofunctionalization events after the teleost genome duplication (TGD, or they represent preserved ancestral functions that pre-date the TGD. Here, we provide evolutionary perspective to the molecular fingerprints of skeletal cells and assess the role of genome duplication in generating novel gene functions. We compared the molecular fingerprints of skeletogenic cells in two ray-finned fish: zebrafish (Danio rerio--a teleost--and the spotted gar (Lepisosteus oculatus--a "living fossil" representative of a lineage that diverged from the teleost lineage prior to the TGD (i.e., the teleost sister group. We analyzed developing embryos for expression of the structural collagen genes col1a2, col2a1, col10a1, and col11a2 in well-formed cartilage and bone, and studied expression of skeletal regulators, including the transcription factor genes sox9 and runx2, during mesenchymal condensation. Results Results provided no evidence for the evolution of novel functions among gene duplicates in zebrafish compared to the gar outgroup, but our findings shed light on the evolution of the osteoblast. Zebrafish and gar chondrocytes both expressed col10a1 as they matured, but both species' osteoblasts also expressed col10a1, which tetrapod osteoblasts do not express. This novel finding, along with sox9 and col2a1 expression in developing osteoblasts of both zebrafish and gar, demonstrates that osteoblasts of both a teleost and a basally diverging ray-fin fish express components of the supposed chondrocyte molecular fingerprint. Conclusions Our

  10. Loss of Lrp2 in zebrafish disrupts pronephric tubular clearance but not forebrain development.

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    Kur, Esther; Christa, Anna; Veth, Kerry N; Gajera, Chandresh R; Andrade-Navarro, Miguel A; Zhang, Jingjing; Willer, Jason R; Gregg, Ronald G; Abdelilah-Seyfried, Salim; Bachmann, Sebastian; Link, Brian A; Hammes, Annette; Willnow, Thomas E

    2011-06-01

    Low-density lipoprotein receptor-related protein 2 (LRP2) is a multifunctional cell surface receptor conserved from nematodes to humans. In mammals, it acts as regulator of sonic hedgehog and bone morphogenetic protein pathways in patterning of the embryonic forebrain and as a clearance receptor in the adult kidney. Little is known about activities of this LRP in other phyla. Here, we extend the functional elucidation of LRP2 to zebrafish as a model organism of receptor (dys)function. We demonstrate that expression of Lrp2 in embryonic and larval fish recapitulates the patterns seen in mammalian brain and kidney. Furthermore, we studied the consequence of receptor deficiencies in lrp2 and in lrp2b, a homologue unique to fish, using ENU mutagenesis or morpholino knockdown. While receptor-deficient zebrafish suffer from overt renal resorption deficiency, their brain development proceeds normally, suggesting evolutionary conservation of receptor functions in pronephric duct clearance but not in patterning of the teleost forebrain.

  11. Distinct Functions of Different scl Isoforms in Zebrafish Definitive Hematopoietic Stem Cell Initiation and Maintenance

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    Lan, Yahui

    2011-07-01

    The establishment of entire blood system relies on the multi-potent hematopoietic stem cells (HSCs), thus identifying the molecular mechanism in HSC generation is of importance for not only complementing the fundamental knowledge in stem cell biology, but also providing insights to the regenerative therapies. Recent researches have documented the formation of nascent HSCs through a direct transition from ventral aortic endothelium, named as endothelial hematopoietic transition (EHT) process. However, the precise genetic program engaged in this process remains largely elusive. The transcription factor scl plays pivotal and conserved roles in embryonic and adult hematopoiesis from teleosts to mammals. Our lab have previously identified a new truncated scl isoform, scl-beta, which is indispensible for the specification of HSCs in the ventral wall of dorsal aorta (VDA), the zebrafish equivalent of mammalian fetal hematopoietic organ. Here we observe that, by combining time-lapse confocal imaging of transgenic zebrafish and genetic epistasis analysis, scl-beta is expressed in a subset of ventral aortic endothelial cells and critical for their forthcoming transformation to hemogenic endothelium; in contrast, runx1 is required downstream to govern the successful egress of the hemogenic endothelial cells to become naive HSCs. In addition, the traditional known full-length scl-alpha isoform is firstly evidenced to be required for the maintenance or survival of newly formed HSCs in VDA. Collectively our data has established the genetic hierarchy controlling discrete steps in the consecutive process of HSC formation from endothelial cells and further development in VDA.

  12. Identification and analysis of genes involved in bone formation - a genetic approach in zebrafish -

    NARCIS (Netherlands)

    Spoorendonk, K.M.

    2009-01-01

    For many years bone research has been mainly performed in mice, chicken, cell culture systems, or human material from the clinic. In this thesis, we make use of the zebrafish (Danio rerio), a relatively new model system in this field. This small teleost offers possibilities which makes it a great co

  13. Identification and analysis of genes involved in bone formation – a genetic approach in zebrafish

    NARCIS (Netherlands)

    Spoorendonk, K.M.

    2009-01-01

    For many years bone research has been mainly performed in mice, chicken, cell culture systems, or human material from the clinic. In this thesis, we make use of the zebrafish (Danio rerio), a relatively new model system in this field. This small teleost offers possibilities which makes it a great co

  14. Transcriptional Profiling of Caudal Fin Regeneration in Zebrafish

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    Michael Schebesta

    2006-01-01

    Full Text Available Regeneration of severed limbs in adult animals is restricted to urodele amphibians. Mammals, including humans, have very limited regenerative capabilities and even with proper treatment, only the tips of our digits can grow back. Teleost fish can regenerate amputated fins, the evolutionary ancestors of limbs. To elucidate the principles of limb-fin regeneration, we performed an Affymetrix microarray screen on regenerating caudal fins 12, 24, 48, and 72 h post amputation. Approximately 15,000 zebrafish transcripts were analyzed, identifying 829 transcripts as differentially expressed during regeneration. Of those, 563 were up-regulated and 266 were down-regulated. We constructed a comprehensive database containing expression data, functional assignment, and background information from the literature for each differentially expressed transcript. In order to validate our findings, we employed three approaches: (1 microarray expression analysis of genes previously implicated in fin regeneration, (2 RT-PCR analysis of genes newly identified as differentially expressed during regeneration, and (3 in situ hybridization of the up-regulated genes bambi, dlx5A, and her6. Moreover, we show that Smad 1/5/8 proteins, effector molecules of Bmp signaling, are phosphorylated during fin regeneration. Taken together, we provide a comprehensive database of fin regeneration that will serve as an important tool for understanding the molecular mechanisms of regeneration.

  15. Comparison of two chemically-induced colitis-models in adult zebrafish, using optical projection tomography and novel transcriptional markers

    DEFF Research Database (Denmark)

    Haarder, Simon; Kania, Per Walter; Holm, Thomas

    2016-01-01

    of proinflammatory cytokines, acute-phase reactants and metalloprotease 9 in both chemical models, primarily after 72 hours. In comparison, transcription factors and cytokines associated with Th1 and Th17 (Crohn’s) and Th2 (ulcerative colitis) were mainly not affected in this acute setting. However, elevated...... zebrafish. In conclusion, a distinct acute inflammatory reaction was induced in both chemical models. Further, oxazolone and TNBS did not result in clear-cut Th2 and Th1/Th17 pathway signaling at this early timepoint, but the applied molecular tools may provide further insight to the IBD pathogenesis...

  16. Sex-dependent effects of microcystin-LR on hypothalamic-pituitary-gonad axis and gametogenesis of adult zebrafish

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    Liu, Wanjing; Chen, Chuanyue; Chen, Liang; Wang, Li; Li, Jian; Chen, Yuanyuan; Jin, Jienan; Kawan, Atufa; Zhang, Xuezhen

    2016-03-01

    While microcystins (MCs) have been reported to exert reproductive toxicity on fish with a sex-dependent effect, the underlying mechanism has been rarely investigated. In the present study, zebrafish were exposed to 1, 5 and 20 μg/L MC-LR for 30 d. The gonad-somatic index declined in all treated males. 17β-estradiol (E2), testosterone (T), 11-keto testosterone (11-KT) and follicle-stimulating hormone (FSH) levels increased in serum from all treated females, while T, FSH and luteinizing hormone (LH) levels changed in all treated males. Histomorphological observation showed that MC-LR exposure evidently retarded oogenesis and spermatogenesis. Transcriptional changes of 22 genes of the hypothalamic-pituitary-gonad (HPG) axis exhibited sex-specific responses, and the relationship between gene transcriptions and gametogenesis was evaluated by principle component analysis (PCA). Major contributors to PC1 (gnrh2, gnrhr3, ar, lhr, hmgra, hmgrb and cyp19a) were positively correlated with the number of post-vitellogenic oocytes, while PC1 (gnrh2, lhβ, erβ, fshr, cyp11a and 17βhsd) were positively correlated with the number of spermatozoa. The protein levels of 17βHSD and CYP19a were affected in both females and males. In conclusion, this study first investigated the sex-dependent effects of microcystins on fish reproduction and revealed some important molecular biomarkers related to gametogenesis in zebrafish suffered from MC-LR.

  17. Comparative effects of nodularin and microcystin-LR in zebrafish: 2. Uptake and molecular effects in eleuthero-embryos and adult liver with focus on endoplasmic reticulum stress.

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    Faltermann, Susanne; Grundler, Verena; Gademann, Karl; Pernthaler, Jakob; Fent, Karl

    2016-02-01

    . In contrast to adult liver, MC-LR and nodularin did not result in detectable changes of mRNA levels of selected target genes involved in ER-stress in zebrafish eleuthero-embryos, nor was the abundance of transcripts belonging to the MAPK and pro-apoptosis pathways altered. In conclusion, our data indicate that MC-LR and nodularin have similar transcriptional effects. They lead to changes in mRNA levels of genes that suggest induction of ER-stress, and furthermore, lead to increased level of tnfα mRNA in the adult liver, which suggests a novel (transcriptional) mode of action in fish. However, although taken up by eleuthero-embryos, no transcriptional changes induced by these cyanobacterial toxins were detected. This is probably due to action to specific organs such as liver and kidneys that could not be identified by whole-embryo sampling.

  18. Duplication and divergence of fgf8 functions in teleost development and evolution.

    Science.gov (United States)

    Jovelin, Richard; He, Xinjun; Amores, Angel; Yan, Yi-Lin; Shi, Ruihua; Qin, Baifang; Roe, Bruce; Cresko, William A; Postlethwait, John H

    2007-12-15

    Fibroblast growth factors play critical roles in many aspects of embryo patterning that are conserved across broad phylogenetic distances. To help understand the evolution of fibroblast growth factor functions, we identified members of the Fgf8/17/18-subfamily in the three-spine stickleback Gasterosteus aculeatus, and investigated their evolutionary relationships and expression patterns. We found that fgf17b is the ortholog of tetrapod Fgf17, whereas the teleost genes called fgf8 and fgf17a are duplicates of the tetrapod gene Fgf8, and thus should be called fgf8a and fgf8b. Phylogenetic analysis supports the view that the Fgf8/17/18-subfamily expanded during the ray-fin fish genome duplication. In situ hybridization experiments showed that stickleback fgf8 duplicates exhibited common and unique expression patterns, indicating that tissue specialization followed the gene duplication event. Moreover, direct comparison of stickleback and zebrafish embryonic expression patterns of fgf8 co-orthologs suggested lineage-specific independent subfunction partitioning and the acquisition or the loss of ortholog functions. In tetrapods, Fgf8 plays an important role in the apical ectodermal ridge of the developing pectoral appendage. Surprisingly, differences in the expression of fgf8a in the apical ectodermal ridge of the pectoral fin bud in zebrafish and stickleback, coupled with the role of fgf16 and fgf24 in teleost pectoral appendage show that different Fgf genes may play similar roles in limb development in various vertebrates.

  19. Cortisol regulates sodium homeostasis by stimulating the transcription of sodium-chloride transporter (NCC) in zebrafish (Danio rerio).

    Science.gov (United States)

    Lin, Chia-Hao; Hu, Huei-Jyun; Hwang, Pung-Pung

    2016-02-15

    In mammals, sodium/hydrogen exchanger (NHE) and sodium-chloride cotransporter (NCC) are expressed in renal tubules, and exhibit functional redundancy and mutual compensation in Na(+) uptake. In teleosts, the gills of the adult and skin of the embryonic stage function as external kidneys, and ionocytes are responsible for ionoregulation in these tissues. NHE- and NCC-expressing ionocytes mutually cooperate to adjust Na(+) uptake, which is analogous to the activity of the mammalian kidney. Cortisol is a hormone that controls Na(+) uptake through regulating NCC expression and activity in mammals; however, cortisol-mediated control of NCC expression is little understood in non-mammalian vertebrates, such as teleosts. It is essential for our understanding of the evolution of such regulation to determine whether cortisol has a conserved effect on NCC in vertebrates. In the present study, we treated zebrafish embryos with low Na(+) medium (LNa, 0.04 mM Na(+)) for 3 d to stimulate the mRNA expression of nhe3b, ncc, and cyp11b1 (a cortisol-synthesis enzyme) and whole body cortisol level. Exogenous cortisol treatment (20 mg/l, 3 d) resulted in an elevation of whole-body Na(+) content, ncc expression, and the density of ncc-expressing cells in zebrafish larvae. In loss-of-function experiments, microinjection of glucocorticoid receptor (gr) morpholino (MO) suppressed sodium content, ncc expression, and the density of ncc-expressing cells, but injection of mr MO had no such effects. In addition, exogenous cortisol treatment and gr MO injection also altered ncc expression and the density of ncc-expressing cells in gcm2 morphant larvae. Taken together, cortisol and GR appear to regulate Na(+) absorption through stimulating ncc expression and the differentiation of ncc-expressing ionocytes, providing new insights into the actions of cortisol on Na(+) uptake.

  20. Prolactin and teleost ionocytes: new insights into cellular and molecular targets of prolactin in vertebrate epithelia

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    Breves, Jason P.; McCormick, Stephen D.; Karlstrom, Rolf O.

    2014-01-01

    The peptide hormone prolactin is a functionally versatile hormone produced by the vertebrate pituitary. Comparative studies over the last six decades have revealed that a conserved function for prolactin across vertebrates is the regulation of ion and water transport in a variety of tissues including those responsible for whole-organism ion homeostasis. In teleost fishes, prolactin was identified as the “freshwater-adapting hormone”, promoting ion-conserving and water-secreting processes by acting on the gill, kidney, gut and urinary bladder. In mammals, prolactin is known to regulate renal, intestinal, mammary and amniotic epithelia, with dysfunction linked to hypogonadism, infertility, and metabolic disorders. Until recently, our understanding of the cellular mechanisms of prolactin action in fishes has been hampered by a paucity of molecular tools to define and study ionocytes, specialized cells that control active ion transport across branchial and epidermal epithelia. Here we review work in teleost models indicating that prolactin regulates ion balance through action on ion transporters, tight-junction proteins, and water channels in ionocytes, and discuss recent advances in our understanding of ionocyte function in the genetically and embryonically accessible zebrafish (Danio rerio). Given the high degree of evolutionary conservation in endocrine and osmoregulatory systems, these studies in teleost models are contributing novel mechanistic insight into how prolactin participates in the development, function, and dysfunction of osmoregulatory systems across the vertebrate lineage.

  1. 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.

  2. Development and regeneration of the zebrafish maxillary barbel: a novel study system for vertebrate tissue growth and repair.

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    Elizabeth E LeClair

    Full Text Available BACKGROUND: Barbels are integumentary sense organs found in fishes, reptiles and amphibians. The zebrafish, Danio rerio, develops paired nasal and maxillary barbels approximately one month post fertilization. Small in diameter and optically clear, these adult appendages offer a window on the development, maintenance and function of multiple cell types including skin cells, neural-crest derived pigment cells, circulatory vessels, taste buds and sensory nerves. Importantly, barbels in other otophysan fishes (e.g., catfish are known to regenerate; however, this capacity has not been tested in zebrafish. METHODOLOGY/PRINCIPAL FINDINGS: We describe the development of the maxillary barbel in a staged series of wild type and transgenic zebrafish using light microscopy, histology and immunohistochemistry. By imaging transgenic zebrafish containing fluorescently labeled endothelial cells (Tg(fli1a:EGFP, we demonstrate that the barbel contains a long ( approximately 2-3 mm closed-end vessel that we interpret as a large lymphatic. The identity of this vessel was further supported by live imaging of the barbel circulation, extending recent descriptions of the lymphatic system in zebrafish. The maxillary barbel can be induced to regenerate by proximal amputation. After more than 750 experimental surgeries in which approximately 85% of the barbel's length was removed, we find that wound healing is complete within hours, followed by blastema formation ( approximately 3 days, epithelial redifferentiation (3-5 days and appendage elongation. Maximum regrowth occurs within 2 weeks of injury. Although superficially normal, the regenerates are shorter and thicker than the contralateral controls, have abnormally organized mesenchymal cells and extracellular matrix, and contain prominent connective tissue "stumps" at the plane of section--a mode of regeneration more typical of mammalian scarring than other zebrafish appendages. Finally, we show that the maxillary

  3. Adaptation of teleosts to very high salinity

    DEFF Research Database (Denmark)

    Laverty, Gary; Skadhauge, Erik

    2012-01-01

    A number of species of euryhaline teleosts have the remarkable ability to adapt and survive in environments of extreme salinity, up to two or even three times the osmolality of seawater. This review looks at some of the literature describing the adaptive changes that occur, primarily with intesti......A number of species of euryhaline teleosts have the remarkable ability to adapt and survive in environments of extreme salinity, up to two or even three times the osmolality of seawater. This review looks at some of the literature describing the adaptive changes that occur, primarily...

  4. Growth response and expression of muscle growth-related candidate genes in adult zebrafish fed plant and fishmeal protein-based diets.

    Science.gov (United States)

    Ulloa, Pilar E; Peña, Andrea A; Lizama, Carla D; Araneda, Cristian; Iturra, Patricia; Neira, Roberto; Medrano, Juan F

    2013-03-01

    The main objective of this study was to examine the effects of a plant protein- vs. fishmeal-based diet on growth response in a population of 24 families, as well as expression of growth-related genes in the muscle of adult zebrafish (Danio rerio). Each family was split to create two fish populations with similar genetic backgrounds, and the fish were fed either fishmeal (FM diet) or plant protein (PP diet) as the unique protein source in their diets from 35 to 98 days postfertilization (dpf). To understand the effect of the PP diet on gene expression, individuals from three families, representative of the mean weight in both populations, were selected. To understand the effect of familiar variation on gene expression, the same families were evaluated separately. At 98 dpf, growth-related genes Igf1a, Igf2a, mTOR, Pld1a, Mrf4, Myod, Myogenin, and Myostatin1b were evaluated. In males, Myogenin, Mrf4, and Igf2a showed changes attributable to the PP diet. In females, the effect of the PP diet did not modulate the expression in any of the eight genes studied. The effect of familiar variation on gene expression was observed among families. This study shows that PP diet and family variation have effects on gene expression in fish muscle.

  5. Zebrafish as a unique model system in bone research: the power of genetics and in vivo imaging

    NARCIS (Netherlands)

    Spoorendonk, K.M.; Hammond, C.L.; Huitema, L.F.A.; Vanoevelen, J.; Schulte-Merker, S.

    2010-01-01

    For many years bone research has been mainly performed in mice, chicken, cell culture systems or human material from the clinic. In this review, we describe the features of zebrafish (Danio rerio), a relatively new model system in this field. This small teleost offers possibilities which make it a g

  6. The toxicity of a new disinfection by-product, 2,2-dichloroacetamide (DCAcAm), on adult zebrafish (Danio rerio) and its occurrence in the chlorinated drinking water.

    Science.gov (United States)

    Yu, Shilin; Lin, Tao; Chen, Wei; Tao, Hui

    2015-11-01

    The detection method of 2,2-dichloroacetamide (DCAcAm), a new disinfection by-product (DBP) in chlorinated drinking water, was established using a gas chromatograph coupled with a micro-electron capture detector. The chlorinated water samples were taken from ten drinking water treatment plants around Yangtze River or Taihu Lake in China. The concentration of DCAcAm was detected ranging from 0.5 to 1.8μg/L in the waterworks around Yangtze River, and 1.5-2.6μg/L around Taihu Lake. The toxicity of DCAcAm on adult zebrafish was assessed by investigating the metabolism damage with multiple metabolic biomarkers and the accumulation capability with bio-concentration factor. The results showed that DCAcAm could cause the acute metabolism damage and was easily accumulated in zebrafish, and should be extremely cautioned.

  7. Zebrafish: modeling for herpes simplex virus infections.

    Science.gov (United States)

    Antoine, Thessicar Evadney; Jones, Kevin S; Dale, Rodney M; Shukla, Deepak; Tiwari, Vaibhav

    2014-02-01

    For many years, zebrafish have been the prototypical model for studies in developmental biology. In recent years, zebrafish has emerged as a powerful model system to study infectious diseases, including viral infections. Experiments conducted with herpes simplex virus type-1 in adult zebrafish or in embryo models are encouraging as they establish proof of concept with viral-host tropism and possible screening of antiviral compounds. In addition, the presence of human homologs of viral entry receptors in zebrafish such as 3-O sulfated heparan sulfate, nectins, and tumor necrosis factor receptor superfamily member 14-like receptor bring strong rationale for virologists to test their in vivo significance in viral entry in a zebrafish model and compare the structure-function basis of virus zebrafish receptor interaction for viral entry. On the other end, a zebrafish model is already being used for studying inflammation and angiogenesis, with or without genetic manipulations, and therefore can be exploited to study viral infection-associated pathologies. The major advantage with zebrafish is low cost, easy breeding and maintenance, rapid lifecycle, and a transparent nature, which allows visualizing dissemination of fluorescently labeled virus infection in real time either at a localized region or the whole body. Further, the availability of multiple transgenic lines that express fluorescently tagged immune cells for in vivo imaging of virus infected animals is extremely attractive. In addition, a fully developed immune system and potential for receptor-specific knockouts further advocate the use of zebrafish as a new tool to study viral infections. In this review, we focus on expanding the potential of zebrafish model system in understanding human infectious diseases and future benefits.

  8. Mechanisms controlling Pax6 isoform expression in the retina have been conserved between teleosts and mammals.

    Science.gov (United States)

    Lakowski, Jörn; Majumder, Anirban; Lauderdale, James D

    2007-07-15

    The Pax6 gene plays several roles in retinal development, including control of cell proliferation, maintenance of the retinogenic potential of progenitor cells, and cell fate specification. Emerging evidence suggests that these different aspects of Pax6 gene function are mediated by different isoforms of the Pax6 protein; however, relatively little is known about the spatiotemporal expression of Pax6 isoforms in the vertebrate retina. Using bacterial artificial chromosome (BAC) technology, we modified a zebrafish Pax6a BAC such that we could distinguish paired-containing Pax6a transcripts from paired-less Pax6a transcripts. In the zebrafish, the spatial and temporal onset of expression of these transcripts suggests that the paired-less isoform is involved in the cell fate decision leading to the generation of amacrine cells; however, because of limitations associated with transient transgenic analysis, it was not feasible to establish whether this promoter was active in all amacrine cells or in a specific population of amacrine cells. By making mice transgenic for the zebrafish Pax6a BAC reporter transgene, we were able to show that paired-containing and paired-less Pax6a transcripts were differentially expressed in amacrine subpopulations. Our study also directly demonstrates the functional conservation of the regulatory mechanisms governing Pax6 transcription in teleosts and mammals.

  9. Impairment of the cortisol stress response mediated by the hypothalamus-pituitary-interrenal (HPI) axis in zebrafish (Danio rerio) exposed to monocrotophos pesticide.

    Science.gov (United States)

    Zhang, Xiaona; Zhong, Yan; Tian, Hua; Wang, Wei; Ru, Shaoguo

    2015-01-01

    In teleosts, an important component of the stress response is coordinated by the hypothalamic-pituitary-interrenal (HPI) axis. Environmental contaminants might disrupt the stress axis and consequently affect the stress response in fish. To investigate the effect of monocrotophos (MCP) pesticide on the stress response of fish and its potential mechanisms, adult zebrafish (Danio rerio) were exposed to 0, 1, 10, and 100μg/L of a 40% MCP-based pesticide for 21d, after which time fish were subjected to a 3-min air-exposure stressor. Concentrations of the whole-body cortisol were measured by radioimmunoassay and abundances of transcripts of proteins involved in the HPI axis were determined using quantitative real-time PCR. Results showed that 100μg/L of MCP pesticide decreased whole-body cortisol levels of female zebrafish in response to an acute stressor, but without any effect on the cortisol response in males. 100μg/L MCP pesticide reduced POMC and GR expression in the brain, MC2R and P45011β expression in the head kidney, but enhanced 20β-HSD2 expression in the head kidney, suggesting that MCP damaged the HPI axis involving acting at pituitary regulatory levels, inhibiting cortisol synthesis and stimulating cortisol catabolism, or disturbing the negative feedback regulation. Additionally, MCP depressed liver GR transcription but did not affect phosphoenolpyruvate carboxykinase and tyrosine aminotransferase expression in zebrafish, suggesting a role for this pesticide in reducing target tissue responsiveness to cortisol. Considered together, the reduced ability to elevate cortisol levels in response to an acute stress may be an endocrine dysfunction occurring in zebrafish subchronically exposed to MCP pesticide.

  10. CERKL knockdown causes retinal degeneration in zebrafish.

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    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.

  11. Effects of dietary exposure to brominated flame retardant BDE-47 on thyroid condition, gonadal development and growth of zebrafish

    Science.gov (United States)

    Torres, Leticia; Orazio, Carl E.; Peterman, Paul H.; Patino, Reynaldo

    2013-01-01

    Little is known about the effects of brominated flame retardants in teleosts and some of the information currently available is inconsistent. This study examined effects of dietary exposure to 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) on thyroid condition, body mass and size, and gonadal development of zebrafish. Pubertal, 49-day-old (posthatch) fish were fed diets without BDE-47 (control) or with 1, 5 or 25 μg/g BDE-47/diet. Treatments were conducted in triplicate 30-L tanks each containing 50 zebrafish, and 15 fish per treatment (5 per tank) were sampled at days 40, 80 and 120 of exposure. Measurements were taken of body mass, standard length, head depth and head length. Sex (at 40–120 days of exposure), germ cell stage (at 40 days) and thyroid condition (at 120 days; follicular cell height, colloid depletion, angiogenesis) were histologically determined. Whole-body BDE-47 levels at study completion were within the high end of levels reported in environmentally exposed (wild) fishes. Analysis of variance was used to determine differences among treatments at each sampling time. No effects were observed on thyroid condition or germ cell stage in either sex. Reduced head length was observed in females exposed to BDE-47 at 80 days but not at 40 or 120 days. In males, no apparent effects of BDE-47 were observed at 40 and 80 days, but fish exposed to 25 μg/g had lower body mass at 120 days compared to control fish. These observations suggest that BDE-47 at environmentally relevant whole-body concentrations does not affect thyroid condition or pubertal development of zebrafish but does affect growth during the juvenile-to-adult transition, especially in males.

  12. Temporal pattern of loss/persistence of duplicate genes involved in signal transduction and metabolic pathways after teleost-specific genome duplication

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    Sato Yukuto

    2009-06-01

    Full Text Available Abstract Background Recent genomic studies have revealed a teleost-specific third-round whole genome duplication (3R-WGD event occurred in a common ancestor of teleost fishes. However, it is unclear how the genes duplicated in this event were lost or persisted during the diversification of teleosts, and therefore, how many of the duplicated genes contribute to the genetic differences among teleosts. This subject is also important for understanding the process of vertebrate evolution through WGD events. We applied a comparative evolutionary approach to this question by focusing on the genes involved in long-term potentiation, taste and olfactory transduction, and the tricarboxylic acid cycle, based on the whole genome sequences of four teleosts; zebrafish, medaka, stickleback, and green spotted puffer fish. Results We applied a state-of-the-art method of maximum-likelihood phylogenetic inference and conserved synteny analyses to each of 130 genes involved in the above biological systems of human. These analyses identified 116 orthologous gene groups between teleosts and tetrapods, and 45 pairs of 3R-WGD-derived duplicate genes among them. This suggests that more than half [(45×2/(116+45] = 56.5% of the loci, probably more than ten thousand genes, present in a common ancestor of the four teleosts were still duplicated after the 3R-WGD. The estimated temporal pattern of gene loss suggested that, after the 3R-WGD, many (71/116 of the duplicated genes were rapidly lost during the initial 75 million years (MY, whereas on average more than half (27.3/45 of the duplicated genes remaining in the ancestor of the four teleosts (45/116 have persisted for about 275 MY. The 3R-WGD-derived duplicates that have persisted for a long evolutionary periods of time had significantly larger number of interacting partners and longer length of protein coding sequence, implying that they tend to be more multifunctional than the singletons after the 3R-WGD. Conclusion

  13. Arsenic transport by zebrafish aquaglyceroporins

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    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

  14. Functional Assessment of Cardiac Responses of Adult Zebrafish (Danio rerio to Acute and Chronic Temperature Change Using High-Resolution Echocardiography.

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    Ling Lee

    Full Text Available The zebrafish (Danio rerio is an important organism as a model for understanding vertebrate cardiovascular development. However, little is known about adult ZF cardiac function and how contractile function changes to cope with fluctuations in ambient temperature. The goals of this study were to: 1 determine if high resolution echocardiography (HRE in the presence of reduced cardiodepressant anesthetics could be used to accurately investigate the structural and functional properties of the ZF heart and 2 if the effect of ambient temperature changes both acutely and chronically could be determined non-invasively using HRE in vivo. Heart rate (HR appears to be the critical factor in modifying cardiac output (CO with ambient temperature fluctuation as it increases from 78 ± 5.9 bpm at 18°C to 162 ± 9.7 bpm at 28°C regardless of acclimation state (cold acclimated CA- 18°C; warm acclimated WA- 28°C. Stroke volume (SV is highest when the ambient temperature matches the acclimation temperature, though this difference did not constitute a significant effect (CA 1.17 ± 0.15 μL at 18°C vs 1.06 ± 0.14 μl at 28°C; WA 1.10 ± 0.13 μL at 18°C vs 1.12 ± 0.12 μl at 28°C. The isovolumetric contraction time (IVCT was significantly shorter in CA fish at 18°C. The CA group showed improved systolic function at 18°C in comparison to the WA group with significant increases in both ejection fraction and fractional shortening and decreases in IVCT. The decreased early peak (E velocity and early peak velocity / atrial peak velocity (E/A ratio in the CA group are likely associated with increased reliance on atrial contraction for ventricular filling.

  15. Short-term exposure to low concentrations of the synthetic androgen methyltestosterone affects vitellogenin and steroid levels in adult male zebrafish (Danio rerio).

    Science.gov (United States)

    Andersen, Lene; Goto-Kazeto, Rie; Trant, John M; Nash, Jon P; Korsgaard, Bodil; Bjerregaard, Poul

    2006-03-10

    Short-term effects of methyltestosterone (MT) on the endocrine system of adult male zebrafish (Danio rerio) were examined. Males were exposed to 0, 4.5, 6.6, 8.5, 19.8, 35.9, 62.3 ng MT/l and ethinylestradiol (EE2) (26.4 ng/l) for 7 days. Several physiological endpoints that may be affected by endocrine disrupters were analysed, specifically vitellogenin (VTG) concentration, estradiol (E2), testosterone (T), and 11-ketotestosterone (KT) content, brain aromatase activity and gene expression of CYP19A1 and CYP19A2 in the testis. Exposure to the lowest MT concentration (4.5 ng MT/l), and the EE2 increased the concentration of VTG significantly compared to solvent control group. Exposure to higher concentrations of MT did not increase VTG levels. Endogenous KT and T levels decreased significantly in a concentration-dependent manner in response to the MT exposure and the lowest effective concentrations were 6.4 and 8.5 ng MT/l, respectively. The levels of KT and T were also significantly suppressed by EE2 when compared to the solvent control group. Significant decreases in endogenous E2 levels were found in some MT groups but it was not possible to distinguish a simple concentration-response relationship. No effects of MT or EE2 on the brain aromatase activity or on testicular gene expression of CYP19A1 and CYP19A2 were detected. The results show that androgens such as MT can act as endocrine disrupters even at very low concentrations.

  16. Mutations in zebrafish lrp2 result in adult-onset ocular pathogenesis that models myopia and other risk factors for glaucoma.

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    Kerry N Veth

    2011-02-01

    Full Text Available The glaucomas comprise a genetically complex group of retinal neuropathies that typically occur late in life and are characterized by progressive pathology of the optic nerve head and degeneration of retinal ganglion cells. In addition to age and family history, other significant risk factors for glaucoma include elevated intraocular pressure (IOP and myopia. The complexity of glaucoma has made it difficult to model in animals, but also challenging to identify responsible genes. We have used zebrafish to identify a genetically complex, recessive mutant that shows risk factors for glaucoma including adult onset severe myopia, elevated IOP, and progressive retinal ganglion cell pathology. Positional cloning and analysis of a non-complementing allele indicated that non-sense mutations in low density lipoprotein receptor-related protein 2 (lrp2 underlie the mutant phenotype. Lrp2, previously named Megalin, functions as an endocytic receptor for a wide-variety of bioactive molecules including Sonic hedgehog, bone morphogenic protein 4, retinol-binding protein, vitamin D-binding protein, and apolipoprotein E, among others. Detailed phenotype analyses indicated that as lrp2 mutant fish age, many individuals--but not all--develop high IOP and severe myopia with obviously enlarged eye globes. This results in retinal stretch and prolonged stress to retinal ganglion cells, which ultimately show signs of pathogenesis. Our studies implicate altered Lrp2-mediated homeostasis as important for myopia and other risk factors for glaucoma in humans and establish a new genetic model for further study of phenotypes associated with this disease.

  17. Sodium and chloride transport in soft water and hard water acclimated zebrafish (Danio rerio)

    DEFF Research Database (Denmark)

    Boisen, A M Z; Amstrup, J; Novak, I;

    2003-01-01

    While the zebrafish is commonly used for studies of developmental biology and toxicology, very little is known about their osmoregulatory physiology. The present investigation of Na(+) and Cl(-) transport revealed that the zebrafish is able to tolerate extremely low ambient ion concentrations...... and that this is achieved at least in part by a greatly enhanced apparent uptake capacity and affinity for both ions. Zebrafish maintain plasma and whole body electrolyte concentrations similar to most other freshwater teleosts even in deionized water containing only 35 microM NaCl, i.e soft water. We recorded an extremely...... inhibitor was more variable. Differential response of Na(+) uptake to amiloride depending on acclimation medium suggests that different Na(+) transport mechanisms are employed by zebrafish acclimated to soft and hard water....

  18. Expression and functions of ASIC1 in the zebrafish retina.

    Science.gov (United States)

    Liu, Sha; Wang, Mei-Xia; Mao, Cheng-Jie; Cheng, Xiao-Yu; Wang, Chen-Tao; Huang, Jian; Zhong, Zhao-Min; Hu, Wei-Dong; Wang, Fen; Hu, Li-Fang; Wang, Han; Liu, Chun-Feng

    2014-12-12

    It has been demonstrated that acid sensing ionic channels (ASICs) are present in the central and peripheral nervous system of mammals, including the retina. However, it remains unclear whether the zebrafish retina also expresses ASICs. In the present study, the expression and distribution of zasic1 were examined in the retina of zebrafish. Both zasic1 mRNA and protein expressions were detected in the adult zebrafish retina. A wide distribution of ASIC1 in zebrafish retina was confirmed using whole mount in situ hybridization and immunohistochemistry study. Acidosis-induced currents in the isolated retinal ganglion cells (RGCs) were also recorded using whole cell patch clamping. Moreover, blockade of ASICs channel significantly reduced the locomotion of larval zebrafish in response to light exposure. In sum, our data demonstrate the presence of ASIC1 and its possible functional relevance in the retina of zebrafish.

  19. Acquisition of glial cells missing 2 enhancers contributes to a diversity of ionocytes in zebrafish.

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    Takanori Shono

    Full Text Available Glial cells missing 2 (gcm2 encoding a GCM-motif transcription factor is expressed in the parathyroid in amniotes. In contrast, gcm2 is expressed in pharyngeal pouches (a homologous site of the parathyroid, gills, and H(+-ATPase-rich cells (HRCs, a subset of ionocytes on the skin surface of the teleost fish zebrafish. Ionocytes are specialized cells that are involved in osmotic homeostasis in aquatic vertebrates. Here, we showed that gcm2 is essential for the development of HRCs and Na(+-Cl(- co-transporter-rich cells (NCCCs, another subset of ionocytes in zebrafish. We also identified gcm2 enhancer regions that control gcm2 expression in ionocytes of zebrafish. Comparisons of the gcm2 locus with its neighboring regions revealed no conserved elements between zebrafish and tetrapods. Furthermore, We observed gcm2 expression patterns in embryos of the teleost fishes Medaka (Oryzias latipes and fugu (Fugu niphobles, the extant primitive ray-finned fishes Polypterus (Polypterus senegalus and sturgeon (a hybrid of Huso huso × Acipenser ruhenus, and the amphibian Xenopus (Xenopus laevis. Although gcm2-expressing cells were observed on the skin surface of Medaka and fugu, they were not found in Polypterus, sturgeon, or Xenopus. Our results suggest that an acquisition of enhancers for the expression of gcm2 contributes to a diversity of ionocytes in zebrafish during evolution.

  20. The pH sensitivity of Aqp0 channels in tetraploid and diploid teleosts.

    Science.gov (United States)

    Chauvigné, François; Zapater, Cinta; Stavang, Jon Anders; Taranger, Geir Lasse; Cerdà, Joan; Finn, Roderick Nigel

    2015-05-01

    Water homeostasis and the structural integrity of the vertebrate lens is partially mediated by AQP0 channels. Emerging evidence indicates that external pH may be involved in channel gating. Here we show that a tetraploid teleost, the Atlantic salmon, retains 4 aqp0 genes (aqp0a1, -0a2, -0b1, and -0b2), which are highly, but not exclusively, expressed in the lens. Functional characterization reveals that, although each paralog permeates water efficiently, the permeability is respectively shifted to the neutral, alkaline, or acidic pH in Aqp0a1, -0a2, and -0b1, whereas that of Aqp0b2 is not regulated by external pH. Mutagenesis studies demonstrate that Ser(38), His(39), and His(40) residues in the extracellular transmembrane domain of α-helix 2 facing the water pore are critical for the pH modulation of water transport. To validate these findings, we show that both zebrafish Aqp0a and -0b are functional water channels with respective pH sensitivities toward alkaline or acid pH ranges and that an N-terminal allelic variant (Ser(19)) of Aqp0b exists that abolishes water transport in Xenopus laevis oocytes. The data suggest that the alkaline pH sensitivity is a conserved trait in teleost Aqp0 a-type channels, whereas mammalian AQP0 and some teleost Aqp0 b-type channels display an acidic pH permeation preference.

  1. Recent advances in the study of zebrafish extracellular matrix proteins.

    Science.gov (United States)

    Jessen, Jason R

    2015-05-01

    The zebrafish extracellular matrix (ECM) is a dynamic and pleomorphic structure consisting of numerous proteins that together regulate a variety of cellular and morphogenetic events beginning as early as gastrulation. The zebrafish genome encodes a similar complement of ECM proteins as found in other vertebrate organisms including glycoproteins, fibrous proteins, proteoglycans, glycosaminoglycans, and interacting or modifying proteins such as integrins and matrix metalloproteinases. As a genetic model system combined with its amenability to high-resolution microscopic imaging, the zebrafish allows interrogation of ECM protein structure and function in both the embryo and adult. Accumulating data have identified important roles for zebrafish ECM proteins in processes as diverse as cell polarity, migration, tissue mechanics, organ laterality, muscle contraction, and regeneration. In this review, I highlight recently published data on these topics that demonstrate how the ECM proteins fibronectin, laminin, and collagen contribute to zebrafish development and adult homeostasis.

  2. Transient laminin beta 1a Induction Defines the Wound Epidermis during Zebrafish Fin Regeneration.

    Science.gov (United States)

    Chen, Chen-Hui; Merriman, Alexander F; Savage, Jeremiah; Willer, Jason; Wahlig, Taylor; Katsanis, Nicholas; Yin, Viravuth P; Poss, Kenneth D

    2015-08-01

    The first critical stage in salamander or teleost appendage regeneration is creation of a specialized epidermis that instructs growth from underlying stump tissue. Here, we performed a forward genetic screen for mutations that impair this process in amputated zebrafish fins. Positional cloning and complementation assays identified a temperature-sensitive allele of the ECM component laminin beta 1a (lamb1a) that blocks fin regeneration. lamb1a, but not its paralog lamb1b, is sharply induced in a subset of epithelial cells after fin amputation, where it is required to establish and maintain a polarized basal epithelial cell layer. These events facilitate expression of the morphogenetic factors shha and lef1, basolateral positioning of phosphorylated Igf1r, patterning of new osteoblasts, and regeneration of bone. By contrast, lamb1a function is dispensable for juvenile body growth, homeostatic adult tissue maintenance, repair of split fins, or renewal of genetically ablated osteoblasts. fgf20a mutations or transgenic Fgf receptor inhibition disrupt lamb1a expression, linking a central growth factor to epithelial maturation during regeneration. Our findings reveal transient induction of lamb1a in epithelial cells as a key, growth factor-guided step in formation of a signaling-competent regeneration epidermis.

  3. Transient laminin beta 1a Induction Defines the Wound Epidermis during Zebrafish Fin Regeneration.

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    Chen-Hui Chen

    2015-08-01

    Full Text Available The first critical stage in salamander or teleost appendage regeneration is creation of a specialized epidermis that instructs growth from underlying stump tissue. Here, we performed a forward genetic screen for mutations that impair this process in amputated zebrafish fins. Positional cloning and complementation assays identified a temperature-sensitive allele of the ECM component laminin beta 1a (lamb1a that blocks fin regeneration. lamb1a, but not its paralog lamb1b, is sharply induced in a subset of epithelial cells after fin amputation, where it is required to establish and maintain a polarized basal epithelial cell layer. These events facilitate expression of the morphogenetic factors shha and lef1, basolateral positioning of phosphorylated Igf1r, patterning of new osteoblasts, and regeneration of bone. By contrast, lamb1a function is dispensable for juvenile body growth, homeostatic adult tissue maintenance, repair of split fins, or renewal of genetically ablated osteoblasts. fgf20a mutations or transgenic Fgf receptor inhibition disrupt lamb1a expression, linking a central growth factor to epithelial maturation during regeneration. Our findings reveal transient induction of lamb1a in epithelial cells as a key, growth factor-guided step in formation of a signaling-competent regeneration epidermis.

  4. Deriving cell lines from zebrafish embryos and tumors.

    Science.gov (United States)

    Choorapoikayil, Suma; Overvoorde, John; den Hertog, Jeroen

    2013-09-01

    Over the last two decades the zebrafish has emerged as a powerful model organism in science. The experimental accessibility, the broad range of zebrafish mutants, and the highly conserved genetic and biochemical pathways between zebrafish and mammals lifted zebrafish to become one of the most attractive vertebrate models to study gene function and to model human diseases. Zebrafish cell lines are highly attractive to investigate cell biology and zebrafish cell lines complement the experimental tools that are available already. We established a straightforward method to culture cells from a single zebrafish embryo or a single tumor. Here we describe the generation of fibroblast-like cell lines from wild-type and ptenb(-/-) embryos and an endothelial-like cell line from a tumor of an adult ptena(+/-)ptenb(-/-) zebrafish. This protocol can easily be adapted to establish stable cell lines from any mutant or transgenic zebrafish line and the average time to obtain a pro-stable cell line is 3-5 months.

  5. Effects of ethanol exposure on nervous system development in zebrafish.

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    Cole, Gregory J; Zhang, Chengjin; Ojiaku, Princess; Bell, Vanessa; Devkota, Shailendra; Mukhopadhyay, Somnath

    2012-01-01

    Alcohol (ethanol) is a teratogen that adversely affects nervous system development in a wide range of animal species. In humans numerous congenital abnormalities arise as a result of fetal alcohol exposure, leading to a spectrum of disorders referred to as fetal alcohol spectrum disorder (FASD). These abnormalities include craniofacial defects as well as neurological defects that affect a variety of behaviors. These human FASD phenotypes are reproduced in the rodent central nervous system (CNS) following prenatal ethanol exposure. While the study of ethanol effects on zebrafish development has been more limited, several studies have shown that different strains of zebrafish exhibit differential susceptibility to ethanol-induced cyclopia, as well as behavioral deficits. Molecular mechanisms underlying the effects of ethanol on CNS development also appear to be shared between rodent and zebrafish. Thus, zebrafish appear to recapitulate the observed effects of ethanol on human and mouse CNS development, indicating that zebrafish can serve as a complimentary developmental model system to study the molecular basis of FASD. Recent studies examining the effect of ethanol exposure on zebrafish nervous system development are reviewed, with an emphasis on attempts to elucidate possible molecular pathways that may be impacted by developmental ethanol exposure. Recent work from our laboratories supports a role for perturbed extracellular matrix function in the pathology of ethanol exposure during zebrafish CNS development. The use of the zebrafish model to assess the effects of ethanol exposure on adult nervous system function as manifested by changes in zebrafish behavior is also discussed.

  6. In silico and in situ characterization of the zebrafish (Danio rerio gnrh3 (sGnRH gene

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    Husebye Harald

    2002-08-01

    Full Text Available Abstract Background Gonadotropin releasing hormone (GnRH is responsible for stimulation of gonadotropic hormone (GtH in the hypothalamus-pituitary-gonadal axis (HPG. The regulatory mechanisms responsible for brain specificity make the promoter attractive for in silico analysis and reporter gene studies in zebrafish (Danio rerio. Results We have characterized a zebrafish [Trp7, Leu8] or salmon (s GnRH variant, gnrh3. The gene includes a 1.6 Kb upstream regulatory region and displays the conserved structure of 4 exons and 3 introns, as seen in other species. An in silico defined enhancer at -976 in the zebrafish promoter, containing adjacent binding sites for Oct-1, CREB and Sp1, was predicted in 2 mammalian and 5 teleost GnRH promoters. Reporter gene studies confirmed the importance of this enhancer for cell specific expression in zebrafish. Interestingly the promoter of human GnRH-I, known as mammalian GnRH (mGnRH, was shown capable of driving cell specific reporter gene expression in transgenic zebrafish. Conclusions The characterized zebrafish Gnrh3 decapeptide exhibits complete homology to the Atlantic salmon (Salmo salar GnRH-III variant. In silico analysis of mammalian and teleost GnRH promoters revealed a conserved enhancer possessing binding sites for Oct-1, CREB and Sp1. Transgenic and transient reporter gene expression in zebrafish larvae, confirmed the importance of the in silico defined zebrafish enhancer at -976. The capability of the human GnRH-I promoter of directing cell specific reporter gene expression in zebrafish supports orthology between GnRH-I and GnRH-III.

  7. FishNet: an online database of zebrafish anatomy

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    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.

  8. Lipid droplet biology and evolution illuminated by the characterization of a novel perilipin in teleost fish

    Science.gov (United States)

    Granneman, James G; Kimler, Vickie A; Zhang, Huamei; Ye, Xiangqun; Luo, Xixia; Postlethwait, John H; Thummel, Ryan

    2017-01-01

    Perilipin (PLIN) proteins constitute an ancient family important in lipid droplet (LD) formation and triglyceride metabolism. We identified an additional PLIN clade (plin6) that is unique to teleosts and can be traced to the two whole genome duplications that occurred early in vertebrate evolution. Plin6 is highly expressed in skin xanthophores, which mediate red/yellow pigmentation and trafficking, but not in tissues associated with lipid metabolism. Biochemical and immunochemical analyses demonstrate that zebrafish Plin6 protein targets the surface of pigment-containing carotenoid droplets (CD). Protein kinase A (PKA) activation, which mediates CD dispersion in xanthophores, phosphorylates Plin6 on conserved residues. Knockout of plin6 in zebrafish severely impairs the ability of CD to concentrate carotenoids and prevents tight clustering of CD within carotenoid bodies. Ultrastructural and functional analyses indicate that LD and CD are homologous structures, and that Plin6 was functionalized early in vertebrate evolution for concentrating and trafficking pigment. DOI: http://dx.doi.org/10.7554/eLife.21771.001 PMID:28244868

  9. Identification of MicroRNAs in Zebrafish Spermatozoa.

    Science.gov (United States)

    Jia, Kun-Tong; Zhang, Jing; Jia, Peng; Zeng, Lin; Jin, Yilin; Yuan, Yongming; Chen, Jieying; Hong, Yunhan; Yi, Meisheng

    2015-12-01

    MicroRNAs (miRNAs) participate in almost all biological processes. Plenty of evidences show that some testis- or spermatozoa-specific miRNAs play crucial roles in the process of gonad and germ cell development. In this study, the spermatozoa miRNA profiles were investigated through a combination of illumina deep sequencing and bioinformatics analysis in zebrafish. Deep sequencing of small RNAs yielded 11,820,680 clean reads. By mapping to the zebrafish genome, we identified 400 novel and 204 known miRNAs that could be grouped into 104 families. Furthermore, we selected the six highest expressions of known miRNAs to detect their expression patterns in different tissues by stem-loop quantitative real-time polymerase chain reaction. We found that among the six miRNAs, dre-miR-202-5p displayed specific and high expression in zebrafish spermatozoa and testis. Fluorescence in situ hybridization analysis indicated that dre-miR-202-5p was predominantly expressed in all kind of germ cells at different spermatogenetic stages, including spermatogonia and spermatozoa, but barely expressed in the germ cells in the ovary. This sex-biased expression pattern suggests that dre-miR-202-5p might be related to spermatogenesis and the functioning of spermatozoa. The identification of miRNAs in zebrafish spermatozoa and germ cells offers new insights into the spermatogenesis and spermatozoa in the teleost and other vertebrates.

  10. Antarctic teleost immunoglobulins: more extreme, more interesting.

    Science.gov (United States)

    Coscia, Maria Rosaria; Varriale, Sonia; Giacomelli, Stefano; Oreste, Umberto

    2011-11-01

    We have investigated the immunoglobulin molecule and the genes encoding it in teleosts living in the Antarctic seas at the constant temperature of -1.86 °C. The majority of Antarctic teleosts belong to the suborder Notothenioidei (Perciformes), which includes only a few non-Antarctic species. Twenty-one Antarctic and two non-Antarctic Notothenioid species were included in our studies. We sequenced immunoglobulin light chains in two species and μ heavy chains, partially or totally, in twenty species. In the case of heavy chain, genomic DNA and the cDNA encoding the secreted and the membrane form were analyzed. From one species, Trematomus bernacchii, a spleen cDNA library was constructed to evaluate the diversity of VH gene segments. T. bernacchii IgM, purified from the serum and bile, was characterized. Homology Modelling and Molecular Dynamics were used to determine the molecular structure of T. bernacchii and Chionodraco hamatus immunoglobulin domains. This paper sums up the previous results and broadens them with the addition of unpublished data.

  11. Expression of gdnf and nos in adult zebrafish brain during the regeneration after spinal cord injury%成年斑马鱼脊髓损伤修复中脑gdnf 和nos 基因的表达

    Institute of Scientific and Technical Information of China (English)

    谢琳; 房萍; 林金飞; 潘洪超; 张帆; 申延琴

    2013-01-01

    成年斑马鱼(Danio rerio)具有很强的脊髓损伤后自主修复的能力,但目前其机制不明.为了研究斑马鱼中脑组织对脊髓再生的影响,文章应用成年斑马鱼脊髓损伤模型,采用实时定量PCR 方法和原位杂交技术,检测了斑马鱼脑中胶质细胞源性神经营养因子(gdnf)和一氧化氮合酶(nos)基因在脊髓损伤后4 h、12 h、6 d、11 d的表达情况,展示了这两种基因在斑马鱼脑内不同核团的动态表达变化.结果显示,成年斑马鱼脊髓损伤后,神经营养因子gdnf 基因在损伤急性期(4 h、12 h)和神经修复期(6 d、11 d)于斑马鱼脑内呈现显著性升高(P<0.05),而一氧化氮合酶基因nos 的表达于损伤急性期显著性升高 (P<0.05),随后下降,并在修复期 (11 d)显著降低(P<0.05).这表明,脊髓损伤后,高表达gdnf 基因同时低表达nos 基因的脑环境给脊髓损伤提供了良好的神经再生微环境,从而可能促进轴突的再生长及运动能力的恢复.%Recently, it is unclear about the mechanism of notable regenerated ability of adult zebrafish after spinal cord injury. To investigate the effects of brain on restoration from spinal cord injury, adult zebrafish spinal cord injury model was built and brain samples were dissected at different time points after the injury. Real-time quantitative PCR and in situ hybridization were applied to reveal the dynamics of glial cell line-derived neurotrophic factor (gdnf) and nitric oxide synthases (nos) mRNA expression in various regions of zebrafish brain. The results showed that, compared to sham group at each time points separately, the expression of gdnf mRNA in adult zebrafish brain during both acute phase (4 h and 12 h) and chronic phase of neuroregeneration (6 d and 11d) increased significantly (P<0.05). The expression of nos mRNA in zebrafish brain enhanced during acute phase, and then reduced to the level lower than the sham group during the chronic phase of neuroregeneration

  12. Ftr82 Is Critical for Vascular Patterning during Zebrafish Development

    Directory of Open Access Journals (Sweden)

    Hsueh-Wei Chang

    2017-01-01

    Full Text Available Cellular components and signaling pathways are required for the proper growth of blood vessels. Here, we report for the first time that a teleost-specific gene ftr82 (finTRIM family, member 82 plays a critical role in vasculature during zebrafish development. To date, there has been no description of tripartite motif proteins (TRIM in vascular development, and the role of ftr82 is unknown. In this study, we found that ftr82 mRNA is expressed during the development of vessels, and loss of ftr82 by morpholino (MO knockdown impairs the growth of intersegmental vessels (ISV and caudal vein plexus (CVP, suggesting that ftr82 plays a critical role in promoting ISV and CVP growth. We showed the specificity of ftr82 MO by analyzing ftr82 expression products and expressing ftr82 mRNA to rescue ftr82 morphants. We further showed that the knockdown of ftr82 reduced ISV cell numbers, suggesting that the growth impairment of vessels is likely due to a decrease of cell proliferation and migration, but not cell death. In addition, loss of ftr82 affects the expression of vascular markers, which is consistent with the defect of vascular growth. Finally, we showed that ftr82 likely interacts with vascular endothelial growth factor (VEGF and Notch signaling. Together, we identify teleost-specific ftr82 as a vascular gene that plays an important role for vascular development in zebrafish.

  13. Crypt cells are involved in kin recognition in larval zebrafish

    Science.gov (United States)

    Biechl, Daniela; Tietje, Kristin; Gerlach, Gabriele; Wullimann, Mario F.

    2016-01-01

    Zebrafish larvae imprint on visual and olfactory kin cues at day 5 and 6 postfertilization, respectively, resulting in kin recognition later in life. Exposure to non-kin cues prevents imprinting and kin recognition. Imprinting depends on MHC class II related signals and only larvae sharing MHC class II alleles can imprint on each other. Here, we analyzed which type of olfactory sensory neuron (OSN) detects kin odor. The single teleost olfactory epithelium harbors ciliated OSNs carrying OR and TAAR gene family receptors (mammals: main olfactory epithelium) and microvillous OSNs with V1R and V2R gene family receptors (mammals: vomeronasal organ). Additionally, teleosts exhibit crypt cells which possess microvilli and cilia. We used the activity marker pERK (phosphorylated extracellular signal regulated kinase) after stimulating 9 day old zebrafish larvae with either non-kin conspecific or food odor. While food odor activated both ciliated and microvillous OSNs, only the latter were activated by conspecific odor, crypt cells showed no activation to both stimuli. Then, we tested imprinted and non-imprinted larvae (full siblings) for kin odor detection. We provide the first direct evidence that crypt cells, and likely a subpopulation of microvillous OSNs, but not ciliated OSNs, play a role in detecting a kin odor related signal. PMID:27087508

  14. Characterization of snakehead rhabdovirus infection in zebrafish (Danio rerio).

    Science.gov (United States)

    Phelan, Peter E; Pressley, Meagan E; Witten, P Eckhard; Mellon, Mark T; Blake, Sharon; Kim, Carol H

    2005-02-01

    The zebrafish, Danio rerio, has become recognized as a valuable model for the study of development, genetics, and toxicology. Recently, the zebrafish has been recognized as a useful model for infectious disease and immunity. In this study, the pathogenesis and antiviral immune response of zebrafish to experimental snakehead rhabdovirus (SHRV) infection was characterized. Zebrafish 24 h postfertilization to 30 days postfertilization were susceptible to infection by immersion in 10(6) 50% tissue culture infective doses (TCID50) of SHRV/ml, and adult zebrafish were susceptible to infection by intraperitoneal (i.p.) injection of 10(5) TCID50 of SHRV/ml. Mortalities exceeded 40% in infected fish, and clinical presentation of infection included petechial hemorrhaging, redness of the abdomen, and erratic swim behavior. Virus reisolation and reverse transcription-PCR analysis of the viral nucleocapsid gene confirmed the presence of SHRV. Histological sections of moribund embryonic and juvenile fish revealed necrosis of the pharyngeal epithelium and liver, in addition to congestion of the swim bladder by cell debris. Histopathology in adult fish injected i.p. was confined to the site of injection. The antiviral response in zebrafish was monitored by quantitative real-time PCR analysis of zebrafish interferon (IFN) and Mx expression. IFN and Mx levels were elevated in zebrafish exposed to SHRV, although expression and intensity differed with age and route of infection. This study is the first to examine the pathogenesis of SHRV infection in zebrafish. Furthermore, this study is the first to describe experimental infection of zebrafish embryos with a viral pathogen, which will be important for future experiments involving targeted gene disruption and forward genetic screens.

  15. Cadmium potentiates toxicity of cypermethrin in zebrafish.

    Science.gov (United States)

    Yang, Ye; Ye, Xiaoqing; He, Buyuan; Liu, Jing

    2016-02-01

    Co-occurrence of pesticides such as synthetic pyrethroids and metals in aquatic ecosystems raises concerns over their combined ecological effects. Cypermethrin, 1 of the top 5 synthetic pyrethroids in use, has been extensively detected in surface water. Cadmium (Cd) has been recognized as 1 of the most toxic metals and is a common contaminant in the aquatic system. However, little information is available regarding their joint toxicity. In the present study, combined toxicity of cypermethrin and Cd and the underlying mechanisms were investigated. Zebrafish embryos and adults were exposed to the individual contaminant or binary mixtures. Co-exposure to cypermethrin and Cd produced synergistic effects on the occurrence of crooked body, pericardial edema, and noninflation of swim bladder. The addition of Cd significantly potentiated cypermethrin-induced spasms and caused more oxidative stress in zebrafish larvae. Cypermethrin-mediated induction of transcription levels and catalytic activities of cytochrome P450 (CYP) enzyme were significantly down-regulated by Cd in both zebrafish larvae and adults. Chemical analytical data showed that in vitro elimination of cypermethrin by CYP1A1 was inhibited by Cd. The addition of Cd caused an elevation of in vivo cypermethrin residue levels in the mixture-exposed adult zebrafish. These results suggest that the enhanced toxicity of cypermethrin in the presence of Cd results from the inhibitory effects of Cd on CYP-mediated biotransformation of this pesticide. The authors' findings provide a deeper understanding of the mechanistic basis accounting for the joint toxicity of cypermethrin and Cd.

  16. 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.

  17. Adult Neurogenesis in Fish.

    Science.gov (United States)

    Ganz, Julia; Brand, Michael

    2016-07-01

    Teleost fish have a remarkable neurogenic and regenerative capacity in the adult throughout the rostrocaudal axis of the brain. The distribution of proliferation zones shows a remarkable conservation, even in distantly related teleost species, suggesting a common teleost ground plan of proliferation zones. There are different progenitor populations in the neurogenic niches-progenitors positive for radial glial markers (dorsal telencephalon, hypothalamus) and progenitors with neuroepithelial-like characteristics (ventral telencephalon, optic tectum, cerebellum). Definition of these progenitors has allowed studying their role in normal growth of the adult brain, but also when challenged following a lesion. From these studies, important roles have emerged for intrinsic mechanisms and extrinsic signals controlling the activation of adult neurogenesis that enable regeneration of the adult brain to occur, opening up new perspectives on rekindling regeneration also in the context of the mammalian brain.

  18. Neuroendocrinology of reproduction in teleost fish.

    Science.gov (United States)

    Zohar, Yonathan; Muñoz-Cueto, José Antonio; Elizur, Abigail; Kah, Olivier

    2010-02-01

    This review aims at synthesizing the most relevant information regarding the neuroendocrine circuits controlling reproduction, mainly gonadotropin release, in teleost fish. In teleosts, the pituitary receives a more or less direct innervation by neurons sending projections to the vicinity of the pituitary gonadotrophs. Among the neurotransmitters and neuropeptides released by these nerve endings are gonadotrophin-releasing hormones (GnRH) and dopamine, acting as stimulatory and inhibitory factors (in many but not all fish) on the liberation of LH and to a lesser extent that of FSH. The activity of the corresponding neurons depends on a complex interplay between external and internal factors that will ultimately influence the triggering of puberty and sexual maturation. Among these factors are sex steroids and other peripheral hormones and growth factors, but little is known regarding their targets. However, very recently a new actor has entered the field of reproductive physiology. KiSS1, first known as a tumor suppressor called metastin, and its receptor GPR54, are now central to the regulation of GnRH, and consequently LH and FSH secretion in mammals. The KiSS system is notably viewed as instrumental in integrating both environmental cues and metabolic signals and passing this information onto the reproductive axis. In fish, there are two KiSS genes, KiSS1 and KiSS2, expressed in neurons of the preoptic area and mediobasal hypothalamus. Pionneer studies indicate that KiSS and GPR54 expression seem to be activated at puberty. Although precise information as to the physiological effects of KiSS1 in fish, notably on GnRH neurons and gonadotropin release, is still limited, KiSS neurons may emerge as the "gatekeeper" of puberty and reproduction in fish as in mammals.

  19. The GC-heterogeneity of teleost fishes

    Directory of Open Access Journals (Sweden)

    Gautier Christian

    2008-12-01

    Full Text Available Abstract Background One of the most striking features of mammalian and birds chromosomes is the variation in the guanine-cytosine (GC content that occurs over scales of hundreds of kilobases to megabases; this is known as the "isochore" structure. Among other vertebrates the presence of isochores depends upon the taxon; isochore are clearly present in Crocodiles and turtles but fish genome seems very homogeneous on GC content. This has suggested a unique isochore origin after the divergence between Sarcopterygii and Actinopterygii, but before that between Sauropsida and mammals. However during more than 30 years of analysis, isochore characteristics have been studied and many important biological properties have been associated with the isochore structure of human genomes. For instance, the genes are more compact and their density is highest in GC rich isochores. Results This paper shows in teleost fish genomes the existence of "GC segmentation" sharing some of the characteristics of isochores although teleost fish genomes presenting a particular homogeneity in CG content. The entire genomes of T nigroviridis and D rerio are now available, and this has made it possible to check whether a mosaic structure associated with isochore properties can be found in these fishes. In this study, hidden Markov models were trained on fish genes (T nigroviridis and D rerio which were classified by using the isochore class of their human orthologous. A clear segmentation of these genomes was detected. Conclusion The GC content is an excellent indicator of isochores in heterogeneous genomes as mammals. The segmentation we obtained were well correlated with GC content and other properties associated to GC content such as gene density, the number of exons per gene and the length of introns. Therefore, the GC content is the main property that allows the detection of isochore but more biological properties have to be taken into account. This method allows detecting

  20. Husbandry of zebrafish, Danio rerio, and the cortisol stress response.

    Science.gov (United States)

    Pavlidis, Michail; Digka, Nikoletta; Theodoridi, Antonia; Campo, Aurora; Barsakis, Konstantinos; Skouradakis, Gregoris; Samaras, Athanasios; Tsalafouta, Alexandra

    2013-12-01

    The effect of common husbandry conditions (crowding, social environment, water quality, handling, and background color) on the cortisol stress response in adult zebrafish, Danio rerio, was investigated to check the usefulness of zebrafish as a model organism in aquaculture research. In addition, a noninvasive methodology for assessing stress was evaluated. Zebrafish showed a fast cortisol response with high values at 30 min that returned to basal levels within 2 h of poststress. There was a significant positive correlation between trunk cortisol concentrations and the free water cortisol rate (r(2)=0.829-0.850, pzebrafish. It is concluded that adult laboratory zebrafish had a preference for a transparent or black background aquarium, at a number of 10 individuals per 2 L of available water volume, to express their normal behavior and avoid increased cortisol stress reaction.

  1. Cardiac Hypertrophy Involves both Myocyte Hypertrophy and Hyperplasia in Anemic Zebrafish

    OpenAIRE

    Xiaojing Sun; Tiffany Hoage; Ping Bai; Yonghe Ding; Zhenyue Chen; Ruilin Zhang; Wei Huang; Ashad Jahangir; Barry Paw; Yi-Gang Li; Xiaolei Xu

    2009-01-01

    BACKGROUND: An adult zebrafish heart possesses a high capacity of regeneration. However, it has been unclear whether and how myocyte hyperplasia contributes to cardiac remodeling in response to biomechanical stress and whether myocyte hypertrophy exists in the zebrafish. To address these questions, we characterized the zebrafish mutant tr265/tr265, whose Band 3 mutation disrupts erythrocyte formation and results in anemia. Although Band 3 does not express and function in the heart, the chroni...

  2. Advances in the Fc receptor of teleosts%鱼类Fc受体研究

    Institute of Scientific and Technical Information of China (English)

    邵茜; 安利国; 杨桂文

    2009-01-01

    Fc受体是免疫细胞表面一种重要受体分子,通过与免疫球蛋白Fc段结合触发多种生物学功能,是联系体液免疫和细胞免疫的桥梁.部分硬骨鱼中已经发现了Fc受体,在斑马鱼、斑点叉尾鲴和鲤鱼中都克隆到了Fc受体的γ亚基,在鲨鱼和大西洋鲑中证明有能够与免疫球蛋白结合的Fc受体存在,并在斑点叉尾鲴、河豚和虹鳟中存在着类似α亚基的Fc受体.对鱼类Fc受体的发现和研究必将为了解鱼类的免疫机制及免疫进化提供重要的资料.%Fc receptors(FcRs) of immunoglobulins on immune cell play an important role in immune system by providing a link between antibody-antigen complexes and the effect cell. FcRs supply an essential link between humoral and cellular immunity. FcRs have been cloned in teleost fish, the γ subunit of Fc receptor has been cloned from zebrafish, catfish and common carp, while the subunits binding to immunoglobulins have been identified from shark, atlantic salmon, catfish, fugu and rainbow trout. Identification of subunits of the Fc receptors could lead to a farther investigation of the immune mechanism in teleosts and the phylogeny of Fc receptors.

  3. Evolution of Space Dependent Growth in the Teleost Astyanax mexicanus

    OpenAIRE

    2012-01-01

    The relationship between growth rate and environmental space is an unresolved issue in teleosts. While it is known from aquaculture studies that stocking density has a negative relationship to growth, the underlying mechanisms have not been elucidated, primarily because the growth rate of populations rather than individual fish were the subject of all previous studies. Here we investigate this problem in the teleost Astyanax mexicanus, which consists of a sighted surface-dwelling form (surfac...

  4. Lack of plasma kallikrein-kinin system cascade in teleosts.

    Directory of Open Access Journals (Sweden)

    Marty Kwok-Shing Wong

    Full Text Available The kallikrein-kinin system (KKS consists of two major cascades in mammals: "plasma KKS" consisting of high molecular-weight (HMW kininogen (KNG, plasma kallikrein (KLKB1, and bradykinin (BK; and "tissue KKS" consisting of low molecular-weight (LMW KNG, tissue kallikreins (KLKs, and [Lys(0]-BK. Some components of the KKS have been identified in the fishes, but systematic analyses have not been performed, thus this study aims to define the KKS components in teleosts and pave a way for future physiological and evolutionary studies. Through a combination of genomics, molecular, and biochemical methods, we showed that the entire plasma KKS cascade is absent in teleosts. Instead of two KNGs as found in mammals, a single molecular weight KNG was found in various teleosts, which is homologous to the mammalian LMW KNG. Results of molecular phylogenetic and synteny analyses indicated that the all current teleost genomes lack KLKB1, and its unique protein structure, four apple domains and one trypsin domain, could not be identified in any genome or nucleotide databases. We identified some KLK-like proteins in teleost genomes by synteny and conserved domain analyses, which could be the orthologs of tetrapod KLKs. A radioimmunoassay system was established to measure the teleost BK and we found that [Arg(0]-BK is the major circulating form instead of BK, which supports that the teleost KKS is similar to the mammalian tissue KKS. Coincidently, coelacanths are the earliest vertebrate that possess both HMW KNG and KLKB1, which implies that the plasma KKS could have evolved in the early lobe-finned fish and descended to the tetrapod lineage. The co-evolution of HMW KNG and KLKB1 in lobe-finned fish and early tetrapods may mark the emergence of the plasma KKS and a contact activation system in blood coagulation, while teleosts may have retained a single KKS cascade.

  5. WDR55 is a nucleolar modulator of ribosomal RNA synthesis, cell cycle progression, and teleost organ development.

    Directory of Open Access Journals (Sweden)

    Norimasa Iwanami

    2008-08-01

    Full Text Available The thymus is a vertebrate-specific organ where T lymphocytes are generated. Genetic programs that lead to thymus development are incompletely understood. We previously screened ethylnitrosourea-induced medaka mutants for recessive defects in thymus development. Here we report that one of those mutants is caused by a missense mutation in a gene encoding the previously uncharacterized protein WDR55 carrying the tryptophan-aspartate-repeat motif. We find that WDR55 is a novel nucleolar protein involved in the production of ribosomal RNA (rRNA. Defects in WDR55 cause aberrant accumulation of rRNA intermediates and cell cycle arrest. A mutation in WDR55 in zebrafish also leads to analogous defects in thymus development, whereas WDR55-null mice are lethal before implantation. These results indicate that WDR55 is a nuclear modulator of rRNA synthesis, cell cycle progression, and embryonic organogenesis including teleost thymus development.

  6. Identification and quantification of 5α-dihydrotestosterone in the teleost fathead minnow (Pimephales promelas) by gas chromatography-tandem mass spectrometry.

    Science.gov (United States)

    Margiotta-Casaluci, Luigi; Courant, Frédérique; Antignac, Jean-Philippe; Le Bizec, Bruno; Sumpter, John P

    2013-09-15

    The steroid hormone 5α-dihydrotestosterone (DHT) is one of the most physiologically important androgens in male vertebrates, with the exception of teleost fish, in which it is generally assumed that DHT does not play any major physiological role. However, this assumption is challenged by the fact that all the components involved in DHT biosynthesis and action are present and evolutionary conserved in teleost fish. In fact, testosterone (T) is converted into DHT by two isoforms of the enzyme steroid-5-alpha-reductase (5αR), and both 5αRs gene expression and enzymatic activity have been detected in several tissues of different teleost species, which also have an androgen receptor with high binding affinity to DHT. This body of evidence strongly suggest that DHT is synthesised by teleost fish. We investigated this hypothesis using the cyprinid fathead minnow (Pimephales promelas) as the experimental model. The study of the evolutionary and functional conservation of 5αRs in teleost fish was used to support the experimental approach, based on an ultrasensitive gas chromatography-tandem mass spectrometry (GC-MS/MS) method to identify and measure simultaneously T and DHT in fathead minnow biological fluids and tissues. The analyses were performed using plasma samples collected from both male and female adult fish and samples of testicular tissue collected from sexually mature males. Both T and DHT were identified and quantified in all the samples analysed, and in particular, the high concentrations of DHT quantified in the testes suggested that these organs are a likely site of synthesis of DHT in the teleost fathead minnow, as they are in mammals. These results may represent the basis for future studies aimed at elucidating the physiological role, if any, of DHT in teleost fish.

  7. Establishment of Multi-Site Infection Model in Zebrafish Larvae for Studying Staphylococcus aureus Infectious Disease

    Institute of Scientific and Technical Information of China (English)

    Ya-juan Li; Bing Hu

    2012-01-01

    Zebrafish (Danio rerio) is an ideal model for studying the mechanism of infectious disease and the interaction between host and pathogen.As a teleost,zebrafish has developed a complete immune system which is similar to mammals.Moreover,the easy acquirement of large amounts of transparent embryos makes it a good candidate for gene manipulation and drug screening.In a zebrafish infection model,all of the site,timing,and dose of the bacteria microinjection into the embryo are important factors that determine the bacterial infection of host.Here,we established a multi-site infection model in zebrafish larvae of 36 hours post-fertilization (hpf) by microinjecting wild-type or GFP-expressing Staphylococcus aereus (S.aureus) with gradient burdens into different embryo sites including the pericardial cavity (PC),eye,the fourth hindbrain ventricle (4V),yolk circulation valley (YCV),caudal vein (CV),yolk body (YB),and Duct of Cuvier (DC) to resemble human infectious disease.With the combination of GFP-expressing S.aureus and transgenic zebrafish Tg (corola:eGFP; lyz:Dsred) and Tg (lyz:Dsred) lines whose macrophages or neutrophils are fluorescent labeled,we observed the dynamic process of bacterial infection by in vivo multicolored confocal fluorescence imaging.Analyses of zebrafish embryo survival,bacterial proliferation and myeloid cells phagocytosis show that the site- and dose-dependent differences exist in infection of different bacterial entry routes.This work provides a consideration for the future study of pathogenesis and host resistance through selection of multi-site infection model.More interaction mechanisms between pathogenic bacteria virulence factors and the immune responses of zebrafish could be determined through zebrafish multi-site infection model.

  8. Cloning and comparative analyses of the zebrafish Ugt repertoire reveal its evolutionary diversity.

    Directory of Open Access Journals (Sweden)

    Haiyan Huang

    Full Text Available UDP-glucuronosyltransferases (Ugts are a supergene family of phase II drug-metabolizing enzymes that catalyze the conjugation of numerous hydrophobic small molecules with the UDP-glucuronic acid, converting them into hydrophilic molecules. Here, we report the identification and cloning of the complete zebrafish Ugt gene repertoire. We found that the zebrafish genome contains 45 Ugt genes that can be divided into three families: Ugt1, Ugt2, and Ugt5. Both Ugt1 and Ugt2 have two unlinked clusters: a and b. The Ugt1a, Ugt1b, Ugt2a, and Ugt2b clusters each contain variable and constant regions, similar to that of the protocadherin (Pcdh, immunoglobulin (Ig, and T-cell receptor (Tcr clusters. Cloning the full-length coding sequences confirmed that each of the variable exons is separately spliced to the set of constant exons within each zebrafish Ugt cluster. Comparative analyses showed that both a and b clusters of the zebrafish Ugt1 and Ugt2 genes have orthologs in other teleosts, suggesting that they may be resulted from the "fish-specific" whole-genome duplication event. The Ugt5 genes are a novel family of Ugt genes that exist in teleosts and amphibians. Their entire open reading frames are encoded by single large exons. The zebrafish Ugt1, Ugt2, and Ugt5 genes can generate additional transcript diversity through alternative splicing. Based on phylogenetic analyses, we propose that the ancestral tetrapod and teleost Ugt1 clusters contained multiple Ugt1 paralogs. After speciation, these ancestral Ugt1 clusters underwent lineage-specific gene loss and duplication. The ancestral vertebrate Ugt2 cluster also underwent lineage-specific duplication. The intronless Ugt5 open reading frames may be derived from retrotransposition followed by gene duplication. They have been expanded dramatically in teleosts and have become the most abundant Ugt family in these lineages. These findings have interesting implications regarding the molecular evolution of

  9. Genetic determinants of hyaloid and retinal vasculature in zebrafish

    Directory of Open Access Journals (Sweden)

    Hyde David R

    2007-10-01

    Full Text Available Abstract Background The retinal vasculature is a capillary network of blood vessels that nourishes the inner retina of most mammals. Developmental abnormalities or microvascular complications in the retinal vasculature result in severe human eye diseases that lead to blindness. To exploit the advantages of zebrafish for genetic, developmental and pharmacological studies of retinal vasculature, we characterised the intraocular vasculature in zebrafish. Results We show a detailed morphological and developmental analysis of the retinal blood supply in zebrafish. Similar to the transient hyaloid vasculature in mammalian embryos, vessels are first found attached to the zebrafish lens at 2.5 days post fertilisation. These vessels progressively lose contact with the lens and by 30 days post fertilisation adhere to the inner limiting membrane of the juvenile retina. Ultrastructure analysis shows these vessels to exhibit distinctive hallmarks of mammalian retinal vasculature. For example, smooth muscle actin-expressing pericytes are ensheathed by the basal lamina of the blood vessel, and vesicle vacuolar organelles (VVO, subcellular mediators of vessel-retinal nourishment, are present. Finally, we identify 9 genes with cell membrane, extracellular matrix and unknown identity that are necessary for zebrafish hyaloid and retinal vasculature development. Conclusion Zebrafish have a retinal blood supply with a characteristic developmental and adult morphology. Abnormalities of these intraocular vessels are easily observed, enabling application of genetic and chemical approaches in zebrafish to identify molecular regulators of hyaloid and retinal vasculature in development and disease.

  10. Characterization and Expression of the Zebrafish qki Paralogs.

    Science.gov (United States)

    Radomska, Katarzyna J; Sager, Jonathan; Farnsworth, Bryn; Tellgren-Roth, Åsa; Tuveri, Giulia; Peuckert, Christiane; Kettunen, Petronella; Jazin, Elena; Emilsson, Lina S

    2016-01-01

    Quaking (QKI) is an RNA-binding protein involved in post-transcriptional mRNA processing. This gene is found to be associated with several human neurological disorders. Early expression of QKI proteins in the developing mouse neuroepithelium, together with neural tube defects in Qk mouse mutants, suggest the functional requirement of Qk for the establishment of the nervous system. As a knockout of Qk is embryonic lethal in mice, other model systems like the zebrafish could serve as a tool to study the developmental functions of qki. In the present study we sought to characterize the evolutionary relationship and spatiotemporal expression of qkia, qki2, and qkib; zebrafish homologs of human QKI. We found that qkia is an ancestral paralog of the single tetrapod Qk gene that was likely lost during the fin-to-limb transition. Conversely, qkib and qki2 are orthologs, emerging at the root of the vertebrate and teleost lineage, respectively. Both qki2 and qkib, but not qkia, were expressed in the progenitor domains of the central nervous system, similar to expression of the single gene in mice. Despite having partially overlapping expression domains, each gene has a unique expression pattern, suggesting that these genes have undergone subfunctionalization following duplication. Therefore, we suggest the zebrafish could be used to study the separate functions of qki genes during embryonic development.

  11. Characterization and Expression of the Zebrafish qki Paralogs.

    Directory of Open Access Journals (Sweden)

    Katarzyna J Radomska

    Full Text Available Quaking (QKI is an RNA-binding protein involved in post-transcriptional mRNA processing. This gene is found to be associated with several human neurological disorders. Early expression of QKI proteins in the developing mouse neuroepithelium, together with neural tube defects in Qk mouse mutants, suggest the functional requirement of Qk for the establishment of the nervous system. As a knockout of Qk is embryonic lethal in mice, other model systems like the zebrafish could serve as a tool to study the developmental functions of qki. In the present study we sought to characterize the evolutionary relationship and spatiotemporal expression of qkia, qki2, and qkib; zebrafish homologs of human QKI. We found that qkia is an ancestral paralog of the single tetrapod Qk gene that was likely lost during the fin-to-limb transition. Conversely, qkib and qki2 are orthologs, emerging at the root of the vertebrate and teleost lineage, respectively. Both qki2 and qkib, but not qkia, were expressed in the progenitor domains of the central nervous system, similar to expression of the single gene in mice. Despite having partially overlapping expression domains, each gene has a unique expression pattern, suggesting that these genes have undergone subfunctionalization following duplication. Therefore, we suggest the zebrafish could be used to study the separate functions of qki genes during embryonic development.

  12. Manipulation of Fgf and Bmp signaling in teleost fishes suggests potential pathways for the evolutionary origin of multicuspid teeth.

    Science.gov (United States)

    Jackman, William R; Davies, Shelby H; Lyons, David B; Stauder, Caitlin K; Denton-Schneider, Benjamin R; Jowdry, Andrea; Aigler, Sharon R; Vogel, Scott A; Stock, David W

    2013-01-01

    Teeth with two or more cusps have arisen independently from an ancestral unicuspid condition in a variety of vertebrate lineages, including sharks, teleost fishes, amphibians, lizards, and mammals. One potential explanation for the repeated origins of multicuspid teeth is the existence of multiple adaptive pathways leading to them, as suggested by their different uses in these lineages. Another is that the addition of cusps required only minor changes in genetic pathways regulating tooth development. Here we provide support for the latter hypothesis by demonstrating that manipulation of the levels of Fibroblast growth factor (Fgf) or Bone morphogenetic protein (Bmp) signaling produces bicuspid teeth in the zebrafish (Danio rerio), a species lacking multicuspid teeth in its ancestry. The generality of these results for teleosts is suggested by the conversion of unicuspid pharyngeal teeth into bicuspid teeth by similar manipulations of the Mexican Tetra (Astyanax mexicanus). That these manipulations also produced supernumerary teeth in both species supports previous suggestions of similarities in the molecular control of tooth and cusp number. We conclude that despite their apparent complexity, the evolutionary origin of multicuspid teeth is positively constrained, likely requiring only slight modifications of a pre-existing mechanism for patterning the number and spacing of individual teeth.

  13. Comparative epigenomics in distantly related teleost species identifies conserved cis-regulatory nodes active during the vertebrate phylotypic period

    Science.gov (United States)

    Tena, Juan J.; González-Aguilera, Cristina; Fernández-Miñán, Ana; Vázquez-Marín, Javier; Parra-Acero, Helena; Cross, Joe W.; Rigby, Peter W.J.; Carvajal, Jaime J.; Wittbrodt, Joachim; Gómez-Skarmeta, José L.; Martínez-Morales, Juan R.

    2014-01-01

    The complex relationship between ontogeny and phylogeny has been the subject of attention and controversy since von Baer’s formulations in the 19th century. The classic concept that embryogenesis progresses from clade general features to species-specific characters has often been revisited. It has become accepted that embryos from a clade show maximum morphological similarity at the so-called phylotypic period (i.e., during mid-embryogenesis). According to the hourglass model, body plan conservation would depend on constrained molecular mechanisms operating at this period. More recently, comparative transcriptomic analyses have provided conclusive evidence that such molecular constraints exist. Examining cis-regulatory architecture during the phylotypic period is essential to understand the evolutionary source of body plan stability. Here we compare transcriptomes and key epigenetic marks (H3K4me3 and H3K27ac) from medaka (Oryzias latipes) and zebrafish (Danio rerio), two distantly related teleosts separated by an evolutionary distance of 115–200 Myr. We show that comparison of transcriptome profiles correlates with anatomical similarities and heterochronies observed at the phylotypic stage. Through comparative epigenomics, we uncover a pool of conserved regulatory regions (≈700), which are active during the vertebrate phylotypic period in both species. Moreover, we show that their neighboring genes encode mainly transcription factors with fundamental roles in tissue specification. We postulate that these regulatory regions, active in both teleost genomes, represent key constrained nodes of the gene networks that sustain the vertebrate body plan. PMID:24709821

  14. Zebrafish: an animal model for research in veterinary medicine.

    Science.gov (United States)

    Nowik, N; Podlasz, P; Jakimiuk, A; Kasica, N; Sienkiewicz, W; Kaleczyc, J

    2015-01-01

    The zebrafish (Danio rerio) has become known as an excellent model organism for studies of vertebrate biology, vertebrate genetics, embryonal development, diseases and drug screening. Nevertheless, there is still lack of detailed reports about usage of the zebrafish as a model in veterinary medicine. Comparing to other vertebrates, they can lay hundreds of eggs at weekly intervals, externally fertilized zebrafish embryos are accessible to observation and manipulation at all stages of their development, which makes possible to simplify the research techniques such as fate mapping, fluorescent tracer time-lapse lineage analysis and single cell transplantation. Although zebrafish are only 2.5 cm long, they are easy to maintain. Intraperitoneal and intracerebroventricular injections, blood sampling and measurement of food intake are possible to be carry out in adult zebrafish. Danio rerio is a useful animal model for neurobiology, developmental biology, drug research, virology, microbiology and genetics. A lot of diseases, for which the zebrafish is a perfect model organism, affect aquatic animals. For a part of them, like those caused by Mycobacterium marinum or Pseudoloma neutrophila, Danio rerio is a natural host, but the zebrafish is also susceptible to the most of fish diseases including Itch, Spring viraemia of carp and Infectious spleen and kidney necrosis. The zebrafish is commonly used in research of bacterial virulence. The zebrafish embryo allows for rapid, non-invasive and real time analysis of bacterial infections in a vertebrate host. Plenty of common pathogens can be examined using zebrafish model: Streptococcus iniae, Vibrio anguillarum or Listeria monocytogenes. The steps are taken to use the zebrafish also in fungal research, especially that dealing with Candida albicans and Cryptococcus neoformans. Although, the zebrafish is used commonly as an animal model to study diseases caused by external agents, it is also useful in studies of metabolic

  15. Time-lapse imaging of neural development: zebrafish lead the way into the fourth dimension.

    Science.gov (United States)

    Rieger, Sandra; Wang, Fang; Sagasti, Alvaro

    2011-07-01

    Time-lapse imaging is often the only way to appreciate fully the many dynamic cell movements critical to neural development. Zebrafish possess many advantages that make them the best vertebrate model organism for live imaging of dynamic development events. This review will discuss technical considerations of time-lapse imaging experiments in zebrafish, describe selected examples of imaging studies in zebrafish that revealed new features or principles of neural development, and consider the promise and challenges of future time-lapse studies of neural development in zebrafish embryos and adults.

  16. The mechanism for primordial germ-cell migration is conserved between Japanese eel and zebrafish.

    Directory of Open Access Journals (Sweden)

    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.

  17. NICHD Zebrafish Core

    Data.gov (United States)

    Federal Laboratory Consortium — The core[HTML_REMOVED]s goal is to help researchers of any expertise perform zebrafish experiments aimed at illuminating basic biology and human disease mechanisms,...

  18. 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.

  19. Persistent impaired glucose metabolism in a zebrafish hyperglycemia model.

    Science.gov (United States)

    Capiotti, Katiucia Marques; Antonioli, Régis; Kist, Luiza Wilges; Bogo, Maurício Reis; Bonan, Carla Denise; Da Silva, Rosane Souza

    2014-05-01

    Diabetes mellitus (DM) affects over 10% of the world's population. Hyperglycemia is the main feature for the diagnosis of this disease. The zebrafish (Danio rerio) is an established model organism for the study of various metabolic diseases. In this paper, hyperglycemic zebrafish, when immersed in a 111 mM glucose solution for 14 days, developed increased glycation of proteins from the eyes, decreased mRNA levels of insulin receptors in the muscle, and a reversion of high blood glucose level after treatment with anti-diabetic drugs (glimepiride and metformin) even after 7 days of glucose withdrawal. Additionally, hyperglycemic zebrafish developed an impaired response to exogenous insulin, which was recovered after 7 days of glucose withdrawal. These data suggest that the exposure of adult zebrafish to high glucose concentration is able to induce persistent metabolic changes probably underlined by a hyperinsulinemic state and impaired peripheral glucose metabolism.

  20. Complement system in zebrafish.

    Science.gov (United States)

    Zhang, Shicui; Cui, Pengfei

    2014-09-01

    Zebrafish is recently emerging as a model species for the study of immunology and human diseases. Complement system is the humoral backbone of the innate immune defense, and our knowledge as such in zebrafish has dramatically increased in the recent years. This review summarizes the current research progress of zebrafish complement system. The global searching for complement components in genome database, together with published data, has unveiled the existence of all the orthologues of mammalian complement components identified thus far, including the complement regulatory proteins and complement receptors, in zebrafish. Interestingly, zebrafish complement components also display some distinctive features, such as prominent levels of extrahepatic expression and isotypic diversity of the complement components. Future studies should focus on the following issues that would be of special importance for understanding the physiological role of complement components in zebrafish: conclusive identification of complement genes, especially those with isotypic diversity; analysis and elucidation of function and mechanism of complement components; modulation of innate and adaptive immune response by complement system; and unconventional roles of complement-triggered pathways.

  1. Steroids in teleost fishes: A functional point of view.

    Science.gov (United States)

    Tokarz, Janina; Möller, Gabriele; Hrabě de Angelis, Martin; Adamski, Jerzy

    2015-11-01

    Steroid hormones are involved in the regulation of a variety of processes like embryonic development, sex differentiation, metabolism, immune responses, circadian rhythms, stress response, and reproduction in vertebrates. Teleost fishes and humans show a remarkable conservation in many developmental and physiological aspects, including the endocrine system in general and the steroid hormone related processes in particular. This review provides an overview of the current knowledge about steroid hormone biosynthesis and the steroid hormone receptors in teleost fishes and compares the findings to the human system. The impact of the duplicated genome in teleost fishes on steroid hormone biosynthesis and perception is addressed. Additionally, important processes in fish physiology regulated by steroid hormones, which are most dissimilar to humans, are described. We also give a short overview on the influence of anthropogenic endocrine disrupting compounds on steroid hormone signaling and the resulting adverse physiological effects for teleost fishes. By this approach, we show that the steroidogenesis, hormone receptors, and function of the steroid hormones are reasonably well understood when summarizing the available data of all teleost species analyzed to date. However, on the level of a single species or a certain fish-specific aspect of physiology, further research is needed.

  2. Coupling mechanical deformations and planar cell polarity to create regular patterns in the zebrafish retina.

    Directory of Open Access Journals (Sweden)

    Guillaume Salbreux

    Full Text Available The orderly packing and precise arrangement of epithelial cells is essential to the functioning of many tissues, and refinement of this packing during development is a central theme in animal morphogenesis. The mechanisms that determine epithelial cell shape and position, however, remain incompletely understood. Here, we investigate these mechanisms in a striking example of planar order in a vertebrate epithelium: The periodic, almost crystalline distribution of cone photoreceptors in the adult teleost fish retina. Based on observations of the emergence of photoreceptor packing near the retinal margin, we propose a mathematical model in which ordered columns of cells form as a result of coupling between planar cell polarity (PCP and anisotropic tissue-scale mechanical stresses. This model recapitulates many observed features of cone photoreceptor organization during retinal growth and regeneration. Consistent with the model's predictions, we report a planar-polarized distribution of Crumbs2a protein in cone photoreceptors in both unperturbed and regenerated tissue. We further show that the pattern perturbations predicted by the model to occur if the imposed stresses become isotropic closely resemble defects in the cone pattern in zebrafish lrp2 mutants, in which intraocular pressure is increased, resulting in altered mechanical stress and ocular enlargement. Evidence of interactions linking PCP, cell shape, and mechanical stresses has recently emerged in a number of systems, several of which show signs of columnar cell packing akin to that described here. Our results may hence have broader relevance for the organization of cells in epithelia. Whereas earlier models have allowed only for unidirectional influences between PCP and cell mechanics, the simple, phenomenological framework that we introduce here can encompass a broad range of bidirectional feedback interactions among planar polarity, shape, and stresses; our model thus represents a

  3. A third broad lineage of major histocompatibility complex (MHC) class I in teleost fish; MHC class II linkage and processed genes.

    Science.gov (United States)

    Dijkstra, Johannes Martinus; Katagiri, Takayuki; Hosomichi, Kazuyoshi; Yanagiya, Kazuyo; Inoko, Hidetoshi; Ototake, Mitsuru; Aoki, Takashi; Hashimoto, Keiichiro; Shiina, Takashi

    2007-04-01

    Most of the previously studied teleost MHC class I molecules can be classified into two broad lineages: "U" and "Z/ZE." However, database reports on genes in cyprinid and salmonid fishes show that there is a third major lineage, which lacks detailed analysis so far. We designated this lineage "L" because of an intriguing linkage characteristic. Namely, one zebrafish L locus is closely linked with MHC class II loci, despite the extensively documented nonlinkage of teleost class I with class II. The L lineage consists of highly variable, nonclassical MHC class I genes, and has no apparent orthologues outside teleost fishes. Characteristics that distinguish the L lineage from most other MHC class I are (1) absence of two otherwise highly conserved tryptophan residues W51 and W60 in the alpha1 domain, (2) a low GC content of the alpha1 and alpha2 exons, and (3) an HINLTL motif including a possible glycosylation site in the alpha3 domain. In rainbow trout (Oncorhynchus mykiss) we analyzed several intact L genes in detail, including their genomic organization and transcription pattern. The gene Onmy-LAA is quite different from the genes Onmy-LBA, Onmy-LCA, Onmy-LDA, and Onmy-LEA, while the latter four are similar and categorized as "Onmy-LBA-like." Whereas the Onmy-LAA gene is organized like a canonical MHC class I gene, the Onmy-LBA-like genes are processed and lack all introns except intron 1. Onmy-LAA is predominantly expressed in the intestine, while the Onmy-LBA-like transcripts display a rather homogeneous tissue distribution. To our knowledge, this is the first description of an MHC class I lineage with multiple copies of processed genes, which are intact and transcribed. The present study significantly improves the knowledge of MHC class I variation in teleosts.

  4. A histone demethylase is necessary for regeneration in zebrafish.

    Science.gov (United States)

    Stewart, Scott; Tsun, Zhi-Yang; Izpisua Belmonte, Juan Carlos

    2009-11-24

    Urodele amphibians and teleost fish regenerate amputated body parts via a process called epimorphic regeneration. A hallmark of this phenomenon is the reactivation of silenced developmental regulatory genes that previously functioned during embryonic patterning. We demonstrate that histone modifications silence promoters of numerous genes involved in zebrafish caudal fin regeneration. Silenced developmental regulatory genes contain bivalent me(3)K4/me(3)K27 H3 histone modifications created by the concerted action of Polycomb (PcG) and Trithorax histone methyltransferases. During regeneration, this silent, bivalent chromatin is converted to an active state by loss of repressive me(3)K27 H3 modifications, occurring at numerous genes that appear to function during regeneration. Loss-of-function studies demonstrate a requirement for a me(3)K27 H3 demethylase during fin regeneration. These results indicate that histone modifications at discreet genomic positions may serve as a crucial regulatory event in the initiation of fin regeneration.

  5. Measuring zebrafish turning rate.

    Science.gov (United States)

    Mwaffo, Violet; Butail, Sachit; di Bernardo, Mario; Porfiri, Maurizio

    2015-06-01

    Zebrafish is becoming a popular animal model in preclinical research, and zebrafish turning rate has been proposed for the analysis of activity in several domains. The turning rate is often estimated from the trajectory of the fish centroid that is output by commercial or custom-made target tracking software run on overhead videos of fish swimming. However, the accuracy of such indirect methods with respect to the turning rate associated with changes in heading during zebrafish locomotion is largely untested. Here, we compare two indirect methods for the turning rate estimation using the centroid velocity or position data, with full shape tracking for three different video sampling rates. We use tracking data from the overhead video recorded at 60, 30, and 15 frames per second of zebrafish swimming in a shallow water tank. Statistical comparisons of absolute turning rate across methods and sampling rates indicate that, while indirect methods are indistinguishable from full shape tracking, the video sampling rate significantly influences the turning rate measurement. The results of this study can aid in the selection of the video capture frame rate, an experimental design parameter in zebrafish behavioral experiments where activity is an important measure.

  6. Conserved gene regulation during acute inflammation between zebrafish and mammals

    Science.gov (United States)

    Forn-Cuní, G.; Varela, M.; Pereiro, P.; Novoa, B.; Figueras, A.

    2017-01-01

    Zebrafish (Danio rerio), largely used as a model for studying developmental processes, has also emerged as a valuable system for modelling human inflammatory diseases. However, in a context where even mice have been questioned as a valid model for these analysis, a systematic study evaluating the reproducibility of human and mammalian inflammatory diseases in zebrafish is still lacking. In this report, we characterize the transcriptomic regulation to lipopolysaccharide in adult zebrafish kidney, liver, and muscle tissues using microarrays and demonstrate how the zebrafish genomic responses can effectively reproduce the mammalian inflammatory process induced by acute endotoxin stress. We provide evidence that immune signaling pathways and single gene expression is well conserved throughout evolution and that the zebrafish and mammal acute genomic responses after lipopolysaccharide stimulation are highly correlated despite the differential susceptibility between species to that compound. Therefore, we formally confirm that zebrafish inflammatory models are suited to study the basic mechanisms of inflammation in human inflammatory diseases, with great translational impact potential. PMID:28157230

  7. MEK Inhibitors Reverse cAMP-Mediated Anxiety in Zebrafish

    DEFF Research Database (Denmark)

    Lundegaard, Pia R.; Anastasaki, Corina; Grant, Nicola J.;

    2015-01-01

    Altered phosphodiesterase (PDE)-cyclic AMP (cAMP) activity is frequently associated with anxiety disorders, but current therapies act by reducing neuronal excitability rather than targeting PDE-cAMP-mediated signaling pathways. Here, we report the novel repositioning of anti-cancer MEK inhibitors...... as anxiolytics in a zebrafish model of anxiety-like behaviors. PDE inhibitors or activators of adenylate cyclase cause behaviors consistent with anxiety in larvae and adult zebrafish. Small-molecule screening identifies MEK inhibitors as potent suppressors of cAMP anxiety behaviors in both larvae and adult...... zebrafish, while causing no anxiolytic behavioral effects on their own. The mechanism underlying cAMP-induced anxiety is via crosstalk to activation of the RAS-MAPK signaling pathway. We propose that targeting crosstalk signaling pathways can be an effective strategy for mental health disorders, and advance...

  8. Histopathological alterations in liver anatomy after exposure to chlorpyrifos in zebrafish (Danio rerio)

    OpenAIRE

    Bangeppagari, Manjunatha

    2015-01-01

    Chlorpyrifos is an organophosphate pesticide widely used in agriculture and aquaculture. This study investigated its effects on histopathology of zebrafish (Danio rerio) liver. For this six adult male and six adult female zebrafish were exposed to 200 ?g/L of chlorpyrifos for 24h, 48h, 72h and 96hrs. Chlorpyrifos toxicity on liver histopathological changes were examined by light microscopy. Structural damage spotted in the liver were vacuolization and presence of sinusoid spaces were observed...

  9. Impaired Lymphocytes Development and Xenotransplantation of Gastrointestinal Tumor Cells in Prkdc-Null SCID Zebrafish Model

    Directory of Open Access Journals (Sweden)

    In Hye Jung

    2016-08-01

    Full Text Available Severe combined immunodeficiency (SCID mice have widely been used as hosts for human tumor cell xenograft study. This animal model, however, is labor intensive. As zebrafish is largely emerging as a promising model system for studying human diseases including cancer, developing efficient immunocompromised strains for tumor xenograft study are also demanded in zebrafish. Here, we have created the Prkdc-null SCID zebrafish model which provides the stable immune-deficient background required for xenotransplantation of tumor cell. In this study, the two transcription activator-like effector nucleases that specifically target the exon3 of the zebrafish Prkdc gene were used to induce a frame shift mutation, causing a complete knockout of the gene function. The SCID zebrafish showed susceptibility to spontaneous infection, a well-known phenotype found in the SCID mutation. Further characterization revealed that the SCID zebrafish contained no functional T and B lymphocytes which reflected the phenotypes identified in the mice SCID model. Intraperitoneal injection of human cancer cells into the adult SCID zebrafish clearly showed tumor cell growth forming into a solid mass. Our present data show the suitability of using the SCID zebrafish strain for xenotransplantation experiments, and in vivo monitoring of the tumor cell growth in the zebrafish demonstrates use of the animal model as a new platform of tumor xenograft study.

  10. Elastoidin actinotrichia in Coelacanth fins: a comparison with teleosts.

    Science.gov (United States)

    Geraudie, J; Meunier, F J

    1980-01-01

    In this work, we present the first ultrastructural evidences of actinotrichia in the Coelacanth Latimeria. We describe its actinotrichia with the electron microscope (SEM and TEM) and compare their structure to Teleost actinotrichia. Both elements present similar fine structure, i.e. a periodic cross-striation of 60-65 nm; the plesiomorphic character of actinotrichia is discussed in Osteichthyes.

  11. Dynamics of DNA hydroxymethylation in zebrafish.

    Science.gov (United States)

    Kamstra, Jorke H; Løken, Marianne; Aleström, Peter; Legler, Juliette

    2015-06-01

    During embryonic development in mammals, most of the methylated cytosines in the paternal genome are converted to 5-hydroxymethyldeoxycytidine (hmC), as part of DNA methylation reprogramming. Recent data also suggest tissue-specific functional roles of hmC, perhaps as an epigenetic mark. However, limited data are available on the levels and tissue distribution in zebrafish. In this study, we used high-performance liquid chromatography mass spectrometry to quantify hmC and 5-methyldeoxycytidine (mC) in zebrafish during development and in different tissues of the adult fish. Low levels of mC were found at 0.5 hours postfertilization (hpf) (1-2 cell stage) (1.9%), and increased to 8.4% by 96 hpf, with similar levels observed in different adult tissues. No hmC was detected up to 12 hpf, but levels increased during development from 24 up to 96 hpf (0.23%). In tissues, the highest levels of hmC were found in the brain (0.49%), intermediate levels in muscle (0.13%), liver (0.08%), and intestine (0.06%) and low levels in testis (0.01%), with an inverse correlation between hmC and mC. Our results indicate similar tissue distribution and levels of hmC between zebrafish and mammals, but distinct differences during embryonic development. Although more research is needed, these results support the use of zebrafish as an alternative model in the elucidation of tissue-specific functions of hmC.

  12. Genetic evidence for shared mechanisms of epimorphic regeneration in zebrafish.

    Science.gov (United States)

    Qin, Zhao; Barthel, Linda K; Raymond, Pamela A

    2009-06-09

    In a microarray-based gene profiling analysis of Müller glia-derived retinal stem cells in light-damaged retinas from adult zebrafish, we found that 2 genes required for regeneration of fin and heart tissues in zebrafish, hspd1 (heat shock 60-kDa protein 1) and mps1 (monopolar spindle 1), were up-regulated. Expression of both genes in the neurogenic Müller glia and progenitors was independently verified by quantitative reverse transcriptase PCR and in situ hybridization. Functional analysis of temperature-sensitive mutants of hspd1 and mps1 revealed that both are necessary for Müller glia-based cone photoreceptor regeneration in adult zebrafish retina. In the amputated fin, hspd1 is required for the induction of mesenchymal stem cells and blastema formation, whereas mps1 is required at a later step for rapid cell proliferation and outgrowth. This temporal sequence of hspd1 and mps1 function is conserved in the regenerating retina. Comparison of gene expression profiles from regenerating zebrafish retina, caudal fin, and heart muscle revealed additional candidate genes potentially implicated in injury-induced epimorphic regeneration in diverse zebrafish tissues.

  13. Exploring novel hormones essential for seawater adaptation in teleost fish.

    Science.gov (United States)

    Takei, Yoshio

    2008-05-15

    Marine fish are dehydrated in hyperosmotic seawater (SW), but maintain water balance by drinking surrounding SW if they are capable of excreting the excess ions, particularly Na(+) and Cl(-), absorbed with water by the intestine. An integrative approach is essential for understanding the mechanisms for SW adaptation, in which hormones play pivotal roles. Comparative genomic analyses have shown that hormones that have Na(+)-extruding and vasodepressor properties are greatly diversified in teleost fish. Physiological studies at molecular to organismal levels have revealed that these diversified hormones are much more potent and efficacious in teleost fish than in mammals and are important for survival in SW and for maintenance of low arterial pressure in a gravity-free aquatic environment. This is typified by the natriuretic peptide (NP) family, which is diversified into seven members (ANP, BNP, VNP and CNP1, 2, 3 and 4) and exerts potent hyponatremic and vasodepressor actions in marine fish. Another example is the guanylin family, which consists of three paralogs (guanylin, uroguanylin and renoguanylin), and stimulates Cl(-) secretion into the intestinal lumen and activates the absorptive-type Na-K-2Cl cotransporter by local luminocrine actions. The most recent addition is the adrenomedullin (AM) family, which has five members (AM1, 2, 3, 4 and 5), with AM2 and AM5 showing the most potent or efficacious vasodepressor and osmoregulatory effects among known hormones in teleost fish. Accumulating evidence strongly indicates that members of these diversified hormone families play essential roles in SW adaptation in teleost fish. In this short review, the author has attempted to propose a novel approach for identification of new hormones that are important for SW adaptation using comparative genomic and functional studies. The author has also suggested potential hormone families that are diversified in teleost fish and appear to be involved in SW adaptation through their

  14. Cognitive aging in zebrafish.

    Directory of Open Access Journals (Sweden)

    Lili Yu

    Full Text Available BACKGROUND: Age-related impairments in cognitive functions represent a growing clinical and social issue. Genetic and behavioral characterization of animal models can provide critical information on the intrinsic and environmental factors that determine the deterioration or preservation of cognitive abilities throughout life. METHODOLOGY/PRINCIPAL FINDINGS: Behavior of wild-type, mutant and gamma-irradiated zebrafish (Danio rerio was documented using image-analysis technique. Conditioned responses to spatial, visual and temporal cues were investigated in young, middle-aged and old animals. The results demonstrate that zebrafish aging is associated with changes in cognitive responses to emotionally positive and negative experiences, reduced generalization of adaptive associations, increased stereotypic and reduced exploratory behavior and altered temporal entrainment. Genetic upregulation of cholinergic transmission attenuates cognitive decline in middle-aged achesb55/+ mutants, compared to wild-type siblings. In contrast, the genotoxic stress of gamma-irradiation accelerates the onset of cognitive impairment in young zebrafish. CONCLUSIONS/SIGNIFICANCE: These findings would allow the use of powerful molecular biological resources accumulated in the zebrafish field to address the mechanisms of cognitive senescence, and promote the search for therapeutic strategies which may attenuate age-related cognitive decline.

  15. ZEBRAFISH CHROMOSOME-BANDING

    NARCIS (Netherlands)

    PIJNACKER, LP; FERWERDA, MA

    1995-01-01

    Banding techniques were carried out on metaphase chromosomes of zebrafish (Danio rerio) embryos. The karyotypes with the longest chromosomes consist of 12 metacentrics, 26 submetacentrics, and 12 subtelocentrics (2n = 50). All centromeres are C-band positive. Eight chromosomes have a pericentric C-b

  16. A comparative analysis of glomerulus development in the pronephros of medaka and zebrafish.

    Directory of Open Access Journals (Sweden)

    Koichiro Ichimura

    Full Text Available The glomerulus of the vertebrate kidney links the vasculature to the excretory system and produces the primary urine. It is a component of every single nephron in the complex mammalian metanephros and also in the primitive pronephros of fish and amphibian larvae. This systematic work highlights the benefits of using teleost models to understand the pronephric glomerulus development. The morphological processes forming the pronephric glomerulus are astoundingly different between medaka and zebrafish. (1 The glomerular primordium of medaka - unlike the one of zebrafish - exhibits a C-shaped epithelial layer. (2 The C-shaped primordium contains a characteristic balloon-like capillary, which is subsequently divided into several smaller capillaries. (3 In zebrafish, the bilateral pair of pronephric glomeruli is fused at the midline to form a glomerulus, while in medaka the two parts remain unmerged due to the interposition of the interglomerular mesangium. (4 Throughout pronephric development the interglomerular mesangial cells exhibit numerous cytoplasmic granules, which are reminiscent of renin-producing (juxtaglomerular cells in the mammalian afferent arterioles. Our systematic analysis of medaka and zebrafish demonstrates that in fish, the morphogenesis of the pronephric glomerulus is not stereotypical. These differences need be taken into account in future analyses of medaka mutants with glomerulus defects.

  17. Molecular characterization of prosomeric and intraprosomeric subdivisions of the embryonic zebrafish diencephalon.

    Science.gov (United States)

    Lauter, Gilbert; Söll, Iris; Hauptmann, Giselbert

    2013-04-01

    During development of the early neural tube, positional information provided by signaling gradients is translated into a grid of transverse and longitudinal transcription factor expression domains. Transcription factor specification codes defining distinct histogenetic domains within this grid are evolutionarily conserved across vertebrates and may reflect an underlying common vertebrate bauplan. When compared to the rich body of comparative gene expression studies of tetrapods, there is considerably less comparative data available for teleost fish. We used sensitive multicolor fluorescent in situ hybridization to generate a detailed map of regulatory gene expression domains in the embryonic zebrafish diencephalon. The high resolution of this technique allowed us to resolve abutting and overlapping gene expression of different transcripts. We found that the relative topography of gene expression patterns in zebrafish was highly similar to those of orthologous genes in tetrapods and consistent with a three-prosomere organization of the alar and basal diencephalon. Our analysis further demonstrated a conservation of intraprosomeric subdivisions within prosomeres 1, 2, and 3 (p1, p2, and p3). A tripartition of zebrafish p1 was identified reminiscent of precommissural (PcP), juxtacommissural (JcP), and commissural (CoP) pretectal domains of tetrapods. The constructed detailed diencephalic transcription factor gene expression map further identified molecularly distinct thalamic and prethalamic rostral and caudal domains and a prethalamic eminence histogenetic domain in zebrafish. Our comparative gene expression analysis conformed with the idea of a common bauplan for the diencephalon of anamniote and amniote vertebrates from fish to mammals.

  18. Effect of various antiepileptic drugs in zebrafish PTZ-seizure model

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    P Gupta

    2014-01-01

    Full Text Available Recently zebrafish larvae have emerged as a high-throughput model for screening pharmacological activities. The present study was undertaken to investigate the effect of established anticonvulsants, such as valproic acid, carbamazepine, gabapentin, diazepam, lacosamide and pregabalin against pentylenetetrazole (6 mM seizures in adult zebrafish. Different phases of seizures (increase swim activity, rapid whirlpool-like circling swim behaviour and brief clonus-like seizures leading to loss of posture were elicited in zebrafish on exposure for 15 min to 6 mM pentylenetetrazole. The exposure of zebrafish to an increasing concentration of the anticonvulsants alongside 6 mM pentylenetetrazole showed concentration-dependent elevation of seizure latency against pentylenetetrazole-induced seizures except for pregabalin, which failed to produce any anticonvulsant activity in zebrafish. Moreover the proconvulsant activity of caffeine was also evaluated using suboptimal concentration (4 mM of pentylenetetrazole in adult zebrafish. Decrease in seizure latency of different phases of seizures was observed with increasing concentration of caffeine compared with its respective control group. In view of the above findings, the results of the present study suggested that adult zebrafish produce the expected anticonvulsive and proconvulsive effects and could potentially be used as a screen in future epilepsy research.

  19. Morphogenesis underlying the development of the everted teleost telencephalon

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    Folgueira Mónica

    2012-09-01

    Full Text Available Abstract Background Although the mechanisms underlying brain patterning and regionalization are very much conserved, the morphology of different brain regions is extraordinarily variable across vertebrate phylogeny. This is especially manifest in the telencephalon, where the most dramatic variation is seen between ray-finned fish, which have an everted telencephalon, and all other vertebrates, which have an evaginated telencephalon. The mechanisms that generate these distinct morphologies are not well understood. Results Here we study the morphogenesis of the zebrafish telencephalon from 12 hours post fertilization (hpf to 5 days post fertilization (dpf by analyzing forebrain ventricle formation, evolving patterns of gene and transgene expression, neuronal organization, and fate mapping. Our results highlight two key events in telencephalon morphogenesis. First, the formation of a deep ventricular recess between telencephalon and diencephalon, the anterior intraencephalic sulcus (AIS, effectively creates a posterior ventricular wall to the telencephalic lobes. This process displaces the most posterior neuroepithelial territory of the telencephalon laterally. Second, as telencephalic growth and neurogenesis proceed between days 2 and 5 of development, the pallial region of the posterior ventricular wall of the telencephalon bulges into the dorsal aspect of the AIS. This brings the ventricular zone (VZ into close apposition with the roof of the AIS to generate a narrow ventricular space and the thin tela choroidea (tc. As the pallial VZ expands, the tc also expands over the upper surface of the telencephalon. During this period, the major axis of growth and extension of the pallial VZ is along the anteroposterior axis. This second step effectively generates an everted telencephalon by 5 dpf. Conclusion Our description of telencephalic morphogenesis challenges the conventional model that eversion is simply due to a laterally directed outfolding of

  20. Apoptosis in thymus of teleost fish.

    Science.gov (United States)

    Romano, Nicla; Ceccarelli, Giuseppina; Caprera, Cecilia; Caccia, Elisabetta; Baldassini, Maria Rosaria; Marino, Giovanna

    2013-08-01

    The presence and distribution of apoptotic cells during thymus development and in adult were studied by in situ end-labelling of fragmented DNA in three temperate species carp (Cyprinus carpio), sea bass (Dicentrarchus labrax) and dusky grouper (Epinephelus marginatus) and in the adult thymus of three Antarctic species belonging to the genus Trematomus spp. During thymus development some few isolated apoptotic cell (AC) firstly appeared in the central-external part of the organ (carp: 5 days ph; sea bass: 35 days ph grouper: 43 days ph). Initially the cells were isolated and then increased in number and aggregated in small groups in the outer-cortical region of the thymus larvae. The high density of apoptotic cells was observed in the junction between cortex and medulla from its appearance (border between cortex and medulla, BCM). ACs decreased in number in juveniles and adult as well as the ACs average diameter. In late juveniles and in adulthood, the apoptosis were restricted to the cortex. In Antarctic species the thymus is highly adapted to low temperature (high vascularisation to effort the circulation of glycoproteins enriched plasma and strongly compact parenchyma). The apoptosis process was more extended (4-7 fold) as compare with the thymus of temperate species, even if the distribution of ACs was similar in all examined species. Data suggested a common process of T lymphocyte negative-selection in BCM of thymus during the ontogeny. The selection process seems to be still active in adult polar fish, but restricted mainly in the cortex zone.

  1. 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...

  2. Brain derived neurotrophic factor in the retina of the teleost N. furzeri.

    Science.gov (United States)

    Gatta, Claudia; Castaldo, Luciana; Cellerino, Alessandro; de Girolamo, Paolo; Lucini, Carla; D'Angelo, Livia

    2014-07-01

    BDNF plays an important role in the development and maintenance of visual circuitries in the retina and brain visual centers. In adulthood, BDNF signaling is involved in neural protection and regeneration of retina. In this survey, we investigated the expression of BDNF in the retina of adult Nothobranchius furzeri, a teleost fish employed for age research. After describing the retina of N. furzeri and confirming that the structure is organized in layers as in all vertebrates, we have studied the localization of BDNF mRNA and protein throughout the retinal layers. BDNF mRNA is detectable in all layers, whereas the protein is lacking in the photoreceptors. The occurrence of BDNF provides new insights on its role in the retina, particularly in view of age-related disease of retina.

  3. Diversity of trypanorhynch metacestodes in teleost fishes from coral reefs off eastern Australia and New Caledonia

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    Beveridge Ian

    2014-01-01

    Full Text Available Trypanorhynch metacestodes were examined from teleosts from coral reefs in eastern Australia and from New Caledonia. From over 12,000 fishes examined, 33 named species of trypanorhynchs were recovered as well as three species of tentacularioids which are described but not named. Host-parasite and parasite-host lists are provided, including more than 100 new host records. Lacistorhynchoid and tentacularioid taxa predominated with fewer otobothrioid and gymnorhynchoids. Five species, Callitetrarhynchus gracilis, Floriceps minacanthus, Pseudotobothrium dipsacum, Pseudolacistorhynchus heroniensis and Ps. shipleyi, were particularly common and exhibited low host specificity. Limited data suggested a higher diversity of larval trypanorhynchs in larger piscivorous fish families. Several fish families surveyed extensively (Blenniidae, Chaetodontidae, Gobiidae, Kyphosidae and Scaridae yielded no trypanorhynch larvae. The overall similarity between the fauna of the Great Barrier Reef and New Caledonia was 45%. Where available, information on the adult stages in elasmobranchs has been included.

  4. Genetic evidence for shared mechanisms of epimorphic regeneration in zebrafish

    OpenAIRE

    Qin, Zhao; Barthel, Linda K.; Raymond, Pamela A.

    2009-01-01

    In a microarray-based gene profiling analysis of Müller glia-derived retinal stem cells in light-damaged retinas from adult zebrafish, we found that 2 genes required for regeneration of fin and heart tissues in zebrafish, hspd1 (heat shock 60-kDa protein 1) and mps1 (monopolar spindle 1), were up-regulated. Expression of both genes in the neurogenic Müller glia and progenitors was independently verified by quantitative reverse transcriptase PCR and in situ hybridization. Functional analysis o...

  5. Transgenic overexpression of cdx1b induces metaplastic changes of gene expression in zebrafish esophageal squamous epithelium.

    Science.gov (United States)

    Hu, Bo; Chen, Hao; Liu, Xiuping; Zhang, Chengjin; Cole, Gregory J; Lee, Ju-Ahng; Chen, Xiaoxin

    2013-06-01

    Cdx2 has been suggested to play an important role in Barrett's esophagus or intestinal metaplasia (IM) in the esophagus. To investigate whether transgenic overexpression of cdx1b, the functional equivalent of mammalian Cdx2 in zebrafish, may lead to IM of zebrafish esophageal squamous epithelium, a transgenic zebrafish system was developed by expressing cdx1b gene under the control of zebrafish keratin 5 promoter (krt5p). Gene expression in the esophageal squamous epithelium of wild-type and transgenic zebrafish was analyzed by Affymetrix microarray and confirmed by in situ hybridization. Morphology, mucin expression, cell proliferation, and apoptosis were analyzed by hematoxylin & eosin (HE) staining, Periodic acid Schiff (PAS) Alcian blue staining, proliferating cell nuclear antigen (PCNA) immunohistochemical staining, and TUNEL assay as well. cdx1b was found to be overexpressed in the nuclei of esophageal squamous epithelial cells of the transgenic zebrafish. Ectopic expression of cdx1b disturbed the development of this epithelium in larval zebrafish and induced metaplastic changes in gene expression in the esophageal squamous epithelial cells of adult zebrafish, that is, up-regulation of intestinal differentiation markers and down-regulation of squamous differentiation markers. However, cdx1b failed to induce histological IM, or to modulate cell proliferation and apoptosis in the squamous epithelium of adult transgenic zebrafish.

  6. N-cadherin is required for the polarized cell behaviors that drive neurulation in the zebrafish.

    Science.gov (United States)

    Hong, Elim; Brewster, Rachel

    2006-10-01

    Through the direct analysis of cell behaviors, we address the mechanisms underlying anterior neural tube morphogenesis in the zebrafish and the role of the cell adhesion molecule N-cadherin (N-cad) in this process. We demonstrate that although the mode of neurulation differs at the morphological level between amphibians and teleosts, the underlying cellular mechanisms are conserved. Contrary to previous reports, the zebrafish neural plate is a multi-layered structure, composed of deep and superficial cells that converge medially while undergoing radial intercalation, to form a single cell-layered neural tube. Time-lapse recording of individual cell behaviors reveals that cells are polarized along the mediolateral axis and exhibit protrusive activity. In N-cad mutants, both convergence and intercalation are blocked. Moreover, although N-cad-depleted cells are not defective in their ability to form protrusions, they are unable to maintain them stably. Taken together, these studies uncover key cellular mechanisms underlying neural tube morphogenesis in teleosts, and reveal a role for cadherins in promoting the polarized cell behaviors that underlie cellular rearrangements and shape the vertebrate embryo.

  7. Skeletogenic fate of zebrafish cranial and trunk neural crest.

    Directory of Open Access Journals (Sweden)

    Erika Kague

    Full Text Available The neural crest (NC is a major contributor to the vertebrate craniofacial skeleton, detailed in model organisms through embryological and genetic approaches, most notably in chick and mouse. Despite many similarities between these rather distant species, there are also distinct differences in the contribution of the NC, particularly to the calvariae of the skull. Lack of information about other vertebrate groups precludes an understanding of the evolutionary significance of these differences. Study of zebrafish craniofacial development has contributed substantially to understanding of cartilage and bone formation in teleosts, but there is currently little information on NC contribution to the zebrafish skeleton. Here, we employ a two-transgene system based on Cre recombinase to genetically label NC in the zebrafish. We demonstrate NC contribution to cells in the cranial ganglia and peripheral nervous system known to be NC-derived, as well as to a subset of myocardial cells. The indelible labeling also enables us to determine NC contribution to late-forming bones, including the calvariae. We confirm suspected NC origin of cartilage and bones of the viscerocranium, including cartilages such as the hyosymplectic and its replacement bones (hymandibula and symplectic and membranous bones such as the opercle. The cleithrum develops at the border of NC and mesoderm, and as an ancestral component of the pectoral girdle was predicted to be a hybrid bone composed of both NC and mesoderm tissues. However, we find no evidence of a NC contribution to the cleithrum. Similarly, in the vault of the skull, the parietal bones and the caudal portion of the frontal bones show no evidence of NC contribution. We also determine a NC origin for caudal fin lepidotrichia; the presumption is that these are derived from trunk NC, demonstrating that these cells have the ability to form bone during normal vertebrate development.

  8. Identification and Expression Analysis of the Complete Family of Zebrafish pkd Genes

    Science.gov (United States)

    England, Samantha J.; Campbell, Paul C.; Banerjee, Santanu; Swanson, Annika J.; Lewis, Katharine E.

    2017-01-01

    Polycystic kidney disease (PKD) proteins are trans-membrane proteins that have crucial roles in many aspects of vertebrate development and physiology, including the development of many organs as well as left–right patterning and taste. They can be divided into structurally-distinct PKD1-like and PKD2-like proteins and usually one PKD1-like protein forms a heteromeric polycystin complex with a PKD2-like protein. For example, PKD1 forms a complex with PKD2 and mutations in either of these proteins cause Autosomal Dominant Polycystic Kidney Disease (ADPKD), which is the most frequent potentially-lethal single-gene disorder in humans. Here, we identify the complete family of pkd genes in zebrafish and other teleosts. We describe the genomic locations and sequences of all seven genes: pkd1, pkd1b, pkd1l1, pkd1l2a, pkd1l2b, pkd2, and pkd2l1. pkd1l2a/pkd1l2b are likely to be ohnologs of pkd1l2, preserved from the whole genome duplication that occurred at the base of the teleosts. However, in contrast to mammals and cartilaginous and holostei fish, teleosts lack pkd2l2, and pkdrej genes, suggesting that these have been lost in the teleost lineage. In addition, teleost, and holostei fish have only a partial pkd1l3 sequence, suggesting that this gene may be in the process of being lost in the ray-finned fish lineage. We also provide the first comprehensive description of the expression of zebrafish pkd genes during development. In most structures we detect expression of one pkd1-like gene and one pkd2-like gene, consistent with these genes encoding a heteromeric protein complex. For example, we found that pkd2 and pkd1l1 are expressed in Kupffer's vesicle and pkd1 and pkd2 are expressed in the developing pronephros. In the spinal cord, we show that pkd1l2a and pkd2l1 are co-expressed in KA cells. We also identify potential co-expression of pkd1b and pkd2 in the floor-plate. Interestingly, and in contrast to mouse, we observe expression of all seven pkd genes in regions

  9. Building neurophenomics in zebrafish: Effects of prior testing stress and test batteries.

    Science.gov (United States)

    Song, Cai; Yang, Lei; Wang, JiaJia; Chen, Peirong; Li, Shaomin; Liu, Yingcong; Nguyen, Michael; Kaluyeva, Aleksandra; Kyzar, Evan J; Gaikwad, Siddharth; Kalueff, Allan V

    2016-09-15

    The zebrafish (Danio rerio) is a promising model organism for neurophenomics - a new field of neuroscience linking neural phenotypes to various genetic and environmental factors. However, the effects of prior experimental manipulations on zebrafish performance in different behavioral paradigms remain unclear. Here, we examine the influence of selected stressful procedures and test batteries on adult zebrafish anxiety-like behaviors in two commonly used models - the novel tank (NTT) and the light-dark box (LDB) tests. While no overt behavioral differences between outbred short-fin wild-type (WT) and mutant 'pink' glowfish were seen in both tests under baseline (control) conditions, an acute severe stressor (a 30-min car transportation) detected significantly lower mutant fish anxiety-like behavior in these tests. In contrast, WT zebrafish showed no overt NTT or LDB responses following a mild stressor (5-min 40-Wt light) exposure, also showing no differences in batteries of NTT and LDB run immediately one after another, or with a 1-day interval. Collectively, these findings suggest that zebrafish may be relatively less sensitive (e.g., than other popular species, such as rodents) to the test battery effect, and show that stronger stressors may be needed (to complement low-to-moderate stress aquatic screens) to better reveal phenotypical variance in zebrafish assays. Strengthening the value of zebrafish models in neurophenotyping research, this study indicates the potential of using more test batteries and a wider spectrum of pre-test stressors in zebrafish behavioral assays.

  10. Spatial and temporal control of transgene expression in zebrafish.

    Directory of Open Access Journals (Sweden)

    Alexander A Akerberg

    Full Text Available Transgenic zebrafish research has provided valuable insights into gene functions and cell behaviors directing vertebrate development, physiology, and disease models. Most approaches use constitutive transgene expression and therefore do not provide control over the timing or levels of transgene induction. We describe an inducible gene expression system that uses new tissue-specific zebrafish transgenic lines that express the Gal4 transcription factor fused to the estrogen-binding domain of the human estrogen receptor. We show these Gal4-ERT driver lines confer rapid, tissue-specific induction of UAS-controlled transgenes following tamoxifen exposure in both embryos and adult fish. We demonstrate how this technology can be used to define developmental windows of gene function by spatiotemporal-controlled expression of constitutively active Notch1 in embryos. Given the array of existing UAS lines, the modular nature of this system will enable many previously intractable zebrafish experiments.

  11. CALCIFIED ECTODERMAL COLLAGENS OF SHARK TOOTH ENAMEL AND TELEOST SCALE.

    Science.gov (United States)

    MOSS, M L; JONES, S J; PIEZ, K A

    1964-08-28

    Amino acid analysis of protein from the enamel of shark teeth and from teleost scales shows the presence of collagens which can be classified chemically as ectodermal. This finding, together with results from a histological examination of the development of these tissues, constitutes strong evidence that both proteins are derived from the ectoderm, like the enamel of higher vertebrates. Since both are calcified, calcification cannot be a specific property of collagens of mesodermal origin alone.

  12. Cholinergic and adrenergic influence on the teleost heart in vivo.

    Science.gov (United States)

    Axelsson, M; Ehrenström, F; Nilsson, S

    1987-01-01

    The tonical cholinergic and adrenergic influence on the heart rate was investigated in vivo in seven species of marine teleosts (pollack, Pollachius pollachius; cuckoo wrasse, Labrus mixtus; ballan wrasse, Labrus berggylta; five-bearded rockling, Ciliata mustela; tadpole fish, Raniceps raninus; eel-pout, Zoarces viviparus and short-spined sea scorpion, Myoxocephalus scor pius) during rest and, in two of the species (P. pollachius and L. mixtus), also during moderate swimming exercise in a Blazka-type swim tunnel. Ventral aortic blood pressure and heart rate were recorded via a catheter implanted in an afferent branchial artery, and the influence of the cholinergic and adrenergic tonus on the heart rate was assessed by injection of atropine and sotalol respectively. During rest the adrenergic tonus was higher than the cholinergic tonus in all species except L. berggylta, where the reverse was true. In P. pollachius and L. mixtus, exercise appeared to produce a lowering of the cholinergic tonus on the heart and, possibly, a slight increase of the adrenergic tonus. The nature of the adrenergic tonus (humoral or neural) is not clear, but the low plasma concentrations of catecholamines both during rest and exercise could be interpreted in favour of a mainly neural adrenergic tonus on the teleost heart. These experiments are compatible with the view that both a cholinergic inhibitory tonus and an adrenergic excitatory tonus are general features in the control of the teleost heart in vivo, both at rest and during moderate swimming exercise.

  13. Immunoglobulin genes and their transcriptional control in teleosts.

    Science.gov (United States)

    Hikima, Jun-ichi; Jung, Tae-Sung; Aoki, Takashi

    2011-09-01

    Immunoglobulin (Ig), which exists only in jawed vertebrates, is one of the most important molecules in adaptive immunity. In the last two decades, many teleost Ig genes have been identified by in silico data mining from the enormous gene and EST databases of many fish species. In this review, the organization of Ig gene segments, the expressed Ig isotypes and their transcriptional controls are discussed. The Ig heavy chain (IgH) locus in teleosts encodes the variable (V), the diversity (D), the joining (J) segments and three different isotypic constant (C) regions including Cμ, Cδ, and Cζ/τ genes, and is organized as a "translocon" type like the IgH loci of higher vertebrates. In contrast, the Ig light (L) chain locus is arranged in a "multicluster" or repeating set of VL, JL, and CL segments. The IgL chains have four isotypes; two κ L1/G and L3/F), σ (L2) and λ. The transcription of IgH genes in teleosts is regulated by a VH promoter and the Eμ3' enhancer, which both function in a B cell-specific manner. The location of the IgH locus, structure and transcriptional function of the Eμ3' enhancer are important to our understanding of the evolutional changes that have occurred in the IgH gene locus.

  14. Central Pathways Integrating Metabolism and Reproduction in Teleosts

    Directory of Open Access Journals (Sweden)

    Md eShahjahan

    2014-03-01

    Full Text Available Energy balance plays an important role in the control of reproduction. However, the cellular and molecular mechanisms connecting the two systems are not well understood especially in teleosts. The hypothalamus plays a crucial role in the regulation of both energy balance and reproduction, and contains a number of neuropeptides, including gonadotropin-releasing hormone (GnRH, orexin, neuropeptide-Y (NPY, ghrelin, pituitary adenylate cyclase-activating polypeptide (PACAP, α-melanocyte stimulating hormone (α-MSH, melanin-concentrating hormone (MCH, cholecystokinin (CCK, 26RFa, nesfatin, kisspeptin, and gonadotropin-inhibitory hormone (GnIH. These neuropeptides are involved in the control of energy balance and reproduction either directly or indirectly. On the other hand, synthesis and release of these hypothalamic neuropeptides are regulated by metabolic signals from the gut and the adipose tissue. Furthermore, neurons producing these neuropeptides interact with each other, providing neuronal basis of the link between energy balance and reproduction. This review summarizes the advances made in our understanding of the physiological roles of the hypothalamic neuropeptides in energy balance and reproduction in teleosts, and discusses how they interact with GnRH, kisspeptin, and pituitary gonadotropins to control reproduction in teleosts.

  15. Zebrafish Craniofacial Development: A Window into Early Patterning.

    Science.gov (United States)

    Mork, Lindsey; Crump, Gage

    2015-01-01

    The formation of the face and skull involves a complex series of developmental events mediated by cells derived from the neural crest, endoderm, mesoderm, and ectoderm. Although vertebrates boast an enormous diversity of adult facial morphologies, the fundamental signaling pathways and cellular events that sculpt the nascent craniofacial skeleton in the embryo have proven to be highly conserved from fish to man. The zebrafish Danio rerio, a small freshwater cyprinid fish from eastern India, has served as a popular model of craniofacial development since the 1990s. Unique strengths of the zebrafish model include a simplified skeleton during larval stages, access to rapidly developing embryos for live imaging, and amenability to transgenesis and complex genetics. In this chapter, we describe the anatomy of the zebrafish craniofacial skeleton; its applications as models for the mammalian jaw, middle ear, palate, and cranial sutures; the superior imaging technology available in fish that has provided unprecedented insights into the dynamics of facial morphogenesis; the use of the zebrafish to decipher the genetic underpinnings of craniofacial biology; and finally a glimpse into the most promising future applications of zebrafish craniofacial research.

  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 ex

  17. CD4-Transgenic Zebrafish Reveal Tissue-Resident Th2- and Regulatory T Cell-like Populations and Diverse Mononuclear Phagocytes.

    Science.gov (United States)

    Dee, Christopher T; Nagaraju, Raghavendar T; Athanasiadis, Emmanouil I; Gray, Caroline; Fernandez Del Ama, Laura; Johnston, Simon A; Secombes, Christopher J; Cvejic, Ana; Hurlstone, Adam F L

    2016-11-01

    CD4(+) T cells are at the nexus of the innate and adaptive arms of the immune system. However, little is known about the evolutionary history of CD4(+) T cells, and it is unclear whether their differentiation into specialized subsets is conserved in early vertebrates. In this study, we have created transgenic zebrafish with vibrantly labeled CD4(+) cells allowing us to scrutinize the development and specialization of teleost CD4(+) leukocytes in vivo. We provide further evidence that CD4(+) macrophages have an ancient origin and had already emerged in bony fish. We demonstrate the utility of this zebrafish resource for interrogating the complex behavior of immune cells at cellular resolution by the imaging of intimate contacts between teleost CD4(+) T cells and mononuclear phagocytes. Most importantly, we reveal the conserved subspecialization of teleost CD4(+) T cells in vivo. We demonstrate that the ancient and specialized tissues of the gills contain a resident population of il-4/13b-expressing Th2-like cells, which do not coexpress il-4/13a Additionally, we identify a contrasting population of regulatory T cell-like cells resident in the zebrafish gut mucosa, in marked similarity to that found in the intestine of mammals. Finally, we show that, as in mammals, zebrafish CD4(+) T cells will infiltrate melanoma tumors and obtain a phenotype consistent with a type 2 immune microenvironment. We anticipate that this unique resource will prove invaluable for future investigation of T cell function in biomedical research, the development of vaccination and health management in aquaculture, and for further research into the evolution of adaptive immunity.

  18. CD4-Transgenic Zebrafish Reveal Tissue-Resident Th2- and Regulatory T Cell–like Populations and Diverse Mononuclear Phagocytes

    Science.gov (United States)

    Dee, Christopher T.; Nagaraju, Raghavendar T.; Athanasiadis, Emmanouil I.; Gray, Caroline; Fernandez del Ama, Laura; Johnston, Simon A.; Secombes, Christopher J.

    2016-01-01

    CD4+ T cells are at the nexus of the innate and adaptive arms of the immune system. However, little is known about the evolutionary history of CD4+ T cells, and it is unclear whether their differentiation into specialized subsets is conserved in early vertebrates. In this study, we have created transgenic zebrafish with vibrantly labeled CD4+ cells allowing us to scrutinize the development and specialization of teleost CD4+ leukocytes in vivo. We provide further evidence that CD4+ macrophages have an ancient origin and had already emerged in bony fish. We demonstrate the utility of this zebrafish resource for interrogating the complex behavior of immune cells at cellular resolution by the imaging of intimate contacts between teleost CD4+ T cells and mononuclear phagocytes. Most importantly, we reveal the conserved subspecialization of teleost CD4+ T cells in vivo. We demonstrate that the ancient and specialized tissues of the gills contain a resident population of il-4/13b–expressing Th2-like cells, which do not coexpress il-4/13a. Additionally, we identify a contrasting population of regulatory T cell–like cells resident in the zebrafish gut mucosa, in marked similarity to that found in the intestine of mammals. Finally, we show that, as in mammals, zebrafish CD4+ T cells will infiltrate melanoma tumors and obtain a phenotype consistent with a type 2 immune microenvironment. We anticipate that this unique resource will prove invaluable for future investigation of T cell function in biomedical research, the development of vaccination and health management in aquaculture, and for further research into the evolution of adaptive immunity. PMID:27694495

  19. Proteomics of early zebrafish embryos

    Directory of Open Access Journals (Sweden)

    Heisenberg Carl-Philipp

    2006-01-01

    Full Text Available Abstract Background Zebrafish (D. rerio has become a powerful and widely used model system for the analysis of vertebrate embryogenesis and organ development. While genetic methods are readily available in zebrafish, protocols for two dimensional (2D gel electrophoresis and proteomics have yet to be developed. Results As a prerequisite to carry out proteomic experiments with early zebrafish embryos, we developed a method to efficiently remove the yolk from large batches of embryos. This method enabled high resolution 2D gel electrophoresis and improved Western blotting considerably. Here, we provide detailed protocols for proteomics in zebrafish from sample preparation to mass spectrometry (MS, including a comparison of databases for MS identification of zebrafish proteins. Conclusion The provided protocols for proteomic analysis of early embryos enable research to be taken in novel directions in embryogenesis.

  20. Somatic mutagenesis with a Sleeping Beauty transposon system leads to solid tumor formation in zebrafish.

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    Maura McGrail

    Full Text Available Large-scale sequencing of human cancer genomes and mouse transposon-induced tumors has identified a vast number of genes mutated in different cancers. One of the outstanding challenges in this field is to determine which genes, when mutated, contribute to cellular transformation and tumor progression. To identify new and conserved genes that drive tumorigenesis we have developed a novel cancer model in a distantly related vertebrate species, the zebrafish, Danio rerio. The Sleeping Beauty (SB T2/Onc transposon system was adapted for somatic mutagenesis in zebrafish. The carp ß-actin promoter was cloned into T2/Onc to create T2/OncZ. Two transgenic zebrafish lines that contain large concatemers of T2/OncZ were isolated by injection of linear DNA into the zebrafish embryo. The T2/OncZ transposons were mobilized throughout the zebrafish genome from the transgene array by injecting SB11 transposase RNA at the 1-cell stage. Alternatively, the T2/OncZ zebrafish were crossed to a transgenic line that constitutively expresses SB11 transposase. T2/OncZ transposon integration sites were cloned by ligation-mediated PCR and sequenced on a Genome Analyzer II. Between 700-6800 unique integration events in individual fish were mapped to the zebrafish genome. The data show that introduction of transposase by transgene expression or RNA injection results in an even distribution of transposon re-integration events across the zebrafish genome. SB11 mRNA injection resulted in neoplasms in 10% of adult fish at ∼10 months of age. T2/OncZ-induced zebrafish tumors contain many mutated genes in common with human and mouse cancer genes. These analyses validate our mutagenesis approach and provide additional support for the involvement of these genes in human cancers. The zebrafish T2/OncZ cancer model will be useful for identifying novel and conserved genetic drivers of human cancers.

  1. Zebrafish (Danio rerio) bioassay for visceral toxicosis of catfish and botulinum neurotoxin serotype E.

    Science.gov (United States)

    Chatla, Kamalakar; Gaunt, Patricia; Petrie-Hanson, Lora; Hohn, Claudia; Ford, Lorelei; Hanson, Larry

    2014-03-01

    Visceral toxicosis of catfish (VTC), a sporadic disease of cultured channel catfish (Ictalurus punctatus) often with high mortality, is caused by botulinum neurotoxin serotype E (BoNT/E). Presumptive diagnosis of VTC is based on characteristic clinical signs and lesions, and the production of these signs and mortality after sera from affected fish is administered to sentinel catfish. The diagnosis is confirmed if the toxicity is neutralized with BoNT/E antitoxin. Because small catfish are often unavailable, the utility of adult zebrafish (Danio rerio) was evaluated in BoNT/E and VTC bioassays. Channel catfish and zebrafish susceptibilities were compared using trypsin-activated BoNT/E in a 96-hr trial by intracoelomically administering 0, 1.87, 3.7, 7.5, 15, or 30 pg of toxin per gram of body weight (g-bw) of fish. All of the zebrafish died at the 7.5 pg/g-bw and higher, while the catfish died at the 15 pg/g-bw dose and higher. To test the bioassay, sera from VTC-affected fish or control sera were intracoelomically injected at a dose of 10 µl per zebrafish and 20 µl/g-bw for channel catfish. At 96 hr post-injection, 78% of the zebrafish and 50% of the catfish receiving VTC sera died, while no control fish died. When the VTC sera were preincubated with BoNT/E antitoxin, they became nontoxic to zebrafish. Histology of zebrafish injected with either VTC serum or BoNT/E demonstrated renal necrosis. Normal catfish serum was toxic to larval zebrafish in immersion exposures, abrogating their utility in VTC bioassays. The results demonstrate bioassays using adult zebrafish for detecting BoNT/E and VTC are sensitive and practical.

  2. Automatic zebrafish heartbeat detection and analysis for zebrafish embryos.

    Science.gov (United States)

    Pylatiuk, Christian; Sanchez, Daniela; Mikut, Ralf; Alshut, Rüdiger; Reischl, Markus; Hirth, Sofia; Rottbauer, Wolfgang; Just, Steffen

    2014-08-01

    A fully automatic detection and analysis method of heartbeats in videos of nonfixed and nonanesthetized zebrafish embryos is presented. This method reduces the manual workload and time needed for preparation and imaging of the zebrafish embryos, as well as for evaluating heartbeat parameters such as frequency, beat-to-beat intervals, and arrhythmicity. The method is validated by a comparison of the results from automatic and manual detection of the heart rates of wild-type zebrafish embryos 36-120 h postfertilization and of embryonic hearts with bradycardia and pauses in the cardiac contraction.

  3. Role of G protein-coupled estrogen receptor 1, GPER, in inhibition of oocyte maturation by endogenous estrogens in zebrafish.

    Science.gov (United States)

    Pang, Yefei; Thomas, Peter

    2010-06-15

    Estrogen inhibition of oocyte maturation (OM) and the role of GPER (formerly known as GPR30) were investigated in zebrafish. Estradiol-17beta (E2) and G-1, a GPER-selective agonist, bound to zebrafish oocyte membranes suggesting the presence of GPER which was confirmed by immunocytochemistry using a specific GPER antibody. Incubation of follicle-enclosed oocytes with an aromatase inhibitor, ATD, and enzymatic and manual removal of the ovarian follicle cell layers significantly increased spontaneous OM which was partially reversed by co-treatment with either 100 nM E2 or G-1. Incubation of denuded oocytes with the GPER antibody blocked the inhibitory effects of estrogens on OM, whereas microinjection of estrogen receptor alpha (ERalpha) antisense oligonucleotides into the oocytes was ineffective. The results suggest that endogenous estrogens produced by the follicle cells inhibit or delay spontaneous maturation of zebrafish oocytes and that this estrogen action is mediated through GPER. Treatment with E2 and G-1 also attenuated the stimulatory effect of the teleost maturation-inducing steroid, 17,20beta-dihyroxy-4-pregnen-3-one (DHP), on OM. Moreover, E2 and G-1 down-regulated the expression of membrane progestin receptor alpha (mPRalpha), the intermediary in DHP induction of OM. Conversely DHP treatment caused a >50% decline in GPER mRNA levels. The results suggest that estrogens and GPER are critical components of the endocrine system controlling the onset of OM in zebrafish. A model is proposed for the dual control of the onset of oocyte maturation in teleosts by estrogens and progestins acting through GPER and mPRalpha, respectively, at different stages of oocyte development.

  4. The Effect of Zeaxanthin on the Visual Acuity of Zebrafish.

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    Eric A Saidi

    Full Text Available Oral supplementation of carotenoids such as zeaxanthin or lutein which naturally occur in human retina have been shown to improve vision and prevent progression of damage to advanced AMD in some studies. The zebrafish eye shares many physiological similarities with the human eye and is increasingly being used as model for vision research. We hypothesized that injection of zeaxanthin into the zebrafish eye would improve the visual acuity of the zebrafish over time. Visual acuity, calculated in cycles per degree, was measured in adult zebrafish to establish a consistent baseline using the optokinetic response. Zeaxanthin dissolved into phosphate buffered saline (PBS or PBS only was injected into the anterior chamber of the right and left eyes of the Zebrafish. Visual acuities were measured at 1 week and 3, 8 and 12 weeks post-injection to compare to baseline values. Repeated measures ANOVA was used to compare visual acuities between fish injected with PBS and zeaxanthin. A significant improvement in visual acuity, 14% better than before the injection (baseline levels, was observed one week after injection with zeaxanthin (p = 0.04. This improvement peaked at more than 30% for some fish a few weeks after the injection and improvement in vision persisted at 3 weeks after injection (p = 0.006. The enhanced visual function was not significantly better than baseline at 8 weeks (p = 0.19 and returned to baseline levels 12 weeks after the initial injection (p = 0.50. Zeaxanthin can improve visual acuity in zebrafish eyes. Further studies are required to develop a better understanding of the role zeaxanthin and other carotenoids play during normal visual function.

  5. The Effect of Zeaxanthin on the Visual Acuity of Zebrafish.

    Science.gov (United States)

    Saidi, Eric A; Davey, Pinakin Gunvant; Cameron, D Joshua

    2015-01-01

    Oral supplementation of carotenoids such as zeaxanthin or lutein which naturally occur in human retina have been shown to improve vision and prevent progression of damage to advanced AMD in some studies. The zebrafish eye shares many physiological similarities with the human eye and is increasingly being used as model for vision research. We hypothesized that injection of zeaxanthin into the zebrafish eye would improve the visual acuity of the zebrafish over time. Visual acuity, calculated in cycles per degree, was measured in adult zebrafish to establish a consistent baseline using the optokinetic response. Zeaxanthin dissolved into phosphate buffered saline (PBS) or PBS only was injected into the anterior chamber of the right and left eyes of the Zebrafish. Visual acuities were measured at 1 week and 3, 8 and 12 weeks post-injection to compare to baseline values. Repeated measures ANOVA was used to compare visual acuities between fish injected with PBS and zeaxanthin. A significant improvement in visual acuity, 14% better than before the injection (baseline levels), was observed one week after injection with zeaxanthin (p = 0.04). This improvement peaked at more than 30% for some fish a few weeks after the injection and improvement in vision persisted at 3 weeks after injection (p = 0.006). The enhanced visual function was not significantly better than baseline at 8 weeks (p = 0.19) and returned to baseline levels 12 weeks after the initial injection (p = 0.50). Zeaxanthin can improve visual acuity in zebrafish eyes. Further studies are required to develop a better understanding of the role zeaxanthin and other carotenoids play during normal visual function.

  6. Zebrafish pancreas development.

    Science.gov (United States)

    Tiso, Natascia; Moro, Enrico; Argenton, Francesco

    2009-11-27

    An accurate understanding of the molecular events governing pancreas development can have an impact on clinical medicine related to diabetes, obesity and pancreatic cancer, diseases with a high impact in public health. Until 1996, the main animal models in which pancreas formation and differentiation could be studied were mouse and, for some instances related to early development, chicken and Xenopus. Zebrafish has penetrated this field very rapidly offering a new model of investigation; by joining functional genomics, genetics and in vivo whole mount visualization, Danio rerio has allowed large scale and fine multidimensional analysis of gene functions during pancreas formation and differentiation.

  7. The zebrafish progranulin gene family and antisense transcripts

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    Baranowski David

    2005-11-01

    Full Text Available Abstract Background Progranulin is an epithelial tissue growth factor (also known as proepithelin, acrogranin and PC-cell-derived growth factor that has been implicated in development, wound healing and in the progression of many cancers. The single mammalian progranulin gene encodes a glycoprotein precursor consisting of seven and one half tandemly repeated non-identical copies of the cystine-rich granulin motif. A genome-wide duplication event hypothesized to have occurred at the base of the teleost radiation predicts that mammalian progranulin may be represented by two co-orthologues in zebrafish. Results The cDNAs encoding two zebrafish granulin precursors, progranulins-A and -B, were characterized and found to contain 10 and 9 copies of the granulin motif respectively. The cDNAs and genes encoding the two forms of granulin, progranulins-1 and -2, were also cloned and sequenced. Both latter peptides were found to be encoded by precursors with a simplified architecture consisting of one and one half copies of the granulin motif. A cDNA encoding a chimeric progranulin which likely arises through the mechanism of trans-splicing between grn1 and grn2 was also characterized. A non-coding RNA gene with antisense complementarity to both grn1 and grn2 was identified which may have functional implications with respect to gene dosage, as well as in restricting the formation of the chimeric form of progranulin. Chromosomal localization of the four progranulin (grn genes reveals syntenic conservation for grna only, suggesting that it is the true orthologue of mammalian grn. RT-PCR and whole-mount in situ hybridization analysis of zebrafish grns during development reveals that combined expression of grna and grnb, but not grn1 and grn2, recapitulate many of the expression patterns observed for the murine counterpart. This includes maternal deposition, widespread central nervous system distribution and specific localization within the epithelial

  8. The evolution and appearance of C3 duplications in fish originate an exclusive teleost c3 gene form with anti-inflammatory activity.

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    Gabriel Forn-Cuní

    Full Text Available The complement system acts as a first line of defense and promotes organism homeostasis by modulating the fates of diverse physiological processes. Multiple copies of component genes have been previously identified in fish, suggesting a key role for this system in aquatic organisms. Herein, we confirm the presence of three different previously reported complement c3 genes (c3.1, c3.2, c3.3 and identify five additional c3 genes (c3.4, c3.5, c3.6, c3.7, c3.8 in the zebrafish genome. Additionally, we evaluate the mRNA expression levels of the different c3 genes during ontogeny and in different tissues under steady-state and inflammatory conditions. Furthermore, while reconciling the phylogenetic tree with the fish species tree, we uncovered an event of c3 duplication common to all teleost fishes that gave rise to an exclusive c3 paralog (c3.7 and c3.8. These paralogs showed a distinct ability to regulate neutrophil migration in response to injury compared with the other c3 genes and may play a role in maintaining the balance between inflammatory and homeostatic processes in zebrafish.

  9. Nicotinic involvement in memory function in zebrafish.

    Science.gov (United States)

    Levin, Edward D; Chen, Elaine

    2004-01-01

    Zebrafish are an emerging model for the study of the molecular mechanisms of brain function. To conduct studies of the neural bases of behavior in zebrafish, we must understand the behavioral function of zebrafish and how it is altered by perturbations of brain function. This study determined nicotine actions on memory function in zebrafish. With the methods that we have developed to assess memory in zebrafish using delayed spatial alternation (DSA), we determined the dose effect function of acute nicotine on memory function in zebrafish. As in rodents and primates, low nicotine doses improve memory in zebrafish, while high nicotine doses have diminished effect and can impair memory. This study shows that nicotine affects memory function in zebrafish much like in rats, mice, monkeys and humans. Now, zebrafish can be used to help understand the molecular mechanisms crucial to nicotine effects on memory.

  10. Genome-wide Gene Expression Profiling of Acute Metal Exposures in Male Zebrafish

    Science.gov (United States)

    2014-10-23

    and methods Experimental overview Fig. 1 shows the design scheme. Fish exposures Briefly, adult,male zebrafishwere exposed to concentrations of each...before and after exposures. Initially 25 adult (6–9 months) presumptive male zebrafish were selected per condition to ensure that 20 male fish were...evaluated in whole adult male zebra ???sh following acute 24 h high dose exposure to three metals with known human health risks. Male adult zebra ???sh

  11. Antibiotic toxicity and absorption in zebrafish using liquid chromatography-tandem mass spectrometry.

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    Fan Zhang

    Full Text Available Evaluation of drug toxicity is necessary for drug safety, but in vivo drug absorption is varied; therefore, a rapid, sensitive and reliable method for measuring drugs is needed. Zebrafish are acceptable drug toxicity screening models; we used these animals with a liquid chromatography-tandem mass spectrometry (LC-MS/MS method in a multiple reaction monitoring mode to quantify drug uptake in zebrafish to better estimate drug toxicity. Analytes were recovered from zebrafish homogenate by collecting supernatant. Measurements were confirmed for drugs in the range of 10-1,000 ng/mL. Four antibiotics with different polarities were tested to explore any correlation of drug polarity, absorption, and toxicity. Zebrafish at 3 days post-fertilization (dpf absorbed more drug than those at 6 h post-fertilization (hpf, and different developmental periods appeared to be differentially sensitive to the same compound. By observing abnormal embryos and LD50 values, zebrafish embryos at 6 hpf were considered to be suitable for evaluating embryotoxicity. Also, larvae at 3 dpf were adapted to measure acute drug toxicity in adult mammals. Thus, we can exploit zebrafish to study drug toxicity and can reliably quantify drug uptake with LC-MS/MS. This approach will be helpful for future studies of toxicology in zebrafish.

  12. Mind the fish: zebrafish as a model in cognitive social neuroscience.

    Science.gov (United States)

    Oliveira, Rui F

    2013-01-01

    Understanding how the brain implements social behavior on one hand, and how social processes feedback on the brain to promote fine-tuning of behavioral output according to changes in the social environment is a major challenge in contemporary neuroscience. A critical step to take this challenge successfully is finding the appropriate level of analysis when relating social to biological phenomena. Given the enormous complexity of both the neural networks of the brain and social systems, the use of a cognitive level of analysis (in an information processing perspective) is proposed here as an explanatory interface between brain and behavior. A conceptual framework for a cognitive approach to comparative social neuroscience is proposed, consisting of the following steps to be taken across different species with varying social systems: (1) identification of the functional building blocks of social skills; (2) identification of the cognitive mechanisms underlying the previously identified social skills; and (3) mapping these information processing mechanisms onto the brain. Teleost fish are presented here as a group of choice to develop this approach, given the diversity of social systems present in closely related species that allows for planned phylogenetic comparisons, and the availability of neurogenetic tools that allows the visualization and manipulation of selected neural circuits in model species such as the zebrafish. Finally, the state-of-the art of zebrafish social cognition and of the tools available to map social cognitive abilities to neural circuits in zebrafish are reviewed.

  13. Mind the fish: zebrafish as a model in cognitive social neuroscience

    Directory of Open Access Journals (Sweden)

    Rui F Oliveira

    2013-08-01

    Full Text Available Understanding how the brain implements social behavior on one hand, and how social processes feedback on the brain to promote fine-tuning of behavioural output according to changes in the social environment is a major challenge in contemporary neuroscience. A critical step to take this challenge successfully is finding the appropriate level of analysis when relating social to biological phenomena. Given the enormous complexity of both the neural networks of the brain and social systems, the use of a cognitive level of analysis (in an information processing perspective is proposed here as an explanatory interface between brain and behavior. A conceptual framework for a cognitive approach to comparative social neuroscience is proposed, consisting of the following steps to be taken across different species with varying social systems: (1 identification of the functional building blocks of social skills; (2 identification of the cognitive mechanisms underlying the previously identified social skills; and (3 mapping these information processing mechanisms onto the brain. Teleost fish are presented here as a group of choice to develop this approach, given the diversity of social systems present in closely related species that allows for planned phylogenetic comparisons, and the availability of neurogenetic tools that allows the visualization and manipulation of selected neural circuits in model species such as the zebrafish. Finally, the state-of-the art of zebrafish social cognition and of the tools available to map social cognitive abilities to neural circuits in zebrafish are reviewed.

  14. Identification of IL-34 in teleost fish: differential expression of rainbow trout IL-34, MCSF1 and MCSF2, ligands of the MCSF receptor.

    Science.gov (United States)

    Wang, Tiehui; Kono, Tomoya; Monte, Milena M; Kuse, Haruka; Costa, Maria M; Korenaga, Hiroki; Maehr, Tanja; Husain, Mansourah; Sakai, Masahiro; Secombes, Christopher J

    2013-04-01

    The mononuclear phagocyte system is composed of monocytes, macrophages and dendritic cells and has crucial roles in inflammation, autoimmunity, infection, cancer, organ transplantation and in maintaining organismal homeostasis. Interleukin-34 (IL-34) and macrophage colony stimulating factor (MCSF), both signalling through the MCSF receptor, regulate the mononuclear phagocyte system. A single IL-34 and MCSF gene are present in tetrapods. Two types of MCSF exist in teleost fish which is resulted from teleost-wide whole genome duplication. In this report, we first identified and sequence analysed six IL-34 genes in five teleost fish, rainbow trout, fugu, Atlantic salmon, catfish and zebrafish. The fish IL-34 molecules had a higher identity within fish group but low identities to IL-34s from birds (27.2-33.8%) and mammals (22.2-31.4%). However, they grouped with tetrapod IL-34 molecules in phylogenetic tree analysis, had a similar 7 exon/6 intron gene organisation, and genes in the IL-34 loci were syntenically conserved. In addition, the regions of the four main helices, along with a critical N-glycosylation site were well conserved. Taken together these data suggest that the teleost IL-34 genes described in this report are orthologues of tetrapod IL-34. Comparative expression study of the three trout MCSFR ligands revealed that IL-34, MCSF1 and MCSF2 are differentially expressed in tissues and cell lines. The expression of MCSF1 and MCSF2 showed great variance in different tissues and cell lines, suggesting a role in the differentiation and maintenance of specific macrophage lineages in specific locations. The relatively high levels of IL-34 expression across different tissues suggests a homeostatic role of IL-34 for the macrophage lineage in fish. One striking observation in the present study was the lack of induction of MCSF1 and MCSF2 expression but the quick induction of IL-34 expression by PAMPs and inflammatory cytokines in cell lines and primary head kidney

  15. Three nuclear and two membrane estrogen receptors in basal teleosts, Anguilla sp.: Identification, evolutionary history and differential expression regulation

    DEFF Research Database (Denmark)

    Lafont, Anne Gaëlle; Rousseau, Karine; Tomkiewicz, Jonna;

    2016-01-01

    teleosts. Phylogeny and synteny analyses suggest that they result from teleost whole genome duplication (3R). In contrast to conserved 3R-duplicated ESR2 and GPER, one of 3R-duplicated ESR1 has been lost shortly after teleost emergence. Quantitative PCRs revealed that the five receptors are all widely...

  16. Antimicrobial Peptides as Mediators of Innate Immunity in Teleosts

    Directory of Open Access Journals (Sweden)

    Barbara A. Katzenback

    2015-09-01

    Full Text Available Antimicrobial peptides (AMPs have been identified throughout the metazoa suggesting their evolutionarily conserved nature and their presence in teleosts is no exception. AMPs are short (18–46 amino acids, usually cationic, amphipathic peptides. While AMPs are diverse in amino acid sequence, with no two AMPs being identical, they collectively appear to have conserved functions in the innate immunity of animals towards the pathogens they encounter in their environment. Fish AMPs are upregulated in response to pathogens and appear to have direct broad-spectrum antimicrobial activity towards both human and fish pathogens. However, an emerging role for AMPs as immunomodulatory molecules has become apparent—the ability of AMPs to activate the innate immune system sheds light onto the multifaceted capacity of these small peptides to combat pathogens through direct and indirect means. Herein, this review focuses on the role of teleost AMPs as modulators of the innate immune system and their regulation in response to pathogens or other exogenous molecules. The capacity to regulate AMP expression by exogenous factors may prove useful in modulating AMP expression in fish to prevent disease, particularly in aquaculture settings where crowded conditions and environmental stress pre-dispose these fish to infection.

  17. Neutrophil Development, Migration, and Function in Teleost Fish

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    Jeffrey J. Havixbeck

    2015-11-01

    Full Text Available It is now widely recognized that neutrophils are sophisticated cells that are critical to host defense and the maintenance of homeostasis. In addition, concepts such as neutrophil plasticity are helping to define the range of phenotypic profiles available to cells in this group and the physiological conditions that contribute to their differentiation. Herein, we discuss key features of the life of a teleost neutrophil including their development, migration to an inflammatory site, and contributions to pathogen killing and the control of acute inflammation. The potent anti-microbial mechanisms elicited by these cells in bony fish are a testament to their long-standing evolutionary contributions in host defense. In addition, recent insights into their active roles in the control of inflammation prior to induction of apoptosis highlight their importance to the maintenance of host integrity in these early vertebrates. Overall, our goal is to summarize recent progress in our understanding of this cell type in teleost fish, and to provide evolutionary context for the contributions of this hematopoietic lineage in host defense and an efficient return to homeostasis following injury or infection.

  18. Antimicrobial Peptides as Mediators of Innate Immunity in Teleosts.

    Science.gov (United States)

    Katzenback, Barbara A

    2015-09-25

    Antimicrobial peptides (AMPs) have been identified throughout the metazoa suggesting their evolutionarily conserved nature and their presence in teleosts is no exception. AMPs are short (18-46 amino acids), usually cationic, amphipathic peptides. While AMPs are diverse in amino acid sequence, with no two AMPs being identical, they collectively appear to have conserved functions in the innate immunity of animals towards the pathogens they encounter in their environment. Fish AMPs are upregulated in response to pathogens and appear to have direct broad-spectrum antimicrobial activity towards both human and fish pathogens. However, an emerging role for AMPs as immunomodulatory molecules has become apparent-the ability of AMPs to activate the innate immune system sheds light onto the multifaceted capacity of these small peptides to combat pathogens through direct and indirect means. Herein, this review focuses on the role of teleost AMPs as modulators of the innate immune system and their regulation in response to pathogens or other exogenous molecules. The capacity to regulate AMP expression by exogenous factors may prove useful in modulating AMP expression in fish to prevent disease, particularly in aquaculture settings where crowded conditions and environmental stress pre-dispose these fish to infection.

  19. Embryonic senescence and laminopathies in a progeroid zebrafish model.

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    Eriko Koshimizu

    Full Text Available BACKGROUND: Mutations that disrupt the conversion of prelamin A to mature lamin A cause the rare genetic disorder Hutchinson-Gilford progeria syndrome and a group of laminopathies. Our understanding of how A-type lamins function in vivo during early vertebrate development through aging remains limited, and would benefit from a suitable experimental model. The zebrafish has proven to be a tractable model organism for studying both development and aging at the molecular genetic level. Zebrafish show an array of senescence symptoms resembling those in humans, which can be targeted to specific aging pathways conserved in vertebrates. However, no zebrafish models bearing human premature senescence currently exist. PRINCIPAL FINDINGS: We describe the induction of embryonic senescence and laminopathies in zebrafish harboring disturbed expressions of the lamin A gene (LMNA. Impairments in these fish arise in the skin, muscle and adipose tissue, and sometimes in the cartilage. Reduced function of lamin A/C by translational blocking of the LMNA gene induced apoptosis, cell-cycle arrest, and craniofacial abnormalities/cartilage defects. By contrast, induced cryptic splicing of LMNA, which generates the deletion of 8 amino acid residues lamin A (zlamin A-Δ8, showed embryonic senescence and S-phase accumulation/arrest. Interestingly, the abnormal muscle and lipodystrophic phenotypes were common in both cases. Hence, both decrease-of-function of lamin A/C and gain-of-function of aberrant lamin A protein induced laminopathies that are associated with mesenchymal cell lineages during zebrafish early development. Visualization of individual cells expressing zebrafish progerin (zProgerin/zlamin A-Δ37 fused to green fluorescent protein further revealed misshapen nuclear membrane. A farnesyltransferase inhibitor reduced these nuclear abnormalities and significantly prevented embryonic senescence and muscle fiber damage induced by zProgerin. Importantly, the adult

  20. Effect of acute ethanol administration on zebrafish tail-beat motion.

    Science.gov (United States)

    Bartolini, Tiziana; Mwaffo, Violet; Butail, Sachit; Porfiri, Maurizio

    2015-11-01

    Zebrafish is becoming a species of choice in neurobiological and behavioral studies of alcohol-related disorders. In these efforts, the activity of adult zebrafish is typically quantified using indirect activity measures that are either scored manually or identified automatically from the fish trajectory. The analysis of such activity measures has produced important insight into the effect of acute ethanol exposure on individual and social behavior of this vertebrate species. Here, we leverage a recently developed tracking algorithm that reconstructs fish body shape to investigate the effect of acute ethanol administration on zebrafish tail-beat motion in terms of amplitude and frequency. Our results demonstrate a significant effect of ethanol on the tail-beat amplitude as well as the tail-beat frequency, both of which were found to robustly decrease for high ethanol concentrations. Such a direct measurement of zebrafish motor functions is in agreement with evidence based on indirect activity measures, offering a complementary perspective in behavioral screening.

  1. Mitragynine attenuates withdrawal syndrome in morphine-withdrawn zebrafish.

    Directory of Open Access Journals (Sweden)

    Beng-Siang Khor

    Full Text Available A major obstacle in treating drug addiction is the severity of opiate withdrawal syndrome, which can lead to unwanted relapse. Mitragynine is the major alkaloid compound found in leaves of Mitragyna speciosa, a plant widely used by opiate addicts to mitigate the harshness of drug withdrawal. A series of experiments was conducted to investigate the effect of mitragynine on anxiety behavior, cortisol level and expression of stress pathway related genes in zebrafish undergoing morphine withdrawal phase. Adult zebrafish were subjected to two weeks chronic morphine exposure at 1.5 mg/L, followed by withdrawal for 24 hours prior to tests. Using the novel tank diving tests, we first showed that morphine-withdrawn zebrafish display anxiety-related swimming behaviors such as decreased exploratory behavior and increased erratic movement. Morphine withdrawal also elevated whole-body cortisol levels, which confirms the phenotypic stress-like behaviors. Exposing morphine-withdrawn fish to mitragynine however attenuates majority of the stress-related swimming behaviors and concomitantly lower whole-body cortisol level. Using real-time PCR gene expression analysis, we also showed that mitragynine reduces the mRNA expression of corticotropin releasing factor receptors and prodynorphin in zebrafish brain during morphine withdrawal phase, revealing for the first time a possible link between mitragynine's ability to attenuate anxiety during opiate withdrawal with the stress-related corticotropin pathway.

  2. Determinism and stochasticity during maturation of the zebrafish antibody repertoire

    Science.gov (United States)

    Jiang, Ning; Weinstein, Joshua A.; Penland, Lolita; White, Richard A.; Fisher, Daniel S.; Quake, Stephen R.

    2011-01-01

    It is thought that the adaptive immune system of immature organisms follows a more deterministic program of antibody creation than is found in adults. We used high-throughput sequencing to characterize the diversifying antibody repertoire in zebrafish over five developmental time points. We found that the immune system begins in a highly stereotyped state with preferential use of a small number of V (variable) D (diverse) J (joining) gene segment combinations, but that this stereotypy decreases dramatically as the zebrafish mature, with many of the top VDJ combinations observed in 2-wk-old zebrafish virtually disappearing by 1 mo. However, we discovered that, in the primary repertoire, there are strong correlations in VDJ use that increase with zebrafish maturity, suggesting that VDJ recombination involves a level of deterministic programming that is unexpected. This stereotypy is masked by the complex diversification processes of antibody maturation; the variation and lack of correlation in full repertoires between individuals appears to be derived from randomness in clonal expansion during the affinity maturation process. These data provide a window into the mechanisms of VDJ recombination and diversity creation and allow us to better understand how the adaptive immune system achieves diversity. PMID:21393572

  3. Chemokine-guided angiogenesis directs coronary vasculature formation in zebrafish.

    Science.gov (United States)

    Harrison, Michael R M; Bussmann, Jeroen; Huang, Ying; Zhao, Long; Osorio, Arthela; Burns, C Geoffrey; Burns, Caroline E; Sucov, Henry M; Siekmann, Arndt F; Lien, Ching-Ling

    2015-05-26

    Interruption of the coronary blood supply severely impairs heart function with often fatal consequences for patients. However, the formation and maturation of these coronary vessels is not fully understood. Here we provide a detailed analysis of coronary vessel development in zebrafish. We observe that coronary vessels form in zebrafish by angiogenic sprouting of arterial cells derived from the endocardium at the atrioventricular canal. Endothelial cells express the CXC-motif chemokine receptor Cxcr4a and migrate to vascularize the ventricle under the guidance of the myocardium-expressed ligand Cxcl12b. cxcr4a mutant zebrafish fail to form a vascular network, whereas ectopic expression of Cxcl12b ligand induces coronary vessel formation. Importantly, cxcr4a mutant zebrafish fail to undergo heart regeneration following injury. Our results suggest that chemokine signaling has an essential role in coronary vessel formation by directing migration of endocardium-derived endothelial cells. Poorly developed vasculature in cxcr4a mutants likely underlies decreased regenerative potential in adults.

  4. Molecular cloning and analysis of zebrafish voltage-gated sodium channel beta subunit genes: implications for the evolution of electrical signaling in vertebrates

    Directory of Open Access Journals (Sweden)

    Zhong Tao P

    2007-07-01

    Full Text Available Abstract Background Action potential generation in excitable cells such as myocytes and neurons critically depends on voltage-gated sodium channels. In mammals, sodium channels exist as macromolecular complexes that include a pore-forming alpha subunit and 1 or more modulatory beta subunits. Although alpha subunit genes have been cloned from diverse metazoans including flies, jellyfish, and humans, beta subunits have not previously been identified in any non-mammalian species. To gain further insight into the evolution of electrical signaling in vertebrates, we investigated beta subunit genes in the teleost Danio rerio (zebrafish. Results We identified and cloned single zebrafish gene homologs for beta1-beta3 (zbeta1-zbeta3 and duplicate genes for beta4 (zbeta4.1, zbeta4.2. Sodium channel beta subunit loci are similarly organized in fish and mammalian genomes. Unlike their mammalian counterparts, zbeta1 and zbeta2 subunit genes display extensive alternative splicing. Zebrafish beta subunit genes and their splice variants are differentially-expressed in excitable tissues, indicating tissue-specific regulation of zbeta1-4 expression and splicing. Co-expression of the genes encoding zbeta1 and the zebrafish sodium channel alpha subunit Nav1.5 in Chinese Hamster Ovary cells increased sodium current and altered channel gating, demonstrating functional interactions between zebrafish alpha and beta subunits. Analysis of the synteny and phylogeny of mammalian, teleost, amphibian, and avian beta subunit and related genes indicated that all extant vertebrate beta subunits are orthologous, that beta2/beta4 and beta1/beta3 share common ancestry, and that beta subunits are closely related to other proteins sharing the V-type immunoglobulin domain structure. Vertebrate sodium channel beta subunit genes were not identified in the genomes of invertebrate chordates and are unrelated to known subunits of the para sodium channel in Drosophila. Conclusion The

  5. 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.

  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. Comprehensive profiling of zebrafish hepatic proximal promoter CpG island methylation and its modification during chemical carcinogenesis

    Directory of Open Access Journals (Sweden)

    Gong Zhiyuan

    2011-01-01

    Full Text Available Abstract Background DNA methylation is an epigenetic mechanism associated with regulation of gene expression and it is modulated during chemical carcinogenesis. The zebrafish is increasingly employed as a human disease model; however there is a lack of information on DNA methylation in zebrafish and during fish tumorigenesis. Results A novel CpG island tiling array containing 44,000 probes, in combination with immunoprecipitation of methylated DNA, was used to achieve the first comprehensive methylation profiling of normal adult zebrafish liver. DNA methylation alterations were detected in zebrafish liver tumors induced by the environmental carcinogen 7, 12-dimethylbenz(aanthracene. Genes significantly hypomethylated in tumors were associated particularly with proliferation, glycolysis, transcription, cell cycle, apoptosis, growth and metastasis. Hypermethylated genes included those associated with anti-angiogenesis and cellular adhesion. Of 49 genes that were altered in expression within tumors, and which also had appropriate CpG islands and were co-represented on the tiling array, approximately 45% showed significant changes in both gene expression and methylation. Conclusion The functional pathways containing differentially methylated genes in zebrafish hepatocellular carcinoma have also been reported to be aberrantly methylated during tumorigenesis in humans. These findings increase the confidence in the use of zebrafish as a model for human cancer in addition to providing the first comprehensive mapping of DNA methylation in the normal adult zebrafish liver.

  8. Zebrafish brain lipid characterization and quantification by ¹H nuclear magnetic resonance spectroscopy and MALDI-TOF mass spectrometry.

    Science.gov (United States)

    van Amerongen, Yvonne F; Roy, Upasana; Spaink, Herman P; de Groot, Huub J M; Huster, Daniel; Schiller, Jürgen; Alia, A

    2014-06-01

    Lipids play an important role in many neurodegenerative diseases, such as Parkinson's disease, Alzheimer's disease, and Huntington's disease. Zebrafish models for these diseases have been recently developed. The detailed brain lipid composition of the adult zebrafish is not known, and therefore, the representativeness of these models cannot be properly evaluated. In this study, we characterized the total lipid composition of healthy adult zebrafish using (1)H nuclear magnetic resonance spectroscopy. A close resemblance of the zebrafish brain composition is shown in comparison to the human brain. Moreover, several lipids involved in the pathogenesis of neurodegenerative diseases (i.e., cholesterol, phosphatidylcholine, docosahexaenoic acid, and further, polyunsaturated fatty acids) are detected and quantified. These lipids might represent useful biomarkers in future research toward human therapies. Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry coupled with high-performance thin-layer chromatography was used for further characterization of zebrafish brain lipids. Our results show that the lipid composition of the zebrafish brain is rather similar to the human brain and thus confirms that zebrafish represents a good model for studying various brain diseases.

  9. Phylogenetic consistencies among chondrichthyan and teleost fishes in their bioaccumulation of multiple trace elements from seawater

    Energy Technology Data Exchange (ETDEWEB)

    Jeffree, Ross A., E-mail: R.Jeffree@iaea.org [IAEA Marine Environment Laboratories, 4, Quai Antoine 1er, MC 98000 (Monaco); Oberhansli, Francois; Teyssie, Jean-Louis [IAEA Marine Environment Laboratories, 4, Quai Antoine 1er, MC 98000 (Monaco)

    2010-07-15

    Multi-tracer experiments determined the accumulation from seawater of selected radioactive trace elements (Mn-54, Co-60, Zn-65, Cs-134, Am-241, Cd-109, Ag-110m, Se-75 and Cr-51) by three teleost and three chondrichthyan fish species to test the hypothesis that these phylogenetic groups have different bioaccumulation characteristics, based on previously established contrasts between the carcharhiniform chondrichthyan Scyliorhinus canicula (dogfish) and the pleuronectiform teleost Psetta maxima (turbot). Discriminant function analysis on whole body: water concentration factors (CFs) separated dogfish and turbot in two independent experiments. Classification functions grouped the perciform teleosts, seabream (Sparus aurata) and seabass (Dicentrarchus labrax), with turbot and grouped the chondrichthyans, undulate ray (Raja undulata; Rajiformes) and spotted torpedo (Torpedo marmorata; Torpediniformes), with dogfish, thus supporting our hypothesis. Hierarchical classificatory, multi-dimensional scaling and similarity analyses based on the CFs for the nine radiotracers, also separated all three teleosts (that aggregated lower in the hierarchy) from the three chondrichthyan species. The three chondrichthyans were also more diverse amongst themselves compared to the three teleosts. Particular trace elements that were more important in separating teleosts and chondrichthyans were Cs-134 that was elevated in teleosts and Zn-65 that was elevated in chondrichthyans, these differences being due to their differential rates of uptake rather than loss. Chondrichthyans were also higher in Cr-51, Co-60, Ag-110m and Am-241, whereas teleosts were higher only in Mn-54. These contrasts in bioaccumulation patterns between teleosts and chondrichthyans are interpreted in the context of both proximate causes of underlying differences in physiology and anatomy, as well as the ultimate cause of their evolutionary divergence over more than 500 million years before present (MyBP). Our results

  10. Md1 and Rp105 regulate innate immunity and viral resistance in zebrafish.

    Science.gov (United States)

    Candel, Sergio; Sepulcre, María P; Espín-Palazón, Raquel; Tyrkalska, Sylwia D; de Oliveira, Sofía; Meseguer, José; Mulero, Victoriano

    2015-06-01

    TLR4 was the first TLR family member identified in mammals and is responsible for the activation of the immune response by bacterial LPS. Later, MD1 and RP105 were shown to form complexes that directly interact with the MD2-TLR4 complex, acting as physiological negative regulators of LPS signaling. Despite the general conservation of various TLR families from fish to mammals, several differences can be appreciated, such as the high tolerance of fish to LPS, the absence of the crucial accessory molecules Md2 and Cd14 for Tlr4 signaling in fish, the absence of Tlr4 in some fish species, and the confirmation that LPS does not signal through Tlr4 in zebrafish. The present study has identified the Rp105 and Md1 homologs in zebrafish, confirming (i) Rp105 and Tlr4 evolved from a common ancestor before the divergence between fish and tetrapods and (ii) the presence of Md1 in teleost fish and the lack of Md2, suggesting that the divergence of these accessory molecules occurred in the tetrapod lineage. Biochemical and functional studies indicate that Md1 binds both Rp105 and Tlr4 in zebrafish. Genetic inhibition of zebrafish Md1 and Rp105 reveals that Md1 or Rp105 deficiency impairs the expression of genes encoding pro-inflammatory and antiviral molecules, leading to increased susceptibility to viral infection. These results shed light on the evolutionary history of Md1 and Rp105 and uncover a previously unappreciated function of these molecules in the regulation of innate immunity.

  11. Stress and fear responses in the teleost pallium

    DEFF Research Database (Denmark)

    Silva, Patricia Isabel da Mota E.; Martins, C.I.M.; Khan, Uniza Wahid;

    2015-01-01

    combining skin extract with other challenges are needed to reveal neuroendocrine effects associated with this predator cue. Confinement stress resulted in an elevation of cortisol and serotonin (5-hydroxytryptamine, 5-HT) metabolism in both Dl and Dm. A similar tendency was observed in fish exposed...... been found in the teleost telencephalon. The dorsolateral (Dl) and dorsomedial (Dm) regions of the pallium are thought to perform hippocampus and amygdala-like functions respectively. To what degree these regions are involved in the neuroendocrine responses to stress and predator cues however remains...... largely unknown. In the present study the involvement of Dl and Dm in such responses was investigated by exposing Nile tilapia (Oreochromis niloticus) to a standardized confinement stress and to skin extract from conspecifics. Nile tilapia develops a characteristic anticipatory behaviour to hand feeding...

  12. Neurobehavioral impairments produced by developmental lead exposure persisted for generations in zebrafish (Danio rerio).

    Science.gov (United States)

    Xu, Xiaojuan; Weber, Daniel; Burge, Rebekah; VanAmberg, Kelsey

    2016-01-01

    The zebrafish has become a useful animal model for studying the effects of environmental contaminants on neurobehavioral development due to its ease of breeding, high number of eggs per female, short generation times, and a well-established avoidance conditioning paradigm. Using avoidance conditioning as the behavioral paradigm, the present study investigated the effects of embryonic exposure to lead (Pb) on learning in adult zebrafish and the third (F3) generation of those fish. In Experiment 1, adult zebrafish that were developmentally exposed to 0.0, 0.1, 1.0 or 10.0μM Pb (2-24h post fertilization) as embryos were trained and tested for avoidance responses. The results showed that adult zebrafish hatched from embryos exposed to 0.0 or 0.1μM Pb learned avoidance responses during training and displayed significantly increased avoidance responses during testing, while those hatched from embryos exposed to 1.0 or 10.0μM Pb displayed no significant increases in avoidance responses from training to testing. In Experiment 2, the F3 generation of zebrafish that were developmentally exposed to an identical exposure regimen as in Experiment 1 were trained and tested for avoidance responses. The results showed that the F3 generation of zebrafish developmentally exposed as embryos to 0.0 or 0.1μM Pb learned avoidance responses during training and displayed significantly increased avoidance responses during testing, while the F3 generation of zebrafish developmentally exposed as embryos to 1.0 or 10.0μM Pb displayed no significant changes in avoidance responses from training to testing. Thus, developmental Pb exposure produced learning impairments that persisted for at least three generations, demonstrating trans-generational effects of embryonic exposure to Pb.

  13. Evolution of space dependent growth in the teleost Astyanax mexicanus.

    Directory of Open Access Journals (Sweden)

    Natalya D Gallo

    Full Text Available The relationship between growth rate and environmental space is an unresolved issue in teleosts. While it is known from aquaculture studies that stocking density has a negative relationship to growth, the underlying mechanisms have not been elucidated, primarily because the growth rate of populations rather than individual fish were the subject of all previous studies. Here we investigate this problem in the teleost Astyanax mexicanus, which consists of a sighted surface-dwelling form (surface fish and several blind cave-dwelling (cavefish forms. Surface fish and cavefish are distinguished by living in spatially contrasting environments and therefore are excellent models to study the effects of environmental size on growth. Multiple controlled growth experiments with individual fish raised in confined or unconfined spaces showed that environmental size has a major impact on growth rate in surface fish, a trait we have termed space dependent growth (SDG. In contrast, SDG has regressed to different degrees in the Pachón and Tinaja populations of cavefish. Mating experiments between surface and Pachón cavefish show that SDG is inherited as a dominant trait and is controlled by multiple genetic factors. Despite its regression in blind cavefish, SDG is not affected when sighted surface fish are raised in darkness, indicating that vision is not required to perceive and react to environmental space. Analysis of plasma cortisol levels showed that an elevation above basal levels occurred soon after surface fish were exposed to confined space. This initial cortisol peak was absent in Pachón cavefish, suggesting that the effects of confined space on growth may be mediated partly through a stress response. We conclude that Astyanax reacts to confined spaces by exhibiting SDG, which has a genetic component and shows evolutionary regression during adaptation of cavefish to confined environments.

  14. Evolution of space dependent growth in the teleost Astyanax mexicanus.

    Science.gov (United States)

    Gallo, Natalya D; Jeffery, William R

    2012-01-01

    The relationship between growth rate and environmental space is an unresolved issue in teleosts. While it is known from aquaculture studies that stocking density has a negative relationship to growth, the underlying mechanisms have not been elucidated, primarily because the growth rate of populations rather than individual fish were the subject of all previous studies. Here we investigate this problem in the teleost Astyanax mexicanus, which consists of a sighted surface-dwelling form (surface fish) and several blind cave-dwelling (cavefish) forms. Surface fish and cavefish are distinguished by living in spatially contrasting environments and therefore are excellent models to study the effects of environmental size on growth. Multiple controlled growth experiments with individual fish raised in confined or unconfined spaces showed that environmental size has a major impact on growth rate in surface fish, a trait we have termed space dependent growth (SDG). In contrast, SDG has regressed to different degrees in the Pachón and Tinaja populations of cavefish. Mating experiments between surface and Pachón cavefish show that SDG is inherited as a dominant trait and is controlled by multiple genetic factors. Despite its regression in blind cavefish, SDG is not affected when sighted surface fish are raised in darkness, indicating that vision is not required to perceive and react to environmental space. Analysis of plasma cortisol levels showed that an elevation above basal levels occurred soon after surface fish were exposed to confined space. This initial cortisol peak was absent in Pachón cavefish, suggesting that the effects of confined space on growth may be mediated partly through a stress response. We conclude that Astyanax reacts to confined spaces by exhibiting SDG, which has a genetic component and shows evolutionary regression during adaptation of cavefish to confined environments.

  15. Conserved structure and expression of hsp70 paralogs in teleost fishes

    DEFF Research Database (Denmark)

    Metzger, David C.H.; Hansen, Jakob Hemmer; Schulte, Patricia M.

    2016-01-01

    The cytosolic 70 KDa heat shock proteins (Hsp70s) are widely used as biomarkers of environmental stress in ecological and toxicological studies in fish. Here we analyze teleost genome sequences to show that two genes encoding inducible hsp70s (hsp70-1 and hsp70-2) are likely present in all teleost...... fish. Phylogenetic and synteny analyses indicate that hsp70-1 and hsp70-2 are distinct paralogs that originated prior to the diversification of the teleosts. The promoters of both genes contain a TATA box and conserved heat shock elements (HSEs), but unlike mammalian HSP70s, both genes contain...

  16. Optokinetic behavior is reversed in achiasmatic mutant zebrafish larvae.

    Science.gov (United States)

    Rick, J M; Horschke, I; Neuhauss, S C

    2000-05-18

    The vertebrate optokinetic nystagmus (OKN) is a compensatory oculomotor behavior that is evoked by movement of the visual environment. It functions to stabilize visual images on the retina. The OKN can be experimentally evoked by rotating a drum fitted with stripes around the animal and has been studied extensively in many vertebrate species, including teleosts. This simple behavior has earlier been used to screen for mutations affecting visual system development in the vertebrate model organism zebrafish. In such a screen, we have found a significant number of homozygous belladonna (bel) mutant larvae to be defective in the correct execution of the OKN [1]. We now show that about 40% of homozygous bel larvae display a curious reversal of the OKN upon visual stimulation. Monocular stimulation leads to primary activation of ipsilateral eye movements in larvae that behave like the wild type. In contrast, affected larvae display contralateral activation of eye movements upon monocular stimulation. Anatomical analysis of retinal ganglion cell axon projections reveal a morphological basis for the observed behavioral defect. All animals with OKN reversal are achiasmatic. Further behavioral examination of affected larvae show that OKN-reversed animals execute this behavior in a stimulus-velocity-independent manner. Our data support a parsimonious model of optokinetic reversal by the opening of a controlling feedback loop at the level of the optic chiasm that is solely responsible for the observed behavioral abnormality in mutant belladonna larvae.

  17. Development of the lateral line canal system through a bone remodeling process in zebrafish.

    Science.gov (United States)

    Wada, Hironori; Iwasaki, Miki; Kawakami, Koichi

    2014-08-01

    The lateral line system of teleost fish is composed of mechanosensory receptors (neuromasts), comprising superficial receptors and others embedded in canals running under the skin. Canal diameter and size of the canal neuromasts are correlated with increasing body size, thus providing a very simple system to investigate mechanisms underlying the coordination between organ growth and body size. Here, we examine the development of the trunk lateral line canal system in zebrafish. We demonstrated that trunk canals originate from scales through a bone remodeling process, which we suggest is essential for the normal growth of canals and canal neuromasts. Moreover, we found that lateral line cells are required for the formation of canals, suggesting the existence of mutual interactions between the sensory system and surrounding connective tissues.

  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. Embryonic Development: Chicken and Zebrafish

    Directory of Open Access Journals (Sweden)

    Veerle M. Darras

    2011-01-01

    Full Text Available Chicken and zebrafish are two model species regularly used to study the role of thyroid hormones in vertebrate development. Similar to mammals, chickens have one thyroid hormone receptor α (TRα and one TRβ gene, giving rise to three TR isoforms: TRα, TRβ2, and TRβ0, the latter with a very short amino-terminal domain. Zebrafish also have one TRβ gene, providing two TRβ1 variants. The zebrafish TRα gene has been duplicated, and at least three TRα isoforms are expressed: TRαA1-2 and TRαB are very similar, while TRαA1 has a longer carboxy-terminal ligand-binding domain. All these TR isoforms appear to be functional, ligand-binding receptors. As in other vertebrates, the different chicken and zebrafish TR isoforms have a divergent spatiotemporal expression pattern, suggesting that they also have distinct functions. Several isoforms are expressed from the very first stages of embryonic development and early chicken and zebrafish embryos respond to thyroid hormone treatment with changes in gene expression. Future studies in knockdown and mutant animals should allow us to link the different TR isoforms to specific processes in embryonic development.

  20. Retro-orbital blood acquisition facilitates circulating microRNA measurement in zebrafish with paracetamol hepatotoxicity.

    Science.gov (United States)

    Vliegenthart, Adriaan D B; Starkey Lewis, Philip; Tucker, Carl S; Del Pozo, Jorge; Rider, Sebastein; Antoine, Daniel J; Dubost, Valérie; Westphal, Magdalena; Moulin, Pierre; Bailey, Matthew A; Moggs, Jonathan G; Goldring, Chris E; Park, B Kevin; Dear, James W

    2014-06-01

    Paracetamol is the commonest cause of acute liver failure in the Western world and biomarkers are needed that report early hepatotoxicity. The liver-enriched microRNA (miRNA), miR-122, is a promising biomarker currently being qualified in humans. For biomarker development and drug toxicity screening, the zebrafish has advantages over rodents; however, blood acquisition in this model remains technically challenging. We developed a method for collecting blood from the adult zebrafish by retro-orbital (RO) bleeding and compared it to the commonly used lateral incision method. The RO technique was more reliable in terms of the blood yield and minimum amount per fish. This new RO technique was used in a zebrafish model of paracetamol toxicity. Paracetamol induced dose-dependent increases in liver cell necrosis, serum alanine transaminase activity, and mortality. In situ hybridization localized expression of miR-122 to the cytoplasm of zebrafish hepatocytes. After collection by RO bleeding, serum miR-122 could be measured and this miRNA was substantially increased by paracetamol 24 h after exposure, an increase that was prevented by delayed (3 h poststart of paracetamol exposure) treatment with acetylcysteine. In summary, collection of blood by RO bleeding facilitated measurement of miR-122 in a zebrafish model of paracetamol hepatotoxicity. The zebrafish represents a new species for measurement of circulating miRNA biomarkers that are translational and can bridge between fish and humans.

  1. Genome-wide gene expression profiling of acute metal exposures in male zebrafish

    Directory of Open Access Journals (Sweden)

    Christine E. Baer

    2014-12-01

    Full Text Available To capture global responses to metal poisoning and mechanistic insights into metal toxicity, gene expression changes were evaluated in whole adult male zebrafish following acute 24 h high dose exposure to three metals with known human health risks. Male adult zebrafish were exposed to nickel chloride, cobalt chloride or sodium dichromate at concentrations corresponding to their respective 96 h LC20, LC40 and LC60 (i.e. 96 h concentrations at which 20%, 40% and 60% lethality is expected, respectively. Histopathology was performed on a subset of metal-exposed zebrafish to phenotypically anchor transcriptional changes associated with each metal exposure. Here we describe in detail the contents and quality controls for the gene expression and other data associated with the study published by Hussainzada and colleagues in BMC Pharmacology and Toxicology (Hussainzada et al., 2014 with the data uploaded to Gene Expression Omnibus (accession number GSE50648.

  2. Genome-wide gene expression profiling of acute metal exposures in male zebrafish

    Science.gov (United States)

    Baer, Christine E.; Ippolito, Danielle L.; Hussainzada, Naissan; Lewis, John A.; Jackson, David A.; Stallings, Jonathan D.

    2014-01-01

    To capture global responses to metal poisoning and mechanistic insights into metal toxicity, gene expression changes were evaluated in whole adult male zebrafish following acute 24 h high dose exposure to three metals with known human health risks. Male adult zebrafish were exposed to nickel chloride, cobalt chloride or sodium dichromate at concentrations corresponding to their respective 96 h LC20, LC40 and LC60 (i.e. 96 h concentrations at which 20%, 40% and 60% lethality is expected, respectively). Histopathology was performed on a subset of metal-exposed zebrafish to phenotypically anchor transcriptional changes associated with each metal exposure. Here we describe in detail the contents and quality controls for the gene expression and other data associated with the study published by Hussainzada and colleagues in BMC Pharmacology and Toxicology (Hussainzada et al., 2014) with the data uploaded to Gene Expression Omnibus (accession number GSE50648). PMID:26484131

  3. Mapping of zebrafish research: a global outlook.

    Science.gov (United States)

    Kinth, Priyamvadah; Mahesh, Gopalakrishnan; Panwar, Yatish

    2013-12-01

    On the basis of analysis of 17,151 records on zebrafish identified from Zebrafish Information Network: the zebrafish model organism database and Web of Science, the research performance on this model organism has been evaluated. The earliest research work on zebrafish as reflected in the databases goes back to 1951. After a rather slow growth till the 1980s, research on zebrafish gained momentum in the 1990s. Analysis shows a rapid and consistent increase in the publication output with 226 publications in the year 1996, to 1929 publications in the year 2012. The prominent areas of zebrafish research, journals, and leading authors as reflected from the research output have been identified. USA is the most productive country with 8196 articles. The most frequently used keywords were also determined to gain insights about the research trends and some of the commonly used keywords other than zebrafish and Danio rerio are development, retina, and gene expression.

  4. Myelopoiesis during Zebrafish Early Development

    Institute of Scientific and Technical Information of China (English)

    Jin Xu; Linsen Du; Zilong Wen

    2012-01-01

    Myelopoiesis is the process of producing all types of myeloid cells including monocytes/macrophages and granulocytes.Myeloid cells are known to manifest a wide spectrum of activities such as immune surveillance and tissue remodeling.Irregularities in myeloid cell development and their function are known to associate with the onset and the progression of a variety of human disorders such as leukemia.In the past decades,extensive studies have been carried out in various model organisms to elucidate the molecular mechanisms underlying myelopoiesis with the hope that these efforts will yield knowledge translatable into therapies for related diseases.Zebrafish has recently emerged as a prominent animal model for studying myelopoiesis,especially during early embryogenesis,largely owing to its unique properties such as transparent embryonic body and external development.This review introduces the methodologies used in zebrafish research and focuses on the recent research progresses of zebrafish myelopoiesis.

  5. Multilevel assessment of ivermectin effects using different zebrafish life stages.

    Science.gov (United States)

    Oliveira, Rhaul; Grisolia, Cesar K; Monteiro, Marta S; Soares, Amadeu M V M; Domingues, Inês

    2016-09-01

    Several studies have shown high toxicity of the veterinary pharmaceutical ivermectin (a semisynthetic avermectin) for aquatic invertebrates however, few data is found for fish species. The present study evaluated the toxicity of ivermectin, to embryos, juveniles, and adults of zebrafish at different levels of biological organization including developmental, behavioural and biochemical. Toxicity tests were performed based on OECD protocols and mortality and behavioural changes were assed for all stages. Biochemical responses were assessed in adults and embryos and included cholinesterases (ChEs), catalase (CAT) (only in embryos), glutathione-S-Transferase (GST), lactate dehydrogenase (LDH) and vitellogenin (VTG) like proteins (only in embryos). Genotoxicity was evaluated in adults. Results showed a higher sensitivity of juvenile and adults of zebrafish (96h-LC10 values of 14.0 and 55.4μg/L, respectively). For embryos a 96h-LC10 of 147.1μg/L was calculated, moreover developmental anomalies and hatching inhibition were observed only at high concentrations (>400μg/L), whereas biochemical and behavioural responses occurred at lower concentrations (Ivermectin did not show to be genotoxic for adult fish. The species sensitivity distribution analysis, based on fish and invertebrate species, indicated a Hazardous Concentration for 5% of the population (HC5) value of 0.057μg/L; suggesting high sensitivity of both groups to ivermectin and a high risk of this compound to aquatic ecosystems.

  6. Glucocorticoid activity detected by in vivo zebrafish assay and in vitro glucocorticoid receptor bioassay at environmental relevant concentrations.

    Science.gov (United States)

    Chen, Qiyu; Jia, Ai; Snyder, Shane A; Gong, Zhiyuan; Lam, Siew Hong

    2016-02-01

    Glucocorticoids are pharmaceutical contaminants of emerging concern due to their incomplete removal during wastewater treatment, increased presence in aquatic environment and their biological potency. The zebrafish is a popular model for aquatic toxicology and environmental risk assessment. This study aimed to determine if glucocorticoids at environmental concentrations would perturb expression of selected glucocorticoid-responsive genes in zebrafish and to investigate their potentials as an in vivo zebrafish assay in complementing in vitro glucocorticoid receptor bioassay. The relative expression of eleven glucocorticoid-responsive genes in zebrafish larvae and liver of adult male zebrafish exposed to three representative glucocorticoids (dexamethasone, prednisolone and triamcinolone) was determined. The expression of pepck, baiap2 and pxr was up-regulated in zebrafish larvae and the expression of baiap2, pxr and mmp-2 was up-regulated in adult zebrafish exposed to glucocorticoids at concentrations equivalent to total glucocorticoids reported in environmental samples. The responsiveness of the specific genes were sufficiently robust in zebrafish larvae exposed to a complex environmental sample detected with in vitro glucocorticoid activity equivalent to 478 pM dexamethasone (DEX-EQ) and confirmed to contain low concentration (0.2 ng/L or less) of the targeted glucocorticoids, and possibly other glucocorticoid-active compounds. The findings provided in vivo relevance to the in vitro glucocorticoid activity and suggested that the environmental sample can perturb glucocorticoid-responsive genes in its original, or half the diluted, concentration as may be found in the environment. The study demonstrated the important complementary roles of in vivo zebrafish and in vitro bioassays coupled with analytical chemistry in monitoring environmental glucocorticoid contaminants.

  7. Divergence of zebrafish and mouse lymphatic cell fate specification pathways

    DEFF Research Database (Denmark)

    van Impel, Andreas; Zhao, Zhonghua; Hermkens, Dorien M A;

    2014-01-01

    . Murine Prox1-null embryos lack lymphatic structures, and sustained expression of Prox1 is indispensable for the maintenance of lymphatic cell fate even at adult stages, highlighting the unique importance of this gene for the lymphatic lineage. Whether this pre-eminent role of Prox1 within the lymphatic...... vasculature is conserved in other vertebrate classes has remained unresolved, mainly owing to the lack of availability of loss-of-function mutants. Here, we re-examine the role of Prox1a in zebrafish lymphangiogenesis. First, using a transgenic reporter line, we show that prox1a is initially expressed...... that the functionally related transcription factors Coup-TFII and Sox18 are also dispensable for lymphangiogenesis. Together, these findings suggest that lymphatic commitment in zebrafish and mice is controlled in fundamentally different ways....

  8. Characterization of the laminin gene family and evolution in zebrafish.

    Science.gov (United States)

    Sztal, Tamar; Berger, Silke; Currie, Peter D; Hall, Thomas E

    2011-02-01

    Laminins are essential components of all basement membranes and are fundamental to tissue development and homeostasis. Humans possess at least 16 different heterotrimeric laminin complexes formed through different combinations of alpha, beta, and gamma chains. Individual chains appear to exhibit unique expression patterns, leading to the notion that overlap between expression domains governs the constitution of complexes found within particular tissues. However, the spatial and temporal expression of laminin genes has not been comprehensively analyzed in any vertebrate model to date. Here, we describe the tissue-specific expression patterns of all laminin genes in the zebrafish, throughout embryonic development and into the "post-juvenile" period, which is representative of the adult body form. In addition, we present phylogenetic and microsynteny analyses, which demonstrate that the majority of our zebrafish sequences are orthologous to human laminin genes. Together, these data represent a fundamental resource for the study of vertebrate laminins.

  9. 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.

  10. The complement system in teleost fish: progress of post-homolog-hunting researches.

    Science.gov (United States)

    Nakao, Miki; Tsujikura, Masakazu; Ichiki, Satoko; Vo, Tam K; Somamoto, Tomonori

    2011-12-01

    Studies on the complement system of bony fish are now finishing a stage of homologue-hunting identification of the components, unveiling existence of almost all the orthologues of mammalian complement components in teleost. Genomic and transcriptomic data for several teleost species have contributed much for the homologue-hunting research progress. Only an exception is identification of orthologues of mammalian complement regulatory proteins and complement receptors. It is of particular interest that teleost complement components often exist as multiple isoforms with possible functional divergence. This review summarizes research progress of teleost complement system following the molecular identification and sequence analysis of the components. The findings of extensive expression analyses of the complement components with special emphasis of their prominent extrahepatic expression, acute-phase response to immunostimulation and various microbial infections, and ontogenic development including maternal transfer are discussed to infer teleost-specific functions of the complement system. Importance of the protein level characterization of the complement components is also emphasized, especially for understanding of the isotypic diversity of the components, a unique feature of teleost complement system.

  11. Comparative Evolution of Duplicated Ddx3 Genes in Teleosts: Insights from Japanese Flounder, Paralichthys olivaceus.

    Science.gov (United States)

    Wang, Zhongkai; Liu, Wei; Song, Huayu; Wang, Huizhen; Liu, Jinxiang; Zhao, Haitao; Du, Xinxin; Zhang, Quanqi

    2015-06-24

    Following the two rounds of whole-genome duplication that occurred during deuterostome evolution, a third genome duplication event occurred in the stem lineage of ray-finned fishes. This teleost-specific genome duplication is thought to be responsible for the biological diversification of ray-finned fishes. DEAD-box polypeptide 3 (DDX3) belongs to the DEAD-box RNA helicase family. Although their functions in humans have been well studied, limited information is available regarding their function in teleosts. In this study, two teleost Ddx3 genes were first identified in the transcriptome of Japanese flounder (Paralichthys olivaceus). We confirmed that the two genes originated from teleost-specific genome duplication through synteny and phylogenetic analysis. Additionally, comparative analysis of genome structure, molecular evolution rate, and expression pattern of the two genes in Japanese flounder revealed evidence of subfunctionalization of the duplicated Ddx3 genes in teleosts. Thus, the results of this study reveal novel insights into the evolution of the teleost Ddx3 genes and constitute important groundwork for further research on this gene family.

  12. Novel Insights into the Genetic Controls of Primitive and Definitive Hematopoiesis from Zebrafish Models

    Directory of Open Access Journals (Sweden)

    Raman Sood

    2012-01-01

    Full Text Available Hematopoiesis is a dynamic process where initiation and maintenance of hematopoietic stem cells, as well as their differentiation into erythroid, myeloid and lymphoid lineages, are tightly regulated by a network of transcription factors. Understanding the genetic controls of hematopoiesis is crucial as perturbations in hematopoiesis lead to diseases such as anemia, thrombocytopenia, or cancers, including leukemias and lymphomas. Animal models, particularly conventional and conditional knockout mice, have played major roles in our understanding of the genetic controls of hematopoiesis. However, knockout mice for most of the hematopoietic transcription factors are embryonic lethal, thus precluding the analysis of their roles during the transition from embryonic to adult hematopoiesis. Zebrafish are an ideal model organism to determine the function of a gene during embryonic-to-adult transition of hematopoiesis since bloodless zebrafish embryos can develop normally into early larval stage by obtaining oxygen through diffusion. In this review, we discuss the current status of the ontogeny and regulation of hematopoiesis in zebrafish. By providing specific examples of zebrafish morphants and mutants, we have highlighted the contributions of the zebrafish model to our overall understanding of the roles of transcription factors in regulation of primitive and definitive hematopoiesis.

  13. Endocannabinoids affect the reproductive functions in teleosts and amphibians.

    Science.gov (United States)

    Cottone, E; Guastalla, A; Mackie, K; Franzoni, M F

    2008-04-16

    Following the discovery in the brain of the bonyfish Fugu rubripes of two genes encoding for type 1 cannabinoid receptors (CB1A and CB1B), investigations on the phylogeny of these receptors have indicated that the cannabinergic system is highly conserved. Among the multiple functions modulated by cannabinoids/endocannabinoids through the CB1 receptors one of the more investigated is the mammalian reproduction. Therefore, since studies performed in animal models other than mammals might provide further insight into the biology of these signalling molecules, the major aim of the present paper was to review the comparative data pointing toward the endocannabinoid involvement in the reproductive control of non-mammalian vertebrates, namely bonyfish and amphibians. The expression and distribution of CB1 receptors were investigated in the CNS and gonads of two teleosts, Pelvicachromis pulcher and Carassius auratus as well as in the anuran amphibians Xenopus laevis and Rana esculenta. In general the large diffusion of neurons targeted by cannabinoids in both fish and amphibian forebrain indicate endocannabinoids as pivotal local messengers in several neural circuits involved in either sensory integrative activities, like the olfactory processes (in amphibians) and food response (in bonyfish), or neuroendocrine machinery (in both). By using immunohistochemistry for CB1 and GnRH-I, the codistribution of the two signalling molecules was found in the fish basal telencephalon and preoptic area, which are key centers for gonadotropic regulation in all vertebrates. A similar topographical codistribution was observed also in the septum of the telencephalon in Rana esculenta and Xenopus laevis. Interestingly, the double standard immunofluorescence on the same brain section, aided with a laser confocal microscope, showed that in anurans a subset of GnRH-I neurons exhibited also the CB1 immunostaining. The fact that CB1-LI-IR was found indeed in the FSH gonadotrophs of the Xenopus

  14. 利用斑马鱼成鱼建立致幻类化合物行为评价模型%Adult zebrafish as a model organism for assessing the effects of hallucinogenic drugs on behaviors

    Institute of Scientific and Technical Information of China (English)

    颜慧; 苏瑞斌; 宫泽辉

    2014-01-01

    Aims To establish several behavioral paradigms to characterize the psychotropic effects of hallucinogens which ze-brafish was utilized as a model animal, and then to investigate the effects of potent hallucinogenic drugs on these models. Methods With the video record and track system, the behavior was recorded and quantified automatically. In the experiments, the bottom dwelling test, social behavior and mirror test were performed to test the hallucinogenic effects of drugs. Metham-phetamine (METH, 2 mg·L-1) and ketamine (20 mg·L-1) were selected as experimental challenges. The 30 min pre-treat-ment time was chosen based on our prior experience in zebrafish models. Results Compared to the normal group, in dwelling test, acute exposure of zebrafish to METH and ketamine de-creased transitions significantly, and in mirror reflection test, the drug-treated fish changed the preference for mirror zone, and ex-hibited aggressive for their mirror images. The pretreatment of METH and ketamine significantly reduced the contact durations, and the ketamine inhibited the contact frequency each other, the results indicated that the social interaction of zebrafish was im-paired. Conclusion The results confirm high sensitivity of ze-brafish models to hallucinogenic compounds with complex behav-ioral and physiological effects.%目的:利用斑马鱼这一新型模式动物,建立致幻类化合物成鱼行为测试模型,并考察模型有效性。方法利用斑马鱼行为视频跟踪分析系统自动记录动物行为参数,以栖底性、镜像反射及社会交互行为作为评测指标,采用甲基苯丙胺浓度2 mg·L-1、氯胺酮浓度20 mg·L-1,分别急性暴露30 min后进行行为测试。结果与正常组相比,在成鱼自发活动方面,甲基苯丙胺与氯胺酮急性暴露对斑马鱼的运动距离无明显影响,而使垂直方向的穿越次数明显下降;在镜像反射方面,给予不同化合物均使斑马鱼在中央区的停留时间明显增

  15. Toxicity of tetrabromobisphenol A (TBBPA) in zebrafish (Danio rerio) in a partial life-cycle test

    NARCIS (Netherlands)

    Kuiper, R.V.; Brandhof, Van den E.J.; Leonards, P.E.G.; Ven, van der L.T.M.; Wester, P.W.; Vos, J.G.

    2007-01-01

    Toxicological effects of the widely used flame retardant, tetrabromobisphenol A (TBBPA) were assessed in a partial life-cycle test with zebrafish (Danio rerio). Exposure of adult fish during 30 days to water-borne TBBPA in nominal concentrations ranging from 0 (control) to 1.5 ¿M was followed by exp

  16. Toxicity of tetrabromobisphenol A (TBBPA) in zebrafish (Danio rerio) in a partial life-cycle test.

    NARCIS (Netherlands)

    Kuiper, R V; Brandhof, E J van den; Leonards, P E G; Ven, L T M van der; Wester, P W; Vos, J G

    2006-01-01

    Toxicological effects of the widely used flame retardant, tetrabromobisphenol A (TBBPA) were assessed in a partial life-cycle test with zebrafish (Danio rerio). Exposure of adult fish during 30 days to water-borne TBBPA in nominal concentrations ranging from 0 (control) to 1.5 muM was followed by ex

  17. Carbon Quantum Dots for Zebrafish Fluorescence Imaging

    Science.gov (United States)

    Kang, Yan-Fei; Li, Yu-Hao; Fang, Yang-Wu; Xu, Yang; Wei, Xiao-Mi; Yin, Xue-Bo

    2015-07-01

    Carbon quantum dots (C-QDs) are becoming a desirable alternative to metal-based QDs and dye probes owing to their high biocompatibility, low toxicity, ease of preparation, and unique photophysical properties. Herein, we describe fluorescence bioimaging of zebrafish using C-QDs as probe in terms of the preparation of C-QDs, zebrafish husbandry, embryo harvesting, and introduction of C-QDs into embryos and larvae by soaking and microinjection. The multicolor of C-QDs was validated with their imaging for zebrafish embryo. The distribution of C-QDs in zebrafish embryos and larvae were successfully observed from their fluorescence emission. the bio-toxicity of C-QDs was tested with zebrafish as model and C-QDs do not interfere to the development of zebrafish embryo. All of the results confirmed the high biocompatibility and low toxicity of C-QDs as imaging probe. The absorption, distribution, metabolism and excretion route (ADME) of C-QDs in zebrafish was revealed by their distribution. Our work provides the useful information for the researchers interested in studying with zebrafish as a model and the applications of C-QDs. The operations related zebrafish are suitable for the study of the toxicity, adverse effects, transport, and biocompatibility of nanomaterials as well as for drug screening with zebrafish as model.

  18. Regulation of zebrafish CYP3A65 transcription by AHR2

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Chin-Teng; Chung, Hsin-Yu; Su, Hsiao-Ting; Tseng, Hua-Pin [Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China); Tzou, Wen-Shyong [Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China); Center of Excellence for Marine Bioenvironment and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China); Hu, Chin-Hwa, E-mail: chhu@mail.ntou.edu.tw [Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China); Center of Excellence for Marine Bioenvironment and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan (China)

    2013-07-15

    CYP3A proteins are the most abundant CYPs in the liver and intestines, and they play a pivotal role in drug metabolism. In mammals, CYP3A genes are induced by various xenobiotics through processes mediated by PXR. We previously identified zebrafish CYP3A65 as a CYP3A ortholog that is constitutively expressed in gastrointestinal tissues, and is upregulated by treatment with dexamethasone, rifampicin or tetrachlorodibenzo-p-dioxin (TCDD). However, the underlying mechanism of TCDD-mediated CYP3A65 transcription is unclear. Here we generated two transgenic zebrafish, Tg(CYP3A65S:EGFP) and Tg(CYP3A65L:EGFP), which contain 2.1 and 5.4 kb 5′ flanking sequences, respectively, of the CYP3A65 gene upstream of EGFP. Both transgenic lines express EGFP in larval gastrointestinal tissues in a pattern similar to that of the endogenous CYP3A65 gene. Moreover, EGFP expression can be significantly induced by TCDD exposure during the larval stage. In addition, EGFP expression can be stimulated by kynurenine, a putative AHR ligand produced during tryptophan metabolism. AHRE elements in the upstream regulatory region of the CYP3A65 gene are indispensible for basal and TCDD-induced transcription. Furthermore, the AHR2 DNA and ligand-binding domains are required to mediate effective CYP3A65 transcription. AHRE sequences are present in the promoters of many teleost CYP3 genes, but not of mammalian CYP3 genes, suggesting that AHR/AHR2-mediated transcription is likely a common regulatory mechanism for teleost CYP3 genes. It may also reflect the different environments that terrestrial and aquatic organisms encounter. - Highlights: • Tg(CYP3A65:EGFP) and CYP3A65 exhibits identical expression pattern. • CYP3A65 can be significantly induced by TCDD or kynurenine. • The AHRE elements are required to mediate CYP3A65 transcription. • The AHR2 DNA and ligand-binding domains are required for CYP3A65 transcription. • AHRE elements are present in many teleost CYP3 genes, but not in

  19. Stimulatory effect of intracerebroventricular administration of orexin A on food intake in the zebrafish, Danio rerio.

    Science.gov (United States)

    Yokobori, Eri; Kojima, Kenji; Azuma, Morio; Kang, Ki Sung; Maejima, Sho; Uchiyama, Minoru; Matsuda, Kouhei

    2011-07-01

    Orexin is a potent orexigenic neuropeptide implicated in feeding regulation of mammals. However, except for the case of goldfish, the involvement of orexin in the feeding behavior of teleost fish has not well been studied. Therefore, we investigated the role of orexin on food intake using a zebrafish (Danio rerio) model. We examined the effect of feeding status on orexin-like immunoreactivity and the expression level of orexin transcript in the brain. The number of neuronal cells showing orexin-like immunoreactivity in the hypothalamic region, including the posterior tuberal nucleus, was significantly increased in fish fasted for 7days. Orexin precursor mRNA levels in the brain obtained from fish fasted for 7 days were higher than those in fish that had been fed normally. We then investigated the effect of intracerebroventricular (ICV) administration of orexin A on food intake. Cumulative food intake was significantly increased by ICV administration of orexin A (at 0.3 and 3 pmol/g body weight, BW) during a 60-min observation period after treatment. The orexin A-induced orexigenic action (at 0.3 pmol/g BW) was blocked by treatment with an orexin receptor antagonist, SB334867, at 10 pmol/g BW. These results indicate that orexin A acts as feeding regulator in the zebrafish.

  20. Zebrafish and medaka: model organisms for a comparative developmental approach of brain asymmetry.

    Science.gov (United States)

    Signore, Iskra A; Guerrero, Néstor; Loosli, Felix; Colombo, Alicia; Villalón, Aldo; Wittbrodt, Joachim; Concha, Miguel L

    2009-04-12

    Comparison between related species is a successful approach to uncover conserved and divergent principles of development. Here, we studied the pattern of epithalamic asymmetry in zebrafish (Danio rerio) and medaka (Oryzias latipes), two related teleost species with 115-200 Myr of independent evolution. We found that these species share a strikingly conserved overall pattern of asymmetry in the parapineal-habenular-interpeduncular system. Nodal signalling exhibits comparable spatial and temporal asymmetric expressions in the presumptive epithalamus preceding the development of morphological asymmetries. Neuroanatomical asymmetries consist of left-sided asymmetric positioning and connectivity of the parapineal organ, enlargement of neuropil in the left habenula compared with the right habenula and segregation of left-right habenular efferents along the dorsoventral axis of the interpeduncular nucleus. Despite the overall conservation of asymmetry, we observed heterotopic changes in the topology of parapineal efferent connectivity, heterochronic shifts in the timing of developmental events underlying the establishment of asymmetry and divergent degrees of canalization of embryo laterality. We offer new tools for developmental time comparison among species and propose, for each of these transformations, novel hypotheses of ontogenic mechanisms that explain interspecies variations that can be tested experimentally. Together, these findings highlight the usefulness of zebrafish and medaka as comparative models to study the developmental mechanisms of epithalamic asymmetry in vertebrates.

  1. Occurrence and origin of sensitivity toward difenoconazole in zebrafish (Danio reio) during different life stages.

    Science.gov (United States)

    Mu, Xiyan; Chai, Tingting; Wang, Kai; Zhang, Jie; Zhu, Lizhen; Li, Xuefeng; Wang, Chengju

    2015-03-01

    We report here an investigation of the mechanisms contributing to the divergent sensitivity toward the triazole fungicide difenoconazole of zebrafish (Danio reio) during different life stages. Adult and embryonic zebrafish were exposed to three different concentrations of difenoconazole (0.01, 0.5 and 1.0mg/L). The death rate, bioaccumulation of difenoconazole, oxidative stress parameters and transcription of related genes were tested at 4 and 8 days post-exposure (dpe). The death rate for adult zebrafish was much higher than that of the embryos at an exposure concentration of 1.0mg/L at both 4 and 8 dpe. The concentrations of difenoconazole in both the embryos and adult fish were similar, except for the group exposed to 0.01mg/L difenoconazole. A decrease in antioxidant enzyme activities was observed in both the embryos and the livers of adult fish after exposure to difenoconazole. Significant lipid peroxidation was found in the livers of adult fish in all exposure groups at 8 dpe, but was not observed in the treated embryos. The gene transcription response of the embryos toward difenoconazole was different from that in the livers of adult fish at 4 dpe. At 8 dpe, the modification in the transcription of the tested genes in the embryos and adult fish was similar, except for the genes related to the synthesis of sterols.

  2. Social Plasticity Relies on Different Neuroplasticity Mechanisms across the Brain Social Decision-Making Network in Zebrafish

    Science.gov (United States)

    Teles, Magda C.; Cardoso, Sara D.; Oliveira, Rui F.

    2016-01-01

    Social living animals need to adjust the expression of their behavior to their status within the group and to changes in social context and this ability (social plasticity) has an impact on their Darwinian fitness. At the proximate level social plasticity must rely on neuroplasticity in the brain social decision-making network (SDMN) that underlies the expression of social behavior, such that the same neural circuit may underlie the expression of different behaviors depending on social context. Here we tested this hypothesis in zebrafish by characterizing the gene expression response in the SDMN to changes in social status of a set of genes involved in different types of neural plasticity: bdnf, involved in changes in synaptic strength; npas4, involved in contextual learning and dependent establishment of GABAergic synapses; neuroligins (nlgn1 and nlgn2) as synaptogenesis markers; and genes involved in adult neurogenesis (wnt3 and neurod). Four social phenotypes were experimentally induced: Winners and Losers of a real-opponent interaction; Mirror-fighters, that fight their own image in a mirror and thus do not experience a change in social status despite the expression of aggressive behavior; and non-interacting fish, which were used as a reference group. Our results show that each social phenotype (i.e., Winners, Losers, and Mirror-fighters) present specific patterns of gene expression across the SDMN, and that different neuroplasticity genes are differentially expressed in different nodes of the network (e.g., BDNF in the dorsolateral telencephalon, which is a putative teleost homolog of the mammalian hippocampus). Winners expressed unique patterns of gene co-expression across the SDMN, whereas in Losers and Mirror-fighters the co-expression patterns were similar in the dorsal regions of the telencephalon and in the supracommissural nucleus of the ventral telencephalic area, but differents in the remaining regions of the ventral telencephalon. These results

  3. Characterization of glutathione-S-transferases in zebrafish (Danio rerio).

    Science.gov (United States)

    Glisic, Branka; Mihaljevic, Ivan; Popovic, Marta; Zaja, Roko; Loncar, Jovica; Fent, Karl; Kovacevic, Radmila; Smital, Tvrtko

    2015-01-01

    Glutathione-S-transferases (GSTs) are one of the key enzymes that mediate phase II of cellular detoxification. The aim of our study was a comprehensive characterization of GSTs in zebrafish (Danio rerio) as an important vertebrate model species frequently used in environmental research. A detailed phylogenetic analysis of GST superfamily revealed 27 zebrafish gst genes. Further insights into the orthology relationships between human and zebrafish GSTs/Gsts were obtained by the conserved synteny analysis. Expression of gst genes in six tissues (liver, kidney, gills, intestine, brain and gonads) of adult male and female zebrafish was determined using qRT-PCR. Functional characterization was performed on 9 cytosolic Gst enzymes after overexpression in E. coli and subsequent protein purification. Enzyme kinetics was measured for GSH and a series of model substrates. Our data revealed ubiquitously high expression of gstp, gstm (except in liver), gstr1, mgst3a and mgst3b, high expression of gsto2 in gills and ovaries, gsta in intestine and testes, gstt1a in liver, and gstz1 in liver, kidney and brain. All zebrafish Gsts catalyzed the conjugation of GSH to model GST substrates 1-chloro-2,4-dinitrobenzene (CDNB) and monochlorobimane (MCB), apart from Gsto2 and Gstz1 that catalyzed GSH conjugation to dehydroascorbate (DHA) and dichloroacetic acid (DCA), respectively. Affinity toward CDNB varied from 0.28 mM (Gstp2) to 3.69 mM (Gstm3), while affinity toward MCB was in the range of 5 μM (Gstt1a) to 250 μM (Gstp1). Affinity toward GSH varied from 0.27 mM (Gstz1) to 4.45 mM (Gstt1a). Turnover number for CDNB varied from 5.25s(-1) (Gstt1a) to 112s(-1) (Gstp2). Only Gst Pi enzymes utilized ethacrynic acid (ETA). We suggest that Gstp1, Gstp2, Gstt1a, Gstz1, Gstr1, Mgst3a and Mgst3b have important role in the biotransformation of xenobiotics, while Gst Alpha, Mu, Pi, Zeta and Rho classes are involved in the crucial physiological processes. In summary, this study provides the

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

    DEFF Research Database (Denmark)

    Korbut, Rozalia; Mehrdana, Foojan; Kania, Per Walter;

    2016-01-01

    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...... 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....... 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...

  5. Analysis of nicastrin gene phylogeny and expression in zebrafish.

    Science.gov (United States)

    Lim, Anne; Moussavi Nik, Seyyed Hani; Ebrahimie, Esmaeil; Lardelli, Michael

    2015-06-01

    NICASTRIN is a component of the aspartyl protease γ-secretase complex which is involved in intramembranous cleavage of type I transmembrane proteins, notably the Notch receptor proteins and the AMYLOID BETA A4 PRECURSOR PROTEIN (APP). This study aimed to characterize the orthologue of the human NICASTRIN (NCSTN) gene in zebrafish, an advantageous model organism for the study of human disease. Zebrafish Nicastrin protein was predicted to possess the conserved glutamate 333 residue and DYIGS motif of human NCSTN that are important for substrate recognition/processing in γ-secretase. Quantitative real-time RT-PCR revealed the profile of relative zebrafish nicastrin (ncstn) transcript levels in embryos at different times during development and in adult tissues. The analysis of synteny conservation revealed local rearrangements of ncstn and another gene, mpz, relative to copa, and pex19. In situ hybridization showed higher relative levels of ncstn transcripts in the developing brain and otic vesicles of embryos at 24 and 48 h post fertilization, respectively. Our observations are consistent with a role for Ncstn protein in Notch signaling within the proliferative ventricular zone of the developing central nervous system.

  6. Histological Characterization of the Dicer1 Mutant Zebrafish Retina

    Directory of Open Access Journals (Sweden)

    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.

  7. Zebrafish Social Behavior in the Wild.

    Science.gov (United States)

    Suriyampola, Piyumika S; Shelton, Delia S; Shukla, Rohitashva; Roy, Tamal; Bhat, Anuradha; Martins, Emília P

    2016-02-01

    Wild zebrafish exhibit a wide range of behavior. We found abundant wild zebrafish in flowing rivers and still water, in large, tightly-knit groups of hundreds of individuals, as well as in small, loose shoals. In two still-water populations, zebrafish were quite small in body size, common, and in tight groups of up to 22 fish. As in earlier laboratory studies, these zebrafish exhibited very low levels of aggression. In slowly flowing water in central India, zebrafish were relatively rare and gathered in small shoals (4-12 fish), often with other small fish, such as Rasbora daniconius. These stream zebrafish were larger in body size (27 mm TL) and much more aggressive than those in still water. In a second river population with much faster flowing water, zebrafish were abundant and again relatively large (21 mm TL). These zebrafish occurred in very large (up to 300 individuals) and tightly-knit (nearest-neighbor distances up to 21 mm) groups that exhibited collective rheotaxis and almost no aggression. This remarkable variation in social behavior of wild zebrafish offers an opportunity for future studies of behavioral genetics, development, and neuroscience.

  8. Bioconcentration of pesticides in zebrafish eleutheroembryos (Danio rerio).

    Science.gov (United States)

    El-Amrani, S; Pena-Abaurrea, M; Sanz-Landaluze, J; Ramos, L; Guinea, J; Cámara, C

    2012-05-15

    The feasibility of a bioaccumulation test based on the use of zebrafish eleutheroembryos as an alternative to adult-individual-based approaches for REACH application has been evaluated for three test compounds, chlorpyrifos, dicofol and atrazine. Following the OECD 305 guidelines, zebrafish eleutheroembryos (72 h after hatching, hpf) were separately exposed to the investigated pesticides at two nominal concentrations below 1% of its corresponding LC(50). The uptake experiments lasted for 48 h. Then, the exposure medium was replaced by a non-contaminated medium for depuration experiments (up to 72 h). Zebrafish eleutheroembryos (larvae 144 hpf, i.e. at the end of the depuration step) and their corresponding exposure media was sampled at ten different times during each experiment and the concentration of the investigated pesticide determined in both the organisms and in the exposure medium. The experimentally determined pesticide accumulation profiles in the eleutheroembryos demonstrated that atrazine has a very fast accumulation kinetic, reaching steady sate (SS) within 24h. Chlorpyrifos and dicofol did not reach the SS within the 48-h uptake experiments although they exhibit higher accumulations than the former pesticide. Two toxicokinetic models were used to calculate the bioconcentration factor (BCF) of the studied pesticide in zebrafish eleutheroembryos. In the former, the BCF was calculated under SS conditions (BCF(SS)). The second was used when the compounds did not reach the SS during the uptake experiment (BCF(k)). Log BCF values of 3.55 and 3.84 for chlorpyrifos; 0.6 and 1.17 for atrazine, and 3.90 for dicofol were experimentally calculated at selected exposure concentrations. These values have been compared with those reported in related bioaccumulation studies and official databases.

  9. Forkhead transcription factor foxe1 regulates chondrogenesis in zebrafish.

    Science.gov (United States)

    Nakada, Chisako; Iida, Atsumi; Tabata, Yoko; Watanabe, Sumiko

    2009-12-15

    Forkhead transcription factor (Fox) e1 is a causative gene for Bamforth-Lazarus syndrome, which is characterized by hypothyroidism and cleft palate. Applying degenerate polymerase chain reaction using primers specific for the conserved forkhead domain, we identified zebrafish foxe1 (foxe1). Foxe1 is expressed in the thyroid, pharynx, and pharyngeal skeleton during development; strongly expressed in the gill and weakly expressed in the brain, eye, and heart in adult zebrafish. A loss of function of foxe1 by morpholino antisense oligo (MO) exhibited abnormal craniofacial development, shortening of Meckel's cartilage and the ceratohyals, and suppressed chondrycytic proliferation. However, at 27 hr post fertilization, the foxe1 MO-injected embryos showed normal dlx2, hoxa2, and hoxb2 expression, suggesting that the initial steps of pharyngeal skeletal development, including neural crest migration and specification of the pharyngeal arch occurred normally. In contrast, at 2 dpf, a severe reduction in the expression of sox9a, colIIaI, and runx2b, which play roles in chondrocytic proliferation and differentiation, was observed. Interestingly, fgfr2 was strongly upregulated in the branchial arches of the foxe1 MO-injected embryos. Unlike Foxe1-null mice, normal thyroid development in terms of morphology and thyroid-specific marker expression was observed in foxe1 MO-injected zebrafish embryos. Taken together, our results indicate that Foxe1 plays an important role in chondrogenesis during development of the pharyngeal skeleton in zebrafish, probably through regulation of fgfr2 expression. Furthermore, the roles reported for FOXE1 in mammalian thyroid development may have been acquired during evolution.

  10. Macondo crude oil from the Deepwater Horizon oil spill disrupts specific developmental processes during zebrafish embryogenesis

    Directory of Open Access Journals (Sweden)

    de Soysa T Yvanka

    2012-05-01

    Full Text Available Abstract Background The Deepwater Horizon disaster was the largest marine oil spill in history, and total vertical exposure of oil to the water column suggests it could impact an enormous diversity of ecosystems. The most vulnerable organisms are those encountering these pollutants during their early life stages. Water-soluble components of crude oil and specific polycyclic aromatic hydrocarbons have been shown to cause defects in cardiovascular and craniofacial development in a variety of teleost species, but the developmental origins of these defects have yet to be determined. We have adopted zebrafish, Danio rerio, as a model to test whether water accumulated fractions (WAF of the Deepwater Horizon oil could impact specific embryonic developmental processes. While not a native species to the Gulf waters, the developmental biology of zebrafish has been well characterized and makes it a powerful model system to reveal the cellular and molecular mechanisms behind Macondo crude toxicity. Results WAF of Macondo crude oil sampled during the oil spill was used to treat zebrafish throughout embryonic and larval development. Our results indicate that the Macondo crude oil causes a variety of significant defects in zebrafish embryogenesis, but these defects have specific developmental origins. WAF treatments caused defects in craniofacial development and circulatory function similar to previous reports, but we extend these results to show they are likely derived from an earlier defect in neural crest cell development. Moreover, we demonstrate that exposure to WAFs causes a variety of novel deformations in specific developmental processes, including programmed cell death, locomotor behavior, sensory and motor axon pathfinding, somitogenesis and muscle patterning. Interestingly, the severity of cell death and muscle phenotypes decreased over several months of repeated analysis, which was correlated with a rapid drop-off in the aromatic and alkane

  11. Evidence for a divergence in function between two glucocorticoid receptors from a basal teleost

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    Li Yi

    2012-08-01

    Full Text Available Abstract Background Duplicated glucocorticoid receptors (GR are present in most teleost fish. The evolutionary advantage of retaining two GRs is unclear, as no subtype specific functional traits or physiological roles have been defined. To identify factors driving the retention of duplicate GRs in teleosts, the current study examined GRs in representatives of two basal ray-finned fish taxa that emerged either side of the teleost lineage whole genome duplication event (WGD event, the acipenseriform, Acipenser ruthenus, (pre-WGD and the osteoglossimorph, Pantodon buchholzi, (post-WGD. Results The study identified a single GR in A. ruthenus (ArGR and two GRs in P. buchholzi (PbGR1 and PbGR2. Phylogenetic analyses showed that ArGR formed a distinct branch separate from the teleosts GRs. The teleost GR lineage was subdivded into two sublineages, each of which contained one of the two P. buchholzi GRs. ArGR, PbGR1 and PbGR2 all possess the unique 9 amino acid insert between the zinc-fingers of the DNA-binding domain that is present in one of the teleost GR lineages (GR1, but not the other (GR2. A splice variant of PbGR2 produces an isoform that lacked these 9 amino acids (PbGR2b. Cortisol stimulated transactivation activity of ArGR, PbGR2b and PbGR1 in vitro; with PbGR2b and PbGR1, the glucocorticoid 11-deoxycortisol was a more potent agonist than cortisol. The hormone sensitivity of PbGR2b and PbGR1 differed in the transactivation assay, with PbGR2b having lower EC50 values and greater fold induction. Conclusions The difference in transactivation activity sensitivity between duplicated GRs of P. buchholzi suggests potential functional differences between the paralogs emerged early in the teleost lineage. Given the pleiotropic nature of GR function in vertebrates, this finding is in accordance with the hypothesis that duplicated GRs were potentially retained through subfunctionalisation followed by gene sharing. A 9 amino acid insert in the DNA

  12. Fibroblast growth factor (Fgf) signaling pathway regulates liver homeostasis in zebrafish.

    Science.gov (United States)

    Tsai, Su-Mei; Liu, Da-Wei; Wang, Wen-Pin

    2013-04-01

    In mammals, fibroblast growth factor (FGF) signaling controls liver specification and regulates the metabolism of lipids, cholesterol, and bile acids. FGF signaling also promotes hepatocyte proliferation, and helps detoxify hepatotoxin during liver regeneration after partial hepatectomy. However, the function of Fgf in zebrafish liver is not yet well understood, specifically for postnatal homeostasis. The current study analyzed the expression of fgf receptors (fgfrs) in the liver of zebrafish. We then investigated the function of Fgf signaling in the zebrafish liver by expressing a dominant-negative Fgf receptor in hepatocytes (lfabp:dnfgfr1-egfp, lf:dnfr). Histological analysis showed that our genetic intervention resulted in a small liver size with defected medial expansion of developing livers in transgenic (Tg) larvae. Morphologically, the liver lobe of lf:dnfr adult fish was shorter than that of control. Ballooning degeneration of hepatocytes was observed in fish as young as 3 months. Further examination revealed the development of hepatic steatosis and cholestasis. In adult Tg fish, we unexpectedly observed increased liver-to-body-weight ratios, with higher percentages of proliferating hepatocytes. Considering all these findings, we concluded that as in mammals, in adult zebrafish the metabolism of lipid and bile acids in the liver are regulated by Fgf signaling. Disruption of the Fgf signal-mediated metabolism might indirectly affect hepatocyte proliferation.

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

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    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. The spotted gar genome illuminates vertebrate evolution and facilitates human-teleost comparisons.

    Science.gov (United States)

    Braasch, Ingo; Gehrke, Andrew R; Smith, Jeramiah J; Kawasaki, Kazuhiko; Manousaki, Tereza; Pasquier, Jeremy; Amores, Angel; Desvignes, Thomas; Batzel, Peter; Catchen, Julian; Berlin, Aaron M; Campbell, Michael S; Barrell, Daniel; Martin, Kyle J; Mulley, John F; Ravi, Vydianathan; Lee, Alison P; Nakamura, Tetsuya; Chalopin, Domitille; Fan, Shaohua; Wcisel, Dustin; Cañestro, Cristian; Sydes, Jason; Beaudry, Felix E G; Sun, Yi; Hertel, Jana; Beam, Michael J; Fasold, Mario; Ishiyama, Mikio; Johnson, Jeremy; Kehr, Steffi; Lara, Marcia; Letaw, John H; Litman, Gary W; Litman, Ronda T; Mikami, Masato; Ota, Tatsuya; Saha, Nil Ratan; Williams, Louise; Stadler, Peter F; Wang, Han; Taylor, John S; Fontenot, Quenton; Ferrara, Allyse; Searle, Stephen M J; Aken, Bronwen; Yandell, Mark; Schneider, Igor; Yoder, Jeffrey A; Volff, Jean-Nicolas; Meyer, Axel; Amemiya, Chris T; Venkatesh, Byrappa; Holland, Peter W H; Guiguen, Yann; Bobe, Julien; Shubin, Neil H; Di Palma, Federica; Alföldi, Jessica; Lindblad-Toh, Kerstin; Postlethwait, John H

    2016-04-01

    To connect human biology to fish biomedical models, we sequenced the genome of spotted gar (Lepisosteus oculatus), whose lineage diverged from teleosts before teleost genome duplication (TGD). The slowly evolving gar genome has conserved in content and size many entire chromosomes from bony vertebrate ancestors. Gar bridges teleosts to tetrapods by illuminating the evolution of immunity, mineralization and development (mediated, for example, by Hox, ParaHox and microRNA genes). Numerous conserved noncoding elements (CNEs; often cis regulatory) undetectable in direct human-teleost comparisons become apparent using gar: functional studies uncovered conserved roles for such cryptic CNEs, facilitating annotation of sequences identified in human genome-wide association studies. Transcriptomic analyses showed that the sums of expression domains and expression levels for duplicated teleost genes often approximate the patterns and levels of expression for gar genes, consistent with subfunctionalization. The gar genome provides a resource for understanding evolution after genome duplication, the origin of vertebrate genomes and the function of human regulatory sequences.

  15. Comparative study of enzymatic antioxidants in muscle of elasmobranch and teleost fishes.

    Science.gov (United States)

    Vélez-Alavez, Marcela; De Anda-Montañez, Juan A; Galván-Magaña, Felipe; Zenteno-Savín, Tania

    2015-09-01

    Exercise may cause an imbalance between pro-oxidants and antioxidants. In skeletal muscle, oxygen flow can increase considerably during vigorous exercise. The antioxidant system in athletes contributes to neutralize the concomitant rise in reactive oxygen species (ROS) production. The objective of this study was to compare the antioxidant system in muscle of three species of elasmobranchs and teleosts, considering differences in swimming capacity among species within each group and evolutionary differences between the two groups. Muscle samples were collected from elasmobranchs (Isurus oxyrinchus, Prionace glauca, Mustelus henlei) and teleosts (Totoaba macdonaldi, Kajikia audax and Coryphaena hippurus) in the coast of the Baja California peninsula, Mexico. The enzymatic activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione S-transferase (GST) was determined by spectrophotometry. The activity of the antioxidant enzymes CAT, GPx and GST was higher in elasmobranchs, as a group, than in teleosts. In fish species with high swimming capacities, P. glauca, K. audax and C. hippurus, antioxidant enzyme activity was higher in comparison with species with lower swimming capacities, M. henlei and T. macdonaldi. It is possible that antioxidant enzymes, particularly SOD, GPx and GST, contribute to avoidance of oxidative damage in teleost and elasmobranch species with higher swimming capacities. The antioxidant enzyme activities in fish appear to depend mainly on their swimming capacity and life style rather than the evolutionary group (elasmobranchs, teleosts).

  16. Subfunction partitioning, the teleost radiation and the annotation of the human genome.

    Science.gov (United States)

    Postlethwait, John; Amores, Angel; Cresko, William; Singer, Amy; Yan, Yi-Lin

    2004-10-01

    Half of all vertebrate species are teleost fish. What accounts for this explosion of biodiversity? Recent evidence and advances in evolutionary theory suggest that genomic features could have played a significant role in the teleost radiation. This review examines evidence for an ancient whole-genome duplication (tetraploidization) event that probably occurred just before the teleost radiation. The partitioning of ancestral subfunctions between gene copies arising from this duplication could have contributed to the genetic isolation of populations, to lineage-specific diversification of developmental programs, and ultimately to phenotypic variation among teleost fish. Beyond its importance for understanding mechanisms that generate biodiversity, the partitioning of subfunctions between teleost co-orthologs of human genes can facilitate the identification of tissue-specific conserved noncoding regions and can simplify the analysis of ancestral gene functions obscured by pleiotropy or haploinsufficiency. Applying these principles on a genomic scale can accelerate the functional annotation of the human genome and understanding of the roles of human genes in health and disease.

  17. The spotted gar genome illuminates vertebrate evolution and facilitates human-to-teleost comparisons

    Science.gov (United States)

    Braasch, Ingo; Gehrke, Andrew R.; Smith, Jeramiah J.; Kawasaki, Kazuhiko; Manousaki, Tereza; Pasquier, Jeremy; Amores, Angel; Desvignes, Thomas; Batzel, Peter; Catchen, Julian; Berlin, Aaron M.; Campbell, Michael S.; Barrell, Daniel; Martin, Kyle J.; Mulley, John F.; Ravi, Vydianathan; Lee, Alison P.; Nakamura, Tetsuya; Chalopin, Domitille; Fan, Shaohua; Wcisel, Dustin; Cañestro, Cristian; Sydes, Jason; Beaudry, Felix E. G.; Sun, Yi; Hertel, Jana; Beam, Michael J.; Fasold, Mario; Ishiyama, Mikio; Johnson, Jeremy; Kehr, Steffi; Lara, Marcia; Letaw, John H.; Litman, Gary W.; Litman, Ronda T.; Mikami, Masato; Ota, Tatsuya; Saha, Nil Ratan; Williams, Louise; Stadler, Peter F.; Wang, Han; Taylor, John S.; Fontenot, Quenton; Ferrara, Allyse; Searle, Stephen M. J.; Aken, Bronwen; Yandell, Mark; Schneider, Igor; Yoder, Jeffrey A.; Volff, Jean-Nicolas; Meyer, Axel; Amemiya, Chris T.; Venkatesh, Byrappa; Holland, Peter W. H.; Guiguen, Yann; Bobe, Julien; Shubin, Neil H.; Di Palma, Federica; Alföldi, Jessica; Lindblad-Toh, Kerstin; Postlethwait, John H.

    2016-01-01

    To connect human biology to fish biomedical models, we sequenced the genome of spotted gar (Lepisosteus oculatus), whose lineage diverged from teleosts before the teleost genome duplication (TGD). The slowly evolving gar genome conserved in content and size many entire chromosomes from bony vertebrate ancestors. Gar bridges teleosts to tetrapods by illuminating the evolution of immunity, mineralization, and development (e.g., Hox, ParaHox, and miRNA genes). Numerous conserved non-coding elements (CNEs, often cis-regulatory) undetectable in direct human-teleost comparisons become apparent using gar: functional studies uncovered conserved roles of such cryptic CNEs, facilitating annotation of sequences identified in human genome-wide association studies. Transcriptomic analyses revealed that the sum of expression domains and levels from duplicated teleost genes often approximate patterns and levels of gar genes, consistent with subfunctionalization. The gar genome provides a resource for understanding evolution after genome duplication, the origin of vertebrate genomes, and the function of human regulatory sequences. PMID:26950095

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

    DEFF Research Database (Denmark)

    Kamei, Hiroyasu; Lu, Ling; Jiao, Shuang;

    2008-01-01

    Background: 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. Methodology/Principal Findings: 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...

  19. Developmental role of acetylcholinesterase in impulse control in zebrafish

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    Matthew O Parker

    2015-10-01

    Full Text Available Cellular and molecular processes that mediate individual variability in impulsivity, a key behavioural component of many neuropsychiatric disorders, are poorly understood. Zebrafish heterozygous for a nonsense mutation in Ache (achesb55/+ showed lower levels of impulsivity in a 5-choice serial reaction time task (5-CSRTT than wild type and ache+/+. Assessment of expression of cholinergic (nAChR, serotonergic (5-HT and dopamine (DR receptor mRNA in both adult and larval (9dpf achesb55/+ revealed significant downregulation of Chrna2, Chrna5 and Drd2 mRNA in achesb55/+ larvae, but no differences in adults. Acute exposure to cholinergic agonist/antagonists had no effect on impulsivity, supporting the hypothesis that behavioural effects observed in adults were due to lasting impact of developmental alterations in cholinergic and dopaminergic signalling. This shows the cross-species role of cholinergic signalling during brain development in impulsivity, and suggests zebrafish may be a useful model for the role of cholinergic pathways as a target for therapeutic advances in addiction medicine.

  20. Atrazine and its degradates have little effect on the corticosteroid stress response in the zebrafish.

    Science.gov (United States)

    Van Der Kraak, Glen; Matsumoto, Jacquie; Kim, Myoungwoo; Hosmer, Alan J

    2015-04-01

    The present study examined the effects of atrazine on basal and forced swimming induced changes in whole body cortisol content in adult zebrafish. Zebrafish were exposed to graded concentrations of atrazine or the atrazine degradates deisopropylatrazine (DIA), deethylatrazine (DEA) and diamino-s-chlorotriazine (DACT) for up to 10 days. Some fish were sampled for the measurement of whole body cortisol levels under basal conditions while others were sampled after being subjected to a 20 min swimming challenge in order to quantify stress induced cortisol levels. In one experiment, zebrafish were subjected to two bouts of forced swimming 3h apart to test whether prior atrazine exposure affects the ability of the fish to respond appropriately to a repeated stressor. The results demonstrated that controls not exposed to atrazine and zebrafish exposed to atrazine or the atrazine degradates at nominal concentrations of up to 100 μg/L consistently exhibited increased whole body cortisol content in response to the swimming challenge. Separate analyses revealed few changes in basal or stress induced cortisol levels following atrazine exposure. Overall, these data suggest that atrazine and some of its degradates at the concentrations tested have minimal effects on the cortisol mediated stress response in the zebrafish.

  1. The comparison of methods for measuring oxidative stress in zebrafish brains.

    Science.gov (United States)

    Moussavi Nik, Seyyed Hani; Croft, Kevin; Mori, Trevor A; Lardelli, Michael

    2014-06-01

    The zebrafish is a versatile model organism with the potential to contribute to our understanding of the molecular pathological mechanisms underlying Alzheimer's disease (AD). An early characteristic of AD brain pathology is lipid peroxidation resulting from oxidative stress. However, changes in lipid peroxidation have not yet been assessed in zebrafish brains, and an earlier attempt to observe changes in F₂-isoprostane levels in the brains of zebrafish exposed to hypoxia was unsuccessful. In this article, we examine the utility of various assays of lipid peroxidation and more general assays of intracellular oxidative stress to detect the changes in oxidative stress in the brains of adult zebrafish exposed to hypoxia or explanted into a sodium azide solution for chemical mimicry of hypoxia. Levels of F₂-isoprostanes and F₄-neuroprostanes were low and variable in zebrafish brains such that statistically significant changes due to hypoxia or chemical mimicry of hypoxia could not be observed. However, measurement of lipid hydroperoxides did reveal significant changes in lipid peroxidation under these conditions, while analyses of catalase gene expression and an assay based on 2',7'-dicholorofluorescein oxidation also revealed changes in oxidative stress levels.

  2. Development of an Animal Model for Alcoholic Liver Disease in Zebrafish.

    Science.gov (United States)

    Lin, Jiun-Nong; Chang, Lin-Li; Lai, Chung-Hsu; Lin, Kai-Jen; Lin, Mei-Fang; Yang, Chih-Hui; Lin, Hsi-Hsun; Chen, Yen-Hsu

    2015-08-01

    Alcoholic liver disease (ALD) continues to be a major cause of liver-related morbidity and mortality worldwide. To date, no zebrafish animal model has demonstrated the characteristic manifestations of ALD in the setting of chronic alcohol exposure. The aim of this study was to develop a zebrafish animal model for ALD. Male adult zebrafish were housed in a 1% (v/v) ethanol solution up to 3 months. A histopathological study showed the characteristic features of alcoholic liver steatosis and steatohepatitis in the early stages of alcohol exposure, including fat droplet accumulation, ballooning degeneration of the hepatocytes, and Mallory body formation. As the exposure time increased, collagen deposition in the extracellular matrix was observed by Sirius red staining and immunofluorescence staining. Finally, anaplastic hepatocytes with pleomorphic nuclei were arranged in trabecular patterns and formed nodules in the zebrafish liver. Over the time course of 1% ethanol exposure, upregulations of lipogenesis, fibrosis, and tumor-related genes were also revealed by semiquantitative and quantitative real-time reverse transcription-polymerase chain reaction. As these data reflect characteristic liver damage by alcohol in humans, this zebrafish animal model may serve as a powerful tool to study the pathogenesis and treatment of ALD and its related disorders in humans.

  3. Effects of β-endosulfan on the growth and reproduction of zebrafish (Danio rerio).

    Science.gov (United States)

    Han, Zhihua; Jiao, Shaojun; Kong, Deyang; Shan, Zhengjun; Zhang, Xiaowei

    2011-11-01

    Because of persistent organic pollution in aquatic environments, the widely used organochlorine pesticide endosulfan, which is a potential endocrine disruptor, is expected to pose a significant risk to aquatic organisms. In the present study, we explored the potential endocrine-disrupting risk of β-endosulfan by investigating its effect on the growth, reproduction, plasma vitellogenin, and organ histology of adult zebrafish. We found that, although β-endosulfan did not significantly affect the growth of zebrafish, it greatly decreased the hatching rate, even at a concentration as low as 10 ng/L. Interestingly, the decrease of the hatching rate was highly correlated with pathological alterations of the testes. Additionally, the values of the gonadosomatic index were significantly reduced in female zebrafish treated with 200 ng/L β-endosulfan, which was also closely associated with ovarian histological changes. More importantly, a significant increase in the level of vitellogenin was observed in all male fish treated with β-endosulfan. Based on these findings, we conclude that β-endosulfan severely affects the reproductive function of zebrafish and the synthesis of vitellogenin in the liver, and thus, β-endosulfan has a serious endocrine disruption function in zebrafish.

  4. Dorsomorphin promotes survival and germline competence of zebrafish spermatogonial stem cells in culture.

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    Ten-Tsao Wong

    Full Text Available Zebrafish spermatogonial cell cultures were established from Tg(piwil1:neo;Tg(piwil1:DsRed transgenic fish using a zebrafish ovarian feeder cell line (OFC3 that was engineered to express zebrafish Lif, Fgf2 and Gdnf. Primary cultures, initiated from testes, were treated with G418 to eliminate the somatic cells and select for the piwil1:neo expressing spermatogonia. Addition of dorsomorphin, a Bmp type I receptor inhibitor, prolonged spermatogonial stem cell (SSC survival in culture and enhanced germline transmission of the SSCs following transplantation into recipient larvae. In contrast, dorsomorphin inhibited the growth and survival of zebrafish female germline stem cells (FGSCs in culture. In the presence of dorsomorphin, the spermatogonia continued to express the germ-cell markers dazl, dnd, nanos3, vasa and piwil1 and the spermatogonial markers plzf and sox17 for at least six weeks in culture. Transplantation experiments revealed that 6 week-old spermatogonial cell cultures maintained in the presence of dorsomorphin were able to successfully colonize the gonad in 18% of recipient larvae and produce functional gametes in the resulting adult chimeric fish. Germline transmission was not successful when the spermatogonia were cultured 6 weeks in the absence of dorsomorphin before transplantation. The results indicate that Bmp signaling is detrimental to SSCs but required for the survival of zebrafish FGSCs in culture. Manipulation of Bmp signaling could provide a strategy to optimize culture conditions of germline stem cells from other species.

  5. Expression and function on embryonic development of lissencephaly-1 genes in zebrafish

    Institute of Scientific and Technical Information of China (English)

    Chengfu Sun; Mafei Xu; Zhen Xing; Zhili Wu; Yiping Li; Tsaiping Li; Mujun Zhao

    2009-01-01

    Lissencephaly is a severe disease characterized by brain malformation. The main causative gene of lissencephaly is LIS1. Mutation or deletion of LIS1 leads to prolifer-ation and migration deficiency of neurons in brain devel-opment. However, little is known about its biological function in embryonic development. In this article, we identified the expression patterns of zebrafish LIS1 gene and investigated its function in embryonic development. We demonstrated that zebrafish consisted of two LIS1 genes, LIS1a and LIS1b. Bioinformatics analysis revealed that LIS1 genes were conserved in evolution both in protein sequences and genomic structures. The expression patterns of zebrafish LIS1a and LIS1b showed that both transcripts were ubiquitously expressed at all embryonic developmental stages and in adult tissues examined. At the protein level, the LIS1 products mainly exist in brain tissue and in embryos at early stages as shown by western blotting analysis. The whole-mount immunostaining data showed that LIS1 proteins were distributed all over the embryos from 1-cell stage to 5 day post-fertilization. Knockdown of LIS1 protein expression through morpholino antisense oligonucleotides resulted in many developmental deficiencies in zebrafish, including brain malformation, circulation abnormality, and body curl. Taken together, our study suggested that zebrafish LIS1 plays a very important role in embryonic development.

  6. Upregulation of leukemia inhibitory factor (LIF during the early stage of optic nerve regeneration in zebrafish.

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    Kazuhiro Ogai

    Full Text Available Fish retinal ganglion cells (RGCs can regenerate their axons after optic nerve injury, whereas mammalian RGCs normally fail to do so. Interleukin 6 (IL-6-type cytokines are involved in cell differentiation, proliferation, survival, and axon regrowth; thus, they may play a role in the regeneration of zebrafish RGCs after injury. In this study, we assessed the expression of IL-6-type cytokines and found that one of them, leukemia inhibitory factor (LIF, is upregulated in zebrafish RGCs at 3 days post-injury (dpi. We then demonstrated the activation of signal transducer and activator of transcription 3 (STAT3, a downstream target of LIF, at 3-5 dpi. To determine the function of LIF, we performed a LIF knockdown experiment using LIF-specific antisense morpholino oligonucleotides (LIF MOs. LIF MOs, which were introduced into zebrafish RGCs via a severed optic nerve, reduced the expression of LIF and abrogated the activation of STAT3 in RGCs after injury. These results suggest that upregulated LIF drives Janus kinase (Jak/STAT3 signaling in zebrafish RGCs after nerve injury. In addition, the LIF knockdown impaired axon sprouting in retinal explant culture in vitro; reduced the expression of a regeneration-associated molecule, growth-associated protein 43 (GAP-43; and delayed functional recovery after optic nerve injury in vivo. In this study, we comprehensively demonstrate the beneficial role of LIF in optic nerve regeneration and functional recovery in adult zebrafish.

  7. Myotonia congenita-associated mutations in chloride channel-1 affect zebrafish body wave swimming kinematics.

    Directory of Open Access Journals (Sweden)

    Wei Cheng

    Full Text Available Myotonia congenita is a human muscle disorder caused by mutations in CLCN1, which encodes human chloride channel 1 (CLCN1. Zebrafish is becoming an increasingly useful model for human diseases, including muscle disorders. In this study, we generated transgenic zebrafish expressing, under the control of a muscle specific promoter, human CLCN1 carrying mutations that have been identified in human patients suffering from myotonia congenita. We developed video analytic tools that are able to provide precise quantitative measurements of movement abnormalities in order to analyse the effect of these CLCN1 mutations on adult transgenic zebrafish swimming. Two new parameters for body-wave kinematics of swimming reveal changes in body curvature and tail offset in transgenic zebrafish expressing the disease-associated CLCN1 mutants, presumably due to their effect on muscle function. The capability of the developed video analytic tool to distinguish wild-type from transgenic zebrafish could provide a useful asset to screen for compounds that reverse the disease phenotype, and may be applicable to other movement disorders besides myotonia congenita.

  8. Myotonia Congenita-Associated Mutations in Chloride Channel-1 Affect Zebrafish Body Wave Swimming Kinematics

    Science.gov (United States)

    Cheng, Wei; Tian, Jing; Burgunder, Jean-Marc; Hunziker, Walter; Eng, How-Lung

    2014-01-01

    Myotonia congenita is a human muscle disorder caused by mutations in CLCN1, which encodes human chloride channel 1 (CLCN1). Zebrafish is becoming an increasingly useful model for human diseases, including muscle disorders. In this study, we generated transgenic zebrafish expressing, under the control of a muscle specific promoter, human CLCN1 carrying mutations that have been identified in human patients suffering from myotonia congenita. We developed video analytic tools that are able to provide precise quantitative measurements of movement abnormalities in order to analyse the effect of these CLCN1 mutations on adult transgenic zebrafish swimming. Two new parameters for body-wave kinematics of swimming reveal changes in body curvature and tail offset in transgenic zebrafish expressing the disease-associated CLCN1 mutants, presumably due to their effect on muscle function. The capability of the developed video analytic tool to distinguish wild-type from transgenic zebrafish could provide a useful asset to screen for compounds that reverse the disease phenotype, and may be applicable to other movement disorders besides myotonia congenita. PMID:25083883

  9. Advances in the Study of Heart Development and Disease Using Zebrafish

    Science.gov (United States)

    Brown, Daniel R.; Samsa, Leigh Ann; Qian, Li; Liu, Jiandong

    2016-01-01

    Animal models of cardiovascular disease are key players in the translational medicine pipeline used to define the conserved genetic and molecular basis of disease. Congenital heart diseases (CHDs) are the most common type of human birth defect and feature structural abnormalities that arise during cardiac development and maturation. The zebrafish, Danio rerio, is a valuable vertebrate model organism, offering advantages over traditional mammalian models. These advantages include the rapid, stereotyped and external development of transparent embryos produced in large numbers from inexpensively housed adults, vast capacity for genetic manipulation, and amenability to high-throughput screening. With the help of modern genetics and a sequenced genome, zebrafish have led to insights in cardiovascular diseases ranging from CHDs to arrhythmia and cardiomyopathy. Here, we discuss the utility of zebrafish as a model system and summarize zebrafish cardiac morphogenesis with emphasis on parallels to human heart diseases. Additionally, we discuss the specific tools and experimental platforms utilized in the zebrafish model including forward screens, functional characterization of candidate genes, and high throughput applications. PMID:27335817

  10. Mechanism of TiO2 nanoparticle-induced neurotoxicity in zebrafish (Danio rerio).

    Science.gov (United States)

    Sheng, Lei; Wang, Ling; Su, Mingyu; Zhao, Xiaoyang; Hu, Renping; Yu, Xiaohong; Hong, Jie; Liu, Dong; Xu, Bingqing; Zhu, Yunting; Wang, Han; Hong, Fashui

    2016-02-01

    Zebrafish (Danio rerio) has been used historically for evaluating the toxicity of environmental and aqueous toxicants, and there is an emerging literature reporting toxic effects of manufactured nanoparticles (NPs) in zebrafish embryos. Few researches, however, are focused on the neurotoxicity on adult zebrafish after subchronic exposure to TiO2 NPs. This study was designed to evaluate the morphological changes, alterations of neurochemical contents, and expressions of memory behavior-related genes in zebrafish brains caused by exposures to 5, 10, 20, and 40 μg/L TiO2 NPs for 45 consecutive days. Our data indicated that spatial recognition memory and levels of norepinephrine, dopamine, and 5-hydroxytryptamine were significantly decreased and NO levels were markedly elevated, and over proliferation of glial cells, neuron apoptosis, and TiO2 NP aggregation were observed after low dose exposures of TiO2 NPs. Furthermore, the low dose exposures of TiO2 NPs significantly activated expressions of C-fos, C-jun, and BDNF genes, and suppressed expressions of p38, NGF, CREB, NR1, NR2ab, and GluR2 genes. These findings imply that low dose exposures of TiO2 NPs may result in the brain damages in zebrafish, provide a developmental basis for evaluating the neurotoxicity of subchronic exposure, and raise the caution of aquatic application of TiO2 NPs.

  11. Endocrine pancreas development in zebrafish.

    Science.gov (United States)

    Tehrani, Zahra; Lin, Shuo

    2011-10-15

    Type 1 diabetes results from the autoimmune destruction of insulin-producing pancreatic β cells. Current efforts to cure diabetes are aimed at replenishing damaged cells by generating a new supply of β cells in vitro. The most promising strategy for achieving this goal is to differentiate embryonic stem (ES) cells by sequentially exposing them to signaling molecules that they would normally encounter in vivo. This approach requires a thorough understanding of the temporal sequence of the signaling events underlying pancreatic β-cell induction during embryonic development. The zebrafish system has emerged as a powerful tool in the study of pancreas development. In this review, we provide a temporal summary of pancreas development in zebrafish with a special focus on the formation of pancreatic β cells.

  12. Deriving cell lines from zebrafish embryos and tumors

    NARCIS (Netherlands)

    Choorapoikayil, S.; Overvoorde, J.; den Hertog, J.

    2013-01-01

    Over the last two decades the zebrafish has emerged as a powerful model organism in science. The experimental accessibility, the broad range of zebrafish mutants, and the highly conserved genetic and biochemical pathways between zebrafish and mammals lifted zebrafish to become one of the most attrac

  13. In vivo induction of oocyte maturation and ovulation in zebrafish.

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    Toshinobu Tokumoto

    Full Text Available The maturation of fish oocytes is a well-characterized system induced by progestins via non-genomic actions. In a previous study, we demonstrated that diethylstilbestrol (DES, a non-steroidal estrogen, induces fish oocyte maturation via the membrane progestin receptor (mPR. Here, we attempted to evaluate the effect of DES as an environmental endocrine disrupting chemical (EDC upon fish oocyte maturation using live zebrafish. DES triggered oocyte maturation within several hours in vivo when administrated directly into the surrounding water. The natural teleost maturation-inducing hormone, 17alpha, 20beta-dihydroxy-4-pregnen-3-one (17,20beta-DHP also induced oocyte maturation in vivo. Steroids such as testosterone, progesterone or 17alpha-hydroxyprogesterone were also effective in vivo. Further studies indicated that externally applied 17,20beta-DHP even induced ovulation. In contrast to 17,20beta -DHP, DES induced maturation but not ovulation. Theoretically this assay system provides a means to distinguish pathways involved in the induction of ovulation, which are known to be induced by genomic actions from the pathway normally involved in the induction of oocyte maturation, a typical non-genomic action-dependent pathway. In summary, we have demonstrated the effect of EDCs on fish oocyte maturation in vivo. To address the effects, we have explored a conceptually new approach to distinguish between the genomic and non-genomic actions induced by steroids. The assay can be applied to screens of progestin-like effects upon oocyte maturation and ovulation for small molecules of pharmacological agents or EDCs.

  14. Expression of GPR177 (Wntless/Evi/Sprinter), a Highly Conserved Wnt-Transport Protein, in Rat Tissues, Zebrafish Embryos, and Cultured Human Cells

    OpenAIRE

    Jin, Jay; Morse, Megan; Frey, Colleen; Petko, Jessica; Levenson, Robert

    2010-01-01

    GPR177 is an evolutionarily conserved transmembrane protein necessary for Wnt protein secretion. Little is currently known, however, regarding expression of GPR177, especially in vertebrate species. We have developed an antiserum against GPR177, and used it to examine expression of GPR177 in human tissue culture cells, adult mouse and rat tissues, as well as developing zebrafish embryos. In rodents, GPR177 is expressed in virtually all tissue types and brain regions examined. In zebrafish, GP...

  15. Identification and functional characterization of cardiac pacemaker cells in zebrafish.

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    Federico Tessadori

    Full Text Available In the mammalian heart a conduction system of nodes and conducting cells generates and transduces the electrical signals evoking myocardial contractions. Specialized pacemaker cells initiating and controlling cardiac contraction rhythmicity are localized in an anatomically identifiable structure of myocardial origin, the sinus node. We previously showed that in mammalian embryos sinus node cells originate from cardiac progenitors expressing the transcription factors T-box transcription factor 3 (Tbx3 and Islet-1 (Isl1. Although cardiac development and function are strikingly conserved amongst animal classes, in lower vertebrates neither structural nor molecular distinguishable components of a conduction system have been identified, questioning its evolutionary origin. Here we show that zebrafish embryos lacking the LIM/homeodomain-containing transcription factor Isl1 display heart rate defects related to pacemaker dysfunction. Moreover, 3D reconstructions of gene expression patterns in the embryonic and adult zebrafish heart led us to uncover a previously unidentified, Isl1-positive and Tbx2b-positive region in the myocardium at the junction of the sinus venosus and atrium. Through their long interconnecting cellular protrusions the identified Isl1-positive cells form a ring-shaped structure. In vivo labeling of the Isl1-positive cells by transgenic technology allowed their isolation and electrophysiological characterization, revealing their unique pacemaker activity. In conclusion we demonstrate that Isl1-expressing cells, organized as a ring-shaped structure around the venous pole, hold the pacemaker function in the adult zebrafish heart. We have thereby identified an evolutionary conserved, structural and molecular distinguishable component of the cardiac conduction system in a lower vertebrate.

  16. Müller glia: Stem cells for generation and regeneration of retinal neurons in teleost fish.

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    Lenkowski, Jenny R; Raymond, Pamela A

    2014-05-01

    Adult zebrafish generate new neurons in the brain and retina throughout life. Growth-related neurogenesis allows a vigorous regenerative response to damage, and fish can regenerate retinal neurons, including photoreceptors, and restore functional vision following photic, chemical, or mechanical destruction of the retina. Müller glial cells in fish function as radial-glial-like neural stem cells. During adult growth, Müller glial nuclei undergo sporadic, asymmetric, self-renewing mitotic divisions in the inner nuclear layer to generate a rod progenitor that migrates along the radial fiber of the Müller glia into the outer nuclear layer, proliferates, and differentiates exclusively into rod photoreceptors. When retinal neurons are destroyed, Müller glia in the immediate vicinity of the damage partially and transiently dedifferentiate, re-express retinal progenitor and stem cell markers, re-enter the cell cycle, undergo interkinetic nuclear migration (characteristic of neuroepithelial cells), and divide once in an asymmetric, self-renewing division to generate a retinal progenitor. This daughter cell proliferates rapidly to form a compact neurogenic cluster surrounding the Müller glia; these multipotent retinal progenitors then migrate along the radial fiber to the appropriate lamina to replace missing retinal neurons. Some aspects of the injury-response in fish Müller glia resemble gliosis as observed in mammals, and mammalian Müller glia exhibit some neurogenic properties, indicative of a latent ability to regenerate retinal neurons. Understanding the specific properties of fish Müller glia that facilitate their robust capacity to generate retinal neurons will inform and inspire new clinical approaches for treating blindness and visual loss with regenerative medicine.

  17. Rdh10a Provides a Conserved Critical Step in the Synthesis of Retinoic Acid during Zebrafish Embryogenesis.

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    Enrico D'Aniello

    Full Text Available The first step in the conversion of vitamin A into retinoic acid (RA in embryos requires retinol dehydrogenases (RDHs. Recent studies have demonstrated that RDH10 is a critical core component of the machinery that produces RA in mouse and Xenopus embryos. If the conservation of Rdh10 function in the production of RA extends to teleost embryos has not been investigated. Here, we report that zebrafish Rdh10a deficient embryos have defects consistent with loss of RA signaling, including anteriorization of the nervous system and enlarged hearts with increased cardiomyocyte number. While knockdown of Rdh10a alone produces relatively mild RA deficient phenotypes, Rdh10a can sensitize embryos to RA deficiency and enhance phenotypes observed when Aldh1a2 function is perturbed. Moreover, excess Rdh10a enhances embryonic sensitivity to retinol, which has relatively mild teratogenic effects compared to retinal and RA treatment. Performing Rdh10a regulatory expression analysis, we also demonstrate that a conserved teleost rdh10a enhancer requires Pax2 sites to drive expression in the eyes of transgenic embryos. Altogether, our results demonstrate that Rdh10a has a conserved requirement in the first step of RA production within vertebrate embryos.

  18. Expression profiles for six zebrafish genes during gonadal sex differentiation

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    Rasmussen Lene J

    2008-06-01

    Full Text Available Abstract Background The mechanism of sex determination in zebrafish is largely unknown and neither sex chromosomes nor a sex-determining gene have been identified. This indicates that sex determination in zebrafish is mediated by genetic signals from autosomal genes. The aim of this study was to determine the precise timing of expression of six genes previously suggested to be associated with sex differentiation in zebrafish. The current study investigates the expression of all six genes in the same individual fish with extensive sampling dates during sex determination and -differentiation. Results In the present study, we have used quantitative real-time PCR to investigate the expression of ar, sox9a, dmrt1, fig alpha, cyp19a1a and cyp19a1b during the expected sex determination and gonadal sex differentiation period. The expression of the genes expected to be high in males (ar, sox9a and dmrt1a and high in females (fig alpha and cyp19a1a was segregated in two groups with more than 10 times difference in expression levels. All of the investigated genes showed peaks in expression levels during the time of sex determination and gonadal sex differentiation. Expression of all genes was investigated on cDNA from the same fish allowing comparison of the high and low expressers of genes that are expected to be highest expressed in either males or females. There were 78% high or low expressers of all three "male" genes (ar, sox9a and dmrt1 in the investigated period and 81% were high or low expressers of both "female" genes (fig alpha and cyp19a1a. When comparing all five genes with expected sex related expression 56% show expression expected for either male or female. Furthermore, the expression of all genes was investigated in different tissue of adult male and female zebrafish. Conclusion In zebrafish, the first significant peak in gene expression during the investigated period (2–40 dph was dmrt1 at 10 dph which indicates involvement of this gene

  19. High-resolution tissue Doppler imaging of the zebrafish heart during its regeneration.

    Science.gov (United States)

    Huang, Chih-Chung; Su, Ta-Han; Shih, Cho-Chiang

    2015-02-01

    The human heart cannot regenerate after injury, whereas the adult zebrafish can fully regenerate its heart even after 20% of the ventricle is amputated. Many studies have begun to reveal the cellular and molecular mechanisms underlying this regenerative process, which have exciting implications for human cardiac diseases. However, the dynamic functions of the zebrafish heart during regeneration are not yet understood. This study established a high-resolution echocardiography for tissue Doppler imaging (TDI) of the zebrafish heart to explore the cardiac functions during different regeneration phases. Experiments were performed on AB-line adult zebrafish (n=40) in which 15% of the ventricle was surgically removed. An 80-MHz ultrasound TDI based on color M-mode imaging technology was employed. The cardiac flow velocities and patterns from both the ventricular chamber and myocardium were measured at different regeneration phases relative to the day of amputation. The peak velocities of early diastolic inflow, early diastolic myocardial motion, late diastolic myocardial motion, early diastolic deceleration slope, and heart rate were increased at 3 days after the myocardium amputation, but these parameters gradually returned to close to their baseline values for the normal heart at 7 days after amputation. The peak velocities of late diastolic inflow, ventricular systolic outflow, and systolic myocardial motion did not significantly differ during the heart regeneration.

  20. Single epicardial cell transcriptome sequencing identifies Caveolin 1 as an essential factor in zebrafish heart regeneration.

    Science.gov (United States)

    Cao, Jingli; Navis, Adam; Cox, Ben D; Dickson, Amy L; Gemberling, Matthew; Karra, Ravi; Bagnat, Michel; Poss, Kenneth D

    2016-01-15

    In contrast to mammals, adult zebrafish have a high capacity to regenerate damaged or lost myocardium through proliferation of cardiomyocytes spared from damage. The epicardial sheet covering the heart is activated by injury and aids muscle regeneration through paracrine effects and as a multipotent cell source, and has received recent attention as a target in cardiac repair strategies. Although it is recognized that epicardium is required for muscle regeneration and itself has high regenerative potential, the extent of cellular heterogeneity within epicardial tissue is largely unexplored. Here, we performed transcriptome analysis on dozens of epicardial lineage cells purified from zebrafish harboring a transgenic reporter for the pan-epicardial gene tcf21. Hierarchical clustering analysis suggested the presence of at least three epicardial cell subsets defined by expression signatures. We validated many new pan-epicardial and epicardial markers by alternative expression assays. Additionally, we explored the function of the scaffolding protein and main component of caveolae, caveolin 1 (cav1), which was present in each epicardial subset. In BAC transgenic zebrafish, cav1 regulatory sequences drove strong expression in ostensibly all epicardial cells and in coronary vascular endothelial cells. Moreover, cav1 mutant zebrafish generated by genome editing showed grossly normal heart development and adult cardiac anatomy, but displayed profound defects in injury-induced cardiomyocyte proliferation and heart regeneration. Our study defines a new platform for the discovery of epicardial lineage markers, genetic tools, and mechanisms of heart regeneration.

  1. PhOTO zebrafish: a transgenic resource for in vivo lineage tracing during development and regeneration.

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    William P Dempsey

    Full Text Available BACKGROUND: Elucidating the complex cell dynamics (divisions, movement, morphological changes, etc. underlying embryonic development and adult tissue regeneration requires an efficient means to track cells with high fidelity in space and time. To satisfy this criterion, we developed a transgenic zebrafish line, called PhOTO, that allows photoconvertible optical tracking of nuclear and membrane dynamics in vivo. METHODOLOGY: PhOTO zebrafish ubiquitously express targeted blue fluorescent protein (FP Cerulean and photoconvertible FP Dendra2 fusions, allowing for instantaneous, precise targeting and tracking of any number of cells using Dendra2 photoconversion while simultaneously monitoring global cell behavior and morphology. Expression persists through adulthood, making the PhOTO zebrafish an excellent tool for studying tissue regeneration: after tail fin amputation and photoconversion of a ∼100 µm stripe along the cut area, marked differences seen in how cells contribute to the new tissue give detailed insight into the dynamic process of regeneration. Photoconverted cells that contributed to the regenerate were separated into three distinct populations corresponding to the extent of cell division 7 days after amputation, and a subset of cells that divided the least were organized into an evenly spaced, linear orientation along the length of the newly regenerating fin. CONCLUSIONS/SIGNIFICANCE: PhOTO zebrafish have wide applicability for lineage tracing at the systems-level in the early embryo as well as in the adult, making them ideal candidate tools for future research in development, traumatic injury and regeneration, cancer progression, and stem cell behavior.

  2. Pesticide- and sex steroid analogue-induced endocrine disruption differentially targets hypothalamo-hypophyseal-gonadal system during gametogenesis in teleosts - A review.

    Science.gov (United States)

    Senthilkumaran, Balasubramanian

    2015-08-01

    Pesticide-induced endocrine disruption often mimics sex steroidal action resulting in physiological functional disarray of hypothalamo-hypophyseal-gonadal (HHG) system at multiple levels. Among various group of pesticides, organochlorine and organophosphate family of pesticides are known to impart sex steroidal mimicking activity with slightly higher resemblance to estrogens when compared to androgenic action. This review will highlight the effects of organochlorine (for e.g. endosulfan) and organophosphate (for e.g. malathion) pesticides in comparison with sex-steroid analogue-induced changes on HHG axis during gametogenesis in few teleost fish models. Interestingly, the effects of these compounds have produced differential effects in juveniles and adults which also vary based on exposure dosage and duration. Further, the treatments had caused at times sexually dimorphic effects indicating that the action of these compounds bring out serious implications in sexual development. A comprehensive overview has been provided by considering all these aspects to recognize the adverse impacts of pesticide-induced endocrine disruption with special reference to endosulfan and malathion as those had been applied even today or used before for controlling agricultural pests in several Asian countries including India. This review also compares the effects of sex-steroid analogues where in sex reversal to reproductive dysfunction is evident, which may imply the extent of sexual plasticity in teleosts compared to other vertebrates.

  3. Evolution history of duplicated smad3 genes in teleost: insights from Japanese flounder, Paralichthys olivaceus

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    Xinxin Du

    2016-09-01

    Full Text Available Following the two rounds of whole-genome duplication (WGD during deuterosome evolution, a third genome duplication occurred in the ray-fined fish lineage and is considered to be responsible for the teleost-specific lineage diversification and regulation mechanisms. As a receptor-regulated SMAD (R-SMAD, the function of SMAD3 was widely studied in mammals. However, limited information of its role or putative paralogs is available in ray-finned fishes. In this study, two SMAD3 paralogs were first identified in the transcriptome and genome of Japanese flounder (Paralichthys olivaceus. We also explored SMAD3 duplication in other selected species. Following identification, genomic structure, phylogenetic reconstruction, and synteny analyses performed by MrBayes and online bioinformatic tools confirmed that smad3a/3b most likely originated from the teleost-specific WGD. Additionally, selection pressure analysis and expression pattern of the two genes performed by PAML and quantitative real-time PCR (qRT-PCR revealed evidence of subfunctionalization of the two SMAD3 paralogs in teleost. Our results indicate that two SMAD3 genes originate from teleost-specific WGD, remain transcriptionally active, and may have likely undergone subfunctionalization. This study provides novel insights to the evolution fates of smad3a/3b and draws attentions to future function analysis of SMAD3 gene family.

  4. Cytochrome P450 20A1 in zebrafish: Cloning, regulation and potential involvement in hyperactivity disorders

    Science.gov (United States)

    Kubota, Akira; O'Meara, Conor M.; Lamb, David C.; Tanguay, Robert L.; Goldstone, Jared V.

    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 liver, heart, gonads, spleen and brain, as well as 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. PMID:26853319

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

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    Hiroyasu Kamei

    Full Text Available BACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: 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

  6. Miniaturized embryo array for automated trapping, immobilization and microperfusion of zebrafish embryos.

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    Jin Akagi

    Full Text Available Zebrafish (Danio rerio has recently emerged as a powerful experimental model in drug discovery and environmental toxicology. Drug discovery screens performed on zebrafish embryos mirror with a high level of accuracy the tests usually performed on mammalian animal models, and fish embryo toxicity assay (FET is one of the most promising alternative approaches to acute ecotoxicity testing with adult fish. Notwithstanding this, automated in-situ analysis of zebrafish embryos is still deeply in its infancy. This is mostly due to the inherent limitations of conventional techniques and the fact that metazoan organisms are not easily susceptible to laboratory automation. In this work, we describe the development of an innovative miniaturized chip-based device for the in-situ analysis of zebrafish embryos. We present evidence that automatic, hydrodynamic positioning, trapping and long-term immobilization of single embryos inside the microfluidic chips can be combined with time-lapse imaging to provide real-time developmental analysis. Our platform, fabricated using biocompatible polymer molding technology, enables rapid trapping of embryos in low shear stress zones, uniform drug microperfusion and high-resolution imaging without the need of manual embryo handling at various developmental stages. The device provides a highly controllable fluidic microenvironment and post-analysis eleuthero-embryo stage recovery. Throughout the incubation, the position of individual embryos is registered. Importantly, we also for first time show that microfluidic embryo array technology can be effectively used for the analysis of anti-angiogenic compounds using transgenic zebrafish line (fli1a:EGFP. The work provides a new rationale for rapid and automated manipulation and analysis of developing zebrafish embryos at a large scale.

  7. Identification of Chemical Inhibitors of β-Catenin-Driven Liver Tumorigenesis in Zebrafish.

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    Kimberley J Evason

    2015-07-01

    Full Text Available Hepatocellular carcinoma (HCC is one of the most lethal human cancers. The search for targeted treatments has been hampered by the lack of relevant animal models for the genetically diverse subsets of HCC, including the 20-40% of HCCs that are defined by activating mutations in the gene encoding β-catenin. To address this chemotherapeutic challenge, we created and characterized transgenic zebrafish expressing hepatocyte-specific activated β-catenin. By 2 months post fertilization (mpf, 33% of transgenic zebrafish developed HCC in their livers, and 78% and 80% of transgenic zebrafish showed HCC at 6 and 12 mpf, respectively. As expected for a malignant process, transgenic zebrafish showed significantly decreased mean adult survival compared to non-transgenic control siblings. Using this novel transgenic model, we screened for druggable pathways that mediate β-catenin-induced liver growth and identified two c-Jun N-terminal kinase (JNK inhibitors and two antidepressants (one tricyclic antidepressant, amitriptyline, and one selective serotonin reuptake inhibitor that suppressed this phenotype. We further found that activated β-catenin was associated with JNK pathway hyperactivation in zebrafish and in human HCC. In zebrafish larvae, JNK inhibition decreased liver size specifically in the presence of activated β-catenin. The β-catenin-specific growth-inhibitory effect of targeting JNK was conserved in human liver cancer cells. Our other class of hits, antidepressants, has been used in patient treatment for decades, raising the exciting possibility that these drugs could potentially be repurposed for cancer treatment. In support of this proposal, we found that amitriptyline decreased tumor burden in a mouse HCC model. Our studies implicate JNK inhibitors and antidepressants as potential therapeutics for β-catenin-induced liver tumors.

  8. Animal Models of Tuberculosis: Zebrafish

    Science.gov (United States)

    van Leeuwen, Lisanne M.; van der Sar, Astrid M.; Bitter, Wilbert

    2015-01-01

    Over the past decade the zebrafish (Danio rerio) has become an attractive new vertebrate model organism for studying mycobacterial pathogenesis. The combination of medium-throughput screening and real-time in vivo visualization has allowed new ways to dissect host pathogenic interaction in a vertebrate host. Furthermore, genetic screens on the host and bacterial sides have elucidated new mechanisms involved in the initiation of granuloma formation and the importance of a balanced immune response for control of mycobacterial pathogens. This article will highlight the unique features of the zebrafish–Mycobacterium marinum infection model and its added value for tuberculosis research. PMID:25414379

  9. Clinically approved iron chelators influence zebrafish mortality, hatching morphology and cardiac function.

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    Jasmine L Hamilton

    Full Text Available Iron chelation therapy using iron (III specific chelators such as desferrioxamine (DFO, Desferal, deferasirox (Exjade or ICL-670, and deferiprone (Ferriprox or L1 are the current standard of care for the treatment of iron overload. Although each chelator is capable of promoting some degree of iron excretion, these chelators are also associated with a wide range of well documented toxicities. However, there is currently very limited data available on their effects in developing embryos. In this study, we took advantage of the rapid development and transparency of the zebrafish embryo, Danio rerio to assess and compare the toxicity of iron chelators. All three iron chelators described above were delivered to zebrafish embryos by direct soaking and their effects on mortality, hatching and developmental morphology were monitored for 96 hpf. To determine whether toxicity was specific to embryos, we examined the effects of chelator exposure via intra peritoneal injection on the cardiac function and gene expression in adult zebrafish. Chelators varied significantly in their effects on embryo mortality, hatching and morphology. While none of the embryos or adults exposed to DFO were negatively affected, ICL -treated embryos and adults differed significantly from controls, and L1 exerted toxic effects in embryos alone. ICL-670 significantly increased the mortality of embryos treated with doses of 0.25 mM or higher and also affected embryo morphology, causing curvature of larvae treated with concentrations above 0.5 mM. ICL-670 exposure (10 µL of 0.1 mM injection also significantly increased the heart rate and cardiac output of adult zebrafish. While L1 exposure did not cause toxicity in adults, it did cause morphological defects in embryos at 0.5 mM. This study provides first evidence on iron chelator toxicity in early development and will help to guide our approach on better understanding the mechanism of iron chelator toxicity.

  10. Identification, Phylogeny, and Function of fabp2 Paralogs in Two Non-Model Teleost Fish Species.

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    Kaitetzidou, Elisavet; Chatzifotis, Stavros; Antonopoulou, Efthimia; Sarropoulou, Elena

    2015-10-01

    Intestinal fatty-acid-binding protein (IFABP or FABP2) is a cytosolic transporter of long-chain fatty acids, which is mainly expressed in cells of intestinal tissue. Fatty acids in teleosts are an important source of energy for growth, reproduction, and swimming and a main ingredient in the yolk sac of embryos and larvae. The fabp2 paralogs, fabp2a and fabp2b, were identified for 26 teleost fish species including the paralogs for the two non-model teleost fish species, namely the gilthead sea bream (Sparus aurata) and the European sea bass (Dicentrarchus labrax). Despite the high similarity of fabp2 paralogs, as well as the identical organization in four exons, paralogs were mapped to different chromosomes/linkage groups supporting the hypothesis that the identified transcripts are true paralogs originating from a single ancestor gene after genome duplication. This was also confirmed by phylogenetic analysis using fabp2 sequences of 26 teleosts and by synteny analysis carried out with ten teleosts. Differential expression analysis of the gilthead sea bream and European sea bass fabp2 paralogs in the intestine after fasting and refeeding experiment further revealed their altered implication in metabolism. Additional expression studies in seven developmental stages of the two species detected fabp2 paralogs relatively early in the embryonic development as well as possible complementary or separated roles of the paralogs. The identification and characterization of the two fabp2 paralogs will contribute significantly to the understanding of the fabp2 evolution as well as of the divergences in fatty acid metabolism.

  11. 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.

  12. Learning and Memory in Zebrafish Larvae

    Directory of Open Access Journals (Sweden)

    Adam Christopher Roberts

    2013-08-01

    Full Text Available 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.

  13. 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.

  14. Zebrafish tracking using convolutional neural networks

    Science.gov (United States)

    XU, Zhiping; Cheng, Xi En

    2017-01-01

    Keeping identity for a long term after occlusion is still an open problem in the video tracking of zebrafish-like model animals, and accurate animal trajectories are the foundation of behaviour analysis. We utilize the highly accurate object recognition capability of a convolutional neural network (CNN) to distinguish fish of the same congener, even though these animals are indistinguishable to the human eye. We used data augmentation and an iterative CNN training method to optimize the accuracy for our classification task, achieving surprisingly accurate trajectories of zebrafish of different size and age zebrafish groups over different time spans. This work will make further behaviour analysis more reliable. PMID:28211462

  15. Zebrafish tracking using convolutional neural networks

    Science.gov (United States)

    Xu, Zhiping; Cheng, Xi En

    2017-02-01

    Keeping identity for a long term after occlusion is still an open problem in the video tracking of zebrafish-like model animals, and accurate animal trajectories are the foundation of behaviour analysis. We utilize the highly accurate object recognition capability of a convolutional neural network (CNN) to distinguish fish of the same congener, even though these animals are indistinguishable to the human eye. We used data augmentation and an iterative CNN training method to optimize the accuracy for our classification task, achieving surprisingly accurate trajectories of zebrafish of different size and age zebrafish groups over different time spans. This work will make further behaviour analysis more reliable.

  16. Nitric oxide permits hypoxia-induced lymphatic perfusion by controlling arterial-lymphatic conduits in zebrafish and glass catfish

    DEFF Research Database (Denmark)

    Dahl Ejby Jensen, Lasse; Cao, Renhai; Hedlund, Eva-Maria;

    2009-01-01

    and lymphatic perfusion. Here we show in the adult zebrafish and glass catfish (Kryptopterus bicirrhis) that blood-lymphatic conduits directly connect arterial vessels to the lymphatic system. Under hypoxic conditions, arterial-lymphatic conduits (ALCs) became highly dilated and linearized by NO...

  17. Diazinon toxicity: effect on protein and nucleic acid metabolism in the liver of zebrafish, Brachydanio rerio (Cyprinidae).

    Science.gov (United States)

    Ansari, B A; Kumar, K

    1988-09-15

    Four-month old adult siblings of zebrafish were exposed to four concentrations of diazinon for up to 168 h. DNA, RNA, protein and total free amino acid content were monitored in the liver. The DNA, RNA and protein contents were significantly reduced, whereas the amino acid content was significantly enhanced. All these changes showed dose- as well as time-dependent response.

  18. An Expressed Sequence Tag (EST-enriched genetic map of turbot (Scophthalmus maximus: a useful framework for comparative genomics across model and farmed teleosts

    Directory of Open Access Journals (Sweden)

    Bouza Carmen

    2012-07-01

    Full Text Available Abstract Background The turbot (Scophthalmus maximus is a relevant species in European aquaculture. The small turbot genome provides a source for genomics strategies to use in order to understand the genetic basis of productive traits, particularly those related to sex, growth and pathogen resistance. Genetic maps represent essential genomic screening tools allowing to localize quantitative trait loci (QTL and to identify candidate genes through comparative mapping. This information is the backbone to develop marker-assisted selection (MAS programs in aquaculture. Expressed sequenced tag (EST resources have largely increased in turbot, thus supplying numerous type I markers suitable for extending the previous linkage map, which was mostly based on anonymous loci. The aim of this study was to construct a higher-resolution turbot genetic map using EST-linked markers, which will turn out to be useful for comparative mapping studies. Results A consensus gene-enriched genetic map of the turbot was constructed using 463 SNP and microsatellite markers in nine reference families. This map contains 438 markers, 180 EST-linked, clustered at 24 linkage groups. Linkage and comparative genomics evidences suggested additional linkage group fusions toward the consolidation of turbot map according to karyotype information. The linkage map showed a total length of 1402.7 cM with low average intermarker distance (3.7 cM; ~2 Mb. A global 1.6:1 female-to-male recombination frequency (RF ratio was observed, although largely variable among linkage groups and chromosome regions. Comparative sequence analysis revealed large macrosyntenic patterns against model teleost genomes, significant hits decreasing from stickleback (54% to zebrafish (20%. Comparative mapping supported particular chromosome rearrangements within Acanthopterygii and aided to assign unallocated markers to specific turbot linkage groups. Conclusions The new gene-enriched high-resolution turbot

  19. Reverse Genetic Approaches in Zebrafish

    Institute of Scientific and Technical Information of China (English)

    Peng Huang; Zuoyan Zhu; Shuo Lin; Bo Zhang

    2012-01-01

    Zebrafish (Danio rerio) is a well-established vertebrate animal model.A comprehensive collection of reverse genetics tools has been developed for studying gene function in this useful organism.Morpholino is the most widely used reagent to knock down target gene expression post-transcriptionally.For a long time,targeted genome modification has been heavily relied on large-scale traditional forward genetic screens,such as ENU (N-ethyl-N-nitrosourea) mutagenesis derived TILLING (Targeting Induced Local Lesions IN Genomes)strategy and pseudo-typed retrovirus mediated insertional mutagenesis.Recently,engineered endonucleases,including ZFNs (zinc finger nucleases) and TALENs (transcription activator-like effector nucleases),provide new and efficient strategies to directly generate sitespecific indel mutations by inducing double strand breaks in target genes.Here we summarize the major reverse genetic approaches for loss-of-function studies used and emerging in zebrafish,including strategies based on genome-wide mutagenesis and methods for sitespecific gene targeting.Future directions and expectations will also be discussed.

  20. Potential of zebrafish as a model for exploring the role of the amygdala in emotional memory and motivational behavior.

    Science.gov (United States)

    Perathoner, Simon; Cordero-Maldonado, Maria Lorena; Crawford, Alexander D

    2016-06-01

    Emotion is a key aspect of behavior, enabling humans and animals to assign either positive or negative values to sensory inputs and thereby to make appropriate decisions. Classical experiments in mammalian models, mainly in primates and rodents, have shown that the amygdala is essential for appetitive and aversive associative processing and that dysfunction of this brain region leads to various psychiatric conditions, including depression, generalized anxiety disorder, panic disorder, phobias, autism, and posttraumatic stress disorder. In the past 2 decades, the zebrafish (Danio rerio; Cyprinidae) has emerged as a versatile, reliable vertebrate model organism for the in vivo study of development, gene function, and numerous aspects of human pathologies. Small size, high fecundity, rapid external development, transparency, genetic tractability, and high genetic and physiologic homology with humans are among the factors that have contributed to the success with this small fish in different biomedical research areas. Recent findings indicate that, despite the anatomical differences in the brain structure of teleosts and tetrapods, fish possess a structure homologous to the mammalian amygdala, a hypothesis that is supported by the expression of molecular markers, analyses of neuronal projections in different brain areas, and behavioral studies. This Review summarizes this evidence and highlights a number of relevant bioassays in zebrafish to study emotional memory and motivational behavior.

  1. Serotonin Promotes Development and Regeneration of Spinal Motor Neurons in Zebrafish.

    Science.gov (United States)

    Barreiro-Iglesias, Antón; Mysiak, Karolina S; Scott, Angela L; Reimer, Michell M; Yang, Yujie; Becker, Catherina G; Becker, Thomas

    2015-11-01

    In contrast to mammals, zebrafish regenerate spinal motor neurons. During regeneration, developmental signals are re-deployed. Here, we show that, during development, diffuse serotonin promotes spinal motor neuron generation from pMN progenitor cells, leaving interneuron numbers unchanged. Pharmacological manipulations and receptor knockdown indicate that serotonin acts at least in part via 5-HT1A receptors. In adults, serotonin is supplied to the spinal cord mainly (90%) by descending axons from the brain. After a spinal lesion, serotonergic axons degenerate caudal to the lesion but sprout rostral to it. Toxin-mediated ablation of serotonergic axons also rostral to the lesion impaired regeneration of motor neurons only there. Conversely, intraperitoneal serotonin injections doubled numbers of new motor neurons and proliferating pMN-like progenitors caudal to the lesion. Regeneration of spinal-intrinsic serotonergic interneurons was unaltered by these manipulations. Hence, serotonin selectively promotes the development and adult regeneration of motor neurons in zebrafish.

  2. Cardiac hypertrophy involves both myocyte hypertrophy and hyperplasia in anemic zebrafish.

    Directory of Open Access Journals (Sweden)

    Xiaojing Sun

    Full Text Available BACKGROUND: An adult zebrafish heart possesses a high capacity of regeneration. However, it has been unclear whether and how myocyte hyperplasia contributes to cardiac remodeling in response to biomechanical stress and whether myocyte hypertrophy exists in the zebrafish. To address these questions, we characterized the zebrafish mutant tr265/tr265, whose Band 3 mutation disrupts erythrocyte formation and results in anemia. Although Band 3 does not express and function in the heart, the chronic anemia imposes a sequential biomechanical stress towards the heart. METHODOLOGY/PRINCIPAL FINDINGS: Hearts of the tr265/tr265 Danio rerio mutant become larger than those of the sibling by week 4 post fertilization and gradually exhibit characteristics of human cardiomyopathy, such as muscular disarray, re-activated fetal gene expression, and severe arrhythmia. At the cellular level, we found both increased individual cardiomyocyte size and increased myocyte proliferation can be detected in week 4 to week 12 tr265/tr265 fish. Interestingly, all tr265/tr265 fish that survive after week-12 have many more cardiomyocytes of smaller size than those in the sibling, suggesting that myocyte hyperplasia allows the long-term survival of these fish. We also show the cardiac hypertrophy process can be recapitulated in wild-type fish using the anemia-inducing drug phenylhydrazine (PHZ. CONCLUSIONS/SIGNIFICANCE: The anemia-induced cardiac hypertrophy models reported here are the first adult zebrafish cardiac hypertrophy models characterized. Unlike mammalian models, both cardiomyocyte hypertrophy and hyperplasia contribute to the cardiac remodeling process in these models, thus allowing the effects of cardiomyocyte hyperplasia on cardiac remodeling to be studied. However, since anemia can induce effects on the heart other than biomechanical, non-anemic zebrafish cardiac hypertrophy models shall be generated and characterized.

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

    Science.gov (United States)

    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.

  4. Osteoclast-like Cells in Early Zebrafish Embryos

    Directory of Open Access Journals (Sweden)

    Faiza Sharif

    2014-06-01

    Full Text Available Objective: Genes involved in bone and tissue remodelling in the vertebrates include matrix metalloproteinase-9 (mmp-9, receptor activator of necrosis factor κ-β (rank, cathepsin-k (Ctsk and tartrate-resistant acid phosphatase (TRAcP. We examine whether these markers are expressed in cells of zebrafish embryos of 1-5 days post fertilization. We also examine adult scales, which are known to contain mature osteoclasts, for comparison. Materials and Methods: In this experimental study, in situ hybrdisation, histochemistry and serial plastic and paraffin sectioning were used to analyse marker expression. Results: We found that mmp-9 mRNA, TRAcP enzyme and Ctsk YFP protein were expressed in haematopoietic tissues and in the cells scattered sparsely in the embryo. Ctsk and rank mRNA were both expressed in the branchial skeleton and in the developing pectoral fin. In these skeletal structures, histology showed that the expressing cells were located around the developing cartilage elements, in the parachondral tissue. In a transgenic zebrafish line with YFP coupled to Ctsk promoter, Ctsk expressing cells were found around pharyngeal skeletal elements. To see whether we could activate osteoclasts, we exposed prim-6 zebrafish embryos to a mixture of 1 μM dexamethasone and 1 μM vitaminutes D3. These compounds, which are known to trigger osteoclastogenensis in cell cultures, lead to an increase in intensity of Ctsk YFP expression around the skeletal elements. Conclusion: Our findings show that cells expressing a range of osteoclast markers are present in early larvae and can be activated by the addition of osteoclastogenic compounds.

  5. Zebrafish Models for Human Acute Organophosphorus Poisoning.

    Science.gov (United States)

    Faria, Melissa; Garcia-Reyero, Natàlia; Padrós, Francesc; Babin, Patrick J; Sebastián, David; Cachot, Jérôme; Prats, Eva; Arick Ii, Mark; Rial, Eduardo; Knoll-Gellida, Anja; Mathieu, Guilaine; Le Bihanic, Florane; Escalon, B Lynn; Zorzano, Antonio; Soares, Amadeu M V M; Raldúa, Demetrio

    2015-10-22

    Terrorist use of organophosphorus-based nerve agents and toxic industrial chemicals against civilian populations constitutes a real threat, as demonstrated by the terrorist attacks in Japan in the 1990 s or, even more recently, in the Syrian civil war. Thus, development of more effective countermeasures against acute organophosphorus poisoning is urgently needed. Here, we have generated and validated zebrafish models for mild, moderate and severe acute organophosphorus poisoning by exposing zebrafish larvae to different concentrations of the prototypic organophosphorus compound chlorpyrifos-oxon. Our results show that zebrafish models mimic most of the pathophysiological mechanisms behind this toxidrome in humans, including acetylcholinesterase inhibition, N-methyl-D-aspartate receptor activation, and calcium dysregulation as well as inflammatory and immune responses. The suitability of the zebrafish larvae to in vivo high-throughput screenings of small molecule libraries makes these models a valuable tool for identifying new drugs for multifunctional drug therapy against acute organophosphorus poisoning.

  6. Telomerase Is Essential for Zebrafish Heart Regeneration

    Directory of Open Access Journals (Sweden)

    Dorota Bednarek

    2015-09-01

    Full Text Available After myocardial infarction in humans, lost cardiomyocytes are replaced by an irreversible fibrotic scar. In contrast, zebrafish hearts efficiently regenerate after injury. Complete regeneration of the zebrafish heart is driven by the strong proliferation response of its cardiomyocytes to injury. Here we show that, after cardiac injury in zebrafish, telomerase becomes hyperactivated, and telomeres elongate transiently, preceding a peak of cardiomyocyte proliferation and full organ recovery. Using a telomerase-mutant zebrafish model, we found that telomerase loss drastically decreases cardiomyocyte proliferation and fibrotic tissue regression after cryoinjury and that cardiac function does not recover. The impaired cardiomyocyte proliferation response is accompanied by the absence of cardiomyocytes with long telomeres and an increased proportion of cardiomyocytes showing DNA damage and senescence characteristics. These findings demonstrate the importance of telomerase function in heart regeneration and highlight the potential of telomerase therapy as a means of stimulating cell proliferation upon myocardial infarction.

  7. Zebrafish YAC, BAC, and PAC genomic libraries.

    Science.gov (United States)

    Amemiya, C T; Zhong, T P; Silverman, G A; Fishman, M C; Zon, L I

    1999-01-01

    Numerous positional cloning projects directed at isolating genes responsible for the myriads of observed developmental defects in the zebrafish are anticipated in the very near future. In this chapter, we have reviewed the YAC, BAC, and PAC large-insert genomic resources available to the zebrafish community. We have discussed how these resources are screened and used in a positional cloning scheme and have pointed out frequently formidable logistical considerations in the approach. Despite being extremely tedious, positional cloning projects in the zebrafish will be comparatively easier to accomplish than in human and mouse, because of unique biological advantages of the zebrafish system. Moreover, the ease and speed at which genes are identified and cloned should rapidly increase as more mapping reagents and information become available, thereby paving the way for meaningful biological studies.

  8. Expression of the zebrafish CD133/prominin1 genes in cellular proliferation zones in the embryonic central nervous system and sensory organs.

    Science.gov (United States)

    McGrail, Maura; Batz, Lindsey; Noack, Kristin; Pandey, Saumya; Huang, Yong; Gu, Xun; Essner, Jeffrey J

    2010-06-01

    The CD133/prominin1 gene encodes a pentamembrane glycoprotein cell surface marker that is expressed in stem cells from neuroepithelial, hematopoietic, and various organ tissues. Here we report the analysis of two zebrafish CD133/prominin1 orthologues, prominin1a and prominin1b. The expression patterns of the zebrafish prominin1a and b genes were analyzed during embryogenesis using whole mount in situ hybridization. prominin1a and b show novel complementary and overlapping patterns of expression in proliferating zones in the developing sensory organs and central nervous system. The expression patterns suggest functional conservation of the zebrafish prominin1 genes. Initial analyses of prominin1a and b in neoplastic tissue show increased expression of both genes in a subpopulation of cells in malignant peripheral nerve sheath tumors in tp53 mutants. Based on these analyses, the zebrafish prominin1 genes will be useful markers for examining proliferating cell populations in adult organs, tissues, and tumors.

  9. Learning and Memory in Zebrafish Larvae

    OpenAIRE

    Adam Christopher Roberts; 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 promisi...

  10. Learning and memory in zebrafish larvae

    OpenAIRE

    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 promisin...

  11. Polygenic sex determination system in zebrafish.

    Directory of Open Access Journals (Sweden)

    Woei Chang Liew

    Full Text Available BACKGROUND: Despite the popularity of zebrafish as a research model, its sex determination (SD mechanism is still unknown. Most cytogenetic studies failed to find dimorphic sex chromosomes and no primary sex determining switch has been identified even though the assembly of zebrafish genome sequence is near to completion and a high resolution genetic map is available. Recent publications suggest that environmental factors within the natural range have minimal impact on sex ratios of zebrafish populations. The primary aim of this study is to find out more about how sex is determined in zebrafish. METHODOLOGY/PRINCIPAL FINDINGS: Using classical breeding experiments, we found that sex ratios across families were wide ranging (4.8% to 97.3% males. On the other hand, repeated single pair crossings produced broods of very similar sex ratios, indicating that parental genotypes have a role in the sex ratio of the offspring. Variation among family sex ratios was reduced after selection for breeding pairs with predominantly male or female offspring, another indication that zebrafish sex is regulated genetically. Further examinations by a PCR-based "blind assay" and array comparative genomic hybridization both failed to find universal sex-linked differences between the male and female genomes. Together with the ability to increase the sex bias of lines by selective breeding, these data suggest that zebrafish is unlikely to utilize a chromosomal sex determination (CSD system. CONCLUSIONS/SIGNIFICANCE: Taken together, our study suggests that zebrafish sex is genetically determined with limited, secondary influences from the environment. As we have not found any sign for CSD in the species, we propose that the zebrafish has a polygenic sex determination system.

  12. Identification of multiple integrin β1 homologs in zebrafish (Danio rerio

    Directory of Open Access Journals (Sweden)

    Boot-Handford Raymond P

    2006-06-01

    Full Text Available Abstract Background Integrins comprise a large family of α,β heterodimeric, transmembrane cell adhesion receptors that mediate diverse essential biological functions. Higher vertebrates possess a single β1 gene, and the β1 subunit associates with a large number of α subunits to form the major class of extracellular matrix (ECM receptors. Despite the fact that the zebrafish (Danio rerio is a rapidly emerging model organism of choice for developmental biology and for models of human disease, little is currently known about β1 integrin sequences and functions in this organism. Results Using RT-PCR, complete coding sequences of zebrafish β1 paralogs were obtained from zebrafish embryos or adult tissues. The results show that zebrafish possess two β1 paralogs (β1–1 and β1–2 that have a high degree of identity to other vertebrate β1 subunits. In addition, a third, more divergent, β1 paralog is present (β1–3, which may have altered ligand-binding properties. Zebrafish also have other divergent β1-like transcripts, which are C-terminally truncated forms lacking the transmembrane and cytoplasmic domains. Together with β1–3 these truncated forms comprise a novel group of β1 paralogs, all of which have a mutation in the ADMIDAS cation-binding site. Phylogenetic and genomic analyses indicate that the duplication that gave rise to β1–1 and β1–2 occurred after the divergence of the tetrapod and fish lineages, while a subsequent duplication of the ancestor of β1–2 may have given rise to β1–3 and an ancestral truncated paralog. A very recent tandem duplication of the truncated β1 paralogs appears to have taken place. The different zebrafish β1 paralogs have varied patterns of temporal expression during development. β1–1 and β1–2 are ubiquitously expressed in adult tissues, whereas the other β1 paralogs generally show more restricted patterns of expression. Conclusion Zebrafish have a large set of integrin β1

  13. Zebrafish circadian clocks: cells that see light.

    Science.gov (United States)

    Tamai, T K; Carr, A J; Whitmore, D

    2005-11-01

    In the classical view of circadian clock organization, the daily rhythms of most organisms were thought to be regulated by a central, 'master' pacemaker, usually located within neural structures of the animal. However, with the results of experiments performed in zebrafish, mammalian cell lines and, more recently, mammalian tissues, this view has changed to one where clock organization is now seen as being highly decentralized. It is clear that clocks exist in the peripheral tissues of animals as diverse as Drosophila, zebrafish and mammals. In the case of Drosophila and zebrafish, these tissues are also directly light-responsive. This light sensitivity and direct clock entrainability is also true for zebrafish cell lines and early-stage embryos. Using luminescent reporter cell lines containing clock gene promoters driving the expression of luciferase and single-cell imaging techniques, we have been able to show how each cell responds rapidly to a single light pulse by being shifted to a common phase, equivalent to the early day. This direct light sensitivity might be related to the requirement for light in these cells to activate the transcription of genes involved in DNA repair. It is also clear that the circadian clock in zebrafish regulates the timing of the cell cycle, demonstrating the wide impact that this light sensitivity and daily rhythmicity has on the biology of zebrafish.

  14. 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.

  15. Roles of inflammatory caspases during processing of zebrafish interleukin-1β in Francisella noatunensis infection

    Science.gov (United States)

    Vojtech, Lucia N.; Scharping, Nichole; Woodson, James C.; Hansen, John D.

    2012-01-01

    The interleukin-1 family of cytokines are essential for the control of pathogenic microbes but are also responsible for devastating autoimmune pathologies. Consequently, tight regulation of inflammatory processes is essential for maintaining homeostasis. In mammals, interleukin-1 beta (IL-1β) is primarily regulated at two levels, transcription and processing. The main pathway for processing IL-1β is the inflammasome, a multiprotein complex that forms in the cytosol and which results in the activation of inflammatory caspase (caspase 1) and the subsequent cleavage and secretion of active IL-1β. Although zebrafish encode orthologs of IL-1β and inflammatory caspases, the processing of IL-1β by activated caspase(s) has never been examined. Here, we demonstrate that in response to infection with the fish-specific bacterial pathogen Francisella noatunensis, primary leukocytes from adult zebrafish display caspase-1-like activity that results in IL-1β processing. Addition of caspase 1 or pancaspase inhibitors considerably abrogates IL-1β processing. As in mammals, this processing event is concurrent with the secretion of cleaved IL-1β into the culture medium. Furthermore, two putative zebrafish inflammatory caspase orthologs, caspase A and caspase B, are both able to cleave IL-1β, but with different specificities. These results represent the first demonstration of processing and secretion of zebrafish IL-1β in response to a pathogen, contributing to our understanding of the evolutionary processes governing the regulation of inflammation.                   

  16. Loss of fibrinogen in zebrafish results in symptoms consistent with human hypofibrinogenemia.

    Directory of Open Access Journals (Sweden)

    Andy H Vo

    Full Text Available Cessation of bleeding after trauma is a necessary evolutionary vertebrate adaption for survival. One of the major pathways regulating response to hemorrhage is the coagulation cascade, which ends with the cleavage of fibrinogen to form a stable clot. Patients with low or absent fibrinogen are at risk for bleeding. While much detailed information is known about fibrinogen regulation and function through studies of humans and mammalian models, bleeding risk in patients cannot always be accurately predicted purely based on fibrinogen levels, suggesting an influence of modifying factors and a need for additional genetic models. The zebrafish has orthologs to the three components of fibrinogen (fga, fgb, and fgg, but it hasn't yet been shown that zebrafish fibrinogen functions to prevent bleeding in vivo. Here we show that zebrafish fibrinogen is incorporated into an induced thrombus, and deficiency results in hemorrhage. An Fgb-eGFP fusion protein is incorporated into a developing thrombus induced by laser injury, but causes bleeding in adult transgenic fish. Antisense morpholino knockdown results in intracranial and intramuscular hemorrhage at 3 days post fertilization. The observed phenotypes are consistent with symptoms exhibited by patients with hypo- and afibrinogenemia. These data demonstrate that zebrafish possess highly conserved orthologs of the fibrinogen chains, which function similarly to mammals through the formation of a fibrin clot.

  17. A simple automated system for appetitive conditioning of zebrafish in their home tanks.

    Science.gov (United States)

    Doyle, Jillian M; Merovitch, Neil; Wyeth, Russell C; Stoyek, Matthew R; Schmidt, Michael; Wilfart, Florentin; Fine, Alan; Croll, Roger P

    2017-01-15

    We describe here an automated apparatus that permits rapid conditioning paradigms for zebrafish. Arduino microprocessors were used to control the delivery of auditory or visual stimuli to groups of adult or juvenile zebrafish in their home tanks in a conventional zebrafish facility. An automatic feeder dispensed precise amounts of food immediately after the conditioned stimuli, or at variable delays for controls. Responses were recorded using inexpensive cameras, with the video sequences analysed with ImageJ or Matlab. Fish showed significant conditioned responses in as few as 5 trials, learning that the conditioned stimulus was a predictor of food presentation at the water surface and at the end of the tank where the food was dispensed. Memories of these conditioned associations persisted for at least 2days after training when fish were tested either as groups or as individuals. Control fish, for which the auditory or visual stimuli were specifically unpaired with food, showed no comparable responses. This simple, low-cost, automated system permits scalable conditioning of zebrafish with minimal human intervention, greatly reducing both variability and labour-intensiveness. It will be useful for studies of the neural basis of learning and memory, and for high-throughput screening of compounds modifying those processes.

  18. Loss of col8a1a function during zebrafish embryogenesis results in congenital vertebral malformations.

    Science.gov (United States)

    Gray, Ryan S; Wilm, Thomas P; Smith, Jeff; Bagnat, Michel; Dale, Rodney M; Topczewski, Jacek; Johnson, Stephen L; Solnica-Krezel, Lilianna

    2014-02-01

    Congenital vertebral malformations (CVM) occur in 1 in 1000 live births and in many cases can cause spinal deformities, such as scoliosis, and result in disability and distress of affected individuals. Many severe forms of the disease, such as spondylocostal dystostosis, are recessive monogenic traits affecting somitogenesis, however the etiologies of the majority of CVM cases remain undetermined. Here we demonstrate that morphological defects of the notochord in zebrafish can generate congenital-type spine defects. We characterize three recessive zebrafish leviathan/col8a1a mutant alleles ((m531, vu41, vu105)) that disrupt collagen type VIII alpha1a (col8a1a), and cause folding of the embryonic notochord and consequently adult vertebral column malformations. Furthermore, we provide evidence that a transient loss of col8a1a function or inhibition of Lysyl oxidases with drugs during embryogenesis was sufficient to generate vertebral fusions and scoliosis in the adult spine. Using periodic imaging of individual zebrafish, we correlate focal notochord defects of the embryo with vertebral malformations (VM) in the adult. Finally, we show that bends and kinks in the notochord can lead to aberrant apposition of osteoblasts normally confined to well-segmented areas of the developing vertebral bodies. Our results afford a novel mechanism for the formation of VM, independent of defects of somitogenesis, resulting from aberrant bone deposition at regions of misshapen notochord tissue.

  19. Maternally inherited npm2 mRNA is crucial for egg developmental competence in zebrafish.

    Science.gov (United States)

    Bouleau, Aurélien; Desvignes, Thomas; Traverso, Juan Martin; Nguyen, Thaovi; Chesnel, Franck; Fauvel, Christian; Bobe, Julien

    2014-08-01

    The molecular mechanisms underlying and determining egg developmental competence remain poorly understood in vertebrates. Nucleoplasmin (Npm2) is one of the few known maternal effect genes in mammals, but this maternal effect has never been demonstrated in nonmammalian species. A link between developmental competence and the abundance of npm2 maternal mRNA in the egg was previously established using a teleost fish model for egg quality. The importance of maternal npm2 mRNA for egg developmental competence remains unknown in any vertebrate species. In the present study, we aimed to characterize the contribution of npm2 maternal mRNA to early developmental success in zebrafish using a knockdown strategy. We report here the oocyte-specific expression of npm2 and maternal inheritance of npm2 mRNA in zebrafish eggs. The knockdown of the protein translated from this maternal mRNA results in developmental arrest before the onset of epiboly and subsequent embryonic death, a phenotype also observed in embryos lacking zygotic transcription. Npm2 knockdown also results in impaired transcription of the first-wave zygotic genes. Our results show that npm2 is also a maternal effect gene in a nonmammalian vertebrate species and that maternally inherited npm2 mRNA is crucial for egg developmental competence. We also show that de novo protein synthesis from npm2 maternal mRNA is critical for developmental success beyond the blastula stage and required for zygotic genome activation. Finally, our results suggest that npm2 maternal mRNA is an important molecular factor of egg quality in fish and possibly in all vertebrates.

  20. Patterning the cone mosaic array in zebrafish retina requires specification of ultraviolet-sensitive cones.

    Directory of Open Access Journals (Sweden)

    Pamela A Raymond

    Full Text Available Cone photoreceptors in teleost fish are organized in precise, crystalline arrays in the epithelial plane of the retina. In zebrafish, four distinct morphological/spectral cone types occupy specific, invariant positions within a regular lattice. The cone lattice is aligned orthogonal and parallel to circumference of the retinal hemisphere: it emerges as cones generated in a germinal zone at the retinal periphery are incorporated as single-cell columns into the cone lattice. Genetic disruption of the transcription factor Tbx2b eliminates most of the cone subtype maximally sensitive to ultraviolet (UV wavelengths and also perturbs the long-range organization of the cone lattice. In the tbx2b mutant, the other three cone types (red, green, and blue cones are specified in the correct proportion, differentiate normally, and acquire normal, planar polarized adhesive interactions mediated by Crumbs 2a and Crumbs 2b. Quantitative image analysis of cell adjacency revealed that the cones in the tbx2b mutant primarily have two nearest neighbors and align in single-cell-wide column fragments that are separated by rod photoreceptors. Some UV cones differentiate at the dorsal retinal margin in the tbx2b mutant, although they are severely dysmorphic and are eventually eliminated. Incorporating loss of UV cones during formation of cone columns at the margin into our previously published mathematical model of zebrafish cone mosaic formation (which uses bidirectional interactions between planar cell polarity proteins and anisotropic mechanical stresses in the plane of the retinal epithelium to generate regular columns of cones parallel to the margin reproduces many features of the pattern disruptions seen in the tbx2b mutant.

  1. Redefining the initiation and maintenance of zebrafish interrenal steroidogenesis by characterizing the key enzyme cyp11a2.

    Science.gov (United States)

    Parajes, Silvia; Griffin, Aliesha; Taylor, Angela E; Rose, Ian T; Miguel-Escalada, Irene; Hadzhiev, Yavor; Arlt, Wiebke; Shackleton, Cedric; Müller, Ferenc; Krone, Nils

    2013-08-01

    Zebrafish are emerging as a model to study steroid hormone action and associated disease. However, steroidogenesis in zebrafish is not well characterized. Mammalian P450 side-chain cleavage enzyme (CYP11A1) catalyzes the first step of steroidogenesis, the conversion of cholesterol to pregnenolone. Previous studies describe an essential role for zebrafish Cyp11a1 during early development. Cyp11a1 has been suggested to be the functional equivalent of mammalian CYP11A1 in the zebrafish interrenal gland (equivalent to the mammalian adrenal), gonad, and brain. However, reported cyp11a1 expression is inconsistent in zebrafish larvae, after active cortisol synthesis commences. Recently a duplicated cyp11a gene, cyp11a2, has been described, which shares an 85% identity with cyp11a1. We aimed to elucidate the specific role of the two cyp11a paralogs. cyp11a1 was expressed from 0 to 48 hours post-fertilization (hpf), whereas cyp11a2 expression started after the development of the interrenal primordium (32 hpf) and was the only paralog in larvae. cyp11a2 is expressed in adult steroidogenic tissues, such as the interrenal, gonads, and brain. In contrast, cyp11a1 was mainly restricted to the gonads. Antisense morpholino knockdown studies confirmed abnormal gastrulation in cyp11a1 morphants. cyp11a2 morphants showed impaired steroidogenesis and a phenotype indicative of metabolic abnormalities. The phenotype was rescued by pregnenolone replacement in cyp11a2 morphants. Thus, we conclude that cyp11a1 is required for early development, whereas cyp11a2 is essential for the initiation and maintenance of zebrafish interrenal steroidogenesis. Importantly, this study highlights the need for a comprehensive characterization of steroidogenesis in zebrafish prior to its implementation as a model organism in translational research of adrenal disease.

  2. Temporally-controlled site-specific recombination in zebrafish.

    Science.gov (United States)

    Hans, Stefan; Kaslin, Jan; Freudenreich, Dorian; Brand, Michael

    2009-01-01

    Conventional use of the site-specific recombinase Cre is a powerful technology in mouse, but almost absent in other vertebrate model organisms. In zebrafish, Cre-mediated recombination efficiency was previously very low. Here we show that using transposon-mediated transgenesis, Cre is in fact highly efficient in this organism. Furthermore, temporal control of recombination can be achieved by using the ligand-inducible CreER(T2). Site-specific recombination only occurs upon administration of the drug tamoxifen (TAM) or its active metabolite, 4-hydroxy-tamoxifen (4-OHT). Cre-mediated recombination is detectable already 4 or 2 hours after administration of TAM or 4-OHT, demonstrating fast recombination kinetics. In addition, low doses of TAM allow mosaic labeling of single cells. Combined, our results show that conditional Cre/lox will be a valuable tool for both, embryonic and adult zebrafish studies. Furthermore, single copy insertion transgenesis of Cre/lox constructs suggest a strategy suitable also for other organisms.

  3. Temporally-controlled site-specific recombination in zebrafish.

    Directory of Open Access Journals (Sweden)

    Stefan Hans

    Full Text Available Conventional use of the site-specific recombinase Cre is a powerful technology in mouse, but almost absent in other vertebrate model organisms. In zebrafish, Cre-mediated recombination efficiency was previously very low. Here we show that using transposon-mediated transgenesis, Cre is in fact highly efficient in this organism. Furthermore, temporal control of recombination can be achieved by using the ligand-inducible CreER(T2. Site-specific recombination only occurs upon administration of the drug tamoxifen (TAM or its active metabolite, 4-hydroxy-tamoxifen (4-OHT. Cre-mediated recombination is detectable already 4 or 2 hours after administration of TAM or 4-OHT, demonstrating fast recombination kinetics. In addition, low doses of TAM allow mosaic labeling of single cells. Combined, our results show that conditional Cre/lox will be a valuable tool for both, embryonic and adult zebrafish studies. Furthermore, single copy insertion transgenesis of Cre/lox constructs suggest a strategy suitable also for other organisms.

  4. Developmental lead exposure causes startle response deficits in zebrafish.

    Science.gov (United States)

    Rice, Clinton; Ghorai, Jugal K; Zalewski, Kathryn; Weber, Daniel N

    2011-10-01

    Lead (Pb(2+)) exposure continues to be an important concern for fish populations. Research is required to assess the long-term behavioral effects of low-level concentrations of Pb(2+) and the physiological mechanisms that control those behaviors. Newly fertilized zebrafish embryos (max) and escape time. With increasing exposure concentrations, a larger number of larvae failed to respond to even the initial tap and, for those that did respond, ceased responding earlier than control larvae. These differences were more pronounced at a frequency of 4 taps/s. (2) Response to a visual stimulus: Fish, exposed as embryos (2-24 hpf) to Pb(2+) (0-10 μM) were tested as adults under low light conditions (≈ 60 μW/m(2)) for visual responses to a rotating black bar. Visual responses were significantly degraded at Pb(2+) concentrations of 30 nM. These data suggest that zebrafish are viable models for short- and long-term sensorimotor deficits induced by acute, low-level developmental Pb(2+) exposures.

  5. A novel zebrafish model to provide mechanistic insights into the inflammatory events in carrageenan-induced abdominal edema.

    Directory of Open Access Journals (Sweden)

    Shi-Ying Huang

    Full Text Available A suitable small animal model may help in the screening and evaluation of new drugs, especially those from natural products, which can be administered at lower dosages, fulfilling an urgent worldwide need. In this study, we explore whether zebrafish could be a model organism for carrageenan-induced abdominal edema. The research results showed that intraperitoneal (i.p. administration of 1.5% λ-carrageenan in a volume of 20 µL significantly increased abdominal edema in adult zebrafish. Levels of the proinflammatory proteins tumor necrosis factor-α (TNF-α and inducible nitric oxide synthase (iNOS were increased in carrageenan-injected adult zebrafish during the development of abdominal edema. An associated enhancement was also observed in the leukocyte marker, myeloperoxidase (MPO. To support these results, we further observed that i.p. methylprednisolone (MP; 1 µg, a positive control, significantly inhibited carrageenan-induced inflammation 24 h after carrageenan administration. Furthermore, i.p. pretreatment with either an anti-TNF-α antibody (1∶5 dilution in a volume of 20 µL or the iNOS-selective inhibitor aminoguanidine (AG; 1 µg inhibited carrageenan-induced abdominal edema in adult zebrafish. This new animal model is uncomplicated, easy to develop, and involves a straightforward inducement of inflammatory edema for the evaluation of small volumes of drugs or test compounds.

  6. Nuclear progesterone receptors are up-regulated by estrogens in neurons and radial glial progenitors in the brain of zebrafish.

    Directory of Open Access Journals (Sweden)

    Nicolas Diotel

    Full Text Available In rodents, there is increasing evidence that nuclear progesterone receptors are transiently expressed in many regions of the developing brain, notably outside the hypothalamus. This suggests that progesterone and/or its metabolites could be involved in functions not related to reproduction, particularly in neurodevelopment. In this context, the adult fish brain is of particular interest, as it exhibits constant growth and high neurogenic activity that is supported by radial glia progenitors. However, although synthesis of neuroprogestagens has been documented recently in the brain of zebrafish, information on the presence of progesterone receptors is very limited. In zebrafish, a single nuclear progesterone receptor (pgr has been cloned and characterized. Here, we demonstrate that this pgr is widely distributed in all regions of the zebrafish brain. Interestingly, we show that Pgr is strongly expressed in radial glial cells and more weakly in neurons. Finally, we present evidence, based on quantitative PCR and immunohistochemistry, that nuclear progesterone receptor mRNA and proteins are upregulated by estrogens in the brain of adult zebrafish. These data document for the first time the finding that radial glial cells are preferential targets for peripheral progestagens and/or neuroprogestagens. Given the crucial roles of radial glial cells in adult neurogenesis, the potential effects of progestagens on their activity and the fate of daughter cells require thorough investigation.

  7. Transient overexpression of adh8a increases allyl alcohol toxicity in zebrafish embryos.

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    Nils Klüver

    Full Text Available Fish embryos are widely used as an alternative model to study toxicity in vertebrates. Due to their complexity, embryos are believed to more resemble an adult organism than in vitro cellular models. However, concerns have been raised with respect to the embryo's metabolic capacity. We recently identified allyl alcohol, an industrial chemical, to be several orders of magnitude less toxic to zebrafish embryo than to adult zebrafish (embryo LC50 = 478 mg/L vs. fish LC50 = 0.28 mg/L. Reports on mammals have indicated that allyl alcohol requires activation by alcohol dehydrogenases (Adh to form the highly reactive and toxic metabolite acrolein, which shows similar toxicity in zebrafish embryos and adults. To identify if a limited metabolic capacity of embryos indeed can explain the low allyl alcohol sensitivity of zebrafish embryos, we compared the mRNA expression levels of Adh isoenzymes (adh5, adh8a, adh8b and adhfe1 during embryo development to that in adult fish. The greatest difference between embryo and adult fish was found for adh8a and adh8b expression. Therefore, we hypothesized that these genes might be required for allyl alcohol activation. Microinjection of adh8a, but not adh8b mRNA led to a significant increase of allyl alcohol toxicity in embryos similar to levels reported for adults (LC50 = 0.42 mg/L in adh8a mRNA-injected embryos. Furthermore, GC/MS analysis of adh8a-injected embryos indicated a significant decline of internal allyl alcohol concentrations from 0.23-58 ng/embryo to levels below the limit of detection (< 4.6 µg/L. Injection of neither adh8b nor gfp mRNA had an impact on internal allyl alcohol levels supporting that the increased allyl alcohol toxicity was mediated by an increase in its metabolization. These results underline the necessity to critically consider metabolic activation in the zebrafish embryo. As demonstrated here, mRNA injection is one useful approach to study the role of candidate enzymes

  8. Can Zebrafish be used to Identify Developmentally Neurotoxic Chemicals

    Science.gov (United States)

    Can Zebrafish be Used to Identify Developmentally Neurotoxic Chemicals? The U.S. Environmental Protection Agency is evaluating methods to screen and prioritize large numbers of chemicals for developmental neurotoxicity. We are exploring behavioral methods using zebrafish by desig...

  9. Neuroblastoma and Its Zebrafish Model.

    Science.gov (United States)

    Zhu, Shizhen; Thomas Look, A

    2016-01-01

    Neuroblastoma, an important developmental tumor arising in the peripheral sympathetic nervous system (PSNS), accounts for approximately 10 % of all cancer-related deaths in children. Recent genomic analyses have identified a spectrum of genetic alterations in this tumor. Amplification of the MYCN oncogene is found in 20 % of cases and is often accompanied by mutational activation of the ALK (anaplastic lymphoma kinase) gene, suggesting their cooperation in tumor initiation and spread. Understanding how complex genetic changes function together in oncogenesis has been a continuing and daunting task in cancer research. This challenge was addressed in neuroblastoma by generating a transgenic zebrafish model that overexpresses human MYCN and activated ALK in the PSNS, leading to tumors that closely resemble human neuroblastoma and new opportunities to probe the mechanisms that underlie the pathogenesis of this tumor. For example, coexpression of activated ALK with MYCN in this model triples the penetrance of neuroblastoma and markedly accelerates tumor onset, demonstrating the interaction of these modified genes in tumor development. Further, MYCN overexpression induces adrenal sympathetic neuroblast hyperplasia, blocks chromaffin cell differentiation, and ultimately triggers a developmentally-timed apoptotic response in the hyperplastic sympathoadrenal cells. In the context of MYCN overexpression, activated ALK provides prosurvival signals that block this apoptotic response, allowing continued expansion and oncogenic transformation of hyperplastic neuroblasts, thus promoting progression to neuroblastoma. This application of the zebrafish model illustrates its value in rational assessment of the multigenic changes that define neuroblastoma pathogenesis and points the way to future studies to identify novel targets for therapeutic intervention.

  10. Ultrastructural and biochemical analysis of epidermal xanthophores and dermal chromatophores of the teleost Sparus aurata.

    Science.gov (United States)

    Ferrer, C; Solano, F; Zuasti, A

    1999-04-01

    We have studied the pigmentary system of the teleost Sparus aurata skin by electron microscopy and chromatographic analysis. Under electron microscopy, we found the dermis to contain the three major types of recognized chromatophores: melanophores, xanthophores and iridophores. Melanophores were more abundant in the dorsal region, whereas the iridophores were more abundant in the ventral region. The most important discovery was that of epidermal xanthophores. Epidermal xanthophores were the only chromatophores in the epidermis, something only found in S aurata and in a teleost species living in the Antartic sea. In contrast, the biochemical analysis did not establish any special characteristics: we found pteridine and flavin pigments located mostly in the pigmented dorsal region. Riboflavin and pterin were two of the most abundant coloured pigment types, but other colourless pigments such as xanthopterin and isoxanthopterin were also detected.

  11. Ultrastructural study of the pituicytes in the pituitary gland of the teleost Diplodus sargus.

    Science.gov (United States)

    Ferrandino, Ida; Grimaldi, Maria Consiglio

    2008-01-31

    An electron microscopic study was performed on the pituitary gland of the Mediterranean teleost fish Diplodus sargus to analyse the morphological characteristics of the pituicytes. In this class of vertebrates, the pituicytes have, like other astroglial cells, a trophic and support function, but they may also play an active role in the release of neurohormones. Most of the pituicytes were of the Dark type. Their shape appeared irregular with long, thin processes protruding from the cellular body. The pituicytes protruded from the neurohypophysis as far as the adenohypophysis. Their cellular bodies were mainly located in the posterior neurohypophysis. In the adenohypophysis, pituicytic processes were intermingled with cells of the pars intermedia and pars distalis, though being more numerous in the former. These processes sometimes surrounded the whole adenohypophyseal cell. This provides further evidence for the possible role of the pituicytes in controlling the release of the pituitary hormones given that, in teleost fishes, there is no distinct portal system or true median eminence.

  12. Sexual phenotype differences in zic2 mRNA abundance in the preoptic area of a protogynous teleost, Thalassoma bifasciatum.

    Science.gov (United States)

    McCaffrey, Katherine; Hawkins, Mary Beth; Godwin, John

    2011-01-01

    The highly conserved members of the zic family of zinc-finger transcription factors are primarily known for their roles in embryonic signaling pathways and regulation of cellular proliferation and differentiation. This study describes sexual phenotype differences in abundances of zic2 mRNA in the preoptic area of the hypothalamus, a region strongly implicated in sexual behavior and function, in an adult teleost, Thalassoma bifasciatum. The bluehead wrasse (Thalassoma bifasciatum) is a valuable model for studying neuroendocrine processes because it displays two discrete male phenotypes, initial phase (IP) males and territorial, terminal phase (TP) males, and undergoes socially-controlled protogynous sex change. Previously generated microarray-based comparisons suggested that zic2 was upregulated in the brains of terminal phase males relative to initial phase males. To further explore this difference, we cloned a 727 bp sequence for neural zic2 from field-collected animals. Riboprobe-based in situ hybridization was employed to localize zic2 signal in adult bluehead brains and assess the relative abundance of brain zic2 mRNA across sexual phenotypes. We found zic2 mRNA expression was extremely abundant in the granular cells of the cerebellum and widespread in other brain regions including in the thalamus, hypothalamus, habenula, torus semicircularis, torus longitudinalis, medial longitudinal fascicle and telencephalic areas. Quantitative autoradiography and phosphorimaging showed zic2 mRNA hybridization signal in the preoptic area of the hypothalamus was significantly higher in terminal phase males relative to both initial phase males and females, and silver grain analysis confirmed this relationship between phenotypes. No significant difference in abundance was found in zic2 signal across phenotypes in the habenula, a brain region not implicated in the control of sexual behavior, or cerebellum.

  13. Sexual phenotype differences in zic2 mRNA abundance in the preoptic area of a protogynous teleost, Thalassoma bifasciatum.

    Directory of Open Access Journals (Sweden)

    Katherine McCaffrey

    Full Text Available The highly conserved members of the zic family of zinc-finger transcription factors are primarily known for their roles in embryonic signaling pathways and regulation of cellular proliferation and differentiation. This study describes sexual phenotype differences in abundances of zic2 mRNA in the preoptic area of the hypothalamus, a region strongly implicated in sexual behavior and function, in an adult teleost, Thalassoma bifasciatum. The bluehead wrasse (Thalassoma bifasciatum is a valuable model for studying neuroendocrine processes because it displays two discrete male phenotypes, initial phase (IP males and territorial, terminal phase (TP males, and undergoes socially-controlled protogynous sex change. Previously generated microarray-based comparisons suggested that zic2 was upregulated in the brains of terminal phase males relative to initial phase males. To further explore this difference, we cloned a 727 bp sequence for neural zic2 from field-collected animals. Riboprobe-based in situ hybridization was employed to localize zic2 signal in adult bluehead brains and assess the relative abundance of brain zic2 mRNA across sexual phenotypes. We found zic2 mRNA expression was extremely abundant in the granular cells of the cerebellum and widespread in other brain regions including in the thalamus, hypothalamus, habenula, torus semicircularis, torus longitudinalis, medial longitudinal fascicle and telencephalic areas. Quantitative autoradiography and phosphorimaging showed zic2 mRNA hybridization signal in the preoptic area of the hypothalamus was significantly higher in terminal phase males relative to both initial phase males and females, and silver grain analysis confirmed this relationship between phenotypes. No significant difference in abundance was found in zic2 signal across phenotypes in the habenula, a brain region not implicated in the control of sexual behavior, or cerebellum.

  14. Delta-8 desaturation activity varies among fatty acyl desaturases of teleost fish: high activity in delta-6 desaturases of marine species.

    Science.gov (United States)

    Monroig, Oscar; Li, Yuanyou; Tocher, Douglas R

    2011-08-01

    The benefits of dietary fish and fish oil are derived from n-3 long-chain polyunsaturated fatty acids (LC-PUFA) that have beneficial effects in a range of human diseases and pathologies such as cardiovascular and other inflammatory disorders, neural development and neurological pathologies. The precursor of n-3 LC-PUFA, 18:3n-3 does not have the same beneficial effects prompting interest in the pathways of endogenous synthesis of LC-PUFA in vertebrates. The LC-PUFA biosynthesis pathway classically involves Δ6 and Δ5 fatty acyl desaturases (Fad), but it was recently shown that Δ6 Fad in mammals also displayed Δ8 activity demonstrating a possible alternative "Δ8-pathway" for the synthesis of LC-PUFA. Our primary hypothesis was that Δ8 desaturase activity would be a common feature of vertebrate Δ6 Fads, and so the aim of the present study was to determine the ability of teleostei Fads for Δ8 desaturation activity. To this end, cDNAs for Fads from a range of freshwater, diadromous and marine teleost fish species were assayed for Δ8 activity in the heterologous yeast expression system. In summary, the present study has demonstrated that Δ8 desaturation activity was also a characteristic of fish orthologs, although the activity varied notably between freshwater/diadromous and marine fish species, with the latter possessing Fads2-like proteins with Δ8 activity far higher than mammalian FADS2. The data showed that, generally, the fish Fad are technically υ-3 desaturases, with new double bonds introduced 3C beyond a pre-existing double bond. However, the ability of zebrafish and rabbitfish Fads, previously characterised as Δ6/Δ5 bifunctional desaturases, to introduce non-methylene interrupted double bonds in 20:3n-3 and 20:2n-6 suggested that a novel combination of regioselectivity modes operates within these enzymes.

  15. Ultrastructural and biochemical analysis of epidermal xanthophores and dermal chromatophores of the teleost Sparus aurata

    OpenAIRE

    Ferrer, C.; Solano, F.; Zuasti, A

    1999-01-01

    We have studied the pigmentary system of the teleost Sparus aurata skin by electron microscopy and chromatographic analysis. Under electron microscopy, we found the dermis to contain the three major types of recognized chromatophores: melanophores, xanthophores and iridophores. Melanophores were more abundant in the dorsal region, whereas the iridophores were more abundant in the ventral region. The most important discovery was that of epidermal xanthophore...

  16. Salinity Regulates Claudin mRNA and Protein Expression in the Teleost Gill

    DEFF Research Database (Denmark)

    Tipsmark, Christian K; Baltzegar, David A; Ozden, Ozkan

    2008-01-01

    The teleost gill carries out NaCl uptake in fresh water (FW) and NaCl excretion in seawater (SW). This transformation with salinity requires close regulation of ion transporter capacity and epithelial permeability. This study investigates the regulation of tight junctional claudins during salinity...... with salinity acclimation and possibly the formation of deeper tight junctions in FW gill. This may reduce ion permeability, which is a critical facet of FW osmoregulation. Key words: osmoregulation, tight junction, tilapia, epithelia....

  17. Sex determination studies in two species of teleost fish, Oreochromis niloticus and Leporinus elongatus

    OpenAIRE

    Baroiller, Jean-Francois; Nakayama, Ichiro; FORESTI,FAUSTO; Chourrout, Daniel

    1996-01-01

    Genetic analyses of sex determination have identified sex chromosomes in many teleost fish species. However, there are several cases for which sex ratios do not fit perfectly with the expectations of heterogametic systems, suggesting the influence of either minor sex determining genes or environmental influences on the process of sex differentiation. The frequent absence of sex chromosome markers makes the identification of minor sex-determining genes very difficult. It is easier to test firs...

  18. One for all and all for one: the importance of shoaling on behavioral and stress responses in zebrafish.

    Science.gov (United States)

    Pagnussat, Natália; Piato, Angelo L; Schaefer, Isabel C; Blank, Martina; Tamborski, Angélica R; Guerim, Laura D; Bonan, Carla D; Vianna, Mônica R M; Lara, Diogo R

    2013-09-01

    Zebrafish has been increasingly used in behavioral studies, but data can present high variability. Most studies have been performed using isolated zebrafish, despite their interactive nature and shoaling behavior. We compared adult zebrafish behavior and cortisol levels after exposure to novelty as well as sensitivity to Pentylenetetrazole (PTZ)-induced seizures in animals tested individually or in groups of three (triplets). In the exploratory behavior task, data from single fish and triplets were not significantly different, but single fish data were more disperse in latency, to enter and time spent in the tank upper part, and crossings. In the light-dark task, time in the light zone and crossings were not different between groups, but latency to enter the dark zone and data variability were. We also observed that the latency to reach stage III seizures induced by PTZ was higher in triplets, but data dispersion was not different from single fish. Finally, cortisol levels of fish individually exposed to a novel environment were higher and more variable than triplets, while both groups had higher levels than unmanipulated animals. Thus, when tested individually, zebrafish are more stressed and present more variable behavior due to disruption of their natural shoal strategies. These features can be beneficial or detrimental depending on study aims and should be considered when designing, analyzing, and interpreting zebrafish behavioral data.

  19. Impairment of social behaviour persists two years after embryonic alcohol exposure in zebrafish: A model of fetal alcohol spectrum disorders.

    Science.gov (United States)

    Fernandes, Yohaan; Rampersad, Mindy; Gerlai, Robert

    2015-10-01

    Zebrafish naturally form social groups called shoals. Previously, we have shown that submerging zebrafish eggs into low concentrations of alcohol (0.00, 0.25, 0.50, 0.75 and 1.00 vol/vol% external bath concentration) during development (24h post-fertilization) for two hours resulted in impaired shoaling response in seven month old young adult zebrafish. Here we investigate whether this embryonic alcohol exposure induced behavioural deficit persists to older age. Zebrafish embryos were exposed either to fresh system water (control) or to 1% alcohol for two hours, 24h after fertilization, and were raised in a high-density tank system. Social behaviour was tested by presenting the experimental fish with a computer animated group of zebrafish images, while automated tracking software measured their behaviour. Control fish were found to respond strongly to animated conspecific images by reducing their distanceand remaining close to the images during image presentation, embryonic alcohol treated fish did not. Our results suggest that the impaired shoaling response of the alcohol exposed fish was not due to altered motor function or visual perception, but likely to a central nervous system alteration affecting social behaviour itself. We found the effects of embryonic alcohol exposure on social behaviour not to diminish with age, a result that demonstrates the deleterious and potentially life-long consequences of exposure to even small amount of alcohol during embryonic development in vertebrates.

  20. 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.

  1. Osmoregulatory actions of the GH/IGF axis in non-salmonid teleosts

    Science.gov (United States)

    Mancera, J.M.; McCormick, S.D.

    1998-01-01

    Salmonid fishes provided the first findings on the influence of the growth hormone (GH)/insulin-like growth factor I (IGF-I) axis on osmoregulation in teleost fishes. Recent studies on non-salmonid species, however, indicate that this physiological action of the GH/IGF-I axis is not restricted to salmonids or anadromous fishes. GH-producing cells in the pituitary of fish acclimated to different salinities show different degrees of activation depending on the species studied. Plasma GH levels either increase or do not change after transfer of fish from freshwater to seawater. Treatment with GH or IGF-I increases salinity tolerance and/or increases gill Na+,K+-ATPase activity of killifish (Fundulus heteroclitus), tilapia (Oreochromis mossambicus and Oreochromis niloticus) and striped bass (Morone saxatilis). As in salmonids, a positive interaction between GH and cortisol for improving hypoosmoregulatory capacity has been described in tilapia (O. mossambicus). Research on the osmoregulatory role of the GH/IGF-I axis is derived from a small number of teleost species. The study of more species with different osmoregulary patterns will be necessary to fully clarify the osmoregulatory role of GH/IGF-I axis in fish. The available data does suggest, however, that the influence of the GH/IGF-I axis on osmoregulation may be a common feature of euryhalinity in teleosts. Copyright (C) 1998 Elsevier Science Inc.

  2. Teleost fish osmoregulation: what have we learned since August Krogh, Homer Smith, and Ancel Keys.

    Science.gov (United States)

    Evans, David H

    2008-08-01

    In the 1930s, August Krogh, Homer Smith, and Ancel Keys knew that teleost fishes were hyperosmotic to fresh water and hyposmotic to seawater, and, therefore, they were potentially salt depleted and dehydrated, respectively. Their seminal studies demonstrated that freshwater teleosts extract NaCl from the environment, while marine teleosts ingest seawater, absorb intestinal water by absorbing NaCl, and excrete the excess salt via gill transport mechanisms. During the past 70 years, their research descendents have used chemical, radioisotopic, pharmacological, cellular, and molecular techniques to further characterize the gill transport mechanisms and begin to study the signaling molecules that modulate these processes. The cellular site for these transport pathways was first described by Keys and is now known as the mitochondrion-rich cell (MRC). The model for NaCl secretion by the marine MRC is well supported, but the model for NaCl uptake by freshwater MRC is more unsettled. Importantly, these ionic uptake mechanisms also appear to be expressed in the marine gill MRC, for acid-base regulation. A large suite of potential endocrine control mechanisms have been identified, and recent evidence suggests that paracrines such as endothelin, nitric oxide, and prostaglandins might also control MRC function.

  3. Cancer and inflammation studies using zebrafish cell lines

    NARCIS (Netherlands)

    He, Shuning

    2010-01-01

    As the zebrafish, Danio rerio, has been increasingly used as an animal model for biomedical research, we aimed to establish zebrafish cell line models for inflammation and cancer studies in this thesis. Several zebrafish cell lines were characterized and their genetic and physiological properties we

  4. Pten function in zebrafish : Anything but a fish story

    NARCIS (Netherlands)

    Stumpf, Miriam; Choorapoikayil, Suma; den Hertog, J.

    2015-01-01

    Zebrafish is an excellent model system for the analysis of gene function. We and others use zebrafish to investigate the function of the tumor suppressor, Pten, in tumorigenesis and embryonic development. Zebrafish have two pten genes, ptena and ptenb. The recently identified N-terminal extension of

  5. Swimming Against the Current: Zebrafish Help Address Educational Challenges.

    Science.gov (United States)

    Pickart, Michael A; Liang, Jennifer; Hutson, Lara; Pierret, Christopher

    2016-08-01

    Zebrafish can be important tools for learning and authentic student research. The broad zebrafish community is rich with examples to improve education for learners of all ages and geographical locales. This special collection of articles is presented with the hope of encouraging readers to reflect on the educational outcomes reported here and to consider new ways zebrafish may engage others to learn and grow.

  6. Sulpiride, but not SCH23390, modifies cocaine-induced conditioned place preference and expression of tyrosine hydroxylase and elongation factor 1α in zebrafish.

    Science.gov (United States)

    Darland, Tristan; Mauch, Justin T; Meier, Ellen M; Hagan, Shannon J; Dowling, John E; Darland, Diane C

    2012-12-01

    Finding genetic polymorphisms and mutations linked to addictive behavior can provide important targets for pharmaceutical and therapeutic interventions. Forward genetic approaches in model organisms such as zebrafish provide a potentially powerful avenue for finding new target genes. In order to validate this use of zebrafish, the molecular nature of its reward system must be characterized. We have previously reported the use of cocaine-induced conditioned place preference (CPP) as a reliable method for screening mutagenized fish for defects in the reward pathway. Here we test if CPP in zebrafish involves the dopaminergic system by co-treating fish with cocaine and dopaminergic antagonists. Sulpiride, a potent D2 receptor (DR2) antagonist, blocked cocaine-induced CPP, while the D1 receptor (DR1) antagonist SCH23390 had no effect. Acute cocaine exposure also induced a rise in the expression of tyrosine hydroxylase (TH), an important enzyme in dopamine synthesis, and a significant decrease in the expression of elongation factor 1α (EF1α), a housekeeping gene that regulates protein synthesis. Cocaine selectively increased the ratio of TH/EF1α in the telencephalon, but not in other brain regions. The cocaine-induced change in TH/EF1α was blocked by co-treatment with sulpiride, but not SCH23390, correlating closely with the action of these drugs on the CPP behavioral response. Immunohistochemical analysis revealed that the drop in EF1α was selective for the dorsal nucleus of the ventral telencephalic area (Vd), a region believed to be the teleost equivalent of the striatum. Examination of TH mRNA and EF1α transcripts suggests that regulation of expression is post-transcriptional, but this requires further examination. These results highlight important similarities and differences between zebrafish and more traditional mammalian model organisms.

  7. Rest mutant zebrafish swim erratically and display atypical spatial preferences.

    Science.gov (United States)

    Moravec, Cara E; Li, Edward; Maaswinkel, Hans; Kritzer, Mary F; Weng, Wei; Sirotkin, Howard I

    2015-05-01

    The Rest/Nrsf transcriptional repressor modulates expression of a large set of neural specific genes. Many of these target genes have well characterized roles in nervous system processes including development, plasticity and synaptogenesis. However, the impact of Rest-mediated transcriptional regulation on behavior has been understudied due in part to the embryonic lethality of the mouse knockout. To investigate the requirement for Rest in behavior, we employed the zebrafish rest mutant to explore a range of behaviors in adults and larva. Adult rest mutants of both sexes showed abnormal behaviors in a novel environment including increased vertical swimming, erratic swimming patterns and a proclivity for the tank walls. Adult males also had diminished reproductive success. At 6 days post fertilization (dpf), rest mutant larva were hypoactive, but displayed normal evoked responses to light and sound stimuli. Overall, these results provide evidence that rest dysfunction produces atypical swimming patterns and preferences in adults, and reduced locomotor activity in larvae. This study provides the first behavioral analysis of rest mutants and reveals specific behaviors that are modulated by Rest.

  8. Widespread roles of microRNAs during zebrafish development and beyond.

    Science.gov (United States)

    Mishima, Yuichiro

    2012-01-01

    MicroRNAs (miRNAs) are a class of small RNAs that are approximately 22 nucleotides in length. Hundreds of miRNA genes are encoded in the animal genome, and each miRNA potentially regulates tens to hundreds of protein-coding transcripts post-transcriptionally. Experimental and bioinformatic approaches have shown widespread regulatory roles for miRNAs in metazoa including roles in cellular homeostasis and human diseases. Since the discoveries of let-7 and lin-4 miRNAs as regulators of developmental timing in Caenorhabditis elegans, functions of miRNAs in the context of animal development have been studied in many model organisms. Although miRNAs are essential to achieve complex developmental processes, the vast majority of animal miRNA functions have yet to be determined. The identification of miRNA-target interactions and the interpretation of their biological significance are often difficult due to the divergent functions of miRNAs in intricate gene regulatory networks. This review summarizes our current knowledge on miRNA functions in vertebrate development by focusing on the progress made in the vertebrate model organism zebrafish (Danio rerio). Studies of miRNA functions in this small teleost highlight several common principles underlying the functions of animal miRNAs.

  9. Hh and Wnt signaling regulate formation of olig2+ neurons in the zebrafish cerebellum.

    Science.gov (United States)

    McFarland, Karen A; Topczewska, Jolanta M; Weidinger, Gilbert; Dorsky, Richard I; Appel, Bruce

    2008-06-01

    The cerebellum, which forms from anterior hindbrain, coordinates motor movements and balance. Sensory input from the periphery is relayed and modulated by cerebellar interneurons, which are organized in layers. The mechanisms that specify the different neurons of the cerebellum and direct its layered organization remain poorly understood. Drawing from investigations of spinal cord, we hypothesized that the embryonic cerebellum is patterned on the dorsoventral axis by opposing morphogens. We tested this using zebrafish. Here we show that expression of olig2, which encodes a bHLH transcription factor, marks a distinct subset of neurons with similarities to eurydendroid neurons, the principal efferent neurons of the teleost cerebellum. In combination with other markers, olig2 reveals a dorsoventral organization of cerebellar neurons in embryos. Disruption of Hedgehog signaling, which patterns the ventral neural tube, produced a two-fold increase in the number of olig2(+) neurons. By contrast, olig2(+) neurons did not develop in embryos deficient for Wnt signaling, which patterns dorsal neural tube, nor did they develop in embryos deficient for both Hedgehog and Wnt signaling. Our data indicate that Hedgehog and Wnt work in opposition across the dorsoventral axis of the cerebellum to regulate formation of olig2(+) neurons. Specifically, we propose that Hedgehog limits the range of Wnt signaling, which is necessary for olig2(+) neuron development.

  10. Selenium status affects selenoprotein expression, reproduction, and F₁ generation locomotor activity in zebrafish (Danio rerio).

    Science.gov (United States)

    Penglase, Sam; Hamre, Kristin; Rasinger, Josef D; Ellingsen, Staale

    2014-06-14

    Se is an essential trace element, and is incorporated into selenoproteins which play important roles in human health. Mammalian selenoprotein-coding genes are often present as paralogues in teleost fish, and it is unclear whether the expression patterns or functions of these fish paralogues reflect their mammalian orthologues. Using the model species zebrafish (Danio rerio; ZF), we aimed to assess how dietary Se affects key parameters in Se metabolism and utilisation including glutathione peroxidase (GPX) activity, the mRNA expression of key Se-dependent proteins (gpx1a, gpx1b, sepp1a and sepp1b), oxidative status, reproductive success and F1 generation locomotor activity. From 27 d until 254 d post-fertilisation, ZF were fed diets with graded levels of Se ranging from deficient ( levels were lowest when dietary Se levels (0·3 mg/kg) resulted in the maximum growth of ZF, and a proposed bimodal mechanism in response to Se status below and above this dietary Se level was identified. The expression of the sepp1 paralogues differed, with only sepp1a responding to Se status. High dietary Se supplementation (30 mg/kg) decreased reproductive success, while the offspring of ZF fed above 0·3 mg Se/kg diet had lower locomotor activity than the other groups. Overall, the novel finding of low selenoprotein expression and activity coinciding with maximum body growth suggests that even small Se-induced variations in redox status may influence cellular growth rates.

  11. Developmental toxicity of cypermethrin in embryo-larval stages of zebrafish.

    Science.gov (United States)

    Shi, Xiangguo; Gu, Aihua; Ji, Guixiang; Li, Yuan; Di, Jing; Jin, Jing; Hu, Fan; Long, Yan; Xia, Yankai; Lu, Chuncheng; Song, Ling; Wang, Shoulin; Wang, Xinru

    2011-10-01

    Cypermethrin, a type II pyrethroid insecticide, is widely used throughout the world in agriculture, forestry, horticulture and homes. Though the neurotoxicity of cypermethrin has been thoroughly studied in adult rodents, little is so far available regarding the developmental toxicity of cypermethrin to fish in early life stages. To explore the potential developmental toxicity of cypermethrin, 4-h post-fertilization (hpf) zebrafish embryos were exposed to various concentrations of cypermethrin (0, 25, 50, 100, 200 and 400 μg L⁻¹) until 96 h. Among a suite of morphological abnormalities, the unique phenotype curvature was observed at concentrations as low as 25 μg L⁻¹. Studies revealed that 400 μg L⁻¹ cypermethrin significantly increased malondialdehyde production. In addition, activity of antioxidative enzymes including superoxide dismutase and catalase were significantly induced in zebrafish larvae in a concentration-dependent manner. To further investigate the toxic effects of cypermethrin on fish, acridine orange (AO) staining was performed at 400 μg L⁻¹ cypermethrin and the result showed notable signs of apoptosis mainly in the nervous system. Cypermethrin also down-regulated ogg1 and increased p53 gene expression as well as the caspase-3 activity. Our results demonstrate that cypermethrin was able to induce oxidative stress and produce apoptosis through the involvement of caspases in zebrafish embryos. In this study, we investigated the developmental toxicity of cypermethrin using zebrafish embryos, which could be helpful in fully understanding the potential mechanisms of cypermethrin exposure during embryogenesis and also suggested that zebrafish could serve as an ideal model for studying developmental toxicity of environmental contaminants.

  12. Copper acutely impairs behavioral function and muscle acetylcholinesterase activity in zebrafish (Danio rerio).

    Science.gov (United States)

    Haverroth, Gabriela M B; Welang, Chariane; Mocelin, Riciéri N; Postay, Daniela; Bertoncello, Kanandra T; Franscescon, Francini; Rosemberg, Denis B; Dal Magro, Jacir; Dalla Corte, Cristiane L

    2015-12-01

    Copper is a heavy metal found at relatively high concentrations in surface waters around the world. Copper is a micronutrient at low concentrations and is essential to several organisms. At higher concentrations copper can become toxic, which reveal the importance of studying the toxic effects of this metal on the aquatic life. Thus, the objective of this study was to evaluate the toxic effects of copper on the behavior and biochemical parameters of zebrafish (Danio rerio). Zebrafish were exposed for 24h at a concentration of 0.006 mg/L Cu. After the exposure period, behavioral profile of animals was recorded through 6 min using two different apparatuses tests: the Novel Tank and the Light-Dark test. After behavioral testing, animals were euthanized with a solution of 250 mg/L of tricaine (MS-222). Brain, muscle, liver and gills were extracted for analysis of parameters related to oxidative stress and accumulation of copper in these tissues. Acetylcholinesterase (AChE) activity was determined in brain and muscle. Results showed acute exposure to copper induces significant changes in behavioral profile of zebrafish by changing locomotion and natural tendency to avoid brightly lit area. On the other hand, there were no significant effects on parameters related to oxidative stress. AChE activity decreased significantly in zebrafish muscle, but there were no significant changes in cerebral AChE activity. Copper levels in tissues did not increase significantly compared to the controls. Taken together, these results indicate that a low concentration of copper can acutely affect behavioral profile of adult zebrafish which could be partially related to an inhibition on muscle AChE activity. These results reinforce the need of additional tests to establishment of safe copper concentrations to aquatic organisms and the importance of behavioral parameters in ecotoxicological studies.

  13. Prolonged hypoxia increases survival even in Zebrafish (Danio rerio showing cardiac arrhythmia.

    Directory of Open Access Journals (Sweden)

    Renate Kopp

    Full Text Available Tolerance towards hypoxia is highly pronounced in zebrafish. In this study even beneficial effects of hypoxia, specifically enhanced survival of zebrafish larvae, could be demonstrated. This effect was actually more pronounced in breakdance mutants, which phenotypically show cardiac arrhythmia. Breakdance mutants (bre are characterized by chronically reduced cardiac output. Despite an about 50% heart rate reduction, they become adults, but survival rate significantly drops to 40%. Normoxic bre animals demonstrate increased hypoxia inducible factor 1 a (Hif-1α expression, which indicates an activated hypoxic signaling pathway. Consequently, cardiovascular acclimation, like cardiac hypertrophy and increased erythrocyte concentration, occurs. Thus, it was hypothesized, that under hypoxic conditions survival might be even more reduced. When bre mutants were exposed to hypoxic conditions, they surprisingly showed higher survival rates than under normoxic conditions and even reached wildtype values. In hypoxic wildtype zebrafish, survival yet exceeded normoxic control values. To specify physiological acclimation, cardiovascular and metabolic parameters were measured before hypoxia started (3 dpf, when the first differences in survival rate occurred (7 dpf and when survival rate plateaued (15 dpf. Hypoxic animals expectedly demonstrated Hif-1α accumulation and consequently enhanced convective oxygen carrying capacity. Moreover, bre animals showed a significantly enhanced heart rate under hypoxic conditions, which reached normoxic wildtype values. This improvement in convective oxygen transport ensured a sufficient oxygen and nutrient supply and was also reflected in the significantly higher mitochondrial activity. The highly optimized energy metabolism observed in hypoxic zebrafish larvae might be decisive for periods of higher energy demand due to organ development, growth and increased activity. However, hypoxia increased survival only during a

  14. Metal uptake and acute toxicity in zebrafish: Common mechanisms across multiple metals

    Energy Technology Data Exchange (ETDEWEB)

    Alsop, Derek, E-mail: alsopde@mcmaster.ca [Department of Biology, McMaster University, 1280 Main St. W., Hamilton, ON L8S 4K1 (Canada); Wood, Chris M. [Department of Biology, McMaster University, 1280 Main St. W., Hamilton, ON L8S 4K1 (Canada)

    2011-10-15

    All metals tested reduced calcium uptake in zebrafish larvae. However, it was whole body sodium loss that was functionally related to toxicity. The zebrafish larvae acute toxicity assay save time, space and resources. - Abstract: Zebrafish larvae (Danio rerio) were used to examine the mechanisms of action and acute toxicities of metals. Larvae had similar physiological responses and sensitivities to waterborne metals as adults. While cadmium and zinc have previously been shown to reduce Ca{sup 2+} uptake, copper and nickel also decreased Ca{sup 2+} uptake, suggesting that the epithelial transport of all these metals is through Ca{sup 2+} pathways. However, exposure to cadmium, copper or nickel for up to 48 h had little or no effect on total whole body Ca{sup 2+} levels, indicating that the reduction of Ca{sup 2+} uptake is not the acute toxic mechanism of these metals. Instead, mortalities were effectively related to whole body Na{sup +}, which decreased up to 39% after 48 h exposures to different metals around their respective 96 h LC50s. Decreases in whole body K{sup +} were also observed, although they were not as pronounced or frequent as Na{sup +} losses. None of the metals tested inhibited Na{sup +} uptake in zebrafish (Na{sup +} uptake was in fact increased with exposure) and the observed losses of Na{sup +}, K{sup +}, Ca{sup 2+} and Mg{sup 2+} were proportional to the ionic gradients between the plasma and water, indicating diffusive ion loss with metal exposure. This study has shown that there is a common pathway for metal uptake and a common mechanism of acute toxicity across groups of metals in zebrafish. The disruption of ion uptake accompanying metal exposure does not appear to be responsible for the acute toxicity of metals, as has been previously suggested, but rather the toxicity is instead due to total ion loss (predominantly Na{sup +}).

  15. Expression patterns of lgr4 and lgr6 during zebrafish development.

    Science.gov (United States)

    Hirose, Kentaro; Shimoda, Nobuyoshi; Kikuchi, Yutaka

    2011-10-01

    Leucine-rich repeat (LRR)-containing G protein-coupled receptors (LGRs) belong to the superfamily of G protein-coupled receptors, and are characterized by the presence of seven transmembrane domains and an extracellular domain that contains a series of LRR motifs. Three Lgr proteins - Lgr4, Lgr5, and Lgr6 - were identified as members of the LGR subfamily. Mouse Lgr4 has been implicated in the formation of various organs through regulation of cell proliferation during development, and Lgr5 and Lgr6 are stem cell markers in the intestine or skin. Although the expression of these three genes has already been characterized in adult mice, their expression profiles during the embryonic and larval development of the organism have not yet been defined. We cloned two zebrafish lgr genes using the zebrafish genomic database. Phylogenetic analyses showed that these two genes are orthologs of mammalian Lgr4 and Lgr6. Zebrafish lgr4 is expressed in the neural plate border, Kupffer's vesicle, neural tube, otic vesicles, midbrain, eyes, forebrain, and brain ventricular zone by 24h post-fertilization (hpf). From 36 to 96hpf, lgr4 expression is detected in the midbrain-hindbrain boundary, otic vesicles, pharyngeal arches, cranial cartilages such as Meckel's cartilages, palatoquadrates, and ceratohyals, cranial cavity, pectoral fin buds, brain ventricular zone, ciliary marginal zone, and digestive organs such as the intestine, liver, and pancreas. In contrast, zebrafish lgr6 is expressed in the notochord, Kupffer's vesicle, the most anterior region of diencephalon, otic vesicles, and the anterior and posterior lateral line primordia by 24hpf. From 48 to 72hpf, lgr6 expression is confined to the anterior and posterior neuromasts, otic vesicles, pharyngeal arches, pectoral fin buds, and cranial cartilages such as Meckel's cartilages, ceratohyals, and trabeculae. Our results provide a basis for future studies aimed at analyzing the functions of zebrafish Lgr4 and Lgr6 in cell