Endogenous retinal neural stem cell reprogramming for neuronal regeneration

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Romain Madelaine

2017-01-01

Full Text Available In humans, optic nerve injuries and associated neurodegenerative diseases are often followed by permanent vision loss. Consequently, an important challenge is to develop safe and effective methods to replace retinal neurons and thereby restore neuronal functions and vision. Identifying cellular and molecular mechanisms allowing to replace damaged neurons is a major goal for basic and translational research in regenerative medicine. Contrary to mammals, the zebrafish has the capacity to fully regenerate entire parts of the nervous system, including retina. This regenerative process depends on endogenous retinal neural stem cells, the Müller glial cells. Following injury, zebrafish Müller cells go back into cell cycle to proliferate and generate new neurons, while mammalian Müller cells undergo reactive gliosis. Recently, transcription factors and microRNAs have been identified to control the formation of new neurons derived from zebrafish and mammalian Müller cells, indicating that cellular reprogramming can be an efficient strategy to regenerate human retinal neurons. Here we discuss recent insights into the use of endogenous neural stem cell reprogramming for neuronal regeneration, differences between zebrafish and mammalian Müller cells, and the need to pursue the identification and characterization of new molecular factors with an instructive and potent function in order to develop theurapeutic strategies for eye diseases.

Small GTPase R-Ras participates in neural tube formation in zebrafish embryonic spinal cord.

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Ohata, Shinya; Uga, Hideko; Okamoto, Hitoshi; Katada, Toshiaki

2018-06-27

Ras related (R-Ras), a small GTPase, is involved in the maintenance of apico-basal polarity in neuroepithelial cells of the zebrafish hindbrain, axonal collapse in cultured murine hippocampal neurons, and maturation of blood vessels in adult mice. However, the role of R-Ras in neural tube formation remains unknown. Using antisense morpholino oligonucleotides (AMOs), we found that in the spinal cord of zebrafish embryos, the lumen was formed bilaterally in rras morphants, whereas it was formed at the midline in control embryos. As AMO can cause off-target effects, we generated rras mutant zebrafish lines using CRISPR/Cas9 technology. Although these rras mutant embryos did not have a bilateral lumen in the spinal cord, the following findings suggest that the phenotype is unlikely due to an off-target effect of rras AMO: 1) The rras morphant phenotype was rescued by an injection of AMO-resistant rras mRNA, and 2) a bilaterally segregated spinal cord was not observed in rras mutant embryos injected with rras AMO. The results suggest that the function of other ras family genes may be redundant in rras mutants. Previous research reported a bilaterally formed lumen in the spinal cord of zebrafish embryos with a mutation in a planar cell polarity (PCP) gene, van gogh-like 2 (vangl2). In the present study, in cultured cells, R-Ras was co-immunoprecipitated with Vangl2 but not with another PCP regulator, Pricke1. Interestingly, the interaction between R-Ras and Vangl2 was stronger in guanine-nucleotide free point mutants of R-Ras than in wild-type or constitutively active (GTP-bound) forms of R-Ras. R-Ras may regulate neural tube formation in cooperation with Vangl2 in the developing zebrafish spinal cord. Copyright © 2018 Elsevier Inc. All rights reserved.

Genetic determinants of hyaloid and retinal vasculature in zebrafish

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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.

A panel of recombinant monoclonal antibodies against zebrafish neural receptors and secreted proteins suitable for wholemount immunostaining.

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Staudt, Nicole; Müller-Sienerth, Nicole; Fane-Dremucheva, Alla; Yusaf, Shahnaz P; Millrine, David; Wright, Gavin J

2015-01-02

Cell surface receptors and secreted proteins play important roles in neural recognition processes, but because their site of action can be a long distance from neuron cell bodies, antibodies that label these proteins are valuable to understand their function. The zebrafish embryo is a popular vertebrate model for neurobiology, but suffers from a paucity of validated antibody reagents. Here, we use the entire ectodomain of neural zebrafish cell surface or secreted proteins expressed in mammalian cells to select monoclonal antibodies to ten different antigens. The antibodies were characterised by Western blotting and the sensitivity of their epitopes to formalin fixation was determined. The rearranged antigen binding regions of the antibodies were amplified and cloned which enabled expression in a recombinant form from a single plasmid. All ten antibodies gave specific staining patterns within formalin-treated embryonic zebrafish brains, demonstrating that this generalised approach is particularly efficient to elicit antibodies that stain native antigen in fixed wholemount tissue. Finally, we show that additional tags can be easily added to the recombinant antibodies for convenient multiplex staining. The antibodies and the approaches described here will help to address the lack of well-defined antibody reagents in zebrafish research. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Knockdown of Zebrafish Blood Vessel Epicardial Substance Results in Incomplete Retinal Lamination

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Yu-Ching Wu

2014-01-01

Full Text Available Cell polarity during eye development determines the normal retinal lamination and differentiation of photoreceptor cells in the retina. In vertebrates, blood vessel epicardial substance (Bves is known to play an important role in the formation and maintenance of the tight junctions essential for epithelial cell polarity. In the current study, we generated a transgenic zebrafish Bves (zbves promoter-EGFP zebrafish line to investigate the expression pattern of Bves in the retina and to study the role of zbves in retinal lamination. Immunostaining with different specific antibodies from retinal cells and transmission electron microscopy were used to identify the morphological defects in normal and Bves knockdown zebrafish. In normal zebrafish, Bves is located at the apical junctions of embryonic retinal neuroepithelia during retinogenesis; later, it is strongly expressed around inner plexiform layer (IPL and retinal pigment epithelium (RPE. In contrast, a loss of normal retinal lamination and cellular polarity was found with undifferentiated photoreceptor cells in Bves knockdown zebrafish. Herein, our results indicated that disruption of Bves will result in a loss of normal retinal lamination.

Knocking Down Snrnp200 Initiates Demorphogenesis of Rod Photoreceptors in Zebrafish

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Yuan Liu

2015-01-01

Full Text Available Purpose. The small nuclear ribonucleoprotein 200 kDa (SNRNP200 gene is a fundamental component for precursor message RNA (pre-mRNA splicing and has been implicated in the etiology of autosomal dominant retinitis pigmentosa (adRP. This study aims to determine the consequences of knocking down Snrnp200 in zebrafish. Methods. Expression of the Snrnp200 transcript in zebrafish was determined via whole mount in situ hybridization. Morpholino oligonucleotide (MO aiming to knock down the expression of Snrnp200 was injected into zebrafish embryos, followed by analyses of aberrant splicing and expression of the U4/U6-U5 tri-small nuclear ribonucleoproteins (snRNPs components and retina-specific transcripts. Systemic changes and retinal phenotypes were further characterized by histological study and immunofluorescence staining. Results. Snrnp200 was ubiquitously expressed in zebrafish. Knocking down Snrnp200 in zebrafish triggered aberrant splicing of the cbln1 gene, upregulation of other U4/U6-U5 tri-snRNP components, and downregulation of a panel of retina-specific transcripts. Systemic defects were found correlated with knockdown of Snrnp200 in zebrafish. Only demorphogenesis of rod photoreceptors was detected in the initial stage, mimicking the disease characteristics of RP. Conclusions. We conclude that knocking down Snrnp200 in zebrafish could alter regular splicing and expression of a panel of genes, which may eventually trigger rod defects.

A novel transgenic zebrafish model for blood-brain and blood-retinal barrier development

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Sugimoto Masahiko

2010-07-01

Full Text Available Abstract Background Development and maintenance of the blood-brain and blood-retinal barrier is critical for the homeostasis of brain and retinal tissue. Despite decades of research our knowledge of the formation and maintenance of the blood-brain (BBB and blood-retinal (BRB barrier is very limited. We have established an in vivo model to study the development and maintenance of these barriers by generating a transgenic zebrafish line that expresses a vitamin D-binding protein fused with enhanced green fluorescent protein (DBP-EGFP in blood plasma, as an endogenous tracer. Results The temporal establishment of the BBB and BRB was examined using this transgenic line and the results were compared with that obtained by injection of fluorescent dyes into the sinus venosus of embryos at various stages of development. We also examined the expression of claudin-5, a component of tight junctions during the first 4 days of development. We observed that the BBB of zebrafish starts to develop by 3 dpf, with expression of claudin-5 in the central arteries preceding it at 2 dpf. The hyaloid vasculature in the zebrafish retina develops a barrier function at 3 dpf, which endows the zebrafish with unique advantages for studying the BRB. Conclusion Zebrafish embryos develop BBB and BRB function simultaneously by 3 dpf, which is regulated by tight junction proteins. The Tg(l-fabp:DBP-EGFP zebrafish will have great advantages in studying development and maintenance of the blood-neural barrier, which is a new application for the widely used vertebrate model.

Up-regulation of DRP-3 long isoform during the induction of neural progenitor cells by glutamate treatment in the ex vivo rat retina

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Tokuda, Kazuhiro, E-mail: r502um@yamaguchi-u.ac.jp [Department of Ophthalmology, Yamaguchi University Graduate School of Medicine, Ube, Yamaguchi (Japan); Department of Biochemistry and Functional Proteomics, Yamaguchi University Graduate School of Medicine, Ube, Yamaguchi (Japan); Kuramitsu, Yasuhiro; Byron, Baron; Kitagawa, Takao [Department of Biochemistry and Functional Proteomics, Yamaguchi University Graduate School of Medicine, Ube, Yamaguchi (Japan); Tokuda, Nobuko [Faculty of Health Sciences, Yamaguchi University Graduate School of Medicine, Ube (Japan); Kobayashi, Daiki; Nagayama, Megumi; Araki, Norie [Department of Tumor Genetics and Biology, Graduate School of Medical Sciences, Kumamoto University, Kumamoto (Japan); Sonoda, Koh-Hei [Department of Ophthalmology, Yamaguchi University Graduate School of Medicine, Ube, Yamaguchi (Japan); Nakamura, Kazuyuki [Department of Biochemistry and Functional Proteomics, Yamaguchi University Graduate School of Medicine, Ube, Yamaguchi (Japan)

2015-08-07

Glutamate has been shown to induce neural progenitor cells in the adult vertebrate retina. However, protein dynamics during progenitor cell induction by glutamate are not fully understood. To identify specific proteins involved in the process, we employed two-dimensional electrophoresis-based proteomics on glutamate untreated and treated retinal ex vivo sections. Rat retinal tissues were incubated with 1 mM glutamate for 1 h, followed by incubation in glutamate-free media for a total of 24 h. Consistent with prior reports, it was found that mitotic cells appeared in the outer nuclear layer without any histological damage. Immunohistological evaluations and immunoblotting confirmed the emergence of neuronal progenitor cells in the mature retina treated with glutamate. Proteomic analysis revealed the up-regulation of dihydropyrimidinase-related protein 3 (DRP-3), DRP-2 and stress-induced-phosphoprotein 1 (STIP1) during neural progenitor cell induction by glutamate. Moreover, mRNA expression of DRP-3, especially, its long isoform, robustly increased in the treated retina compared to that in the untreated retina. These results may indicate that glutamate induces neural progenitor cells in the mature rat retina by up-regulating the proteins which mediate cell mitosis and neurite growth. - Highlights: • Glutamate induced neuronal progenitor cells in the mature rat retina. • Proteomic analysis revealed the up-regulation of DRP-3, DRP-2 and STIP1. • mRNA expression of DRP-3, especially, its long isoform, robustly increased.

Time course Analysis of Gene expression patterns in ZebrafIsh Eye during Optic Nerve Regeneration

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Amy T. Mccurley

2010-01-01

Full Text Available It is well-established that neurons in the adult mammalian central nervous system (CNS are terminally differentiated and, if injured, will be unable to regenerate their connections. In contrast to mammals, zebrafish and other teleosts display a robust neuroregenerative response. Following optic nerve crush (ONX, retinal ganglion cells (RGC regrow their axons to synapse with topographically correct targets in the optic tectum, such that vision is restored in ~21 days. What accounts for these differences between teleostean and mammalian responses to neural injury is not fully understood. A time course analysis of global gene expression patterns in the zebrafish eye after ONX can help to elucidate cellular and molecular mechanisms that contribute to a successful neuroregeneration. To define different phases of regeneration after ONX, alpha tubulin 1 ( tuba1 and growth-associated protein 43 ( gap43 , markers previously shown to correspond to morphophological events, were measured by real time quantitative PCR (qPCR. Microarray analysis was then performed at defined intervals (6 hours, 1, 4, 12, and 21 days post-ONX and compared to SHAM. Results show that optic nerve damage induces multiple, phase-related transcriptional programs, with the maximum number of genes changed and highest fold-change occurring at 4 days. Several functional groups affected by optic nerve regeneration, including cell adhesion, apoptosis, cell cycle, energy metabolism, ion channel activity, and calcium signaling, were identified. Utilizing the whole eye allowed us to identify signaling contributions from the vitreous, immune and glial cells as well as the neural cells of the retina. Comparisons between our dataset and transcriptional profiles from other models of regeneration in zebrafish retina, heart and fin revealed a subset of commonly regulated transcripts, indicating shared mechanisms in different regenerating tissues. Knowledge of gene expression patterns in all

Neural retina-specific Aldh1a1 controls dorsal choroidal vascular development via Sox9 expression in retinal pigment epithelial cells.

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Goto, So; Onishi, Akishi; Misaki, Kazuyo; Yonemura, Shigenobu; Sugita, Sunao; Ito, Hiromi; Ohigashi, Yoko; Ema, Masatsugu; Sakaguchi, Hirokazu; Nishida, Kohji; Takahashi, Masayo

2018-04-03

VEGF secreted from retinal pigment epithelial (RPE) cells is responsible for the choroidal vascular development; however, the molecular regulatory mechanism is unclear. We found that Aldh1a1 -/- mice showed choroidal hypoplasia with insufficient vascularization in the dorsal region, although Aldh1a1, an enzyme that synthesizes retinoic acids (RAs), is expressed in the dorsal neural retina, not in the RPE/choroid complex. The level of VEGF in the RPE/choroid was significantly decreased in Aldh1a1 -/- mice, and RA-dependent enhancement of VEGF was observed in primary RPE cells. An RA-deficient diet resulted in dorsal choroidal hypoplasia, and simple RA treatment of Aldh1a1 -/- pregnant females suppressed choroid hypoplasia in their offspring. We also found downregulation of Sox9 in the dorsal neural retina and RPE of Aldh1a1 -/- mice and RPE-specific disruption of Sox9 phenocopied Aldh1a1 -/- choroidal development. These results suggest that RAs produced by Aldh1a1 in the neural retina directs dorsal choroidal vascular development via Sox9 upregulation in the dorsal RPE cells to enhance RPE-derived VEGF secretion. © 2018, Goto et al.

Histological Characterization of the Dicer1 Mutant Zebrafish Retina

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Saeed Akhtar

2015-01-01

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

Phenothiourea sensitizes zebrafish cranial neural crest and extraocular muscle development to changes in retinoic acid and IGF signaling.

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Brenda L Bohnsack

Full Text Available 1-Phenyl 2-thiourea (PTU is a tyrosinase inhibitor commonly used to block pigmentation and aid visualization of zebrafish development. At the standard concentration of 0.003% (200 µM, PTU inhibits melanogenesis and reportedly has minimal other effects on zebrafish embryogenesis. We found that 0.003% PTU altered retinoic acid and insulin-like growth factor (IGF regulation of neural crest and mesodermal components of craniofacial development. Reduction of retinoic acid synthesis by the pan-aldehyde dehydrogenase inhibitor diethylbenzaldehyde, only when combined with 0.003% PTU, resulted in extraocular muscle disorganization. PTU also decreased retinoic acid-induced teratogenic effects on pharyngeal arch and jaw cartilage despite morphologically normal appearing PTU-treated controls. Furthermore, 0.003% PTU in combination with inhibition of IGF signaling through either morpholino knockdown or pharmacologic inhibition of tyrosine kinase receptor phosphorylation, disrupted jaw development and extraocular muscle organization. PTU in and of itself inhibited neural crest development at higher concentrations (0.03% and had the greatest inhibitory effect when added prior to 22 hours post fertilization (hpf. Addition of 0.003% PTU between 4 and 20 hpf decreased thyroxine (T4 in thyroid follicles in the nasopharynx of 96 hpf embryos. Treatment with exogenous triiodothyronine (T3 and T4 improved, but did not completely rescue, PTU-induced neural crest defects. Thus, PTU should be used with caution when studying zebrafish embryogenesis as it alters the threshold of different signaling pathways important during craniofacial development. The effects of PTU on neural crest development are partially caused by thyroid hormone signaling.

Skeletogenic fate of zebrafish cranial and trunk neural crest.

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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.

cables1 Is Required for Embryonic Neural Development: Molecular, Cellular, and Behavioral Evidence From the Zebrafish

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GROENEWEG, JOLIJN W.; WHITE, YVONNE A.R.; KOKEL, DAVID; PETERSON, RANDALL T.; ZUKERBERG, LAWRENCE R.; BERIN, INNA; RUEDA, BO R.; WOOD, ANTONY W.

2014-01-01

SUMMARY In vitro studies have suggested that the Cables1 gene regulates epithelial cell proliferation, whereas other studies suggest a role in promoting neural differentiation. In efforts to clarify the functions of Cables1 in vivo, we conducted gain- and loss-of-function studies targeting its ortholog (cables1) in the zebrafish embryo. Similar to rodents, zebrafish cables1 mRNA expression is detected most robustly in embryonic neural tissues. Antisense knockdown of cables1 leads to increased numbers of apoptotic cells, particularly in brain tissue, in addition to a distinct behavioral phenotype, characterized by hyperactivity in response to stimulation. Apoptosis and the behavioral abnormality could be rescued by co-expression of a morpholino-resistant cables1 construct. Suppression of p53 expression in cables1 morphants partially rescued both apoptosis and the behavioral phenotype, suggesting that the phenotype of cables1 morphants is due in part to p53-dependent apoptosis. Alterations in the expression patterns of several neural transcription factors were observed in cables1 morphants during early neurulation, suggesting that cables1 is required for early neural differentiation. Ectopic overexpression of cables1 strongly disrupted embryonic morphogenesis, while overexpression of a cables1 mutant lacking the C-terminal cyclin box had little effect, suggesting functional importance of the cyclin box. Lastly, marked reductions in p35, but not Cdk5, were observed in cables1 morphants. Collectively, these data suggest that cables1 is important for neural differentiation during embryogenesis, in a mechanism that likely involves interactions with the Cdk5/p35 kinase pathway. PMID:21268180

The cone-dominant retina and the inner ear of zebrafish express the ortholog of CLRN1, the causative gene of human Usher syndrome type 3A.

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Phillips, Jennifer B; Västinsalo, Hanna; Wegner, Jeremy; Clément, Aurélie; Sankila, Eeva-Marja; Westerfield, Monte

2013-12-01

Clarin-1 (CLRN1) is the causative gene in Usher syndrome type 3A, an autosomal recessive disorder characterized by progressive vision and hearing loss. CLRN1 encodes Clarin-1, a glycoprotein with homology to the tetraspanin family of proteins. Previous cell culture studies suggest that Clarin-1 localizes to the plasma membrane and interacts with the cytoskeleton. Mouse models demonstrate a role for the protein in mechanosensory hair bundle integrity, but the function of Clarin-1 in hearing remains unclear. Even less is known of its role in vision, because the Clrn1 knockout mouse does not exhibit a retinal phenotype and expression studies in murine retinas have provided conflicting results. Here, we describe cloning and expression analysis of the zebrafish clrn1 gene, and report protein localization of Clarin-1 in auditory and visual cells from embryonic through adult stages. We detect clrn1 transcripts as early as 24h post-fertilization, and expression is maintained through adulthood. In situ hybridization experiments show clrn1 transcripts enriched in mechanosensory hair cells and supporting cells of the inner ear and lateral line organ, photoreceptors, and cells of the inner retina. In mechanosensory hair cells, Clarin-1 is polarized to the apical cell body and the synapses. In the retina, Clarin-1 localizes to lateral cell contacts between photoreceptors and is associated with the outer limiting membrane and subapical processes emanating from Müller glial cells. We also find Clarin-1 protein in the outer plexiform, inner nuclear and ganglion cell layers of the retina. Given the importance of Clarin-1 function in the human retina, it is imperative to find an animal model with a comparable requirement. Our data provide a foundation for exploring the role of Clarin-1 in retinal cell function and survival in a diurnal, cone-dominant species. Copyright © 2013 Elsevier B.V. All rights reserved.

Imaging a seizure model in zebrafish with structured illumination light sheet microscopy

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Liu, Yang; Dale, Savannah; Ball, Rebecca; VanLeuven, Ariel J.; Baraban, Scott; Sornborger, Andrew; Lauderdale, James D.; Kner, Peter

2018-02-01

Zebrafish are a promising vertebrate model for elucidating how neural circuits generate behavior under normal and pathological conditions. The Baraban group first demonstrated that zebrafish larvae are valuable for investigating seizure events and can be used as a model for epilepsy in humans. Because of their small size and transparency, zebrafish embryos are ideal for imaging seizure activity using calcium indicators. Light-sheet microscopy is well suited to capturing neural activity in zebrafish because it is capable of optical sectioning, high frame rates, and low excitation intensities. We describe work in our lab to use light-sheet microscopy for high-speed long-time imaging of neural activity in wildtype and mutant zebrafish to better understand the connectivity and activity of inhibitory neural networks when GABAergic signaling is altered in vivo. We show that, with light-sheet microscopy, neural activity can be recorded at 23 frames per second in twocolors for over 10 minutes allowing us to capture rare seizure events in mutants. We have further implemented structured illumination to increase resolution and contrast in the vertical and axial directions during high-speed imaging at an effective frame rate of over 7 frames per second.

Development of neural retina in retinopathy of prematurity

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Yi Luo

2015-10-01

Full Text Available Retinopathy of prematurity(ROPis an important cause of infant blindness and visual impairment in the world, of which main clinical characteristics are peripheral retinal vascular abnormalities, including large non-perfusion area and abnormal neovascularization. Numerous researches have demonstrated that ROP affects the differentiation and maturity of retinal photoreceptor cells, with more significantly effect on rods than cones, and later mostly caused ametropia, strabismus, amblyopia and a series of abnormal visual functions, the specific mechanism remains unclear. After treatments, even the retinal vascular proliferation lesions disappear itself, but the abnormal development of photoreceptor cells and the resulting visual dysfunction will persist. Currently the best evaluation mean of clinical assessment about retinal function is mainly visual electrophysiology, especially flash electroretinogram(f-ERG, which can reflect the whole retinal functional status before ganglion cells, has a unique significance for the evaluation of retinal photoreceptor cells function. In this review, we aims at the development of neural retina(mainly photoreceptor cellsand its related mechanisms, also the visual function changes appeared in the late period about ROP and its mechanisms, guiding us to pursuit better methods for treatment.

The ciliary margin zone of the mammalian retina generates retinal ganglion cells

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Marcucci, Florencia; Murcia-Belmonte, Veronica; Coca, Yaiza; Ferreiro-Galve, Susana; Wang, Qing; Kuwajima, Takaaki; Khalid, Sania; Ross, M. Elizabeth; Herrera, Eloisa; Mason, Carol

2016-01-01

Summary The retina of lower vertebrates grows continuously by integrating new neurons generated from progenitors in the ciliary margin zone (CMZ). Whether the mammalian CMZ provides the neural retina with retinal cells is controversial. Live-imaging of embryonic retina expressing eGFP in the CMZ shows that cells migrate laterally from the CMZ to the neural retina where differentiated retinal ganglion cells (RGCs) reside. As Cyclin D2, a cell-cycle regulator, is enriched in ventral CMZ, we analyzed Cyclin D2−/− mice to test whether the CMZ is a source of retinal cells. Neurogenesis is diminished in Cyclin D2 mutants, leading to a reduction of RGCs in the ventral retina. In line with these findings, in the albino retina, the decreased production of ipsilateral RGCs is correlated with fewer Cyclin D2+ cells. Together, these results implicate the mammalian CMZ as a neurogenic site that produces RGCs and whose proper generation depends on Cyclin D2 activity. PMID:28009286

Selective retina therapy (SRT); Selektive Retina-Therapie (SRT)

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Brinkmann, R.; Birngruber, R. [Luebeck Univ. (Germany). Inst. fuer Biomedizinische Optik; Medizinisches Laserzentrum Luebeck GmbH, Luebeck (Germany)

2007-07-01

Selective Retina Therapy (SRT) is a new and very gentle laser method developed at the Medical Laser Center Luebeck. It is currently investigated clinically in order to treat retinal disorders associated with a decreased function of the retinal pigment epithelium (RPE). SRT is designed to selectively effect the RPE while sparing the neural retina and the photoreceptors as well as the choroid. Aim of the therapy is the rejuvenation of the RPE in the treated areas, which should ideally lead to a long term metabolic increase at the chorio-retinal junction. In contrast to conventional laser photocoagulation, which is associated with a complete thermal necrosis of the treated site, SRT completely retains full vision. This paper reviews the methods and mechanisms behind selective RPE effects and reports the first clinical results. An online dosimetry technique to visualize the ophthalmoscopically invisible effects is introduced. (orig.)

The Neural Retina in Retinopathy of Prematurity

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Hansen, Ronald M.; Moskowitz, Anne; Akula, James D.; Fulton, Anne B.

2016-01-01

Retinopathy of prematurity (ROP) is a neurovascular disease that affects prematurely born infants and is known to have significant long term effects on vision. We conducted the studies described herein not only to learn more about vision but also about the pathogenesis of ROP. The coincidence of ROP onset and rapid developmental elongation of the rod photoreceptor outer segments motivated us to consider the role of the rods in this disease. We used noninvasive electroretinographic (ERG), psychophysical, and retinal imaging procedures to study the function and structure of the neurosensory retina. Rod photoreceptor and post-receptor responses are significantly altered years after the preterm days during which ROP is an active disease. The alterations include persistent rod dysfunction, and evidence of compensatory remodeling of the post-receptor retina is found in ERG responses to full-field stimuli and in psychophysical thresholds that probe small retinal regions. In the central retina, both Mild and Severe ROP delay maturation of parafoveal scotopic thresholds and are associated with attenuation of cone mediated multifocal ERG responses, significant thickening of post-receptor retinal laminae, and dysmorphic cone photoreceptors. These results have implications for vision and control of eye growth and refractive development and suggest future research directions. These results also lead to a proposal for noninvasive management using light that may add to the currently invasive therapeutic armamentarium against ROP. PMID:27671171

Mechanisms underlying metabolic and neural defects in zebrafish and human multiple acyl-CoA dehydrogenase deficiency (MADD.

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Yuanquan Song

2009-12-01

Full Text Available In humans, mutations in electron transfer flavoprotein (ETF or electron transfer flavoprotein dehydrogenase (ETFDH lead to MADD/glutaric aciduria type II, an autosomal recessively inherited disorder characterized by a broad spectrum of devastating neurological, systemic and metabolic symptoms. We show that a zebrafish mutant in ETFDH, xavier, and fibroblast cells from MADD patients demonstrate similar mitochondrial and metabolic abnormalities, including reduced oxidative phosphorylation, increased aerobic glycolysis, and upregulation of the PPARG-ERK pathway. This metabolic dysfunction is associated with aberrant neural proliferation in xav, in addition to other neural phenotypes and paralysis. Strikingly, a PPARG antagonist attenuates aberrant neural proliferation and alleviates paralysis in xav, while PPARG agonists increase neural proliferation in wild type embryos. These results show that mitochondrial dysfunction, leading to an increase in aerobic glycolysis, affects neurogenesis through the PPARG-ERK pathway, a potential target for therapeutic intervention.

Immunomodulation-accelerated neuronal regeneration following selective rod photoreceptor cell ablation in the zebrafish retina.

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White, David T; Sengupta, Sumitra; Saxena, Meera T; Xu, Qingguo; Hanes, Justin; Ding, Ding; Ji, Hongkai; Mumm, Jeff S

2017-05-02

Müller glia (MG) function as inducible retinal stem cells in zebrafish, completely repairing the eye after damage. The innate immune system has recently been shown to promote tissue regeneration in which classic wound-healing responses predominate. However, regulatory roles for leukocytes during cellular regeneration-i.e., selective cell-loss paradigms akin to degenerative disease-are less well defined. To investigate possible roles innate immune cells play during retinal cell regeneration, we used intravital microscopy to visualize neutrophil, macrophage, and retinal microglia responses to induced rod photoreceptor apoptosis. Neutrophils displayed no reactivity to rod cell loss. Peripheral macrophage cells responded to rod cell loss, as evidenced by morphological transitions and increased migration, but did not enter the retina. Retinal microglia displayed multiple hallmarks of immune cell activation: increased migration, translocation to the photoreceptor cell layer, proliferation, and phagocytosis of dying cells. To test function during rod cell regeneration, we coablated microglia and rod cells or applied immune suppression and quantified the kinetics of ( i ) rod cell clearance, ( ii ) MG/progenitor cell proliferation, and ( iii ) rod cell replacement. Coablation and immune suppressants applied before cell loss caused delays in MG/progenitor proliferation rates and slowed the rate of rod cell replacement. Conversely, immune suppressants applied after cell loss had been initiated led to accelerated photoreceptor regeneration kinetics, possibly by promoting rapid resolution of an acute immune response. Our findings suggest that microglia control MG responsiveness to photoreceptor loss and support the development of immune-targeted therapeutic strategies for reversing cell loss associated with degenerative retinal conditions.

Direct transdifferentiation in the vertebrate retina.

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Opas, M; Dziak, E

1998-03-01

Transdifferentiation is the process by which differentiated cells alter their identity to become other, distinct cell types. The conversion of neural retina into lens epithelium is one of the most spectacular examples of transdifferentiation. We show that the redirection of cell fate from neural retina to lens and subsequent transdifferentiation is independent of cell replication as it occurs in growth-arrested cell populations. Using DNA ratiometry of individual cells in these cultures we show that, indeed, individual amitotic cells do transdifferentiate. Hence, choice of fate in transdifferentiating cells does not rely on a "community effect" but instead can be categorized as a leadership effect> For lack of overt lens progenitors, and most importantly, for its mitotic independence, we conclude that lens colony formation in vitro does occur by direct transdifferentiation and not by clonal proliferation of progenitor cells.

Learning and memory in zebrafish larvae

Science.gov (United States)

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

2013-01-01

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

microRNA expression in the neural retina: Focus on Müller glia.

Science.gov (United States)

Quintero, Heberto; Lamas, Mónica

2018-03-01

The neural retina hosts a unique specialized type of macroglial cell that not only preserves retinal homeostasis, function, and integrity but also may serve as a source of new neurons during regenerative processes: the Müller cell. Precise microRNA-driven mechanisms of gene regulation impel and direct the processes of Müller glia lineage acquisition from retinal progenitors during development, the triggering of their response to retinal degeneration and, in some cases, Müller cell reprogramming and regenerative events. In this review we survey the recent reports describing, through functional assays, the regulatory role of microRNAs in Müller cell physiology, differentiation potential, and retinal pathology. We discuss also the evidence based on expression analysis that points out the relevance of a Müller glia-specific microRNA signature that would orchestrate these processes. © 2017 Wiley Periodicals, Inc.

Sinup, a novel Siaz-interacting nuclear protein, modulates neural plate formation in the zebrafish embryos

International Nuclear Information System (INIS)

Ro, Hyunju; Won, Minho; Lee, Su-Ui; Kim, Kyoon E.; Huh, Tae-Lin; Kim, Cheol-Hee; Rhee, Myungchull

2005-01-01

Siah, the vertebrate homologue of the Drosophila seven in absentia (sina) gene, is well conserved from Drosophila to mammal and involved in ubiquitination and proteasome-dependent degradation of various target proteins. To identify cellular proteins interacting with Siah, we screened a zebrafish cDNA library with zebrafish Siah (Siaz) as bait in a yeast two-hybrid assay. We identified a cDNA encoding a novel protein composed of 145 amino acids and termed it as Sinup (Siaz-interacting-nuclear-protein). Sinup is a novel nuclear protein that binds to the highly conserved C-terminal protein-interacting domain of Siaz both in vivo and in vitro. During development, sinup transcripts are abundant from the one-cell stage to the early blastula and then markedly diminished, suggesting sinup largely exists as maternal transcripts. sinup overexpression induced lateral expansion of the neural plate and in consequence caused ectopic expression of otx-2 and hoxb1b during the late gastrula stage. In addition, the lateral/paraxial expression of wnt8 at the onset of gastrulation is suppressed by the forced expression of sinup while the expression levels of various dorso-ventral markers are unaffected. In contrast, interfering with sinup functions using sinup morpholino oligonucleotides gradually diminished the anterior neuroectoderm from the posterior region, and resulted in compete loss of hindbrain at the 3-somites stage. Our report suggests that sinup expression should be tightly regulated during early embryonic development for the proper neural plate formation

HB-EGF is necessary and sufficient for Müller glia dedifferentiation and retina regeneration

Science.gov (United States)

Wan, Jin; Ramachandran, Rajesh; Goldman, Daniel

2011-01-01

Summary Müller glia (MG) dedifferentiation into a cycling population of multipotent progenitors is crucial to zebrafish retina regeneration. The mechanisms underlying MG dedifferentiation are unknown. Here we report that heparin-binding epidermal-like growth factor (HB-EGF) is rapidly induced in MG residing at the injury site and that proHB-EGF ectodomain shedding is necessary for retina regeneration. Remarkably, HB-EGF stimulates the formation of multipotent MG-derived progenitors in the uninjured retina. We show that HB-EGF mediates its effects via an EGFR/MAPK signal transduction cascade that regulates the expression of regeneration-associated genes, like ascl1a and pax6b. We also uncover an HB-EGF/Ascl1a/Notch/hb-egfa signaling loop that helps define the zone of injury-responsive MG. Finally, we show that HB-EGF acts upstream of the Wnt/β-catenin signaling cascade that controls progenitor proliferation. These data provide a link between extracellular signaling and regeneration-associated gene expression in the injured retina and suggest strategies for stimulating retina regeneration in mammals. PMID:22340497

Graph theoretical model of a sensorimotor connectome in zebrafish.

Science.gov (United States)

Stobb, Michael; Peterson, Joshua M; Mazzag, Borbala; Gahtan, Ethan

2012-01-01

Mapping the detailed connectivity patterns (connectomes) of neural circuits is a central goal of neuroscience. The best quantitative approach to analyzing connectome data is still unclear but graph theory has been used with success. We present a graph theoretical model of the posterior lateral line sensorimotor pathway in zebrafish. The model includes 2,616 neurons and 167,114 synaptic connections. Model neurons represent known cell types in zebrafish larvae, and connections were set stochastically following rules based on biological literature. Thus, our model is a uniquely detailed computational representation of a vertebrate connectome. The connectome has low overall connection density, with 2.45% of all possible connections, a value within the physiological range. We used graph theoretical tools to compare the zebrafish connectome graph to small-world, random and structured random graphs of the same size. For each type of graph, 100 randomly generated instantiations were considered. Degree distribution (the number of connections per neuron) varied more in the zebrafish graph than in same size graphs with less biological detail. There was high local clustering and a short average path length between nodes, implying a small-world structure similar to other neural connectomes and complex networks. The graph was found not to be scale-free, in agreement with some other neural connectomes. An experimental lesion was performed that targeted three model brain neurons, including the Mauthner neuron, known to control fast escape turns. The lesion decreased the number of short paths between sensory and motor neurons analogous to the behavioral effects of the same lesion in zebrafish. This model is expandable and can be used to organize and interpret a growing database of information on the zebrafish connectome.

Monte Carlo simulation of zinc protoporphyrin fluorescence in the retina

Science.gov (United States)

Chen, Xiaoyan; Lane, Stephen

2010-02-01

We have used Monte Carlo simulation of autofluorescence in the retina to determine that noninvasive detection of nutritional iron deficiency is possible. Nutritional iron deficiency (which leads to iron deficiency anemia) affects more than 2 billion people worldwide, and there is an urgent need for a simple, noninvasive diagnostic test. Zinc protoporphyrin (ZPP) is a fluorescent compound that accumulates in red blood cells and is used as a biomarker for nutritional iron deficiency. We developed a computational model of the eye, using parameters that were identified either by literature search, or by direct experimental measurement to test the possibility of detecting ZPP non-invasively in retina. By incorporating fluorescence into Steven Jacques' original code for multi-layered tissue, we performed Monte Carlo simulation of fluorescence in the retina and determined that if the beam is not focused on a blood vessel in a neural retina layer or if part of light is hitting the vessel, ZPP fluorescence will be 10-200 times higher than background lipofuscin fluorescence coming from the retinal pigment epithelium (RPE) layer directly below. In addition we found that if the light can be focused entirely onto a blood vessel in the neural retina layer, the fluorescence signal comes only from ZPP. The fluorescence from layers below in this second situation does not contribute to the signal. Therefore, the possibility that a device could potentially be built and detect ZPP fluorescence in retina looks very promising.

Graph theoretical model of a sensorimotor connectome in zebrafish.

Directory of Open Access Journals (Sweden)

Michael Stobb

Full Text Available Mapping the detailed connectivity patterns (connectomes of neural circuits is a central goal of neuroscience. The best quantitative approach to analyzing connectome data is still unclear but graph theory has been used with success. We present a graph theoretical model of the posterior lateral line sensorimotor pathway in zebrafish. The model includes 2,616 neurons and 167,114 synaptic connections. Model neurons represent known cell types in zebrafish larvae, and connections were set stochastically following rules based on biological literature. Thus, our model is a uniquely detailed computational representation of a vertebrate connectome. The connectome has low overall connection density, with 2.45% of all possible connections, a value within the physiological range. We used graph theoretical tools to compare the zebrafish connectome graph to small-world, random and structured random graphs of the same size. For each type of graph, 100 randomly generated instantiations were considered. Degree distribution (the number of connections per neuron varied more in the zebrafish graph than in same size graphs with less biological detail. There was high local clustering and a short average path length between nodes, implying a small-world structure similar to other neural connectomes and complex networks. The graph was found not to be scale-free, in agreement with some other neural connectomes. An experimental lesion was performed that targeted three model brain neurons, including the Mauthner neuron, known to control fast escape turns. The lesion decreased the number of short paths between sensory and motor neurons analogous to the behavioral effects of the same lesion in zebrafish. This model is expandable and can be used to organize and interpret a growing database of information on the zebrafish connectome.

Progranulin regulates neurogenesis in the developing vertebrate retina.

Science.gov (United States)

Walsh, Caroline E; Hitchcock, Peter F

2017-09-01

We evaluated the expression and function of the microglia-specific growth factor, Progranulin-a (Pgrn-a) during developmental neurogenesis in the embryonic retina of zebrafish. At 24 hpf pgrn-a is expressed throughout the forebrain, but by 48 hpf pgrn-a is exclusively expressed by microglia and/or microglial precursors within the brain and retina. Knockdown of Pgrn-a does not alter the onset of neurogenic programs or increase cell death, however, in its absence, neurogenesis is significantly delayed-retinal progenitors fail to exit the cell cycle at the appropriate developmental time and postmitotic cells do not acquire markers of terminal differentiation, and microglial precursors do not colonize the retina. Given the link between Progranulin and cell cycle regulation in peripheral tissues and transformed cells, we analyzed cell cycle kinetics among retinal progenitors following Pgrn-a knockdown. Depleting Pgrn-a results in a significant lengthening of the cell cycle. These data suggest that Pgrn-a plays a dual role during nervous system development by governing the rate at which progenitors progress through the cell cycle and attracting microglial progenitors into the embryonic brain and retina. Collectively, these data show that Pgrn-a governs neurogenesis by regulating cell cycle kinetics and the transition from proliferation to cell cycle exit and differentiation. © 2017 The Authors. Developmental Neurobiology Published by Wiley Periodicals, Inc. Develop Neurobiol 77: 1114-1129, 2017. © 2017 The Authors. Developmental Neurobiology Published by Wiley Periodicals, Inc.

Rod photoreceptors express GPR55 in the adult vervet monkey retina

DEFF Research Database (Denmark)

Bouskila, Joseph; Javadi, Pasha; Casanova, Christian

2013-01-01

. Yet, its formal classification is still a matter of debate. CB1R and CB2R expression patterns are well described for rodent and monkey retinas. In the monkey retina, CB1R has been localized in its neural (cone photoreceptor, horizontal, bipolar, amacrine and ganglion cells) and CB2R in glial...

Zebrafish Lacking Circadian Gene per2 Exhibit Visual Function Deficiency

Directory of Open Access Journals (Sweden)

Deng-feng Huang

2018-03-01

Full Text Available The retina has an intrinsic circadian clock, but the importance of this clock for vision is unknown. Zebrafish offer many advantages for studying vertebrate vision and circadian rhythm. Here, we explored the role of zebrafish per2, a light-regulated gene, in visual behavior and the underlying mechanisms. We observed that per2 mutant zebrafish larvae showed decreased contrast sensitivity and visual acuity using optokinetic response (OKR assays. Using a visual motor response (VMR assay, we observed normal OFF responses but abnormal ON responses in mutant zebrafish larvae. Immunofluorescence showed that mutants had a normal morphology of cone photoreceptor cells and retinal organization. However, electron microscopy showed that per2 mutants displayed abnormal and decreased photoreceptor ribbon synapses with arciform density, which resulted in retinal ON pathway defect. We also examined the expression of three cone opsins by quantitative real-time PCR (qRT-PCR, and the expression of long-wave-sensitive opsin (opn1lw and short-wave-sensitive opsin (opn1sw was reduced in mutant zebrafish larvae. qRT-PCR analyses also showed a down-regulation of the clock genes cry1ba and bmal1b in the adult eye of per2 mutant zebrafish. This study identified a mechanism by which a clock gene affects visual function and defined important roles of per2 in retinal information processing.

Vsx2 in the zebrafish retina: restricted lineages through derepression

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Higashijima Shin-ichi

2009-04-01

Full Text Available Abstract Background The neurons in the vertebrate retina arise from multipotent retinal progenitor cells (RPCs. It is not clear, however, which progenitors are multipotent or why they are multipotent. Results In this study we show that the homeodomain transcription factor Vsx2 is initially expressed throughout the retinal epithelium, but later it is downregulated in all but a minor population of bipolar cells and all Müller glia. The Vsx2-negative daughters of Vsx2-positive RPCs divide and give rise to all other cell types in the retina. Vsx2 is a repressor whose targets include transcription factors such as Vsx1, which is expressed in the progenitors of distinct non-Vsx2 bipolars, and the basic helix-loop-helix transcription factor Ath5, which restricts the fate of progenitors to retinal ganglion cells, horizontal cells, amacrine cells and photoreceptors fates. Foxn4, expressed in the progenitors of amacrine and horizontal cells, is also negatively regulated by Vsx2. Conclusion Our data thus suggest Vsx2-positive RPCs are fully multipotent retinal progenitors and that when Vsx2 is downregulated, Vsx2-negative progenitors escape Vsx2 repression and so are able to express factors that restrict lineage potential.

Gene expression changes in the retina following subretinal injection of human neural progenitor cells into a rodent model for retinal degeneration.

Science.gov (United States)

Jones, Melissa K; Lu, Bin; Saghizadeh, Mehrnoosh; Wang, Shaomei

2016-01-01

Retinal degenerative diseases (RDDs) affect millions of people and are the leading cause of vision loss. Although treatment options for RDDs are limited, stem and progenitor cell-based therapies have great potential to halt or slow the progression of vision loss. Our previous studies have shown that a single subretinal injection of human forebrain derived neural progenitor cells (hNPCs) into the Royal College of Surgeons (RCS) retinal degenerate rat offers long-term preservation of photoreceptors and visual function. Furthermore, neural progenitor cells are currently in clinical trials for treating age-related macular degeneration; however, the molecular mechanisms of stem cell-based therapies are largely unknown. This is the first study to analyze gene expression changes in the retina of RCS rats following subretinal injection of hNPCs using high-throughput sequencing. RNA-seq data of retinas from RCS rats injected with hNPCs (RCS(hNPCs)) were compared to sham surgery in RCS (RCS(sham)) and wild-type Long Evans (LE(sham)) rats. Differential gene expression patterns were determined with in silico analysis and confirmed with qRT-PCR. Function, biologic, cellular component, and pathway analyses were performed on differentially expressed genes and investigated with immunofluorescent staining experiments. Analysis of the gene expression data sets identified 1,215 genes that were differentially expressed between RCS(sham) and LE(sham) samples. Additionally, 283 genes were differentially expressed between the RCS(hNPCs) and RCS(sham) samples. Comparison of these two gene sets identified 68 genes with inverse expression (termed rescue genes), including Pdc, Rp1, and Cdc42ep5. Functional, biologic, and cellular component analyses indicate that the immune response is enhanced in RCS(sham). Pathway analysis of the differential expression gene sets identified three affected pathways in RCS(hNPCs), which all play roles in phagocytosis signaling. Immunofluorescent staining

Cadmium affects retinogenesis during zebrafish embryonic development

International Nuclear Information System (INIS)

Hen Chow, Elly Suk; Yu Hui, Michelle Nga; Cheng, Chi Wa; Cheng, Shuk Han

2009-01-01

Ocular malformations are commonly observed in embryos of aquatic species after exposure to toxicants. Using zebrafish embryos as the model organism, we showed that cadmium exposure from sphere stage (4 hpf) to end of segmentation stage (24 hpf) induced microphthalmia in cadmium-treated embryos. Embryos with eye defects were then assessed for visual abilities. Cadmium-exposed embryos were behaviorally blind, showing hyperpigmentation and loss of camouflage response to light. We investigated the cellular basis of the formation of the small eyes phenotype and the induction of blindness by studying retina development and retinotectal projections. Retinal progenitors were found in cadmium-treated embryos albeit in smaller numbers. The number of retinal ganglion cells (RGC), the first class of retinal cells to differentiate during retinogenesis, was reduced, while photoreceptor cells, the last batch of retinal neurons to differentiate, were absent. Cadmium also affected the propagation of neurons in neurogenic waves. The neurons remained in the ventronasal area and failed to spread across the retina. Drastically reduced RGC axons and disrupted optic stalk showed that the optic nerves did not extend from the retina beyond the chiasm into the tectum. Our data suggested that impairment in neuronal differentiation of the retina, disruption in RGC axon formation and absence of cone photoreceptors were the causes of microphthalmia and visual impairment in cadmium-treated embryos

Examination of a Palatogenic Gene Program in Zebrafish

Science.gov (United States)

Swartz, Mary E.; Sheehan-Rooney, Kelly; Dixon, Michael J.; Eberhart, Johann K.

2011-01-01

Human palatal clefting is debilitating and difficult to rectify surgically. Animal models enhance our understanding of palatogenesis and are essential in strategies designed to ameliorate palatal malformations in humans. Recent studies have shown that the zebrafish palate, or anterior neurocranium, is under similar genetic control to the amniote palatal skeleton. We extensively analyzed palatogenesis in zebrafish to determine the similarity of gene expression and function across vertebrates. By 36 hpf palatogenic cranial neural crest cells reside in homologous regions of the developing face compared to amniote species. Transcription factors and signaling molecules regulating mouse palatogenesis are expressed in similar domains during palatogenesis in zebrafish. Functional investigation of a subset of these genes, fgf10a, tgfb2, pax9 and smad5 revealed their necessity in zebrafish palatogenesis. Collectively, these results suggest that the gene regulatory networks regulating palatogenesis may be conserved across vertebrate species, demonstrating the utility of zebrafish as a model for palatogenesis. PMID:22016187

Social learning of an associative foraging task in zebrafish

Science.gov (United States)

Zala, Sarah M.; Määttänen, Ilmari

2013-05-01

The zebrafish ( Danio rerio) is increasingly becoming an important model species for studies on the genetic and neural mechanisms controlling behaviour and cognition. Here, we utilized a conditioned place preference (CPP) paradigm to study social learning in zebrafish. We tested whether social interactions with conditioned demonstrators enhance the ability of focal naïve individuals to learn an associative foraging task. We found that the presence of conditioned demonstrators improved focal fish foraging behaviour through the process of social transmission, whereas the presence of inexperienced demonstrators interfered with the learning of the control focal fish. Our results indicate that zebrafish use social learning for finding food and that this CPP paradigm is an efficient assay to study social learning and memory in zebrafish.

Selective retina therapy (SRT)

International Nuclear Information System (INIS)

Brinkmann, R.; Birngruber, R.

2007-01-01

Selective Retina Therapy (SRT) is a new and very gentle laser method developed at the Medical Laser Center Luebeck. It is currently investigated clinically in order to treat retinal disorders associated with a decreased function of the retinal pigment epithelium (RPE). SRT is designed to selectively effect the RPE while sparing the neural retina and the photoreceptors as well as the choroid. Aim of the therapy is the rejuvenation of the RPE in the treated areas, which should ideally lead to a long term metabolic increase at the chorio-retinal junction. In contrast to conventional laser photocoagulation, which is associated with a complete thermal necrosis of the treated site, SRT completely retains full vision. This paper reviews the methods and mechanisms behind selective RPE effects and reports the first clinical results. An online dosimetry technique to visualize the ophthalmoscopically invisible effects is introduced. (orig.)

FPGA-Based Real Time, Multichannel Emulated-Digital Retina Model Implementation

Directory of Open Access Journals (Sweden)

Zsolt Vörösházi

2009-01-01

Full Text Available The function of the low-level image processing that takes place in the biological retina is to compress only the relevant visual information to a manageable size. The behavior of the layers and different channels of the neuromorphic retina has been successfully modeled by cellular neural/nonlinear networks (CNNs. In this paper, we present an extended, application-specific emulated-digital CNN-universal machine (UM architecture to compute the complex dynamic of this mammalian retina in video real time. The proposed emulated-digital implementation of multichannel retina model is compared to the previously developed models from three key aspects, which are processing speed, number of physical cells, and accuracy. Our primary aim was to build up a simple, real-time test environment with camera input and display output in order to mimic the behavior of retina model implementation on emulated digital CNN by using low-cost, moderate-sized field-programmable gate array (FPGA architectures.

The neurogenetic frontier--lessons from misbehaving zebrafish.

Science.gov (United States)

Burgess, Harold A; Granato, Michael

2008-11-01

One of the central questions in neuroscience is how refined patterns of connectivity in the brain generate and monitor behavior. Genetic mutations can influence neural circuits by disrupting differentiation or maintenance of component neuronal cells or by altering functional patterns of nervous system connectivity. Mutagenesis screens therefore have the potential to reveal not only the molecular underpinnings of brain development and function, but to illuminate the cellular basis of behavior. Practical considerations make the zebrafish an organism of choice for undertaking forward genetic analysis of behavior. The powerful array of experimental tools at the disposal of the zebrafish researcher makes it possible to link molecular function to neuronal properties that underlie behavior. This review focuses on specific challenges to isolating and analyzing behavioral mutants in zebrafish.

Spontaneous activity in the developing mammalian retina: Form and function

Science.gov (United States)

Butts, Daniel Allison

Spontaneous neuronal activity is present in the immature mammalian retina during the initial stages of visual system development, before the retina is responsive to light. This activity consists of bursts of action potentials fired by retinal ganglion cells, and propagates in a wavelike manner across the inner plexiform layer of the retina. Unlike waves in other neural systems, retinal waves have large variability in both their rate and direction of propagation, and individual waves only propagate across small regions of the retina. The unique properties of retinal activity arise from dynamic processes within the developing retina, and produce characteristic spatiotemporal properties. These spatiotemporal properties are of particular interest, since they are believed to play a role in visual system development. This dissertation addresses the complex spatiotemporal patterning of the retinal waves from two different perspectives. First, it proposes how the immature circuitry of the developing retina generates these patterns of activity. In order to reproduce the distinct spatiotemporal properties observed in experiments, a model of the immature retinal circuitry must meet certain requirements, which are satisfied by a coarse-grained model of the developing retina that we propose. Second, this dissertation addresses how the particular spatiotemporal patterning of the retinal waves provides information to the rest of the visual system and, as a result, can be used to guide visual system development. By measuring the properties of this information, we place constraints on the developmental mechanisms that use this activity, and show how the particular spatiotemporal properties of the retinal waves provide this information. Together, this dissertation demonstrates how the apparent complexity of retinal wave patterning can be understood both through the immature circuitry that generates it, and through the developmental mechanisms that may use it. The first three

Zebrafish msxB, msxC and msxE function together to refine the neural-nonneural border and regulate cranial placodes and neural crest development.

Science.gov (United States)

Phillips, Bryan T; Kwon, Hye-Joo; Melton, Colt; Houghtaling, Paul; Fritz, Andreas; Riley, Bruce B

2006-06-15

The zebrafish muscle segment homeobox genes msxB, msxC and msxE are expressed in partially overlapping domains in the neural crest and preplacodal ectoderm. We examined the roles of these msx genes in early development. Disrupting individual msx genes causes modest variable defects, whereas disrupting all three produces a reproducible severe phenotype, suggesting functional redundancy. Neural crest differentiation is blocked at an early stage. Preplacodal development begins normally, but placodes arising from the msx expression domain later show elevated apoptosis and are reduced in size. Cell proliferation is normal in these tissues. Unexpectedly, Msx-deficient embryos become ventralized by late gastrulation whereas misexpression of msxB dorsalizes the embryo. These effects appear to involve Distal-less (Dlx) protein activity, as loss of dlx3b and dlx4b suppresses ventralization in Msx-depleted embryos. At the same time, Msx-depletion restores normal preplacodal gene expression to dlx3b-dlx4b mutants. These data suggest that mutual antagonism between Msx and Dlx proteins achieves a balance of function required for normal preplacodal differentiation and placement of the neural-nonneural border.

Skin too thin? The developing utility of zebrafish skin (neuro)pharmacology for CNS drug discovery research.

Science.gov (United States)

Nguyen, Michael; Poudel, Manoj K; Stewart, Adam Michael; Kalueff, Allan V

2013-09-01

Skin coloration can be affected by many genetic, environmental and pharmacological factors. Zebrafish (Danio rerio) are a useful and versatile model organism in biomedical research due to their genetic tractability, physiological homology to mammals, low cost, reproducibility and high throughput. Zebrafish coloration is mediated by chromatophores - the skin color pigment cells largely controlled by endocrine and neural mechanisms. The characteristic darkening of zebrafish skin is caused by the dispersion (and paling - by aggregation) of melanosomes (pigment-containing organelles), which show high homology to mammalian structures. Various pharmacological agents potently affect zebrafish coloration - the phenotype that often accompanies behavioral effects of the drugs, and may be used for drug discovery. Although zebrafish behavior and skin responses are usually not directly related, they share common regulatory (neural, endocrine) mechanisms, and therefore may be assessed in parallel during psychotropic drug screening. For example, some psychoactive drugs can potently affect zebrafish skin coloration. Can we use this knowledge to refine phenotype-driven psychotropic drug discovery? Here, we present current models using zebrafish skin coloration assays, and discuss how these models may be applied to enhance in vivo CNS drug discovery. Copyright © 2013 Elsevier Inc. All rights reserved.

RETINOBASE: a web database, data mining and analysis platform for gene expression data on retina

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Léveillard Thierry

2008-05-01

Full Text Available Abstract Background The retina is a multi-layered sensory tissue that lines the back of the eye and acts at the interface of input light and visual perception. Its main function is to capture photons and convert them into electrical impulses that travel along the optic nerve to the brain where they are turned into images. It consists of neurons, nourishing blood vessels and different cell types, of which neural cells predominate. Defects in any of these cells can lead to a variety of retinal diseases, including age-related macular degeneration, retinitis pigmentosa, Leber congenital amaurosis and glaucoma. Recent progress in genomics and microarray technology provides extensive opportunities to examine alterations in retinal gene expression profiles during development and diseases. However, there is no specific database that deals with retinal gene expression profiling. In this context we have built RETINOBASE, a dedicated microarray database for retina. Description RETINOBASE is a microarray relational database, analysis and visualization system that allows simple yet powerful queries to retrieve information about gene expression in retina. It provides access to gene expression meta-data and offers significant insights into gene networks in retina, resulting in better hypothesis framing for biological problems that can subsequently be tested in the laboratory. Public and proprietary data are automatically analyzed with 3 distinct methods, RMA, dChip and MAS5, then clustered using 2 different K-means and 1 mixture models method. Thus, RETINOBASE provides a framework to compare these methods and to optimize the retinal data analysis. RETINOBASE has three different modules, "Gene Information", "Raw Data System Analysis" and "Fold change system Analysis" that are interconnected in a relational schema, allowing efficient retrieval and cross comparison of data. Currently, RETINOBASE contains datasets from 28 different microarray experiments performed

TOPOGRAPHIC ORGANIZATION AND SPECIALIZED AREAS IN THE RETINA OF Callopistes palluma: GANGLION CELL LAYER

OpenAIRE

Inzunza, Oscar; Barros B., Zitta; Bravo, Hermes

1998-01-01

In this paper we analyze the topographic distribution and cell body size of neurons (ganglion and displaced amacrine) of layer 8 of the retina in the chilean reptile Callopistes palluma; using whole mount retinaswith nissl stain. Callopistes palluma retina has an area centralis without fovea in which the ganglion cell density amounts 20.000 cells / µm2 while the displaced amacrine neurons is about 7.000 cells / µm2. This neural density decreased gradually towards the peripheral retina. A hor...

The neurogenetic frontier—lessons from misbehaving zebrafish

Science.gov (United States)

Granato, Michael

2008-01-01

One of the central questions in neuroscience is how refined patterns of connectivity in the brain generate and monitor behavior. Genetic mutations can influence neural circuits by disrupting differentiation or maintenance of component neuronal cells or by altering functional patterns of nervous system connectivity. Mutagenesis screens therefore have the potential to reveal not only the molecular underpinnings of brain development and function, but to illuminate the cellular basis of behavior. Practical considerations make the zebrafish an organism of choice for undertaking forward genetic analysis of behavior. The powerful array of experimental tools at the disposal of the zebrafish researcher makes it possible to link molecular function to neuronal properties that underlie behavior. This review focuses on specific challenges to isolating and analyzing behavioral mutants in zebrafish. PMID:18836206

Complex computation in the retina

Science.gov (United States)

Deshmukh, Nikhil Rajiv

Elucidating the general principles of computation in neural circuits is a difficult problem requiring both a tractable model circuit as well as sophisticated measurement tools. This thesis advances our understanding of complex computation in the salamander retina and its underlying circuitry and furthers the development of advanced tools to enable detailed study of neural circuits. The retina provides an ideal model system for neural circuits in general because it is capable of producing complex representations of the visual scene, and both its inputs and outputs are accessible to the experimenter. Chapter 2 describes the biophysical mechanisms that give rise to the omitted stimulus response in retinal ganglion cells described in Schwartz et al., (2007) and Schwartz and Berry, (2008). The extra response to omitted flashes is generated at the input to bipolar cells, and is separable from the characteristic latency shift of the OSR apparent in ganglion cells, which must occur downstream in the circuit. Chapter 3 characterizes the nonlinearities at the first synapse of the ON pathway in response to high contrast flashes and develops a phenomenological model that captures the effect of synaptic activation and intracellular signaling dynamics on flash responses. This work is the first attempt to model the dynamics of the poorly characterized mGluR6 transduction cascade unique to ON bipolar cells, and explains the second lobe of the biphasic flash response. Complementary to the study of neural circuits, recent advances in wafer-scale photolithography have made possible new devices to measure the electrical and mechanical properties of neurons. Chapter 4 reports a novel piezoelectric sensor that facilitates the simultaneous measurement of electrical and mechanical signals in neural tissue. This technology could reveal the relationship between the electrical activity of neurons and their local mechanical environment, which is critical to the study of mechanoreceptors

Bioluminescence Monitoring of Neuronal Activity in Freely Moving Zebrafish Larvae

Science.gov (United States)

Knafo, Steven; Prendergast, Andrew; Thouvenin, Olivier; Figueiredo, Sophie Nunes; Wyart, Claire

2017-01-01

The proof of concept for bioluminescence monitoring of neural activity in zebrafish with the genetically encoded calcium indicator GFP-aequorin has been previously described (Naumann et al., 2010) but challenges remain. First, bioluminescence signals originating from a single muscle fiber can constitute a major pitfall. Second, bioluminescence signals emanating from neurons only are very small. To improve signals while verifying specificity, we provide an optimized 4 steps protocol achieving: 1) selective expression of a zebrafish codon-optimized GFP-aequorin, 2) efficient soaking of larvae in GFP-aequorin substrate coelenterazine, 3) bioluminescence monitoring of neural activity from motor neurons in free-tailed moving animals performing acoustic escapes and 4) verification of the absence of muscle expression using immunohistochemistry. PMID:29130058

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.

Ultrasound-mediated nanoparticle delivery across ex vivo bovine retina after intravitreal injection.

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Huang, Di; Chen, Ying-Shan; Thakur, Sachin S; Rupenthal, Ilva D

2017-10-01

Intravitreal injection is the most common administration route for the treatment of retinal diseases. However, the vitreous and some of the retinal layers themselves act as significant barriers to efficient delivery of drugs administered intravitreally. This study aimed to improve the diffusive mobility of nanoparticles (NPs) in the vitreous and enhance their permeation across the retina after intravitreal injection by application of ultrasound (US). Ex vivo posterior bovine eye cups were used and the vitreous was either left intact or removed gently from the neural retina. Hyaluronic acid coated human serum albumin NPs were administered into the eye cups and continuous US with a frequency of 1MHz, an intensity of 0.5W/cm 2 , and a duration of 30s was applied once or repeatedly via the transscleral route. After pre-determined time points, fluorescence intensities in the vitreous and the retina were analyzed. Short pulses of US significantly improved the diffusive mobility of NPs through the vitreous as well as their penetration across the neural retina into the retinal pigment epithelium and choroid without causing any detectable damage to the ocular tissues. Therefore, transscleral US could be a powerful and safe tool to enhance retinal delivery of intravitreally injected NPs. Copyright © 2017 Elsevier B.V. All rights reserved.

Neural Development in tsc2-Deficient Zebrafish

Science.gov (United States)

2011-10-01

the cells in this region normally give rise to forebrain and retina (Woo and Fraser, 1995). Analyses of chimeric embryos at 7.5 dpf revealed that...white matter. Green color is tissue autofluorescence ; blue is DAPI staining. P, pallium; Po, preoptic region; Sd, dorsal division of subpallium; Tve

Feature Binding in Zebrafish

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P Neri

2012-07-01

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

Mechanisms of cadmium-caused eye hypoplasia and hypopigmentation in zebrafish embryos

International Nuclear Information System (INIS)

Zhang, Ting; Zhou, Xin-Ying; Ma, Xu-Fa; Liu, Jing-Xia

2015-01-01

Highlights: Using high-throughput in situ hybridization screening, we found that genes labeling the neural crest and its derivative pigment cells were sensitive to cadmium toxicity during zebrafish organogenesis, which might contribute to the molecular mechanisms underlying the phenotype defects of head and eye hypoplasia and hypopigmentation in cadmium-exposed embryos. Based on neural crest markers, we identified the doses and times of cadmium exposure that cause damage to the zebrafish organogenesis, and we also found that compounds BIO or RA could neutralize the toxic effects of cadmium. - Abstract: Cadmium-caused head and eye hypoplasia and hypopigmentation has been recognized for a long time, but knowledge of the underlying mechanisms is limited. In this study, we found that high mortality occurred in exposed embryos after 24 hpf, when cadmium (Cd) dosage was above 17.8 μM. Using high-throughput in situ hybridization screening, we found that genes labelling the neural crest and its derivative pigment cells exhibited obviously reduced expression in Cd-exposed embryos from 24 hpf, 2 days earlier than head and eye hypoplasia and hypopigmentation occurred. Moreover, based on expression of crestin, a neural crest marker, we found that embryos before the gastrula stage were more sensitive to cadmium toxicity and that damage caused by Cd on embryogenesis was dosage dependent. In addition, by phenotype observation and detection of neural crest and pigment cell markers, we found that BIO and retinoic acid (RA) could neutralize the toxic effects of Cd on zebrafish embryogenesis. In this study, we first determined that Cd blocked the formation of the neural crest and inhibited specification of pigment cells, which might contribute to the molecular mechanisms underlying the phenotype defects of head and eye hypoplasia and hypopigmentation in Cd-exposed embryos. Moreover, we found that compounds BIO or RA could neutralize the toxic effects of Cd.

Mechanisms of cadmium-caused eye hypoplasia and hypopigmentation in zebrafish embryos

Energy Technology Data Exchange (ETDEWEB)

Zhang, Ting, E-mail: zting@webmail.hzau.edu.cn; Zhou, Xin-Ying, E-mail: 290356082@qq.com; Ma, Xu-Fa, E-mail: xufama@mail.hzau.edu.cn; Liu, Jing-Xia, E-mail: ichliu@mail.hzau.edu.cn

2015-10-15

Highlights: Using high-throughput in situ hybridization screening, we found that genes labeling the neural crest and its derivative pigment cells were sensitive to cadmium toxicity during zebrafish organogenesis, which might contribute to the molecular mechanisms underlying the phenotype defects of head and eye hypoplasia and hypopigmentation in cadmium-exposed embryos. Based on neural crest markers, we identified the doses and times of cadmium exposure that cause damage to the zebrafish organogenesis, and we also found that compounds BIO or RA could neutralize the toxic effects of cadmium. - Abstract: Cadmium-caused head and eye hypoplasia and hypopigmentation has been recognized for a long time, but knowledge of the underlying mechanisms is limited. In this study, we found that high mortality occurred in exposed embryos after 24 hpf, when cadmium (Cd) dosage was above 17.8 μM. Using high-throughput in situ hybridization screening, we found that genes labelling the neural crest and its derivative pigment cells exhibited obviously reduced expression in Cd-exposed embryos from 24 hpf, 2 days earlier than head and eye hypoplasia and hypopigmentation occurred. Moreover, based on expression of crestin, a neural crest marker, we found that embryos before the gastrula stage were more sensitive to cadmium toxicity and that damage caused by Cd on embryogenesis was dosage dependent. In addition, by phenotype observation and detection of neural crest and pigment cell markers, we found that BIO and retinoic acid (RA) could neutralize the toxic effects of Cd on zebrafish embryogenesis. In this study, we first determined that Cd blocked the formation of the neural crest and inhibited specification of pigment cells, which might contribute to the molecular mechanisms underlying the phenotype defects of head and eye hypoplasia and hypopigmentation in Cd-exposed embryos. Moreover, we found that compounds BIO or RA could neutralize the toxic effects of Cd.

Redundant roles of PRDM family members in zebrafish craniofacial development.

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Ding, Hai-Lei; Clouthier, David E; Artinger, Kristin B

2013-01-01

PRDM proteins are evolutionary conserved Zn-Finger transcription factors that share a characteristic protein domain organization. Previous studies have shown that prdm1a is required for the specification and differentiation of neural crest cells in the zebrafish. Here we examine other members of this family, specifically prdm3, 5, and 16, in the differentiation of the zebrafish craniofacial skeleton. prdm3 and prdm16 are strongly expressed in the pharyngeal arches, while prdm5 is expressed specifically in the area of the forming neurocranium. Knockdown of prdm3 and prdm16 results in a reduction in the neural crest markers dlx2a and barx1 and defects in both the viscerocranium and the neurocranium. The knockdown of prdm3 and prdm16 in combination is additive in the neurocranium, but not in the viscerocranium. Injection of sub-optimal doses of prdm1a with prdm3 or prdm16 Morpholinos together leads to more severe phenotypes in the viscerocranium and neurocranium. prdm5 mutants have defects in the neurocranium and prdm1a and prdm5 double mutants also show more severe phenotypes. Overall, our data reveal that prdm3, 5, and 16 are involved in the zebrafish craniofacial development and that prdm1a may interact with prdm3, 5, and 16 in the formation of the craniofacial skeleton in zebrafish. Copyright © 2012 Wiley Periodicals, Inc.

Transcriptomic Analysis Of Purified Embryonic Neural Stem Cells From Zebrafish Embryos Reveals Signalling Pathways Involved In Glycine-dependent Neurogenesis

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Eric eSAMARUT

2016-03-01

Full Text Available How is the initial set of neurons correctly established during the development of the vertebrate central nervous system? In the embryo, glycine and GABA are depolarizing due the immature chloride gradient, which is only reversed to become hyperpolarizing later in post-natal development. We previously showed that glycine regulates neurogenesis via paracrine signalling that promotes calcium transients in neural stem cells (NSCs and their differentiation into interneurons within the spinal cord of the zebrafish embryo. However, the subjacent molecular mechanisms are not yet understood. Our previous work suggests that early neuronal progenitors were not differentiating correctly in the developing spinal cord. As a result, we aimed at identifying the downstream molecular mechanisms involved specifically in NSCs during glycine-dependent embryonic neurogenesis. Using a gfap:GFP transgenic line, we successfully purified NSCs by fluorescence-activated cell sorting (FACS from whole zebrafish embryos and in embryos in which the glycine receptor was knocked down. The strength of this approach is that it focused on the NSC population while tackling the biological issue in an in vivo context in whole zebrafish embryos. After sequencing the transcriptome by RNA-sequencing, we analyzed the genes whose expression was changed upon disruption of glycine signalling and we confirmed the differential expression by independent RTqPCR assay. While over a thousand genes showed altered expression levels, through pathway analysis we identified 14 top candidate genes belonging to five different canonical signalling pathways (signalling by calcium, TGF-beta, sonic hedgehog, Wnt and p53-related apoptosis that are likely to mediate the promotion of neurogenesis by glycine.

Ontogenetic expression of the Otx2 and Crx homeobox genes in the retina of the rat

DEFF Research Database (Denmark)

Rath, Martin F; Morin, Fabrice; Shi, Qiong

2007-01-01

. This confirmed the presence of Otx2 mRNA in both the embryonic retinal pigment epithelium and the developing neural retina. During development, the expression of Otx2 persists in the pigment epithelium, whereas Otx2 expression of the neural retina becomes progressively restricted to the outer nuclear layer......Otx2 and Crx are vertebrate orthologs of the orthodenticle family of homeobox genes, which are involved in retinal development. In this study, the temporal expression patterns of Otx2 and Crx in the rat retina during embryonic and postnatal stages of development were analyzed in detail...... and the outer part of the inner nuclear layer. Immunohistochemistry revealed that Otx2 protein is also present in cell bodies of the ganglion cell layer, which does not contain the Otx2 transcript, suggesting that Otx2 protein is synthesized in cell bodies of the bipolar neurons and then transported...

Zebrafish models for the functional genomics of neurogenetic disorders.

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Kabashi, Edor; Brustein, Edna; Champagne, Nathalie; Drapeau, Pierre

2011-03-01

In this review, we consider recent work using zebrafish to validate and study the functional consequences of mutations of human genes implicated in a broad range of degenerative and developmental disorders of the brain and spinal cord. Also we present technical considerations for those wishing to study their own genes of interest by taking advantage of this easily manipulated and clinically relevant model organism. Zebrafish permit mutational analyses of genetic function (gain or loss of function) and the rapid validation of human variants as pathological mutations. In particular, neural degeneration can be characterized at genetic, cellular, functional, and behavioral levels. Zebrafish have been used to knock down or express mutations in zebrafish homologs of human genes and to directly express human genes bearing mutations related to neurodegenerative disorders such as spinal muscular atrophy, ataxia, hereditary spastic paraplegia, amyotrophic lateral sclerosis (ALS), epilepsy, Huntington's disease, Parkinson's disease, fronto-temporal dementia, and Alzheimer's disease. More recently, we have been using zebrafish to validate mutations of synaptic genes discovered by large-scale genomic approaches in developmental disorders such as autism, schizophrenia, and non-syndromic mental retardation. Advances in zebrafish genetics such as multigenic analyses and chemical genetics now offer a unique potential for disease research. Thus, zebrafish hold much promise for advancing the functional genomics of human diseases, the understanding of the genetics and cell biology of degenerative and developmental disorders, and the discovery of therapeutics. This article is part of a Special Issue entitled Zebrafish Models of Neurological Diseases. Copyright © 2010 Elsevier B.V. All rights reserved.

Optimization of a GCaMP calcium indicator for neural activity imaging.

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Akerboom, Jasper; Chen, Tsai-Wen; Wardill, Trevor J; Tian, Lin; Marvin, Jonathan S; Mutlu, Sevinç; Calderón, Nicole Carreras; Esposti, Federico; Borghuis, Bart G; Sun, Xiaonan Richard; Gordus, Andrew; Orger, Michael B; Portugues, Ruben; Engert, Florian; Macklin, John J; Filosa, Alessandro; Aggarwal, Aman; Kerr, Rex A; Takagi, Ryousuke; Kracun, Sebastian; Shigetomi, Eiji; Khakh, Baljit S; Baier, Herwig; Lagnado, Leon; Wang, Samuel S-H; Bargmann, Cornelia I; Kimmel, Bruce E; Jayaraman, Vivek; Svoboda, Karel; Kim, Douglas S; Schreiter, Eric R; Looger, Loren L

2012-10-03

Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Recent efforts in protein engineering have significantly increased the performance of GECIs. The state-of-the art single-wavelength GECI, GCaMP3, has been deployed in a number of model organisms and can reliably detect three or more action potentials in short bursts in several systems in vivo. Through protein structure determination, targeted mutagenesis, high-throughput screening, and a battery of in vitro assays, we have increased the dynamic range of GCaMP3 by severalfold, creating a family of "GCaMP5" sensors. We tested GCaMP5s in several systems: cultured neurons and astrocytes, mouse retina, and in vivo in Caenorhabditis chemosensory neurons, Drosophila larval neuromuscular junction and adult antennal lobe, zebrafish retina and tectum, and mouse visual cortex. Signal-to-noise ratio was improved by at least 2- to 3-fold. In the visual cortex, two GCaMP5 variants detected twice as many visual stimulus-responsive cells as GCaMP3. By combining in vivo imaging with electrophysiology we show that GCaMP5 fluorescence provides a more reliable measure of neuronal activity than its predecessor GCaMP3. GCaMP5 allows more sensitive detection of neural activity in vivo and may find widespread applications for cellular imaging in general.

Chitosan derived co-spheroids of neural stem cells and mesenchymal stem cells for neural regeneration.

Science.gov (United States)

Han, Hao-Wei; Hsu, Shan-Hui

2017-10-01

Chitosan has been considered as candidate biomaterials for neural applications. The effective treatment of neurodegeneration or injury to the central nervous system (CNS) is still in lack nowadays. Adult neural stem cells (NSCs) represents a promising cell source to treat the CNS diseases but they are limited in number. Here, we developed the core-shell spheroids of NSCs (shell) and mesenchymal stem cells (MSCs, core) by co-culturing cells on the chitosan surface. The NSCs in chitosan derived co-spheroids displayed a higher survival rate than those in NSC homo-spheroids. The direct interaction of NSCs with MSCs in the co-spheroids increased the Notch activity and differentiation tendency of NSCs. Meanwhile, the differentiation potential of MSCs in chitosan derived co-spheroids was significantly enhanced toward neural lineages. Furthermore, NSC homo-spheroids and NSC/MSC co-spheroids derived on chitosan were evaluated for their in vivo efficacy by the embryonic and adult zebrafish brain injury models. The locomotion activity of zebrafish receiving chitosan derived NSC homo-spheroids or NSC/MSC co-spheroids was partially rescued in both models. Meanwhile, the higher survival rate was observed in the group of adult zebrafish implanted with chitosan derived NSC/MSC co-spheroids as compared to NSC homo-spheroids. These evidences indicate that chitosan may provide an extracellular matrix-like environment to drive the interaction and the morphological assembly between NSCs and MSCs and promote their neural differentiation capacities, which can be used for neural regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.

Inner neural retina loss in central retinal artery occlusion.

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Ikeda, Fumiko; Kishi, Shoji

2010-09-01

To report morphologic retinal changes and visual outcomes in acute and chronic central retinal artery occlusion (CRAO). We reviewed ten eyes of ten patients with CRAO (age, 65.3 ± 10.2 years) and measured retinal thicknesses at the central fovea and the perifovea using optical coherence tomography (OCT) over 8 ± 4 months. During the acute phase (within 10 days), the mean inner retinal thicknesses were 148% and 139% of normal values at 1 mm nasal and temporal to the fovea. They decreased to 22% and 11% of normal inner retinal thickness during the chronic phase (3 months or later). The retinal thickness at the perifovea decreased linearly until 3 months but was stable during the chronic phase. In contrast, the foveal thickness increased slightly in the acute phase but was equivalent to the normal level during the chronic phase. As a result of inner retinal atrophy, the foveal pit was shallow during the chronic phase. The final visual acuity was correlated positively with retinal thickness at the perifovea during the chronic CRAO phase. OCT showed that inner retinal necrosis with early swelling and late atrophy occurred in CRAO. The fovea and outer retina appeared to be excluded from ischemic change. The residual inner retina at the perifovea determined the final visual outcomes.

Retina-Inspired Filter.

Science.gov (United States)

Doutsi, Effrosyni; Fillatre, Lionel; Antonini, Marc; Gaulmin, Julien

2018-07-01

This paper introduces a novel filter, which is inspired by the human retina. The human retina consists of three different layers: the Outer Plexiform Layer (OPL), the inner plexiform layer, and the ganglionic layer. Our inspiration is the linear transform which takes place in the OPL and has been mathematically described by the neuroscientific model "virtual retina." This model is the cornerstone to derive the non-separable spatio-temporal OPL retina-inspired filter, briefly renamed retina-inspired filter, studied in this paper. This filter is connected to the dynamic behavior of the retina, which enables the retina to increase the sharpness of the visual stimulus during filtering before its transmission to the brain. We establish that this retina-inspired transform forms a group of spatio-temporal Weighted Difference of Gaussian (WDoG) filters when it is applied to a still image visible for a given time. We analyze the spatial frequency bandwidth of the retina-inspired filter with respect to time. It is shown that the WDoG spectrum varies from a lowpass filter to a bandpass filter. Therefore, while time increases, the retina-inspired filter enables to extract different kinds of information from the input image. Finally, we discuss the benefits of using the retina-inspired filter in image processing applications such as edge detection and compression.

Phenanthrene causes ocular developmental toxicity in zebrafish embryos and the possible mechanisms involved

International Nuclear Information System (INIS)

Huang, Lixing; Wang, Chonggang; Zhang, Youyu; Wu, Meifang; Zuo, Zhenghong

2013-01-01

Highlights: • Phe exposure caused obvious morphological changes in the retina. • Phe exposure caused apoptosis and reduction of cell proliferation in the retina. • Phe causes ocular toxicity might be via the AhR/Zeb1/Mitf/Pax6 signaling pathway. • AhR is a repressor of Zeb1. -- Abstract: Recent studies show that polycyclic aromatic hydrocarbons (PAHs) may be a candidate cause of developmental defects of the retina, but the mechanism is still unclear. We evaluated the mechanism(s) underlying PAH-induced retinal development defects due to exposure to environmental concentrations of Phenanthrene (Phe) in zebrafish. We found that exposure to environmental concentrations of Phe caused obvious morphological changes, developmental retardation, apoptosis, and reduction of cell proliferation in the retina. Our results indicated that Phe could cause visual system developmental defects. Phe exposure up-regulated aryl hydrocarbon receptor (AhR) and microphthalmia-associated transcription factor (Mtif) expression, and down-regulated zinc finger E-box binding homeobox 1 (Zeb1) and paired box 6 (Pax6). Moreover, we demonstrated that AhR was a repressor of Zeb1. We propose that Phe's ocular toxicity is mediated by up-regulating AhR, which then down-regulates Zeb1, in turn inducing Mitf expression while inhibiting Pax6 expression

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

Science.gov (United States)

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

2016-07-01

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

Egfl6 is involved in zebrafish notochord development.

Science.gov (United States)

Wang, Xueqian; Wang, Xin; Yuan, Wei; Chai, Renjie; Liu, Dong

2015-08-01

The epidermal growth factor (EGF) repeat motif defines a superfamily of diverse protein involved in regulating a variety of cellular and physiological processes, such as cell cycle, cell adhesion, proliferation, migration, and neural development. Egfl6, an EGF protein, also named MAGE was first cloned in human tissue. Up to date, the study of zebrafish Egfl6 expression pattern and functional analysis of Egfl6 involved in embryonic development of vertebrate in vivo is thus far lacking. Here we reported that Egfl6 was involved in zebrafish notochord development. It was shown that Egfl6 mRNA was expressed in zebrafish, developing somites, fin epidermis, pharyngeal arches, and hindbrain region. Particularly the secreted Egfl6 protein was significantly accumulated in notochord. Loss of Egfl6 function in zebrafish embryos resulted in curved body with distorted notochord in the posterior trunk. It was observed that expression of all Notch ligand and receptors in notochord of 28 hpf Egfl6 morphants was not affected, except notch2, which was up-regulated. We found that inhibition of Notch signaling by DAPT efficiently rescued notochord developmental defect of Egfl6 deficiency embryos.

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

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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.

Premature aging in telomerase-deficient zebrafish

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Monique Anchelin

2013-09-01

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

Imaging an optogenetic pH sensor reveals that protons mediate lateral inhibition in the retina.

Science.gov (United States)

Wang, Tzu-Ming; Holzhausen, Lars C; Kramer, Richard H

2014-02-01

The reciprocal synapse between photoreceptors and horizontal cells underlies lateral inhibition and establishes the antagonistic center-surround receptive fields of retinal neurons to enhance visual contrast. Despite decades of study, the signal mediating the negative feedback from horizontal cells to cones has remained under debate because the small, invaginated synaptic cleft has precluded measurement. Using zebrafish retinas, we show that light elicits a change in synaptic proton concentration with the correct magnitude, kinetics and spatial dependence to account for lateral inhibition. Light, which hyperpolarizes horizontal cells, causes synaptic alkalinization, whereas activating an exogenously expressed ligand-gated Na(+) channel, which depolarizes horizontal cells, causes synaptic acidification. Whereas acidification was prevented by blocking a proton pump, re-alkalinization was prevented by blocking proton-permeant ion channels, suggesting that distinct mechanisms underlie proton efflux and influx. These findings reveal that protons mediate lateral inhibition in the retina, raising the possibility that protons are unrecognized retrograde messengers elsewhere in the nervous system.

Identification of a locus control region for quadruplicated green-sensitive opsin genes in zebrafish

Science.gov (United States)

Tsujimura, Taro; Chinen, Akito; Kawamura, Shoji

2007-01-01

Duplication of opsin genes has a crucial role in the evolution of visual system. Zebrafish have four green-sensitive (RH2) opsin genes (RH2–1, RH2–2, RH2–3, and RH2–4) arrayed in tandem. They are expressed in the short member of the double cones (SDC) but differ in expression areas in the retina and absorption spectra of their encoding photopigments. The shortest and the second shortest wavelength subtypes, RH2–1 and RH2–2, are expressed in the central-to-dorsal retina. The longer wavelength subtype, RH2–3, is expressed circumscribing the RH2–1/RH2–2 area, and the longest subtype, RH2–4, is expressed further circumscribing the RH2–3 area and mainly occupying the ventral retina. The present report shows that a 0.5-kb region located 15 kb upstream of the RH2 gene array is an essential regulator for their expression. When the 0.5-kb region was deleted from a P1-artificial chromosome (PAC) clone encompassing the four RH2 genes and when one of these genes was replaced with a reporter GFP gene, the GFP expression in SDCs was abolished in the zebrafish to which a series of the modified PAC clones were introduced. Transgenic studies also showed that the 0.5-kb region conferred the SDC-specific expression for promoters of a non-SDC (UV opsin) and a nonretinal (keratin 8) gene. Changing the location of the 0.5-kb region in the PAC clone conferred the highest expression for its proximal gene. The 0.5-kb region was thus designated as RH2-LCR analogous to the locus control region of the L-M opsin genes of primates. PMID:17646658

Distribution and protective function of pituitary adenylate cyclase-activating polypeptide (PACAP in the retina

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Tomoya eNakamachi

2012-11-01

Full Text Available Pituitary adenylate cyclase-activating polypeptide (PACAP, which is found in 27- or 38-amino acid forms, belongs to the VIP/glucagon/secretin family. PACAP and its three receptor subtypes are expressed in neural tissues, with PACAP known to exert a protective effect against several types of neural damage. The retina is considered to be part of the central nervous system, and retinopathy is a common cause of profound and intractable loss of vision. This review will examine the expression and morphological distribution of PACAP and its receptors in the retina, and will summarize the current state of knowledge regarding the protective effect of PACAP against different kinds of retinal damage, such as that identified in association with diabetes, ultraviolet light, hypoxia, optic nerve transection, and toxins. This article will also address PACAP-mediated protective pathways involving retinal glial cells.

The Wnt Co-Receptor Lrp5 Is Required for Cranial Neural Crest Cell Migration in Zebrafish.

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Bernd Willems

Full Text Available During vertebrate neurulation, cranial neural crest cells (CNCCs undergo epithelial to mesenchymal transition (EMT, delaminate from the neural plate border, and migrate as separate streams into different cranial regions. There, they differentiate into distinct parts of the craniofacial skeleton. Canonical Wnt signaling has been shown to be essential for this process at different levels but the involved receptors remained unclear. Here we show that the frizzled co-receptor low-density-lipoprotein (LDL receptor-related protein 5 (Lrp5 plays a crucial role in CNCC migration and morphogenesis of the cranial skeleton. Early during induction and migration of CNCCs, lrp5 is expressed ubiquitously but later gets restricted to CNCC derivatives in the ventral head region besides different regions in the CNS. A knock-down of lrp5 does not interfere with induction of CNCCs but leads to reduced proliferation of premigratory CNCCs. In addition, cell migration is disrupted as CNCCs are found in clusters at ectopic positions in the dorsomedial neuroepithelium after lrp5 knock-down and transient CRISPR/Cas9 gene editing. These migratory defects consequently result in malformations of the craniofacial skeleton. To date, Lrp5 has mainly been associated with bone homeostasis in mammals. Here we show that in zebrafish, lrp5 also controls cell migration during early morphogenetic processes and contributes to shaping the craniofacial skeleton.

Towards Developmental Models of Psychiatric Disorders in Zebrafish

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William Howard James Norton

2013-04-01

Full Text Available Psychiatric disorders are a diverse set of diseases that affect all aspects of mental function including social interaction, thinking, feeling and mood. Although psychiatric disorders place a large economic burden on society, the drugs available to treat them are often palliative with variable efficacy and intolerable side-effects. The development of novel drugs has been hindered by a lack of knowledge about the etiology of these diseases. It is thus necessary to further investigate psychiatric disorders using a combination of human molecular genetics, gene-by-environment studies, in vitro pharmacological and biochemistry experiments, animal models and investigation of the non-biological basis of these diseases, such as environmental effects.Many psychiatric disorders, including autism spectrum disorder, attention-deficit/hyperactivity disorder, mental retardation and schizophrenia can be triggered by alterations to neural development. The zebrafish is a popular model for developmental biology that is increasingly used to study human disease. Recent work has extended this approach to examine psychiatric disorders as well. However, since psychiatric disorders affect complex mental functions that might be human specific, it is not possible to fully model them in fish. In this review, I will propose that the suitability of zebrafish for developmental studies, and the genetic tools available to manipulate them, provide a powerful model to study the roles of genes that are linked to psychiatric disorders during neural development. The relative speed and ease of conducting experiments in zebrafish can be used to address two areas of future research: the contribution of environmental factors to disease onset, and screening for novel therapeutic compounds.

Heparin-Binding EGF-like Growth Factor (HB-EGF) stimulates the proliferation of Müller glia-derived progenitor cells in avian and murine retinas

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Todd, Levi; Volkov, Leo I.; Zelinka, Chris; Squires, Natalie; Fischer, Andy J.

2015-01-01

Müller glia can be stimulated to de-differentiate, proliferate and form Müller glia-derived progenitor cells (MGPCs) that regenerate retinal neurons. In the zebrafish retina, Heparin-Binding EGF-like Growth Factor (HB-EGF) may be one of the key factors that stimulate the formation of proliferating MGPCs. Currently nothing is known about the influence of HB-EGF on the proliferative potential of Müller glia in retinas of birds and rodents. In the chick retina, we found that levels of both hb-egf and egf-receptor are rapidly and transiently up-regulated following NMDA-induced damage. Although intraocular injections of HB-EGF failed to stimulate cell-signaling or proliferation of Müller glia in normal retinas, HB-EGF stimulated proliferation of MGPCs in damaged retinas. By comparison, inhibition of the EGF-receptor (EGFR) decreased the proliferation of MGPCs in damaged retinas. HB-EGF failed to act synergistically with FGF2 to stimulate the formation of MGPCs in the undamaged retina and inhibition of EGF-receptor did not suppress FGF2-mediated formation of MGPCs. In the mouse retina, HB-EGF stimulated the proliferation of Müller glia following NMDA-induced damage. Furthermore, HB-EGF stimulated not only MAPK-signaling in Müller glia/MGPCs, but also activated mTor- and Jak/Stat-signaling. We propose that levels of expression of EGFR are rate-limiting to the responses of Müller glia to HB-EGF and the expression of EGFR can be induced by retinal damage, but not by FGF2-treatment. We conclude that HB-EGF is mitogenic to Müller glia in both chick and mouse retinas, and HB-EGF is an important player in the formation of MGPCs in damaged retinas. PMID:26500021

P3-7: On Prototyping a Visual Prosthesis System with Artificial Retina and Optic Nerve Based on Arrayed Microfibers

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Jian Hong Chen

2012-10-01

Full Text Available The traditional visual prosthesis system combines both a camera and a microelectrode array implanted on the visual neural network including retina, optic nerve, and visual cortex. Here, we introduce a new visual prosthesis system in which an artificial retina and optic nerve are demonstrated. The prototype of optic nerve for image transmission is comprised of arrayed PMMA microfibers with both ends connected with two planes, one functioned as retina for light reception and another attached to visual cortex. The microfibers are drawn from the thin film prepared by PMMA/chlorobenzene solution. Each micro fiber serves as an optical waveguide for the delivery of a single image pixel. It is demonstrated that with proper imaging optics, arrayed micro fibers could be lit as discrete light spots in accordance with the input image. Each micro fiber is expected to function as a stimulation unit for optical neural modulation in a visual prosthesis system.

A homeostatic, chip-based platform for zebrafish larvae immobilization and long-term imaging

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Friedrich, Timo; Zhu, Feng; Wlodkowic, Donald; Kaslin, Jan

2015-12-01

Zebrafish larvae are ideal for toxicology and drug screens due to their transparency, small size and similarity to humans on the genetic level. Using modern imaging techniques, cells and tissues can be dynamically visualised and followed over days in multiple zebrafish. Yet continued imaging experiments require specialized conditions such as: moisture and heat control to maintain specimen homeostasis. Chambers that control the environment are generally very expensive and are not always available for all imaging platforms. A highly customizable mounting configuration with built-in means of controlling temperature and media flow would therefore be a valuable tool for long term imaging experiments. Rapid prototyping using 3D printing is particularly suitable as a production method as it offers high flexibility in design, is widely available and allows a high degree of customizing. We study neural regeneration in zebrafish. Regeneration is limited in humans, but zebrafish recover from neural damage within days. Yet, the underlying regenerative mechanisms remain unclear. We developed an agarose based mounting system that holds the embryos in defined positions along removable strips. Homeostasis and temperature control is ensured by channels circulating buffer and heated water. This allows to image up to 120 larvae simultaneously for more than two days. Its flexibility and the low-volume, high larvae ratio will allow screening of small compound libraries. Taken together, we offer a low cost, highly adaptable solution for long term in-vivo imaging.

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.

2017 Midwest Zebrafish Meeting Report.

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

2017-12-01

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

Multimodality optical coherence tomography and fluorescence confocal scanning laser ophthalmoscopy in a zebrafish model of retinal vascular occlusion and remodeling

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Li, Xiaoyue; Spitz, Kathleen; Bozic, Ivan; Tao, Yuankai K.

2018-02-01

Neovascularization in diabetic retinopathy (DR) and age-related macular degeneration (AMD) result in severe vision-loss and are two of the leading causes of blindness. The structural, metabolic, and vascular changes underlying retinal neovascularization are unknown and, thus, there is an unmet need to identify mechanisms of pathogenesis and novel anti-angiogenic therapies. Zebrafish is a robust ophthalmological model because its retina has comparable structure to the human retina and its fecundity and life-cycle enable development of mutant phenotypes of human pathologies. Here, we perform multimodal imaging with OCT and fluorescence confocal scanning laser ophthalmoscopy (cSLO) to identify changes in retinal structure and function in a zebrafish model of vascular leakage. Transgenic zebrafish with EGFP tagged plasma protein were imaged longitudinally at six time points over two weeks to visualize vascular perfusion changes from diethylaminobenzaldehyde (DEAB) treatment. Complementary contrast from OCT-A perfusion maps and cSLO imaging of plasma protein EGFP shows vascular occlusions posttreatment. cSLO images confirm presence of vessels despite loss of OCT-A signal. Plasma protein EGFP contrast also shows significant changes in vessel structure as compared to baseline images. OCT structural volumes show empty vessel cross-sections confirming non-perfusion. In addition, we present algorithms for automated biometric identification of OCT datasets using OCT-A vascular patterns in the presence of significant vascular perfusion changes. These results establish a framework for large-scale in vivo assays to identify novel anti-angiogenic compounds and understand the mechanisms ofneovascularization associated with retinal ocular pathologies.

The ciliary marginal zone of the zebrafish retina: clonal and time-lapse analysis of a continuously growing tissue.

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Wan, Yinan; Almeida, Alexandra D; Rulands, Steffen; Chalour, Naima; Muresan, Leila; Wu, Yunmin; Simons, Benjamin D; He, Jie; Harris, William A

2016-04-01

Clonal analysis is helping us understand the dynamics of cell replacement in homeostatic adult tissues (Simons and Clevers, 2011). Such an analysis, however, has not yet been achieved for continuously growing adult tissues, but is essential if we wish to understand the architecture of adult organs. The retinas of lower vertebrates grow throughout life from retinal stem cells (RSCs) and retinal progenitor cells (RPCs) at the rim of the retina, called the ciliary marginal zone (CMZ). Here, we show that RSCs reside in a niche at the extreme periphery of the CMZ and divide asymmetrically along a radial (peripheral to central) axis, leaving one daughter in the peripheral RSC niche and the other more central where it becomes an RPC. We also show that RPCs of the CMZ have clonal sizes and compositions that are statistically similar to progenitor cells of the embryonic retina and fit the same stochastic model of proliferation. These results link embryonic and postembryonic cell behaviour, and help to explain the constancy of tissue architecture that has been generated over a lifetime. © 2016. Published by The Company of Biologists Ltd.

Real-time emulation of neural images in the outer retinal circuit.

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Hasegawa, Jun; Yagi, Tetsuya

2008-12-01

We describe a novel real-time system that emulates the architecture and functionality of the vertebrate retina. This system reconstructs the neural images formed by the retinal neurons in real time by using a combination of analog and digital systems consisting of a neuromorphic silicon retina chip, a field-programmable gate array, and a digital computer. While the silicon retina carries out the spatial filtering of input images instantaneously, using the embedded resistive networks that emulate the receptive field structure of the outer retinal neurons, the digital computer carries out the temporal filtering of the spatially filtered images to emulate the dynamical properties of the outer retinal circuits. The emulations of the neural image, including 128 x 128 bipolar cells, are carried out at a frame rate of 62.5 Hz. The emulation of the response to the Hermann grid and a spot of light and an annulus of lights has demonstrated that the system responds as expected by previous physiological and psychophysical observations. Furthermore, the emulated dynamics of neural images in response to natural scenes revealed the complex nature of retinal neuron activity. We have concluded that the system reflects the spatiotemporal responses of bipolar cells in the vertebrate retina. The proposed emulation system is expected to aid in understanding the visual computation in the retina and the brain.

Effect of titanium dioxide nanoparticles on zebrafish embryos and developing retina

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Ya-Jie Wang

2014-12-01

Full Text Available AIM:To investigate the impact of titanium dioxide nanoparticles (TiO2 NPs on embryonic development and retinal neurogenesis. METHODS:The agglomeration and sedimentation of TiO2 NPs solutions at different dilutions were observed, and the ultraviolet-visible spectra of their supernatants were measured. Zebrafish embryos were experimentally exposed to TiO2 NPs until 72h postfertilization (hpf. The retinal neurogenesis and distribution of the microglia were analyzed by immunohistochemistry and whole mount in situ hybridization. RESULTS: The1 mg/L was determined to be an appropriate exposure dose. Embryos exposed to TiO2 NPs had a normal phenotype. The neurogenesis was initiated on time, and ganglion cells, cones and rods were well differentiated at 72 hpf. The expression of fms mRNA and the 4C4 antibody, which were specific to microglia in the central nervous system (CNS, closely resembled their endogenous profile. CONCLUSION:These data demonstrate that short-term exposure to TiO2 NPs at a low dose does not lead to delayed embryonic development or retinal neurotoxicity.

Adaptation in Coding by Large Populations of Neurons in the Retina

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Ioffe, Mark L.

A comprehensive theory of neural computation requires an understanding of the statistical properties of the neural population code. The focus of this work is the experimental study and theoretical analysis of the statistical properties of neural activity in the tiger salamander retina. This is an accessible yet complex system, for which we control the visual input and record from a substantial portion--greater than a half--of the ganglion cell population generating the spiking output. Our experiments probe adaptation of the retina to visual statistics: a central feature of sensory systems which have to adjust their limited dynamic range to a far larger space of possible inputs. In Chapter 1 we place our work in context with a brief overview of the relevant background. In Chapter 2 we describe the experimental methodology of recording from 100+ ganglion cells in the tiger salamander retina. In Chapter 3 we first present the measurements of adaptation of individual cells to changes in stimulation statistics and then investigate whether pairwise correlations in fluctuations of ganglion cell activity change across different stimulation conditions. We then transition to a study of the population-level probability distribution of the retinal response captured with maximum-entropy models. Convergence of the model inference is presented in Chapter 4. In Chapter 5 we first test the empirical presence of a phase transition in such models fitting the retinal response to different experimental conditions, and then proceed to develop other characterizations which are sensitive to complexity in the interaction matrix. This includes an analysis of the dynamics of sampling at finite temperature, which demonstrates a range of subtle attractor-like properties in the energy landscape. These are largely conserved when ambient illumination is varied 1000-fold, a result not necessarily apparent from the measured low-order statistics of the distribution. Our results form a consistent

Transcriptomic analysis across nasal, temporal, and macular regions of human neural retina and RPE/choroid by RNA-Seq

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Whitmore, S. Scott; Wagner, Alex H.; DeLuca, Adam P.; Drack, Arlene V.; Stone, Edwin M.; Tucker, Budd A.; Zeng, Shemin; Braun, Terry A.; Mullins, Robert F.; Scheetz, Todd E.

2014-01-01

Proper spatial differentiation of retinal cell types is necessary for normal human vision. Many retinal diseases, such as Best disease and male germ cell associated kinase (MAK)-associated retinitis pigmentosa, preferentially affect distinct topographic regions of the retina. While much is known about the distribution of cell-types in the retina, the distribution of molecular components across the posterior pole of the eye has not been well-studied. To investigate regional difference in molecular composition of ocular tissues, we assessed differential gene expression across the temporal, macular, and nasal retina and retinal pigment epithelium (RPE)/choroid of human eyes using RNA-Seq. RNA from temporal, macular, and nasal retina and RPE/choroid from four human donor eyes was extracted, poly-A selected, fragmented, and sequenced as 100 bp read pairs. Digital read files were mapped to the human genome and analyzed for differential expression using the Tuxedo software suite. Retina and RPE/choroid samples were clearly distinguishable at the transcriptome level. Numerous transcription factors were differentially expressed between regions of the retina and RPE/choroid. Photoreceptor-specific genes were enriched in the peripheral samples, while ganglion cell and amacrine cell genes were enriched in the macula. Within the RPE/choroid, RPE-specific genes were upregulated at the periphery while endothelium associated genes were upregulated in the macula. Consistent with previous studies, BEST1 expression was lower in macular than extramacular regions. The MAK gene was expressed at lower levels in macula than in extramacular regions, but did not exhibit a significant difference between nasal and temporal retina. The regional molecular distinction is greatest between macula and periphery and decreases between different peripheral regions within a tissue. Datasets such as these can be used to prioritize candidate genes for possible involvement in retinal diseases with

Transcriptomic analysis across nasal, temporal, and macular regions of human neural retina and RPE/choroid by RNA-Seq.

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Whitmore, S Scott; Wagner, Alex H; DeLuca, Adam P; Drack, Arlene V; Stone, Edwin M; Tucker, Budd A; Zeng, Shemin; Braun, Terry A; Mullins, Robert F; Scheetz, Todd E

2014-12-01

Proper spatial differentiation of retinal cell types is necessary for normal human vision. Many retinal diseases, such as Best disease and male germ cell associated kinase (MAK)-associated retinitis pigmentosa, preferentially affect distinct topographic regions of the retina. While much is known about the distribution of cell types in the retina, the distribution of molecular components across the posterior pole of the eye has not been well-studied. To investigate regional difference in molecular composition of ocular tissues, we assessed differential gene expression across the temporal, macular, and nasal retina and retinal pigment epithelium (RPE)/choroid of human eyes using RNA-Seq. RNA from temporal, macular, and nasal retina and RPE/choroid from four human donor eyes was extracted, poly-A selected, fragmented, and sequenced as 100 bp read pairs. Digital read files were mapped to the human genome and analyzed for differential expression using the Tuxedo software suite. Retina and RPE/choroid samples were clearly distinguishable at the transcriptome level. Numerous transcription factors were differentially expressed between regions of the retina and RPE/choroid. Photoreceptor-specific genes were enriched in the peripheral samples, while ganglion cell and amacrine cell genes were enriched in the macula. Within the RPE/choroid, RPE-specific genes were upregulated at the periphery while endothelium associated genes were upregulated in the macula. Consistent with previous studies, BEST1 expression was lower in macular than extramacular regions. The MAK gene was expressed at lower levels in macula than in extramacular regions, but did not exhibit a significant difference between nasal and temporal retina. The regional molecular distinction is greatest between macula and periphery and decreases between different peripheral regions within a tissue. Datasets such as these can be used to prioritize candidate genes for possible involvement in retinal diseases with

Distinct functional and temporal requirements for zebrafish Hdac1 during neural crest-derived craniofacial and peripheral neuron development.

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Myron S Ignatius

Full Text Available The regulation of gene expression is accomplished by both genetic and epigenetic means and is required for the precise control of the development of the neural crest. In hdac1(b382 mutants, craniofacial cartilage development is defective in two distinct ways. First, fewer hoxb3a, dlx2 and dlx3-expressing posterior branchial arch precursors are specified and many of those that are consequently undergo apoptosis. Second, in contrast, normal numbers of progenitors are present in the anterior mandibular and hyoid arches, but chondrocyte precursors fail to terminally differentiate. In the peripheral nervous system, there is a disruption of enteric, DRG and sympathetic neuron differentiation in hdac1(b382 mutants compared to wildtype embryos. Specifically, enteric and DRG-precursors differentiate into neurons in the anterior gut and trunk respectively, while enteric and DRG neurons are rarely present in the posterior gut and tail. Sympathetic neuron precursors are specified in hdac1(b382 mutants and they undergo generic neuronal differentiation but fail to undergo noradrenergic differentiation. Using the HDAC inhibitor TSA, we isolated enzyme activity and temporal requirements for HDAC function that reproduce hdac1(b382 defects in craniofacial and sympathetic neuron development. Our study reveals distinct functional and temporal requirements for zebrafish hdac1 during neural crest-derived craniofacial and peripheral neuron development.

Distinct functional and temporal requirements for zebrafish Hdac1 during neural crest-derived craniofacial and peripheral neuron development.

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Ignatius, Myron S; Unal Eroglu, Arife; Malireddy, Smitha; Gallagher, Glen; Nambiar, Roopa M; Henion, Paul D

2013-01-01

The regulation of gene expression is accomplished by both genetic and epigenetic means and is required for the precise control of the development of the neural crest. In hdac1(b382) mutants, craniofacial cartilage development is defective in two distinct ways. First, fewer hoxb3a, dlx2 and dlx3-expressing posterior branchial arch precursors are specified and many of those that are consequently undergo apoptosis. Second, in contrast, normal numbers of progenitors are present in the anterior mandibular and hyoid arches, but chondrocyte precursors fail to terminally differentiate. In the peripheral nervous system, there is a disruption of enteric, DRG and sympathetic neuron differentiation in hdac1(b382) mutants compared to wildtype embryos. Specifically, enteric and DRG-precursors differentiate into neurons in the anterior gut and trunk respectively, while enteric and DRG neurons are rarely present in the posterior gut and tail. Sympathetic neuron precursors are specified in hdac1(b382) mutants and they undergo generic neuronal differentiation but fail to undergo noradrenergic differentiation. Using the HDAC inhibitor TSA, we isolated enzyme activity and temporal requirements for HDAC function that reproduce hdac1(b382) defects in craniofacial and sympathetic neuron development. Our study reveals distinct functional and temporal requirements for zebrafish hdac1 during neural crest-derived craniofacial and peripheral neuron development.

Expression and knockdown of zebrafish folliculin suggests requirement for embryonic brain morphogenesis.

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Kenyon, Emma J; Luijten, Monique N H; Gill, Harmeet; Li, Nan; Rawlings, Matthew; Bull, James C; Hadzhiev, Yavor; van Steensel, Maurice A M; Maher, Eamonn; Mueller, Ferenc

2016-07-08

Birt-Hogg-Dubé syndrome (BHD) is a dominantly inherited familial cancer syndrome characterised by the development of benign skin fibrofolliculomas, multiple lung and kidney cysts, spontaneous pneumothorax and susceptibility to renal cell carcinoma. BHD is caused by mutations in the gene encoding Folliculin (FLCN). Little is known about what FLCN does in a healthy individual and how best to treat those with BHD. As a first approach to developing a vertebrate model for BHD we aimed to identify the temporal and spatial expression of flcn transcripts in the developing zebrafish embryo. To gain insights into the function of flcn in a whole organism system we generated a loss of function model of flcn by the use of morpholino knockdown in zebrafish. flcn is expressed broadly and upregulated in the fin bud, somites, eye and proliferative regions of the brain of the Long-pec stage zebrafish embryos. Together with knockdown phenotypes, expression analysis suggest involvement of flcn in zebrafish embryonic brain development. We have utilised the zFucci system, an in vivo, whole organism cell cycle assay to study the potential role of flcn in brain development. We found that at the 18 somite stage there was a significant drop in cells in the S-M phase of the cell cycle in flcn morpholino injected embryos with a corresponding increase of cells in the G1 phase. This was particularly evident in the brain, retina and somites of the embryo. Timelapse analysis of the head region of flcn morpholino injected and mismatch control embryos shows the temporal dynamics of cell cycle misregulation during development. In conclusion we show that zebrafish flcn is expressed in a non-uniform manner and is likely required for the maintenance of correct cell cycle regulation during embryonic development. We demonstrate the utilisation of the zFucci system in testing the role of flcn in cell proliferation and suggest a function for flcn in regulating cell proliferation in vertebrate embryonic

Desprendimiento de retina

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L. Jaime Claramunt, Dr.

2010-01-01

El desprendimiento de retina (DR) consiste en la separación entre la retina neurosensorial y el epitelio pigmentario subyacente. Su forma más frecuente es el DR regmatógeno, causado por una rotura en la retina. Se manifiesta generalmente como un defecto en el campo visual o mala visión. Si se pesquisa y trata oportunamente tiene buenas posibilidades de éxito. No obstante, sigue siendo una causa importante de mala visión y ceguera, por lo que su prevención tiene un rol fundamental.

Desprendimiento de retina

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L. Jaime Claramunt, Dr.

2010-11-01

Full Text Available El desprendimiento de retina (DR consiste en la separación entre la retina neurosensorial y el epitelio pigmentario subyacente. Su forma más frecuente es el DR regmatógeno, causado por una rotura en la retina. Se manifiesta generalmente como un defecto en el campo visual o mala visión. Si se pesquisa y trata oportunamente tiene buenas posibilidades de éxito. No obstante, sigue siendo una causa importante de mala visión y ceguera, por lo que su prevención tiene un rol fundamental.

Novel expression patterns of metabotropic glutamate receptor 6 in the zebrafish nervous system.

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Ying-Yu Huang

Full Text Available The metabotropic glutamate receptor 6 (mGluR6 or GRM6 belongs to the class III of the metabotropic glutamate receptor family. It is the only known mGluR that mediates direct synaptic transmission in the nervous system and is thought to mediate the ON-response in the ON-pathway of the vertebrate retina. Phylogenetic and gene structure analysis indicated that the zebrafish genome harbours two mglur6 paralogs, mglur6a and mglur6b. Besides expression in the inner nuclear layer and distinct regions in the brain, both mglur6 paralogs are expressed in ganglion cells of the retina, an expression pattern which can also be observed in the downstream effector molecules gnaoa and gnaob. This unexpected expression pattern is consistent with immunohistological labeling using a peptide antibody specific for the mGluR6b paralog. These expression patterns contradict the existing view that mGluR6 is solely located on ON-bipolar cells where it functions in signal transmission. Consistent with expression in ON-bipolar cells, we report a decreased b-wave amplitude in the electroretinogram after morpholino-based downregulation of mGluR6b, showing a function in the ON response. Our data suggest more widespread functions of mGluR6 mediated signaling in the central nervous system, possibly including sign reversing synapses in the inner retina.

Disruption of the folate pathway in zebrafish causes developmental defects

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Lee Marina S

2012-04-01

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

Comparative proteomic analyses of macular and peripheral retina of cynomolgus monkeys (Macaca fascicularis).

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Okamoto, Haru; Umeda, Shinsuke; Nozawa, Takehiro; Suzuki, Michihiro T; Yoshikawa, Yasuhiro; Matsuura, Etsuko T; Iwata, Takeshi

2010-01-01

The central region of the primate retina is called the macula. The fovea is located at the center of the macula, where the photoreceptors are concentrated to create a neural network adapted for high visual acuity. Damage to the fovea, e.g., by macular dystrophies and age-related macular degeneration, can reduce central visual acuity. The molecular mechanisms leading to these diseases are most likely dependent on the proteins in the macula which differ from those in the peripheral retina in expression level. To investigate whether the distribution of proteins in the macula is different from the peripheral retina, proteomic analyses of tissues from these two regions of cynomolgus monkeys were compared. Two-dimensional gel electrophoresis and mass spectrometry identified 26 proteins that were present only in the macular gel spots. The expression levels of five proteins, cone photoreceptor specific arrestin-C, gamma-synuclein, epidermal fatty acid binding protein, tropomyosin 1alpha chain, and heterogeneous nuclear ribonucleoproteins A2/B1, were significantly higher in the macula than in the peripheral retina. Immunostaining of macula sections by antibodies to each identified protein revealed unique localization in the retina, retinal pigment epithelial cells and the choroidal layer. Some of these proteins were located in cells with higher densities in the macula. We suggest that it will be important to study these proteins to determine their contribution to the pathogenesis and progression of macula diseases.

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

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

2015-06-18

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

Apical Polarity Protein PrkCi Is Necessary for Maintenance of Spinal Cord Precursors in Zebrafish

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Roberts, Randolph K.; Appel, Bruce

2009-01-01

During development, neural precursors divide to produce new precursors and cells that differentiate as neurons and glia. In Drosophila, apicobasal polarity and orientation of the mitotic spindle play important roles in specifying the progeny of neural precursors for different fates. We examined orientation of zebrafish spinal cord precursors using time-lapse imaging and tested the function of protein kinase C, iota (PrkCi), a member of the Par complex of proteins necessary for apicobasal pola...

Zebrafish Adar2 Edits the Q/R site of AMPA receptor Subunit gria2α transcript to ensure normal development of nervous system and cranial neural crest cells.

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I-Chen Li

Full Text Available BACKGROUND: Adar2 deaminates selective adenosines to inosines (A-to-I RNA editing in the double-stranded region of nuclear transcripts. Although the functions of mouse Adar2 and its biologically most important substrate gria2, encoding the GluA2 subunit of AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor, have been extensively studied, the substrates and functions of zebrafish Adar2 remain elusive. METHODS/PRINCIPAL FINDINGS: Expression of Adar2 was perturbed in the adar2 morphant (adar2MO, generated by antisense morpholio oligonucleotides. The Q/R editing of gria2α was reduced in the adar2MO and was enhanced by overexpression of Adar2, demonstrating an evolutionarily conserved activity between zebrafish and mammalian Adar2 in editing the Q/R site of gria2. To delineate the role of Q/R editing of gria2α in the developmental defects observed in the adar2MO, the Q/R editing of gria2α was specifically perturbed in the gria2αQRMO, generated by a morpholio oligonucleotide complementary to the exon complementary sequence (ECS required for the Q/R editing. Analogous to the adar2-deficient and Q/R-editing deficient mice displaying identical neurological defects, the gria2αQRMO and adar2MO displayed identical developmental defects in the nervous system and cranial cartilages. Knockdown p53 abolished apoptosis and partially suppressed the loss of spinal cord motor neurons in these morphants. However, reducing p53 activity neither replenished the brain neuronal populations nor rescued the developmental defects. The expressions of crestin and sox9b in the neural crest cells were reduced in the adar2MO and gria2αQRMO. Overexpressing the edited GluA2αR in the adar2MO restored normal expressions of cresting and sox9b. Moreover, overexpressing the unedited GluA2αQ in the wild type embryos resulted in reduction of crestin and sox9b expressions. These results argue that an elevated GluA2αQ level is sufficient for generating the

Defects in the outer limiting membrane are associated with rosette development in the Nrl-/- retina.

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Michael W Stuck

Full Text Available The neural retinal leucine zipper (Nrl knockout mouse is a widely used model to study cone photoreceptor development, physiology, and molecular biology in the absence of rods. In the Nrl(-/- retina, rods are converted into functional cone-like cells. The Nrl(-/- retina is characterized by large undulations of the outer nuclear layer (ONL commonly known as rosettes. Here we explore the mechanism of rosette development in the Nrl(-/- retina. We report that rosettes first appear at postnatal day (P8, and that the structure of nascent rosettes is morphologically distinct from what is seen in the adult retina. The lumen of these nascent rosettes contains a population of aberrant cells protruding into the subretinal space that induce infolding of the ONL. Morphologically adult rosettes do not contain any cell bodies and are first detected at P15. The cells found in nascent rosettes are photoreceptors in origin but lack inner and outer segments. We show that the adherens junctions between photoreceptors and Müller glia which comprise the retinal outer limiting membrane (OLM are not uniformly formed in the Nrl(-/- retina and thus allow protrusion of a population of developing photoreceptors into the subretinal space where their maturation becomes delayed. These data suggest that the rosettes of the Nrl(-/- retina arise due to defects in the OLM and delayed maturation of a subset of photoreceptors, and that rods may play an important role in the proper formation of the OLM.

Effect of Monocular Deprivation on Rabbit Neural Retinal Cell Densities.

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Mwachaka, Philip Maseghe; Saidi, Hassan; Odula, Paul Ochieng; Mandela, Pamela Idenya

2015-01-01

To describe the effect of monocular deprivation on densities of neural retinal cells in rabbits. Thirty rabbits, comprised of 18 subject and 12 control animals, were included and monocular deprivation was achieved through unilateral lid suturing in all subject animals. The rabbits were observed for three weeks. At the end of each week, 6 experimental and 3 control animals were euthanized, their retinas was harvested and processed for light microscopy. Photomicrographs of the retina were taken and imported into FIJI software for analysis. Neural retinal cell densities of deprived eyes were reduced along with increasing period of deprivation. The percentage of reductions were 60.9% (P < 0.001), 41.6% (P = 0.003), and 18.9% (P = 0.326) for ganglion, inner nuclear, and outer nuclear cells, respectively. In non-deprived eyes, cell densities in contrast were increased by 116% (P < 0.001), 52% (P < 0.001) and 59.6% (P < 0.001) in ganglion, inner nuclear, and outer nuclear cells, respectively. In this rabbit model, monocular deprivation resulted in activity-dependent changes in cell densities of the neural retina in favour of the non-deprived eye along with reduced cell densities in the deprived eye.

Regulation of zebrafish sleep and arousal states: current and prospective approaches

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Cindy N Chiu

2013-04-01

Full Text Available Every day, we shift among various states of sleep and arousal to meet the many demands of our bodies and environment. A central puzzle in neurobiology is how the brain controls these behavioral states, which are essential to an animal’s well-being and survival. Mammalian models have predominated sleep and arousal research, although in the past decade, invertebrate models have made significant contributions to our understanding of the genetic underpinnings of behavioral states. More recently, the zebrafish has emerged as a promising model system for sleep and arousal research. Here we review experimental evidence that the zebrafish, a diurnal vertebrate, exhibits fundamental behavioral and neurochemical characteristics of mammalian sleep and arousal. We also propose how specific advantages of the zebrafish can be harnessed to advance the field. These include tractable genetics to identify and manipulate molecular and cellular regulators of behavioral states, optical transparency to facilitate in vivo observation of neural structure and function, and amenability to high-throughput drug screens to discover novel therapies for neurological disorders.

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

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

2018-04-01

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

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

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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.

Human neural progenitor cells decrease photoreceptor degeneration, normalize opsin distribution and support synapse structure in cultured porcine retina.

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Mollick, Tanzina; Mohlin, Camilla; Johansson, Kjell

2016-09-01

Retinal neurodegenerative disorders like retinitis pigmentosa, age-related macular degeneration, diabetic retinopathy and retinal detachment decrease retinal functionality leading to visual impairment. The pathological events are characterized by photoreceptor degeneration, synaptic disassembly, remodeling of postsynaptic neurons and activation of glial cells. Despite intense research, no effective treatment has been found for these disorders. The current study explores the potential of human neural progenitor cell (hNPC) derived factors to slow the degenerative processes in adult porcine retinal explants. Retinas were cultured for 3 days with or without hNPCs as a feeder layer and investigated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), immunohistochemical, western blot and quantitative real time-polymerase chain reaction (qRT-PCR) techniques. TUNEL showed that hNPCs had the capacity to limit photoreceptor cell death. Among cone photoreceptors, hNPC coculture resulted in better maintenance of cone outer segments and reduced opsin mislocalization. Additionally, maintained synaptic structural integrity and preservation of second order calbindin positive horizontal cells was also observed. However, Müller cell gliosis only seemed to be alleviated in terms of reduced Müller cell density. Our observations indicate that at 3 days of coculture, hNPC derived factors had the capacity to protect photoreceptors, maintain synaptic integrity and support horizontal cell survival. Human neural progenitor cell applied treatment modalities may be an effective strategy to help maintain retinal functionality in neurodegenerative pathologies. Whether hNPCs can independently hinder Müller cell gliosis by utilizing higher concentrations or by combination with other pharmacological agents still needs to be determined. Copyright © 2016 Elsevier B.V. All rights reserved.

Mechanisms of prickle1a function in zebrafish epilepsy and retinal neurogenesis

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Xue Mei

2013-05-01

Epilepsy is a complex neurological disorder characterized by unprovoked seizures. The etiology is heterogeneous with both genetic and environmental causes. Genes that regulate neurotransmitters and ion channels in the central nervous system have been associated with epilepsy. However, a recent screening in human epilepsy patients identified mutations in the PRICKLE1 (PK1 locus, highlighting a potentially novel mechanism underlying seizures. PK1 is a core component of the planar cell polarity network that regulates tissue polarity. Zebrafish studies have shown that Pk1 coordinates cell movement, neuronal migration and axonal outgrowth during embryonic development. Yet how dysfunction of Pk1 relates to epilepsy is unknown. To address the mechanism underlying epileptogenesis, we used zebrafish to characterize Pk1a function and epilepsy-related mutant forms. We show that knockdown of pk1a activity sensitizes zebrafish larva to a convulsant drug. To model defects in the central nervous system, we used the retina and found that pk1a knockdown induces neurite outgrowth defects; yet visual function is maintained. Furthermore, we characterized the functional and biochemical properties of the PK1 mutant forms identified in human patients. Functional analyses demonstrate that the wild-type Pk1a partially suppresses the gene knockdown retinal defects but not the mutant forms. Biochemical analysis reveals increased ubiquitylation of one mutant form and decreased translational efficiency of another mutant form compared with the wild-type Pk1a. Taken together, our results indicate that mutation of human PK1 could lead to defects in neurodevelopment and signal processing, providing insight into seizure predisposition in these patients.

Self-organising aggregates of zebrafish retinal cells for investigating mechanisms of neural lamination.

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Eldred, Megan K; Charlton-Perkins, Mark; Muresan, Leila; Harris, William A

2017-03-15

To investigate the cell-cell interactions necessary for the formation of retinal layers, we cultured dissociated zebrafish retinal progenitors in agarose microwells. Within these wells, the cells re-aggregated within hours, forming tight retinal organoids. Using a Spectrum of Fates zebrafish line, in which all different types of retinal neurons show distinct fluorescent spectra, we found that by 48 h in culture, the retinal organoids acquire a distinct spatial organisation, i.e. they became coarsely but clearly laminated. Retinal pigment epithelium cells were in the centre, photoreceptors and bipolar cells were next most central and amacrine cells and retinal ganglion cells were on the outside. Image analysis allowed us to derive quantitative measures of lamination, which we then used to find that Müller glia, but not RPE cells, are essential for this process. © 2017. Published by The Company of Biologists Ltd.

Molecular Characterization of Notch1 Positive Progenitor Cells in the Developing Retina.

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Galina Dvoriantchikova

Full Text Available The oscillatory expression of Notch signaling in neural progenitors suggests that both repressors and activators of neural fate specification are expressed in the same progenitors. Since Notch1 regulates photoreceptor differentiation and contributes (together with Notch3 to ganglion cell fate specification, we hypothesized that genes encoding photoreceptor and ganglion cell fate activators would be highly expressed in Notch1 receptor-bearing (Notch1+ progenitors, directing these cells to differentiate into photoreceptors or into ganglion cells when Notch1 activity is diminished. To identify these genes, we used microarray analysis to study expression profiles of whole retinas and isolated from them Notch1+ cells at embryonic day 14 (E14 and postnatal day 0 (P0. To isolate Notch1+ cells, we utilized immunomagnetic cell separation. We also used Notch3 knockout (Notch3KO animals to evaluate the contribution of Notch3 signaling in ganglion cell differentiation. Hierarchical clustering of 6,301 differentially expressed genes showed that Notch1+ cells grouped near the same developmental stage retina cluster. At E14, we found higher expression of repressors (Notch1, Hes5 and activators (Dll3, Atoh7, Otx2 of neuronal differentiation in Notch1+ cells compared to whole retinal cell populations. At P0, Notch1, Hes5, and Dll1 expression was significantly higher in Notch1+ cells than in whole retinas. Otx2 expression was more than thirty times higher than Atoh7 expression in Notch1+ cells at P0. We also observed that retinas of wild type animals had only 14% (P < 0.05 more ganglion cells compared to Notch3KO mice. Since this number is relatively small and Notch1 has been shown to contribute to ganglion cell fate specification, we suggested that Notch1 signaling may play a more significant role in RGC development than the Notch3 signaling cascade. Finally, our findings suggest that Notch1+ progenitors--since they heavily express both pro-ganglion cell (Atoh7

Whole-brain activity mapping onto a zebrafish brain atlas.

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

2015-11-01

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

Whole-brain activity mapping onto a zebrafish brain atlas

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

2015-01-01

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

A simple automated system for appetitive conditioning of zebrafish in their home tanks.

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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. Copyright © 2016 Elsevier B.V. All rights reserved.

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

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LeClair, Elizabeth E; Topczewski, Jacek

2010-01-15

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. 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 barbel can regenerate after repeated injury and also in

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

2010-01-01

Full Text Available 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.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 barbel can regenerate after repeated injury and

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

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Orger, Michael B; Portugues, Ruben

2016-01-01

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

Stereotyped Synaptic Connectivity Is Restored during Circuit Repair in the Adult Mammalian Retina.

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Beier, Corinne; Palanker, Daniel; Sher, Alexander

2018-06-04

Proper function of the central nervous system (CNS) depends on the specificity of synaptic connections between cells of various types. Cellular and molecular mechanisms responsible for the establishment and refinement of these connections during development are the subject of an active area of research [1-6]. However, it is unknown if the adult mammalian CNS can form new type-selective synapses following neural injury or disease. Here, we assess whether selective synaptic connections can be reestablished after circuit disruption in the adult mammalian retina. The stereotyped circuitry at the first synapse in the retina, as well as the relatively short distances new neurites must travel compared to other areas of the CNS, make the retina well suited to probing for synaptic specificity during circuit reassembly. Selective connections between short-wavelength sensitive cone photoreceptors (S-cones) and S-cone bipolar cells provides the foundation of the primordial blue-yellow vision, common to all mammals [7-18]. We take advantage of the ground squirrel retina, which has a one-to-one S-cone-to-S-cone-bipolar-cell connection, to test if this connectivity can be reestablished following local photoreceptor loss [8, 19]. We find that after in vivo selective photoreceptor ablation, deafferented S-cone bipolar cells expand their dendritic trees. The new dendrites randomly explore the proper synaptic layer, bypass medium-wavelength sensitive cone photoreceptors (M-cones), and selectively synapse with S-cones. However, non-connected dendrites are not pruned back to resemble unperturbed S-cone bipolar cells. We show, for the first time, that circuit repair in the adult mammalian retina can recreate stereotypic selective wiring. Copyright © 2018 Elsevier Ltd. All rights reserved.

Can Xanthophyll-Membrane Interactions Explain Their Selective Presence in the Retina and Brain?

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Widomska, Justyna; Zareba, Mariusz; Subczynski, Witold Karol

2016-01-01

Epidemiological studies demonstrate that a high dietary intake of carotenoids may offer protection against age-related macular degeneration, cancer and cardiovascular and neurodegenerative diseases. Humans cannot synthesize carotenoids and depend on their dietary intake. Major carotenoids that have been found in human plasma can be divided into two groups, carotenes (nonpolar molecules, such as β-carotene, α-carotene or lycopene) and xanthophylls (polar carotenoids that include an oxygen atom in their structure, such as lutein, zeaxanthin and β-cryptoxanthin). Only two dietary carotenoids, namely lutein and zeaxanthin (macular xanthophylls), are selectively accumulated in the human retina. A third carotenoid, meso-zeaxanthin, is formed directly in the human retina from lutein. Additionally, xanthophylls account for about 70% of total carotenoids in all brain regions. Some specific properties of these polar carotenoids must explain why they, among other available carotenoids, were selected during evolution to protect the retina and brain. It is also likely that the selective uptake and deposition of macular xanthophylls in the retina and brain are enhanced by specific xanthophyll-binding proteins. We hypothesize that the high membrane solubility and preferential transmembrane orientation of macular xanthophylls distinguish them from other dietary carotenoids, enhance their chemical and physical stability in retina and brain membranes and maximize their protective action in these organs. Most importantly, xanthophylls are selectively concentrated in the most vulnerable regions of lipid bilayer membranes enriched in polyunsaturated lipids. This localization is ideal if macular xanthophylls are to act as lipid-soluble antioxidants, which is the most accepted mechanism through which lutein and zeaxanthin protect neural tissue against degenerative diseases. PMID:27030822

Can Xanthophyll-Membrane Interactions Explain Their Selective Presence in the Retina and Brain?

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Justyna Widomska

2016-01-01

Full Text Available Epidemiological studies demonstrate that a high dietary intake of carotenoids may offer protection against age-related macular degeneration, cancer and cardiovascular and neurodegenerative diseases. Humans cannot synthesize carotenoids and depend on their dietary intake. Major carotenoids that have been found in human plasma can be divided into two groups, carotenes (nonpolar molecules, such as β-carotene, α-carotene or lycopene and xanthophylls (polar carotenoids that include an oxygen atom in their structure, such as lutein, zeaxanthin and β-cryptoxanthin. Only two dietary carotenoids, namely lutein and zeaxanthin (macular xanthophylls, are selectively accumulated in the human retina. A third carotenoid, meso-zeaxanthin, is formed directly in the human retina from lutein. Additionally, xanthophylls account for about 70% of total carotenoids in all brain regions. Some specific properties of these polar carotenoids must explain why they, among other available carotenoids, were selected during evolution to protect the retina and brain. It is also likely that the selective uptake and deposition of macular xanthophylls in the retina and brain are enhanced by specific xanthophyll-binding proteins. We hypothesize that the high membrane solubility and preferential transmembrane orientation of macular xanthophylls distinguish them from other dietary carotenoids, enhance their chemical and physical stability in retina and brain membranes and maximize their protective action in these organs. Most importantly, xanthophylls are selectively concentrated in the most vulnerable regions of lipid bilayer membranes enriched in polyunsaturated lipids. This localization is ideal if macular xanthophylls are to act as lipid-soluble antioxidants, which is the most accepted mechanism through which lutein and zeaxanthin protect neural tissue against degenerative diseases.

A programmable artificial retina

International Nuclear Information System (INIS)

Bernard, T.M.; Zavidovique, B.Y.; Devos, F.J.

1993-01-01

An artificial retina is a device that intimately associates an imager with processing facilities on a monolithic circuit. Yet, except for simple environments and applications, analog hardware will not suffice to process and compact the raw image flow from the photosensitive array. To solve this output problem, an on-chip array of bare Boolean processors with halftoning facilities might be used, providing versatility from programmability. By setting the pixel memory size to 3 b, the authors have demonstrated both the technological practicality and the computational efficiency of this programmable Boolean retina concept. Using semi-static shifting structures together with some interaction circuitry, a minimal retina Boolean processor can be built with less than 30 transistors and controlled by as few as 6 global clock signals. The successful design, integration, and test of such a 65x76 Boolean retina on a 50-mm 2 CMOS 2-μm circuit are presented

Measuring mouse retina response near the detection threshold to direct stimulation of photons with sub-poisson statistics

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Tavala, Amir; Dovzhik, Krishna; Schicker, Klaus; Koschak, Alexandra; Zeilinger, Anton

Probing the visual system of human and animals at very low photon rate regime has recently attracted the quantum optics community. In an experiment on the isolated photoreceptor cells of Xenopus, the cell output signal was measured while stimulating it by pulses with sub-poisson distributed photons. The results showed single photon detection efficiency of 29 +/-4.7% [1]. Another behavioral experiment on human suggests a less detection capability at perception level with the chance of 0.516 +/-0.01 (i.e. slightly better than random guess) [2]. Although the species are different, both biological models and experimental observations with classical light stimuli expect that a fraction of single photon responses is filtered somewhere within the retina network and/or during the neural processes in the brain. In this ongoing experiment, we look for a quantitative answer to this question by measuring the output signals of the last neural layer of WT mouse retina using microelectrode arrays. We use a heralded downconversion single-photon source. We stimulate the retina directly since the eye lens (responsible for 20-50% of optical loss and scattering [2]) is being removed. Here, we demonstrate our first results that confirms the response to the sub-poisson distributied pulses. This project was supported by Austrian Academy of Sciences, SFB FoQuS F 4007-N23 funded by FWF and ERC QIT4QAD 227844 funded by EU Commission.

Effect of social isolation on anxiety-related behaviors, cortisol, and monoamines in adult zebrafish.

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Shams, Soaleha; Seguin, Diane; Facciol, Amanda; Chatterjee, Diptendu; Gerlai, Robert

2017-12-01

Social isolation can be used to study behavioral, neural, and hormonal mechanisms that regulate interactions in social animals. Although isolation effects have been reported in social mammals and various fish species, systematic studies with isolated zebrafish are rare. Here, the authors examined behavior (social and nonsocial), physiological stress (whole-body cortisol levels), and neurochemicals (serotonin, dopamine, and their metabolites), following acute and chronic social isolation in adult zebrafish. To observe how isolated fish respond behaviorally to social stimuli, they exposed zebrafish to live conspecifics or animated images after acute (24 hr) or chronic (6 months) social isolation. The authors observed that isolation did not affect locomotor activity, but acute isolation had weak nonsignificant anxiogenic effects in adult zebrafish. They also found that all isolated fish responded to both live and animated social stimuli, and the stress hormone, cortisol was lower in chronically isolated fish. Finally, neurochemical analyses showed that serotonin levels increased when fish were exposed to social stimulus after acute isolation, but its metabolite 5HIAA decreased in response to social stimulus following both acute and chronic isolation. Levels of both dopamine and its metabolite DOPAC were also reduced in fish exposed to social stimulus after acute and chronic isolation. Overall, these results show that isolation in zebrafish is an effective tool to study fundamental mechanisms controlling social interaction at behavioral and physiological levels. (PsycINFO Database Record (c) 2017 APA, all rights reserved).

Generation and characterization of Kctd15 mutations in zebrafish.

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Alison Heffer

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

Cornelia de Lange Syndrome: NIPBL haploinsufficiency downregulates canonical Wnt pathway in zebrafish embryos and patients fibroblasts.

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Pistocchi, A; Fazio, G; Cereda, A; Ferrari, L; Bettini, L R; Messina, G; Cotelli, F; Biondi, A; Selicorni, A; Massa, V

2013-10-17

Cornelia de Lange Syndrome is a severe genetic disorder characterized by malformations affecting multiple systems, with a common feature of severe mental retardation. Genetic variants within four genes (NIPBL (Nipped-B-like), SMC1A, SMC3, and HDAC8) are believed to be responsible for the majority of cases; all these genes encode proteins that are part of the 'cohesin complex'. Cohesins exhibit two temporally separated major roles in cells: one controlling the cell cycle and the other involved in regulating the gene expression. The present study focuses on the role of the zebrafish nipblb paralog during neural development, examining its expression in the central nervous system, and analyzing the consequences of nipblb loss of function. Neural development was impaired by the knockdown of nipblb in zebrafish. nipblb-loss-of-function embryos presented with increased apoptosis in the developing neural tissues, downregulation of canonical Wnt pathway genes, and subsequent decreased Cyclin D1 (Ccnd1) levels. Importantly, the same pattern of canonical WNT pathway and CCND1 downregulation was observed in NIPBL-mutated patient-specific fibroblasts. Finally, chemical activation of the pathway in nipblb-loss-of-function embryos rescued the adverse phenotype and restored the physiological levels of cell death.

Adaptive optics parallel spectral domain optical coherence tomography for imaging the living retina

Science.gov (United States)

Zhang, Yan; Rha, Jungtae; Jonnal, Ravi S.; Miller, Donald T.

2005-06-01

Although optical coherence tomography (OCT) can axially resolve and detect reflections from individual cells, there are no reports of imaging cells in the living human retina using OCT. To supplement the axial resolution and sensitivity of OCT with the necessary lateral resolution and speed, we developed a novel spectral domain OCT (SD-OCT) camera based on a free-space parallel illumination architecture and equipped with adaptive optics (AO). Conventional flood illumination, also with AO, was integrated into the camera and provided confirmation of the focus position in the retina with an accuracy of ±10.3 μm. Short bursts of narrow B-scans (100x560 μm) of the living retina were subsequently acquired at 500 Hz during dynamic compensation (up to 14 Hz) that successfully corrected the most significant ocular aberrations across a dilated 6 mm pupil. Camera sensitivity (up to 94 dB) was sufficient for observing reflections from essentially all neural layers of the retina. Signal-to-noise of the detected reflection from the photoreceptor layer was highly sensitive to the level of cular aberrations and defocus with changes of 11.4 and 13.1 dB (single pass) observed when the ocular aberrations (astigmatism, 3rd order and higher) were corrected and when the focus was shifted by 200 μm (0.54 diopters) in the retina, respectively. The 3D resolution of the B-scans (3.0x3.0x5.7 μm) is the highest reported to date in the living human eye and was sufficient to observe the interface between the inner and outer segments of individual photoreceptor cells, resolved in both lateral and axial dimensions. However, high contrast speckle, which is intrinsic to OCT, was present throughout the AO parallel SD-OCT B-scans and obstructed correlating retinal reflections to cell-sized retinal structures.

A zebrafish model of lethal congenital contracture syndrome 1 reveals Gle1 function in spinal neural precursor survival and motor axon arborization.

Science.gov (United States)

Jao, Li-En; Appel, Bruce; Wente, Susan R

2012-04-01

In humans, GLE1 is mutated in lethal congenital contracture syndrome 1 (LCCS1) leading to prenatal death of all affected fetuses. Although the molecular roles of Gle1 in nuclear mRNA export and translation have been documented, no animal models for this disease have been reported. To elucidate the function of Gle1 in vertebrate development, we used the zebrafish (Danio rerio) model system. gle1 mRNA is maternally deposited and widely expressed. Altering Gle1 using an insertional mutant or antisense morpholinos results in multiple defects, including immobility, small eyes, diminished pharyngeal arches, curved body axis, edema, underdeveloped intestine and cell death in the central nervous system. These phenotypes parallel those observed in LCCS1 human fetuses. Gle1 depletion also results in reduction of motoneurons and aberrant arborization of motor axons. Unexpectedly, the motoneuron deficiency results from apoptosis of neural precursors, not of differentiated motoneurons. Mosaic analyses further indicate that Gle1 activity is required extrinsically in the environment for normal motor axon arborization. Importantly, the zebrafish phenotypes caused by Gle1 deficiency are only rescued by expressing wild-type human GLE1 and not by the disease-linked Fin(Major) mutant form of GLE1. Together, our studies provide the first functional characterization of Gle1 in vertebrate development and reveal its essential role in actively dividing cells. We propose that defective GLE1 function in human LCCS1 results in both neurogenic and non-neurogenic defects linked to the apoptosis of proliferative organ precursors.

Alterations in energy metabolism, neuroprotection and visual signal transduction in the retina of Parkinsonian, MPTP-treated monkeys.

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Laura Campello

Full Text Available Parkinson disease is mainly characterized by the degeneration of dopaminergic neurons in the central nervous system, including the retina. Different interrelated molecular mechanisms underlying Parkinson disease-associated neuronal death have been put forward in the brain, including oxidative stress and mitochondrial dysfunction. Systemic injection of the proneurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP to monkeys elicits the appearance of a parkinsonian syndrome, including morphological and functional impairments in the retina. However, the intracellular events leading to derangement of dopaminergic and other retinal neurons in MPTP-treated animal models have not been so far investigated. Here we have used a comparative proteomics approach to identify proteins differentially expressed in the retina of MPTP-treated monkeys. Proteins were solubilized from the neural retinas of control and MPTP-treated animals, labelled separately with two different cyanine fluorophores and run pairwise on 2D DIGE gels. Out of >700 protein spots resolved and quantified, 36 were found to exhibit statistically significant differences in their expression levels, of at least ± 1.4-fold, in the parkinsonian monkey retina compared with controls. Most of these spots were excised from preparative 2D gels, trypsinized and subjected to MALDI-TOF MS and LC-MS/MS analyses. Data obtained were used for protein sequence database interrogation, and 15 different proteins were successfully identified, of which 13 were underexpressed and 2 overexpressed. These proteins were involved in key cellular functional pathways such as glycolysis and mitochondrial electron transport, neuronal protection against stress and survival, and phototransduction processes. These functional categories underscore that alterations in energy metabolism, neuroprotective mechanisms and signal transduction are involved in MPTP-induced neuronal degeneration in the retina, in similarity to

Effect of monocular deprivation on rabbit neural retinal cell densities

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Philip Maseghe Mwachaka

2015-01-01

Conclusion: In this rabbit model, monocular deprivation resulted in activity-dependent changes in cell densities of the neural retina in favour of the non-deprived eye along with reduced cell densities in the deprived eye.

Corpus vitreum, retina og chorioidea biopsi

DEFF Research Database (Denmark)

Scherfig, Erik Christian Høegh

2002-01-01

oftalmology, biopsy, choroid, corpus vitreum, retina, malignant melanoma, biopsy technic, retinoblastoma......oftalmology, biopsy, choroid, corpus vitreum, retina, malignant melanoma, biopsy technic, retinoblastoma...

Acetylcholine receptors in the human retina

International Nuclear Information System (INIS)

Hutchins, J.B.; Hollyfield, J.G.

1985-01-01

Evidence for a population of acetylcholine (ACh) receptors in the human retina is presented. The authors have used the irreversible ligand 3 H-propylbenzilylcholine mustard ( 3 H-PrBCM) to label muscarinic receptors. 3 H- or 125 I-alpha-bungarotoxin (alpha-BTx) was used to label putative nicotinic receptors. Muscarinic receptors are apparently present in the inner plexiform layer of the retina. Autoradiographic grain densities are reduced in the presence of saturating concentrations of atropine, quinuclidinyl benzilate or scopolamine; this indicates that 3 H-PrBCM binding is specific for a population of muscarinic receptors in the human retina. Binding sites for radiolabeled alpha-BTx are found predominantly in the inner plexiform layer of the retina. Grain densities are reduced in the presence of d-tubocurarine, indicating that alpha-BTx may bind to a pharmacologically relevant nicotinic ACh receptor. This study provides evidence for cholinergic neurotransmission in the human retina

The Analysis of Artificial Retina Organization for Signal Processing

Institute of Scientific and Technical Information of China (English)

WEIHui

2004-01-01

Machine vision is an active branch of artificial intelligence. An important problem in this area is the trade-off among efficiency, accuracy and computation complexity. The human visual system can keep watchfulness to the perimeter of a viewing field while at the same time focus on the center of the field for fine information processing. This mechanism of appropriate assignment of computing resources can reduce the demand for huge and complex hardware structure. Therefore, the design of a computer model based on the biological visual mechanism is an effective approach to resolve problems in machine vision. In this paper, a multi-layer neural model is developed based on the features of receptive field of ganglion in retina to simulate multi-scale perceptive fields of ganglion cell. The neural model can maintain alert on the outer area of the image while capturing and processing more important information in the central part. It may provide valuable inspiration for the implementation of real-time processing and avoidance of huge computation in machine vision.

Mechanisms of social buffering of fear in zebrafish.

Science.gov (United States)

Faustino, Ana I; Tacão-Monteiro, André; Oliveira, Rui F

2017-03-31

Some humans thrive whereas others resign when exposed to threatening situations throughout life. Social support has been identified as an important modulator of these discrepancies in human behaviour, and other social animals also exhibit phenomena in which individuals recover better from aversive events when conspecifics are present - aka social buffering. Here we studied social buffering in zebrafish, by exposing focal fish to an aversive stimulus (alarm substance - AS) either in the absence or presence of conspecific cues. When exposed to AS in the presence of both olfactory (shoal water) and visual (sight of shoal) conspecific cues, focal fish exhibited a lower fear response than when tested alone, demonstrating social buffering in zebrafish. When separately testing each cue's effectiveness, we verified that the visual cue was more effective than the olfactory in reducing freezing in a persistent threat scenario. Finally, we verified that social buffering was independent of shoal size and coincided with a distinct pattern of co-activation of brain regions known to be involved in mammalian social buffering. Thus, this study suggests a shared evolutionary origin for social buffering in vertebrates, bringing new evidence on the behavioural, sensory and neural mechanisms underlying this phenomenon.

Enhancing Understanding of the Visual Cycle by Applying CRISPR/Cas9 Gene Editing in Zebrafish

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Rebecca Ward

2018-04-01

Full Text Available During the vertebrate visual cycle, all-trans-retinal is exported from photoreceptors to the adjacent RPE or Müller glia wherein 11-cis-retinal is regenerated. The 11-cis chromophore is returned to photoreceptors, forming light-sensitive visual pigments with opsin GPCRs. Dysfunction of this process perturbs phototransduction because functional visual pigment cannot be generated. Mutations in visual cycle genes can result in monogenic inherited forms of blindness. Though key enzymatic processes are well characterized, questions remain as to the physiological role of visual cycle proteins in different retinal cell types, functional domains of these proteins in retinoid biochemistry and in vivo pathogenesis of disease mutations. Significant progress is needed to develop effective and accessible treatments for inherited blindness arising from mutations in visual cycle genes. Here, we review opportunities to apply gene editing technology to two crucial visual cycle components, RPE65 and CRALBP. Expressed exclusively in the human RPE, RPE65 enzymatically converts retinyl esters into 11-cis retinal. CRALBP is an 11-cis-retinal binding protein expressed in human RPE and Muller glia. Loss-of-function mutations in either protein results in autosomal recessive forms of blindness. Modeling these human conditions using RPE65 or CRALBP murine knockout models have enhanced our understanding of their biochemical function, associated disease pathogenesis and development of therapeutics. However, rod-dominated murine retinae provide a challenge to assess cone function. The cone-rich zebrafish model is amenable to cost-effective maintenance of a variety of strains. Interestingly, gene duplication in zebrafish resulted in three Rpe65 and two Cralbp isoforms with differential temporal and spatial expression patterns. Functional investigations of zebrafish Rpe65 and Cralbp were restricted to gene knockdown with morpholino oligonucleotides. However, transient

Proteomic interactions in the mouse vitreous-retina complex.

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Jessica M Skeie

Full Text Available Human vitreoretinal diseases are due to presumed abnormal mechanical interactions between the vitreous and retina, and translational models are limited. This study determined whether nonstructural proteins and potential retinal biomarkers were expressed by the normal mouse vitreous and retina.Vitreous and retina samples from mice were collected by evisceration and analyzed by liquid chromatography-tandem mass spectrometry. Identified proteins were further analyzed for differential expression and functional interactions using bioinformatic software.We identified 1,680 unique proteins in the retina and 675 unique proteins in the vitreous. Unbiased clustering identified protein pathways that distinguish retina from vitreous including oxidative phosphorylation and neurofilament cytoskeletal remodeling, whereas the vitreous expressed oxidative stress and innate immunology pathways. Some intracellular protein pathways were found in both retina and vitreous, such as glycolysis and gluconeogenesis and neuronal signaling, suggesting proteins might be shuttled between the retina and vitreous. We also identified human disease biomarkers represented in the mouse vitreous and retina, including carbonic anhydrase-2 and 3, crystallins, macrophage inhibitory factor, glutathione peroxidase, peroxiredoxins, S100 precursors, and von Willebrand factor.Our analysis suggests the vitreous expresses nonstructural proteins that functionally interact with the retina to manage oxidative stress, immune reactions, and intracellular proteins may be exchanged between the retina and vitreous. This novel proteomic dataset can be used for investigating human vitreoretinopathies in mouse models. Validation of vitreoretinal biomarkers for human ocular diseases will provide a critical tool for diagnostics and an avenue for therapeutics.

Proteomic interactions in the mouse vitreous-retina complex.

Science.gov (United States)

Skeie, Jessica M; Mahajan, Vinit B

2013-01-01

Human vitreoretinal diseases are due to presumed abnormal mechanical interactions between the vitreous and retina, and translational models are limited. This study determined whether nonstructural proteins and potential retinal biomarkers were expressed by the normal mouse vitreous and retina. Vitreous and retina samples from mice were collected by evisceration and analyzed by liquid chromatography-tandem mass spectrometry. Identified proteins were further analyzed for differential expression and functional interactions using bioinformatic software. We identified 1,680 unique proteins in the retina and 675 unique proteins in the vitreous. Unbiased clustering identified protein pathways that distinguish retina from vitreous including oxidative phosphorylation and neurofilament cytoskeletal remodeling, whereas the vitreous expressed oxidative stress and innate immunology pathways. Some intracellular protein pathways were found in both retina and vitreous, such as glycolysis and gluconeogenesis and neuronal signaling, suggesting proteins might be shuttled between the retina and vitreous. We also identified human disease biomarkers represented in the mouse vitreous and retina, including carbonic anhydrase-2 and 3, crystallins, macrophage inhibitory factor, glutathione peroxidase, peroxiredoxins, S100 precursors, and von Willebrand factor. Our analysis suggests the vitreous expresses nonstructural proteins that functionally interact with the retina to manage oxidative stress, immune reactions, and intracellular proteins may be exchanged between the retina and vitreous. This novel proteomic dataset can be used for investigating human vitreoretinopathies in mouse models. Validation of vitreoretinal biomarkers for human ocular diseases will provide a critical tool for diagnostics and an avenue for therapeutics.

Social conflict resolution regulated by two dorsal habenular subregions in zebrafish.

Science.gov (United States)

Chou, Ming-Yi; Amo, Ryunosuke; Kinoshita, Masae; Cherng, Bor-Wei; Shimazaki, Hideaki; Agetsuma, Masakazu; Shiraki, Toshiyuki; Aoki, Tazu; Takahoko, Mikako; Yamazaki, Masako; Higashijima, Shin-ichi; Okamoto, Hitoshi

2016-04-01

When animals encounter conflict they initiate and escalate aggression to establish and maintain a social hierarchy. The neural mechanisms by which animals resolve fighting behaviors to determine such social hierarchies remain unknown. We identified two subregions of the dorsal habenula (dHb) in zebrafish that antagonistically regulate the outcome of conflict. The losing experience reduced neural transmission in the lateral subregion of dHb (dHbL)-dorsal/intermediate interpeduncular nucleus (d/iIPN) circuit. Silencing of the dHbL or medial subregion of dHb (dHbM) caused a stronger predisposition to lose or win a fight, respectively. These results demonstrate that the dHbL and dHbM comprise a dual control system for conflict resolution of social aggression. Copyright © 2016, American Association for the Advancement of Science.

Potential teratogenicity of methimazole: exposure of zebrafish embryos to methimazole causes similar developmental anomalies to human methimazole embryopathy.

Science.gov (United States)

Komoike, Yuta; Matsuoka, Masato; Kosaki, Kenjiro

2013-06-01

While methimazole (MMI) is widely used in the therapy for hyperthyroidism, several groups have reported that maternal exposure to MMI results in a variety of congenital anomalies, including choanal and esophageal atresia, iridic and retinal coloboma, and delayed neurodevelopment. Thus, adverse effects of maternal exposure to MMI on fetal development have long been suggested; however, direct evidence for the teratogenicity of MMI has not been presented. Therefore, we studied the effects of MMI on early development by using zebrafish as a model organism. The fertilized eggs of zebrafish were collected immediately after spawning and grown in egg culture water containing MMI at various concentrations. External observation of the embryos revealed that exposure to high concentrations of MMI resulted in loss of pigmentation, hypoplastic hindbrain, turbid tissue in the forebrain, swelling of the notochord, and curly trunk. Furthermore, these effects occurred in a dose-dependent manner. Precise observation of the serial cross-sections of MMI-exposed embryos elucidated delayed development and hypoplasia of the whole brain and spinal cord, narrowing of the pharynx and esophagus, severe disruption of the retina, and aberrant structure of the notochord. These neuronal, pharyngeal, esophageal, and retinal anomalous morphologies have a direct analogy to the congenital anomalies observed in children exposed to MMI in utero. Here, we show the teratogenic effects of MMI on the development of zebrafish and provide the first experimental evidence for the connection between exposure to MMI and human MMI embryopathy. © 2013 Wiley Periodicals, Inc.

A zebrafish model for Waardenburg syndrome type IV reveals diverse roles for Sox10 in the otic vesicle.

Science.gov (United States)

Dutton, Kirsten; Abbas, Leila; Spencer, Joanne; Brannon, Claire; Mowbray, Catriona; Nikaido, Masataka; Kelsh, Robert N; Whitfield, Tanya T

2009-01-01

In humans, mutations in the SOX10 gene are a cause of the auditory-pigmentary disorder Waardenburg syndrome type IV (WS4) and related variants. SOX10 encodes an Sry-related HMG box protein essential for the development of the neural crest; deafness in WS4 and other Waardenburg syndromes is usually attributed to loss of neural-crest-derived melanocytes in the stria vascularis of the cochlea. However, SOX10 is strongly expressed in the developing otic vesicle and so direct roles for SOX10 in the otic epithelium might also be important. Here, we examine the otic phenotype of zebrafish sox10 mutants, a model for WS4. As a cochlea is not present in the fish ear, the severe otic phenotype in these mutants cannot be attributed to effects on this tissue. In zebrafish sox10 mutants, we see abnormalities in all otic placodal derivatives. Gene expression studies indicate deregulated expression of several otic genes, including fgf8, in sox10 mutants. Using a combination of mutant and morphant data, we show that the three sox genes belonging to group E (sox9a, sox9b and sox10) provide a link between otic induction pathways and subsequent otic patterning: they act redundantly to maintain sox10 expression throughout otic tissue and to restrict fgf8 expression to anterior macula regions. Single-cell labelling experiments indicate a small and transient neural crest contribution to the zebrafish ear during normal development, but this is unlikely to account for the strong defects seen in the sox10 mutant. We discuss the implication that the deafness in WS4 patients with SOX10 mutations might reflect a haploinsufficiency for SOX10 in the otic epithelium, resulting in patterning and functional abnormalities in the inner ear.

The neurotoxic effects of N-methyl-N-nitrosourea on the electrophysiological property and visual signal transmission of rat's retina

Energy Technology Data Exchange (ETDEWEB)

Tao, Ye [Department of Ophthalmology, General Hospital of Chinese PLA, Beijing 100853 (China); Chen, Tao [Department of Clinical Aerospace Medicine, Fourth Military Medical University, Xi' an 710032 (China); Liu, Bei [Department of Neurosurgery and Institute for Functional Brain Disorders, Tangdu Hospital, Fourth Military Medical University, Xi' an (China); Yang, Guo Qing [Department of Clinical Aerospace Medicine, Fourth Military Medical University, Xi' an 710032 (China); Peng, Guanghua [Department of Ophthalmology, General Hospital of Chinese PLA, Beijing 100853 (China); Zhang, Hua [Department of Neurosurgery and Institute for Functional Brain Disorders, Tangdu Hospital, Fourth Military Medical University, Xi' an (China); Huang, Yi Fei [Department of Ophthalmology, General Hospital of Chinese PLA, Beijing 100853 (China)

2015-07-01

The neurotoxic effects of N-methyl-N-nitrosourea (MNU) on the inner retinal neurons and related visual signal circuits have not been described in any animal models or human, despite ample morphological evidences about the MNU induced photoreceptor (PR) degeneration. With the helping of MEA (multielectrode array) recording system, we gained the opportunity to systemically explore the neural activities and visual signal pathways of MNU administrated rats. Our MEA research identified remarkable alterations in the electrophysiological properties and firstly provided instructive information about the neurotoxicity of MNU that affects the signal transmission in the inner retina. Moreover, the spatial electrophysiological functions of retina were monitored and found that the focal PRs had different vulnerabilities to the MNU. The MNU-induced PR dysfunction exhibited a distinct spatial- and time-dependent progression. In contrast, the spiking activities of both central and peripheral RGCs altered synchronously in response to the MNU administration. Pharmacological tests suggested that gap junctions played a pivotal role in this homogeneous response of RGCs. SNR analysis of MNU treated retina suggested that the signaling efficiency and fidelity of inner retinal circuits have been ruined by this toxicant, although the microstructure of the inner retina seemed relatively consolidated. The present study provided an appropriate example of MEA investigations on the toxicant induced pathological models and the effects of the pharmacological compounds on neuron activities. The positional MEA information would enrich our knowledge about the pathology of MNU induced RP models, and eventually be instrumental for elucidating the underlying mechanism of human RP. - Highlights: • We systemically explored the neural activities and visual signal pathways of MNU administrated retinas. • The focal photoreceptors had different vulnerabilities to the MNU administration. �

The neurotoxic effects of N-methyl-N-nitrosourea on the electrophysiological property and visual signal transmission of rat's retina

International Nuclear Information System (INIS)

Tao, Ye; Chen, Tao; Liu, Bei; Yang, Guo Qing; Peng, Guanghua; Zhang, Hua; Huang, Yi Fei

2015-01-01

The neurotoxic effects of N-methyl-N-nitrosourea (MNU) on the inner retinal neurons and related visual signal circuits have not been described in any animal models or human, despite ample morphological evidences about the MNU induced photoreceptor (PR) degeneration. With the helping of MEA (multielectrode array) recording system, we gained the opportunity to systemically explore the neural activities and visual signal pathways of MNU administrated rats. Our MEA research identified remarkable alterations in the electrophysiological properties and firstly provided instructive information about the neurotoxicity of MNU that affects the signal transmission in the inner retina. Moreover, the spatial electrophysiological functions of retina were monitored and found that the focal PRs had different vulnerabilities to the MNU. The MNU-induced PR dysfunction exhibited a distinct spatial- and time-dependent progression. In contrast, the spiking activities of both central and peripheral RGCs altered synchronously in response to the MNU administration. Pharmacological tests suggested that gap junctions played a pivotal role in this homogeneous response of RGCs. SNR analysis of MNU treated retina suggested that the signaling efficiency and fidelity of inner retinal circuits have been ruined by this toxicant, although the microstructure of the inner retina seemed relatively consolidated. The present study provided an appropriate example of MEA investigations on the toxicant induced pathological models and the effects of the pharmacological compounds on neuron activities. The positional MEA information would enrich our knowledge about the pathology of MNU induced RP models, and eventually be instrumental for elucidating the underlying mechanism of human RP. - Highlights: • We systemically explored the neural activities and visual signal pathways of MNU administrated retinas. • The focal photoreceptors had different vulnerabilities to the MNU administration. �

Animal models for studying neural crest development: is the mouse different?

Science.gov (United States)

Barriga, Elias H; Trainor, Paul A; Bronner, Marianne; Mayor, Roberto

2015-05-01

The neural crest is a uniquely vertebrate cell type and has been well studied in a number of model systems. Zebrafish, Xenopus and chick embryos largely show consistent requirements for specific genes in early steps of neural crest development. By contrast, knockouts of homologous genes in the mouse often do not exhibit comparable early neural crest phenotypes. In this Spotlight article, we discuss these species-specific differences, suggest possible explanations for the divergent phenotypes in mouse and urge the community to consider these issues and the need for further research in complementary systems. © 2015. Published by The Company of Biologists Ltd.

Regulation of Msx genes by a Bmp gradient is essential for neural crest specification.

Science.gov (United States)

Tribulo, Celeste; Aybar, Manuel J; Nguyen, Vu H; Mullins, Mary C; Mayor, Roberto

2003-12-01

There is evidence in Xenopus and zebrafish embryos that the neural crest/neural folds are specified at the border of the neural plate by a precise threshold concentration of a Bmp gradient. In order to understand the molecular mechanism by which a gradient of Bmp is able to specify the neural crest, we analyzed how the expression of Bmp targets, the Msx genes, is regulated and the role that Msx genes has in neural crest specification. As Msx genes are directly downstream of Bmp, we analyzed Msx gene expression after experimental modification in the level of Bmp activity by grafting a bead soaked with noggin into Xenopus embryos, by expressing in the ectoderm a dominant-negative Bmp4 or Bmp receptor in Xenopus and zebrafish embryos, and also through Bmp pathway component mutants in the zebrafish. All the results show that a reduction in the level of Bmp activity leads to an increase in the expression of Msx genes in the neural plate border. Interestingly, by reaching different levels of Bmp activity in animal cap ectoderm, we show that a specific concentration of Bmp induces msx1 expression to a level similar to that required to induce neural crest. Our results indicate that an intermediate level of Bmp activity specifies the expression of Msx genes in the neural fold region. In addition, we have analyzed the role that msx1 plays on neural crest specification. As msx1 has a role in dorsoventral pattering, we have carried out conditional gain- and loss-of-function experiments using different msx1 constructs fused to a glucocorticoid receptor element to avoid an early effect of this factor. We show that msx1 expression is able to induce all other early neural crest markers tested (snail, slug, foxd3) at the time of neural crest specification. Furthermore, the expression of a dominant negative of Msx genes leads to the inhibition of all the neural crest markers analyzed. It has been previously shown that snail is one of the earliest genes acting in the neural crest

Identification and target prediction of miRNAs specifically expressed in rat neural tissue

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Tu Kang

2009-05-01

Full Text Available Abstract Background MicroRNAs (miRNAs are a large group of RNAs that play important roles in regulating gene expression and protein translation. Several studies have indicated that some miRNAs are specifically expressed in human, mouse and zebrafish tissues. For example, miR-1 and miR-133 are specifically expressed in muscles. Tissue-specific miRNAs may have particular functions. Although previous studies have reported the presence of human, mouse and zebrafish tissue-specific miRNAs, there have been no detailed reports of rat tissue-specific miRNAs. In this study, Home-made rat miRNA microarrays which established in our previous study were used to investigate rat neural tissue-specific miRNAs, and mapped their target genes in rat tissues. This study will provide information for the functional analysis of these miRNAs. Results In order to obtain as complete a picture of specific miRNA expression in rat neural tissues as possible, customized miRNA microarrays with 152 selected miRNAs from miRBase were used to detect miRNA expression in 14 rat tissues. After a general clustering analysis, 14 rat tissues could be clearly classified into neural and non-neural tissues based on the obtained expression profiles with p values Conclusion Our work provides a global view of rat neural tissue-specific miRNA profiles and a target map of miRNAs, which is expected to contribute to future investigations of miRNA regulatory mechanisms in neural systems.

Comparative immunolocalization of the plasma membrane calcium pump and calbindin D28K in chicken retina during embryonic development

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N. Tolosa de Talamoni

2010-05-01

Full Text Available The immunolocalization of the plasma membrane calcium pump (PMCA was studied in 4-week-old chick retina in comparison with calbindin D28K (CaBP immunostaining. We have demonstrated that the monoclonal anti-PMCA antibody 5F10 from human erythrocyte plasma membrane crossreacts with a Ca2+ pump epitope of the cells from the neural retina. The immunolocalization of both proteins was also studied during the embryonic development of the chicken retina. At age 4.5 days, the cells of the retina were faintly immunoreactive to PMCA and CaBP antibodies, but the lack of cellular aggregation and differentiation did not allow discrimination between the two proteins. A clear difference in the localization was seen from the tenth day of development through post-hatching with slight variation. PMCA localized mainly in the outer and inner plexiform layers, in some cells in the ganglion layer, in the nerve fiber layer and slightly in the photoreceptor cells. CaBP was intensely stained in cones, cone pedicles and some amacrine cells. The number of CaBP positive amacrine cells declined after hatching. A few ganglion cells and several nerve fibers were CaBP 333 immunoreactive. The role of these proteins in the early stages of retinal development is unknown, but the results suggest that Ca2+ homeostasis in the retina is well regulated, probably to avoid excessive accumulation of Ca2+, which often leads to neurodegeneration.

Tcf7l1 protects the anterior neural fold from adopting the neural crest fate

Czech Academy of Sciences Publication Activity Database

Mašek, Jan; Machoň, Ondřej; Kořínek, Vladimír; Taketo, M.M.; Kozmik, Zbyněk

2016-01-01

Roč. 143, č. 12 (2016), s. 2206-2216 ISSN 0950-1991 R&D Projects: GA ČR GAP305/12/2042; GA ČR(CZ) GA14-33952S; GA MŠk(CZ) LK11214; GA MŠk LO1419; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:68378050 Keywords : Tcf/Lef * Wnt dignaling * neural crest * forebrain * mouse * zebrafish Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.843, year: 2016

Whole-Retina Reduced Electrophysiological Activity in Mice Bearing Retina-Specific Deletion of Vesicular Acetylcholine Transporter.

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Jake Bedore

Full Text Available Despite rigorous characterization of the role of acetylcholine in retinal development, long-term effects of its absence as a neurotransmitter are unknown. One of the unanswered questions is how acetylcholine contributes to the functional capacity of mature retinal circuits. The current study investigates the effects of disrupting cholinergic signalling in mice, through deletion of vesicular acetylcholine transporter (VAChT in the developing retina, pigmented epithelium, optic nerve and optic stalk, on electrophysiology and structure of the mature retina.A combination of electroretinography, optical coherence tomography imaging and histological evaluation assessed retinal integrity in mice bearing retina- targeted (embryonic day 12.5 deletion of VAChT (VAChTSix3-Cre-flox/flox and littermate controls at 5 and 12 months of age. VAChTSix3-Cre-flox/flox mice did not show any gross changes in nuclear layer cellularity or synaptic layer thickness. However, VAChTSix3-Cre-flox/flox mice showed reduced electrophysiological response of the retina to light stimulus under scotopic conditions at 5 and 12 months of age, including reduced a-wave, b-wave, and oscillatory potential (OP amplitudes and decreased OP peak power and total energy. Reduced a-wave amplitude was proportional to the reduction in b-wave amplitude and not associated with altered a-wave 10%-90% rise time or inner and outer segment thicknesses.This study used a novel genetic model in the first examination of function and structure of the mature mouse retina with disruption of cholinergic signalling. Reduced amplitude across the electroretinogram wave form does not suggest dysfunction in specific retinal cell types and could reflect underlying changes in the retinal and/or extraretinal microenvironment. Our findings suggest that release of acetylcholine by VAChT is essential for the normal electrophysiological response of the mature mouse retina.

Information processing in the outer retina of fish

NARCIS (Netherlands)

Endeman, D.

2017-01-01

The retina translates light into neuronal activity. Thus, it renders visual information of the external environment. The retina can only send a limited amount of information to the brain within a given period. To use this amount optimally, light stimuli are strongly processed in the retina. This

An analog silicon retina with multichip configuration.

Science.gov (United States)

Kameda, Seiji; Yagi, Tetsuya

2006-01-01

The neuromorphic silicon retina is a novel analog very large scale integrated circuit that emulates the structure and the function of the retinal neuronal circuit. We fabricated a neuromorphic silicon retina, in which sample/hold circuits were embedded to generate fluctuation-suppressed outputs in the previous study [1]. The applications of this silicon retina, however, are limited because of a low spatial resolution and computational variability. In this paper, we have fabricated a multichip silicon retina in which the functional network circuits are divided into two chips: the photoreceptor network chip (P chip) and the horizontal cell network chip (H chip). The output images of the P chip are transferred to the H chip with analog voltages through the line-parallel transfer bus. The sample/hold circuits embedded in the P and H chips compensate for the pattern noise generated on the circuits, including the analog communication pathway. Using the multichip silicon retina together with an off-chip differential amplifier, spatial filtering of the image with an odd- and an even-symmetric orientation selective receptive fields was carried out in real time. The analog data transfer method in the present multichip silicon retina is useful to design analog neuromorphic multichip systems that mimic the hierarchical structure of neuronal networks in the visual system.

A single enhancer regulating the differential expression of duplicated red-sensitive opsin genes in zebrafish.

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Taro Tsujimura

2010-12-01

Full Text Available A fundamental step in the evolution of the visual system is the gene duplication of visual opsins and differentiation between the duplicates in absorption spectra and expression pattern in the retina. However, our understanding of the mechanism of expression differentiation is far behind that of spectral tuning of opsins. Zebrafish (Danio rerio have two red-sensitive cone opsin genes, LWS-1 and LWS-2. These genes are arrayed in a tail-to-head manner, in this order, and are both expressed in the long member of double cones (LDCs in the retina. Expression of the longer-wave sensitive LWS-1 occurs later in development and is thus confined to the peripheral, especially ventral-nasal region of the adult retina, whereas expression of LWS-2 occurs earlier and is confined to the central region of the adult retina, shifted slightly to the dorsal-temporal region. In this study, we employed a transgenic reporter assay using fluorescent proteins and P1-artificial chromosome (PAC clones encompassing the two genes and identified a 0.6-kb "LWS-activating region" (LAR upstream of LWS-1, which regulates expression of both genes. Under the 2.6-kb flanking upstream region containing the LAR, the expression pattern of LWS-1 was recapitulated by the fluorescent reporter. On the other hand, when LAR was directly conjugated to the LWS-2 upstream region, the reporter was expressed in the LDCs but also across the entire outer nuclear layer. Deletion of LAR from the PAC clones drastically lowered the reporter expression of the two genes. These results suggest that LAR regulates both LWS-1 and LWS-2 by enhancing their expression and that interaction of LAR with the promoters is competitive between the two genes in a developmentally restricted manner. Sharing a regulatory region between duplicated genes could be a general way to facilitate the expression differentiation in duplicated visual opsins.

Comparison of the neuroinflammatory responses to selective retina therapy and continuous-wave laser photocoagulation in mouse eyes.

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Han, Jung Woo; Choi, Juhye; Kim, Young Shin; Kim, Jina; Brinkmann, Ralf; Lyu, Jungmook; Park, Tae Kwann

2018-02-01

This study investigated microglia and inflammatory cell responses after selective retina therapy (SRT) with microsecond-pulsed laser in comparison to continuous-wave laser photocoagulation (cwPC). Healthy C57BL/6 J mice were treated with either a train of short pulses (SRT; 527-nm, Q-switched, 1.7-μs pulse) or a conventional thermal continuous-wave (532-nm, 100-ms pulse duration) laser. The mice were sacrificed and their eyes were enucleated 1, 3, 7, and 14 days after both laser treatments. Pattern of cell death on retinal section was evaluated by TUNEL assay, and the distribution of activated inflammatory cells and glial cells were observed under immunohistochemistry. Consecutive changes for the expression of cytokines such as IL-1β, TNF-α, and TGF-β were also examined using immunohistochemistry, and compared among each period after quantification by Western blotting. The numbers of TUNEL-positive cells in the retinal pigment epithelium (RPE) layer did not differ in SRT and cwPC lesions, but TUNEL-positive cells in neural retinas were significantly less on SRT. Vague glial cell activation was observed in SRT-treated lesions. The population of inflammatory cells was also significantly decreased after SRT, and the cells were located in the RPE layer and subretinal space. Proinflammatory cytokines, including IL-1β and TNF-α, showed significantly lower levels after SRT; conversely, the level of TGF-β was similar to the cwPC-treated lesion. SRT resulted in selective RPE damage without collateral thermal injury to the neural retina, and apparently produced negligible glial activation. In addition, SRT showed a markedly less inflammatory response than cwPC, which may have important therapeutic implications for several macular diseases.

Distribution of FMRFamide-like immunoreactivity in the brain, retina and nervus terminalis of the sockeye salmon parr, Oncorhynchus nerka.

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Ostholm, T; Ekström, P; Ebbesson, S O

1990-09-01

Neurons displaying FMRFamide(Phe - Met - Arg - Phe - NH2)-like immunoreactivity have recently been implicated in neural plasticity in salmon. We now extend these findings by describing the extent of the FMRF-like immunoreactive (FMRF-IR) system in the brain, retina and olfactory system of sockeye salmon parr using the indirect peroxidase anti-peroxidase technique. FMRF-IR perikarya were found in the periventricular hypothalamus, mesencephalic laminar nucleus, nucleus nervi terminalis and retina (presumed amacrine cells), and along the olfactory nerves. FMRF-IR fibers were distributed throughout the brain with highest densities in the ventral area of the telencephalon, in the medial forebrain bundle, and at the borders between layers III/IV and IV/V in the optic tectum. High densities of immunoreactive fibers were also observed in the area around the torus semicircularis, in the medial hypothalamus, median raphe, ventromedial tegmentum, and central gray. In the retina, immunopositive fibers were localized to the inner plexiform layer, but several fiber elements were also found in the outer plexiform layer. The olfactory system displayed FMRF-IR fibers in the epithelium and along the olfactory nerves. These findings differ from those reported in other species as follows: (i) FMRF-IR cells in the retina have not previously been reported in teleosts; (ii) the presence of FMRF-IR fibers in the outer plexiform layer of the retina is a new finding for any species; (iii) the occurrence of immunopositive cells in the mesencephalic laminar nucleus has to our knowledge not been demonstrated previously.

Distribution and chronotropic effects of serotonin in the zebrafish heart.

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Stoyek, Matthew R; Jonz, Michael G; Smith, Frank M; Croll, Roger P

2017-09-01

Several lines of evidence suggest that serotonin (5-HT) has a regulatory role in cardiovascular function from embryogenesis through adulthood. However, the reported actions of 5-HT are often contradictory and include bradycardia or tachycardia, hypotension or hypertension, and vasodilation or vasoconstriction. Clarifying such cardiac effects requires further research and may benefit from utilizing a model simpler than the mammalian hearts traditionally used in these studies. In the present study, we describe the cardiac distribution and chronotropic responses of 5-HT in the zebrafish heart. A combined anatomical, electrophysiological, and pharmacological approach was used to investigate the involvement of 5-HT pathways, and to compare neural and direct myocardial pathways of biological action. Immunohistochemical methods revealed 5-HT in endocardial cells, glial-like cells, and intracardiac neurons in the atrium. Electrocardiogram (ECG) recordings combined with the administration of pharmacological agents demonstrated that 5-HT acted predominantly through direct myocardial pathways resulting in a reduction of heart rate. Overall, the results of this study contribute significant advances in the establishment of the zebrafish as a new model for studies of the role of 5-HT in autonomic cardiac control. Copyright © 2017 Elsevier B.V. All rights reserved.

Identification and characterization of zebrafish thrombocytes.

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

1999-12-01

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

Hardwiring of fine synaptic layers in the zebrafish visual pathway

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Taylor Michael R

2008-12-01

Full Text Available Abstract Background Neuronal connections are often arranged in layers, which are divided into sublaminae harboring synapses with similar response properties. It is still debated how fine-grained synaptic layering is established during development. Here we investigated two stratified areas of the zebrafish visual pathway, the inner plexiform layer (IPL of the retina and the neuropil of the optic tectum, and determined if activity is required for their organization. Results The IPL of 5-day-old zebrafish larvae is composed of at least nine sublaminae, comprising the connections between different types of amacrine, bipolar, and ganglion cells (ACs, BCs, GCs. These sublaminae were distinguished by their expression of cell type-specific transgenic fluorescent reporters and immunohistochemical markers, including protein kinase Cβ (PKC, parvalbumin (Parv, zrf3, and choline acetyltransferase (ChAT. In the tectum, four retinal input layers abut a laminated array of neurites of tectal cells, which differentially express PKC and Parv. We investigated whether these patterns were affected by experimental disruptions of retinal activity in developing fish. Neither elimination of light inputs by dark rearing, nor a D, L-amino-phosphono-butyrate-induced reduction in the retinal response to light onset (but not offset altered IPL or tectal lamination. Moreover, thorough elimination of chemical synaptic transmission with Botulinum toxin B left laminar synaptic arrays intact. Conclusion Our results call into question a role for activity-dependent mechanisms – instructive light signals, balanced on and off BC activity, Hebbian plasticity, or a permissive role for synaptic transmission – in the synaptic stratification we examined. We propose that genetically encoded cues are sufficient to target groups of neurites to synaptic layers in this vertebrate visual system.

Tlx, an orphan nuclear receptor, regulates cell numbers and astrocyte development in the developing retina.

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Miyawaki, Takaya; Uemura, Akiyoshi; Dezawa, Mari; Yu, Ruth T; Ide, Chizuka; Nishikawa, Shinichi; Honda, Yoshihito; Tanabe, Yasuto; Tanabe, Teruyo

2004-09-15

Tlx belongs to a class of orphan nuclear receptors that underlies many aspects of neural development in the CNS. However, the fundamental roles played by Tlx in the control of eye developmental programs remain elusive. By using Tlx knock-out (KO) mice, we show here that Tlx is expressed by retinal progenitor cells in the neuroblastic layer during the period of retinal layer formation, and it is critical for controlling the generation of appropriate numbers of retinal progenies through the activities of cell cycle-related molecules, cyclin D1 and p27Kip1. Tlx expression is restricted to Müller cells in the mature retina and appears to control their proper development. Furthermore, we show that Tlx is expressed by immature astrocytes that migrate from the optic nerve onto the inner surface of the retina and is required for their generation and maturation, as assessed by honeycomb network formation and expression of R-cadherin, a critical component for vasculogenesis. The impaired astrocyte network formation on the inner retinal surface is accompanied by the loss of vasculogenesis in Tlx KO retinas. Our studies thus indicate that Tlx underlies a fundamental developmental program of retinal organization and controls the generation of the proper numbers of retinal progenies and development of glial cells during the protracted period of retinogenesis.

The retina

DEFF Research Database (Denmark)

van Reyk, David M; Gillies, Mark C; Davies, Michael Jonathan

2003-01-01

A prominent and early feature of the retinopathy of diabetes mellitus is a diffuse increase in vascular permeability. As the disease develops, the development of frank macular oedema may result in vision loss. That reactive oxygen species production is likely to be elevated in the retina, and tha...

The infrared retina

International Nuclear Information System (INIS)

Krishna, Sanjay

2009-01-01

As infrared imaging systems have evolved from the first generation of linear devices to the second generation of small format staring arrays to the present 'third-gen' systems, there is an increased emphasis on large area focal plane arrays (FPAs) with multicolour operation and higher operating temperature. In this paper, we discuss how one needs to develop an increased functionality at the pixel level for these next generation FPAs. This functionality could manifest itself as spectral, polarization, phase or dynamic range signatures that could extract more information from a given scene. This leads to the concept of an infrared retina, which is an array that works similarly to the human eye that has a 'single' FPA but multiple cones, which are photoreceptor cells in the retina of the eye that enable the perception of colour. These cones are then coupled with powerful signal processing techniques that allow us to process colour information from a scene, even with a limited basis of colour cones. Unlike present day multi or hyperspectral systems, which are bulky and expensive, the idea would be to build a poor man's 'infrared colour' camera. We use examples such as plasmonic tailoring of the resonance or bias dependent dynamic tuning based on quantum confined Stark effect or incorporation of avalanche gain to achieve embodiments of the infrared retina.

Progenitor cells from the porcine neural retina express photoreceptor markers after transplantation to the subretinal space of allorecipients

DEFF Research Database (Denmark)

Klassen, Henry; Kiilgaard, Jens Folke; Zahir, Tasneem

2007-01-01

Work in rodents has shown that cultured retinal progenitor cells (RPCs) integrate into the degenerating retina, thus suggesting a potential strategy for treatment of similar degenerative conditions in humans. To demonstrate the relevance of the rodent work to large animals, we derived progenitor...

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

Laser capture microdissection of gonads from juvenile zebrafish

DEFF Research Database (Denmark)

Jørgensen, Anne; Nielsen, John; Morthorst, Jane Ebsen

2009-01-01

was adjusted and optimised to isolate juvenile zebrafish gonads. Results: The juvenile zebrafish gonad is not morphologically distinguishable when using dehydrated cryosections on membrane slides and a specific staining method is necessary to identify the gonads. The protocol setup in this study allows......Background: Investigating gonadal gene expression is important in attempting to elucidate the molecular mechanism of sex determination and differentiation in the model species zebrafish. However, the small size of juvenile zebrafish and correspondingly their gonads complicates this type...... of investigation. Furthermore, the lack of a genetic sex marker in juvenile zebrafish prevents pooling gonads from several individuals. The aim of this study was to establish a method to isolate the gonads from individual juvenile zebrafish allowing future investigations of gonadal gene expression during sex...

3D bioprinting: A new insight into the therapeutic strategy of neural tissue regeneration.

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Hsieh, Fu-Yu; Hsu, Shan-hui

2015-01-01

Acute traumatic injuries and chronic degenerative diseases represent the world's largest unmet medical need. There are over 50 million people worldwide suffering from neurodegenerative diseases. However, there are only a few treatment options available for acute traumatic injuries and neurodegenerative diseases. Recently, 3D bioprinting is being applied to regenerative medicine to address the need for tissues and organs suitable for transplantation. In this commentary, the newly developed 3D bioprinting technique involving neural stem cells (NSCs) embedded in the thermoresponsive biodegradable polyurethane (PU) bioink is reviewed. The thermoresponsive and biodegradable PU dispersion can form gel near 37 °C without any crosslinker. NSCs embedded within the water-based PU hydrogel with appropriate stiffness showed comparable viability and differentiation after printing. Moreover, in the zebrafish embryo neural deficit model, injection of the NSC-laden PU hydrogels promoted the repair of damaged CNS. In addition, the function of adult zebrafish with traumatic brain injury was rescued after implantation of the 3D-printed NSC-laden constructs. Therefore, the newly developed 3D bioprinting technique may offer new possibilities for future therapeutic strategy of neural tissue regeneration.

New tides: using zebrafish to study renal regeneration.

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McCampbell, Kristen K; Wingert, Rebecca A

2014-02-01

Over the past several decades, the zebrafish has become one of the major vertebrate model organisms used in biomedical research. In this arena, the zebrafish has emerged as an applicable system for the study of kidney diseases and renal regeneration. The relevance of the zebrafish model for nephrology research has been increasingly appreciated as the understanding of zebrafish kidney structure, ontogeny, and the response to damage has steadily expanded. Recent studies have documented the amazing regenerative characteristics of the zebrafish kidney, which include the ability to replace epithelial populations after acute injury and to grow new renal functional units, termed nephrons. Here we discuss how nephron composition is conserved between zebrafish and mammals, and highlight how recent findings from zebrafish studies utilizing transgenic technologies and chemical genetics can complement traditional murine approaches in the effort to dissect how the kidney responds to acute damage and identify therapeutics that enhance human renal regeneration. Copyright © 2014 Mosby, Inc. All rights reserved.

Concurrent OCT imaging of stimulus evoked retinal neural activation and hemodynamic responses

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Son, Taeyoon; Wang, Benquan; Lu, Yiming; Chen, Yanjun; Cao, Dingcai; Yao, Xincheng

2017-02-01

It is well established that major retinal diseases involve distortions of the retinal neural physiology and blood vascular structures. However, the details of distortions in retinal neurovascular coupling associated with major eye diseases are not well understood. In this study, a multi-modal optical coherence tomography (OCT) imaging system was developed to enable concurrent imaging of retinal neural activity and vascular hemodynamics. Flicker light stimulation was applied to mouse retinas to evoke retinal neural responses and hemodynamic changes. The OCT images were acquired continuously during the pre-stimulation, light-stimulation, and post-stimulation phases. Stimulus-evoked intrinsic optical signals (IOSs) and hemodynamic changes were observed over time in blood-free and blood regions, respectively. Rapid IOSs change occurred almost immediately after stimulation. Both positive and negative signals were observed in adjacent retinal areas. The hemodynamic changes showed time delays after stimulation. The signal magnitudes induced by light stimulation were observed in blood regions and did not show significant changes in blood-free regions. These differences may arise from different mechanisms in blood vessels and neural tissues in response to light stimulation. These characteristics agreed well with our previous observations in mouse retinas. Further development of the multimodal OCT may provide a new imaging method for studying how retinal structures and metabolic and neural functions are affected by age-related macular degeneration (AMD), glaucoma, diabetic retinopathy (DR), and other diseases, which promises novel noninvasive biomarkers for early disease detection and reliable treatment evaluations of eye diseases.

SINs and SOMs: Neural microcircuits for size tuning in the zebrafish and mouse visual pathway.

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Alison J. Barker

2013-05-01

Full Text Available In many animals, a fast and reliable circuit for discriminating between predator-sized objects and edible (prey-sized objects is necessary for survival. How are receptive fields in visual brain areas organized to extract information about size? Recent studies from the zebrafish optic tectum and the mouse visual cortex suggest de novo shaping of receptive fields by subtypes of inhibitory neurons. Del Bene et al. (2010 describe a population of GABAergic neurons in the zebrafish optic tectum (Superficial Interneurons, SINs that are necessary for size filtering during prey capture. Adesnik et al. (2012 describe a somatostatin-expressing interneuron population (SOMs that confers surround suppression on layer II/III pyramidal cells in mouse V1. Strikingly both the SINs and the SOMs, display size-dependent response properties. Increasing visual stimulus size increases excitatory input to these neurons. Dampening SIN or SOM activity alters tuning of neighboring circuits such that they lose preference for small objects. Both results provide exciting evidence for mechanisms of size filtering in visual circuits. Here we review the roles of the SINs and the SOMs and speculate on the similarity of such spatial filters across species.

Object recognition memory in zebrafish.

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

2016-01-01

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

The importance of Zebrafish in biomedical research.

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Tavares, Bárbara; Santos Lopes, Susana

2013-01-01

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

Zebrafish neurobehavioral phenomics for aquatic neuropharmacology and toxicology research.

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

2016-01-01

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

DNA methylation regulates gabrb2 mRNA expression: developmental variations and disruptions in l-methionine-induced zebrafish with schizophrenia-like symptoms.

Science.gov (United States)

Wang, L; Jiang, W; Lin, Q; Zhang, Y; Zhao, C

2016-11-01

Single nucleotide polymorphisms (SNPs) in the human type A gamma-aminobutyric acid (GABA) receptor β 2 subunit gene (GABRB2) have been associated with schizophrenia and quantitatively correlated with mRNA expression in the postmortem brain tissue of patients with schizophrenia. l-Methionine (MET) administration has been reported to cause a recrudescence of psychotic symptoms in patients with schizophrenia, and similar symptoms have been generated in MET-induced mice. In this study, a zebrafish animal model was used to evaluate the relationship between the gabrb2 mRNA expression and its promoter DNA methylation in developmental and MET-induced schizophrenia-like zebrafish. The results indicated developmental increases in global DNA methylation and decreases in gabrb2 promoter methylation in zebrafish. A significant increase in gabrb2 mRNA levels was observed after GABA was synthesized. Additionally, the MET-triggered schizophrenia-like symptoms in adult zebrafish, involving social withdrawal and cognitive dysfunction analyzed with social interaction and T-maze behavioral tests, were accompanied by significantly increased DNA methylation levels in the global genome and the gabrb2 promoter. Furthermore, the significant correlation between gabrb2 mRNA expression and gabrb2 promoter methylation observed in the developmental stages became non-significant in MET-triggered adult zebrafish. These findings demonstrate that gabrb2 mRNA expression is associated with DNA methylation varies by developmental stage and show that these epigenetic association mechanisms are disrupted in MET-triggered adult zebrafish with schizophrenia-like symptoms. In conclusion, these results provide plausible epigenetic evidence of the GABA A receptor β 2 subunit involvement in the schizophrenia-like behaviors and demonstrate the potential use of zebrafish models in neuropsychiatric research. © 2016 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

Geographic atrophy segmentation in infrared and autofluorescent retina images using supervised learning.

Science.gov (United States)

Devisetti, K; Karnowski, T P; Giancardo, L; Li, Y; Chaum, E

2011-01-01

Geographic Atrophy (GA) of the retinal pigment epithelium (RPE) is an advanced form of atrophic age-related macular degeneration (AMD) and is responsible for about 20% of AMD-related legal blindness in the United States. Two different imaging modalities for retinas, infrared imaging and autofluorescence imaging, serve as interesting complimentary technologies for highlighting GA. In this work we explore the use of neural network classifiers in performing segmentation of GA in registered infrared (IR) and autofluorescence (AF) images. Our segmentation achieved a performance level of 82.5% sensitivity and 92.9% specificity on a per-pixel basis using hold-one-out validation testing. The algorithm, feature extraction, data set and experimental results are discussed and shown.

Episodic-like memory in zebrafish.

Science.gov (United States)

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

2016-11-01

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

Zebrafish swimming in the flow: a particle image velocimetry study

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Violet Mwaffo

2017-11-01

Full Text Available Zebrafish is emerging as a species of choice for the study of a number of biomechanics problems, including balance development, schooling, and neuromuscular transmission. The precise quantification of the flow physics around swimming zebrafish is critical toward a mechanistic understanding of the complex swimming style of this fresh-water species. Although previous studies have elucidated the vortical structures in the wake of zebrafish swimming in placid water, the flow physics of zebrafish swimming against a water current remains unexplored. In an effort to illuminate zebrafish swimming in a dynamic environment reminiscent of its natural habitat, we experimentally investigated the locomotion and hydrodynamics of a single zebrafish swimming in a miniature water tunnel using particle image velocimetry. Our results on zebrafish locomotion detail the role of flow speed on tail beat undulations, heading direction, and swimming speed. Our findings on zebrafish hydrodynamics offer a precise quantification of vortex shedding during zebrafish swimming and demonstrate that locomotory patterns play a central role on the flow physics. This knowledge may help clarify the evolutionary advantage of burst and cruise swimming movements in zebrafish.

Charged particle track reconstruction using artificial neural networks

International Nuclear Information System (INIS)

Glover, C.; Fu, P.; Gabriel, T.; Handler, T.

1992-01-01

This paper summarizes the current state of our research in developing and applying artificial neural network (ANN) algorithm described here is based on a crude model of the retina. It takes as input the coordinates of each charged particle's interaction point (''hit'') in the tracking chamber. The algorithm's output is a set of vectors pointing to other hits that most likely to form a track

Xenotransplantation of human neural progenitor cells to the subretinal space of nonimmunosuppressed pigs

DEFF Research Database (Denmark)

Warfvinge, Karin; Schwartz, Philip H; Kiilgaard, Jens Folke

2011-01-01

To investigate the feasibility of transplanting human neural progenitor cells (hNPCs) to the retina of nonimmunosuppressed pigs, cultured hNPCs were injected into the subretinal space of 5 adult pigs after laser burns were applied to promote donor cell integration. Postoperatively, the retinal ve...

Prey capture in zebrafish larvae serves as a model to study cognitive functions

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Akira eMuto

2013-06-01

Full Text Available Prey capture in zebrafish larvae is an innate behavior which can be observed as early as 4 days post fertilization, the day when they start to swim. This simple behavior apparently involves several neural processes including visual perception, recognition, decision-making, and motor control, and, therefore, serves as a good model system to study cognitive functions underlying natural behaviors in vertebrates. Recent progresses in imaging techniques provided us with a unique opportunity to image neuronal activity in the brain of an intact fish in real-time while the fish perceives a natural prey, paramecium. By expanding this approach, it would be possible to image entire brain areas at a single cell resolution in real-time during prey capture, and identify neuronal circuits important for cognitive functions. Further, activation or inhibition of those neuronal circuits with recently developed optogenetic tools or neurotoxins should shed light on their roles. Thus, we will be able to explore the prey capture in zebrafish larvae more thoroughly at cellular levels, which should establish a basis of understanding of the cognitive function in vertebrates.

Radioadaptive Cytoprotective Pathways in the Mouse Retina

Science.gov (United States)

Zanello, Susana B.; Wotring, V.; Theriot, C.; Ploutz-Snyder, R.; Zhang, Y.; Wu, H.

2010-01-01

Exposure to cosmic radiation implies a risk of tissue degeneration. Radiation retinopathy is a complication of radiotherapy and exhibits common features with other retinopathies and neuropathies. Exposure to a low radiation dose elicits protective cellular events (radioadaptive response), reducing the stress of a subsequent higher dose. To assess the risk of radiation-induced retinal changes and the extent to which a small priming dose reduces this risk, we used a mouse model exposed to a source of Cs-137-gamma radiation. Gene expression profiling of retinas from non-irradiated control C57BL/6J mice (C) were compared to retinas from mice treated with a low 50 mGy dose (LD), a high 6 Gy dose (HD), and a combined treatment of 50 mGy (priming) and 6 Gy (challenge) doses (LHD). Whole retina RNA was isolated and expression analysis for selected genes performed by RTqPCR. Relevant target genes associated with cell death/survival, oxidative stress, cellular stress response and inflammation pathways, were analyzed. Cellular stress response genes were upregulated at 4 hr after the challenge dose in LHD retinas (Sirt1: 1.5 fold, Hsf1: 1.7 fold, Hspa1a: 2.5 fold; Hif1a: 1.8 fold, Bag1: 1.7). A similar trend was observed in LD animals. Most antioxidant enzymes (Hmox1, Sod2, Prdx1, Cygb, Cat1) and inflammatory mediators (NF B, Ptgs2 and Tgfb1) were upregulated in LHD and LD retinas. Expression of the pro-survival gene Bcl2 was upregulated in LD (6-fold) and LHD (4-fold) retinas. In conclusion, cytoprotective gene networks activation in the retina suggests a radioadaptive response to a priming irradiation dose, with mitigation of the deleterious effects of a subsequent high dose exposure. The enhancement of these cytoprotective mechanisms has potential value as a countermeasure to ocular alterations caused by radiation alone or in combination with other factors in spaceflight environments.

Retina image–based optic disc segmentation

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Ching-Lin Wang

2016-05-01

Full Text Available The change of optic disc can be used to diagnose many eye diseases, such as glaucoma, diabetic retinopathy and macular degeneration. Moreover, retinal blood vessel pattern is unique for human beings even for identical twins. It is a highly stable pattern in biometric identification. Since optic disc is the beginning of the optic nerve and main blood vessels in retina, it can be used as a reference point of identification. Therefore, optic disc segmentation is an important technique for developing a human identity recognition system and eye disease diagnostic system. This article hence presents an optic disc segmentation method to extract the optic disc from a retina image. The experimental results show that the optic disc segmentation method can give impressive results in segmenting the optic disc from a retina image.

The Zebrafish Model Organism Database (ZFIN)

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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...

Mechanisms of cadmium-caused eye hypoplasia and hypopigmentation in zebrafish embryos.

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Zhang, Ting; Zhou, Xin-Ying; Ma, Xu-Fa; Liu, Jing-Xia

2015-10-01

Cadmium-caused head and eye hypoplasia and hypopigmentation has been recognized for a long time, but knowledge of the underlying mechanisms is limited. In this study, we found that high mortality occurred in exposed embryos after 24 hpf, when cadmium (Cd) dosage was above 17.8 μM. Using high-throughput in situ hybridization screening, we found that genes labelling the neural crest and its derivative pigment cells exhibited obviously reduced expression in Cd-exposed embryos from 24 hpf, 2 days earlier than head and eye hypoplasia and hypopigmentation occurred. Moreover, based on expression of crestin, a neural crest marker, we found that embryos before the gastrula stage were more sensitive to cadmium toxicity and that damage caused by Cd on embryogenesis was dosage dependent. In addition, by phenotype observation and detection of neural crest and pigment cell markers, we found that BIO and retinoic acid (RA) could neutralize the toxic effects of Cd on zebrafish embryogenesis. In this study, we first determined that Cd blocked the formation of the neural crest and inhibited specification of pigment cells, which might contribute to the molecular mechanisms underlying the phenotype defects of head and eye hypoplasia and hypopigmentation in Cd-exposed embryos. Moreover, we found that compounds BIO or RA could neutralize the toxic effects of Cd. Copyright © 2015 Elsevier B.V. All rights reserved.

Depletion of the Third Complement Component Ameliorates Age-Dependent Oxidative Stress and Positively Modulates Autophagic Activity in Aged Retinas in a Mouse Model

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Dorota Rogińska

2017-01-01

Full Text Available The aim of the study was to investigate the influence of complement component C3 global depletion on the biological structure and function of the aged retina. In vivo morphology (OCT, electrophysiological function (ERG, and the expression of selected oxidative stress-, apoptosis-, and autophagy-related proteins were assessed in retinas of 12-month-old C3-deficient and WT mice. Moreover, global gene expression in retinas was analyzed by RNA arrays. We found that the absence of active C3 was associated with (1 alleviation of the age-dependent decrease in retinal thickness and gradual deterioration of retinal bioelectrical function, (2 significantly higher levels of antioxidant enzymes (catalase and glutathione reductase and the antiapoptotic survivin and Mcl-1/Bak dimer, (3 lower expression of the cellular oxidative stress marker—4HNE—and decreased activity of proapoptotic caspase-3, (4 ameliorated retinal autophagic activity with localization of ubiquitinated protein conjugates commonly along the retinal pigment epithelium (RPE layer, and (5 significantly increased expression of several gene sets associated with maintenance of the physiological functions of the neural retina. Our findings shed light on mechanisms of age-related retinal alterations by identifying C3 as a potential therapeutic target for retinal aging.

Imaging of the peripheral retina

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Marcus Kernt

2013-01-01

Full Text Available The technical progress of the recent years has revolutionized imaging in ophthalmology. Scanning laser ophthalmoscopy (SLO, digital angiography, optical coherence tomography (OCT, and detection of fundus autofluorescence (FAF have fundamentally changed our understanding of numerous retinal and choroidal diseases. Besides the tremendous advances in macular diagnostics, there is more and more evidence that central pathologies are often directly linked to changes in the peripheral retina. This review provides a brief overview on current posterior segment imaging techniques with a special focus on the peripheral retina.

The role of omega-3 long-chain polyunsaturated fatty acids in health and disease of the retina.

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SanGiovanni, John Paul; Chew, Emily Y

2005-01-01

In this work we advance the hypothesis that omega-3 (omega-3) long-chain polyunsaturated fatty acids (LCPUFAs) exhibit cytoprotective and cytotherapeutic actions contributing to a number of anti-angiogenic and neuroprotective mechanisms within the retina. omega-3 LCPUFAs may modulate metabolic processes and attenuate effects of environmental exposures that activate molecules implicated in pathogenesis of vasoproliferative and neurodegenerative retinal diseases. These processes and exposures include ischemia, chronic light exposure, oxidative stress, inflammation, cellular signaling mechanisms, and aging. A number of bioactive molecules within the retina affect, and are effected by such conditions. These molecules operate within complex systems and include compounds classified as eicosanoids, angiogenic factors, matrix metalloproteinases, reactive oxygen species, cyclic nucleotides, neurotransmitters and neuromodulators, pro-inflammatory and immunoregulatory cytokines, and inflammatory phospholipids. We discuss the relationship of LCPUFAs with these bioactivators and bioactive compounds in the context of three blinding retinal diseases of public health significance that exhibit both vascular and neural pathology. How is omega-3 LCPUFA status related to retinal structure and function? Docosahexaenoic acid (DHA), a major dietary omega-3 LCPUFA, is also a major structural lipid of retinal photoreceptor outer segment membranes. Biophysical and biochemical properties of DHA may affect photoreceptor membrane function by altering permeability, fluidity, thickness, and lipid phase properties. Tissue DHA status affects retinal cell signaling mechanisms involved in phototransduction. DHA may operate in signaling cascades to enhance activation of membrane-bound retinal proteins and may also be involved in rhodopsin regeneration. Tissue DHA insufficiency is associated with alterations in retinal function. Visual processing deficits have been ameliorated with DHA supplementation

Simple explant culture of the embryonic chicken retina with long-term preservation of photoreceptors.

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Thangaraj, Gopenath; Greif, Alexander; Layer, Paul G

2011-10-01

Structurally stable in vitro-model systems are indispensible to analyse neural development during embryogenesis, follow cellular differentiation and evaluate neurotoxicological or growth factor effects. Here we describe a three-dimensional, long-term in vitro-culture system of the embryonic chick retina which supports photoreceptor development. Retinal tissue was isolated from E6 chick eye, and cultured as explants by continuous orbital rotation to allow free floatation without any supporting materials. Young stage (E6) immature retinas were cultured for various time periods in order to follow the differentiation of cell types and plexiform layers by immunocytochemical methods. These explants could be cultured for at least 2-3 weeks with remarkable retention of retinal architecture. Interestingly, photoreceptors developed in the absence of pigment epithelium. Electron microscopic studies revealed formation of structures resembling photoreceptor outer segments, a feature not reported previously. Thus, the verification of photoreceptors, Müller cells, inner retinal cells and the inner plexiform layer described in our study establishes this explant culture as a valuable in vivo-like model system. Crown Copyright © 2011. Published by Elsevier Ltd. All rights reserved.

Genomic Organization of Zebrafish microRNAs

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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.

Macrophage–Microbe Interactions: Lessons from the Zebrafish Model

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Nagisa Yoshida

2017-12-01

Full Text Available Macrophages provide front line defense against infections. The study of macrophage–microbe interplay is thus crucial for understanding pathogenesis and infection control. Zebrafish (Danio rerio larvae provide a unique platform to study macrophage–microbe interactions in vivo, from the level of the single cell to the whole organism. Studies using zebrafish allow non-invasive, real-time visualization of macrophage recruitment and phagocytosis. Furthermore, the chemical and genetic tractability of zebrafish has been central to decipher the complex role of macrophages during infection. Here, we discuss the latest developments using zebrafish models of bacterial and fungal infection. We also review novel aspects of macrophage biology revealed by zebrafish, which can potentiate development of new therapeutic strategies for humans.

Whole-Volume Clustering of Time Series Data from Zebrafish Brain Calcium Images via Mixture Modeling.

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Nguyen, Hien D; Ullmann, Jeremy F P; McLachlan, Geoffrey J; Voleti, Venkatakaushik; Li, Wenze; Hillman, Elizabeth M C; Reutens, David C; Janke, Andrew L

2018-02-01

Calcium is a ubiquitous messenger in neural signaling events. An increasing number of techniques are enabling visualization of neurological activity in animal models via luminescent proteins that bind to calcium ions. These techniques generate large volumes of spatially correlated time series. A model-based functional data analysis methodology via Gaussian mixtures is suggested for the clustering of data from such visualizations is proposed. The methodology is theoretically justified and a computationally efficient approach to estimation is suggested. An example analysis of a zebrafish imaging experiment is presented.

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.

Zebrafish: an animal model for research in veterinary medicine.

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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

Multi-organ abnormalities and mTORC1 activation in zebrafish model of multiple acyl-CoA dehydrogenase deficiency.

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Seok-Hyung Kim

2013-06-01

Full Text Available Multiple Acyl-CoA Dehydrogenase Deficiency (MADD is a severe mitochondrial disorder featuring multi-organ dysfunction. Mutations in either the ETFA, ETFB, and ETFDH genes can cause MADD but very little is known about disease specific mechanisms due to a paucity of animal models. We report a novel zebrafish mutant dark xavier (dxa(vu463 that has an inactivating mutation in the etfa gene. dxa(vu463 recapitulates numerous pathological and biochemical features seen in patients with MADD including brain, liver, and kidney disease. Similar to children with MADD, homozygote mutant dxa(vu463 zebrafish have a spectrum of phenotypes ranging from moderate to severe. Interestingly, excessive maternal feeding significantly exacerbated the phenotype. Homozygous mutant dxa(vu463 zebrafish have swollen and hyperplastic neural progenitor cells, hepatocytes and kidney tubule cells as well as elevations in triacylglycerol, cerebroside sulfate and cholesterol levels. Their mitochondria were also greatly enlarged, lacked normal cristae, and were dysfunctional. We also found increased signaling of the mechanistic target of rapamycin complex 1 (mTORC1 with enlarged cell size and proliferation. Treatment with rapamycin partially reversed these abnormalities. Our results indicate that etfa gene function is remarkably conserved in zebrafish as compared to humans with highly similar pathological, biochemical abnormalities to those reported in children with MADD. Altered mTORC1 signaling and maternal nutritional status may play critical roles in MADD disease progression and suggest novel treatment approaches that may ameliorate disease severity.

Imagen de retina de campo ultra-amplio

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Gerardo García-Aguirre

2017-11-01

Conclusión: Las imágenes de campo ultra-amplio han revolucionado la forma en la que estudiamos y entendemos la enfermedad de la retina. A medida que la tecnología para obtenerlas se haga más accesible, formará parte del armamentario de rutina para estudiar las enfermedades de la retina.

Zebrafish: A Versatile Animal Model for Fertility Research

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Jing Ying Hoo

2016-01-01

Full Text Available The utilization of zebrafish in biomedical research is very common in the research world nowadays. Today, it has emerged as a favored vertebrate organism for the research in science of reproduction. There is a significant growth in amount numbers of scientific literature pertaining to research discoveries in reproductive sciences in zebrafish. It has implied the importance of zebrafish in this particular field of research. In essence, the current available literature has covered from the very specific brain region or neurons of zebrafish, which are responsible for reproductive regulation, until the gonadal level of the animal. The discoveries and findings have proven that this small animal is sharing a very close/similar reproductive system with mammals. More interestingly, the behavioral characteristics and along with the establishment of animal courtship behavior categorization in zebrafish have laid an even stronger foundation and firmer reason on the suitability of zebrafish utilization in research of reproductive sciences. In view of the immense importance of this small animal for the development of reproductive sciences, this review aimed at compiling and describing the proximate close similarity of reproductive regulation on zebrafish and human along with factors contributing to the infertility, showing its versatility and its potential usage for fertility research.

Quo natas, Danio?—Recent Progress in Modeling Cancer in Zebrafish

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Stefanie Kirchberger

2017-08-01

Full Text Available Over the last decade, zebrafish has proven to be a powerful model in cancer research. Zebrafish form tumors that histologically and genetically resemble human cancers. The live imaging and cost-effective compound screening possible with zebrafish especially complement classic mouse cancer models. Here, we report recent progress in the field, including genetically engineered zebrafish cancer models, xenotransplantation of human cancer cells into zebrafish, promising approaches toward live investigation of the tumor microenvironment, and identification of therapeutic strategies by performing compound screens on zebrafish cancer models. Given the recent advances in genome editing, personalized zebrafish cancer models are now a realistic possibility. In addition, ongoing automation will soon allow high-throughput compound screening using zebrafish cancer models to be part of preclinical precision medicine approaches.

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

Hypertensive retinopathy identification through retinal fundus image using backpropagation neural network

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Syahputra, M. F.; Amalia, C.; Rahmat, R. F.; Abdullah, D.; Napitupulu, D.; Setiawan, M. I.; Albra, W.; Nurdin; Andayani, U.

2018-03-01

Hypertension or high blood pressure can cause damage of blood vessels in the retina of eye called hypertensive retinopathy (HR). In the event Hypertension, it will cause swelling blood vessels and a decrese in retina performance. To detect HR in patients body, it is usually performed through physical examination of opthalmoscope which is still conducted manually by an ophthalmologist. Certainly, in such a manual manner, takes a ong time for a doctor to detetct HR on aa patient based on retina fundus iamge. To overcome ths problem, a method is needed to identify the image of retinal fundus automatically. In this research, backpropagation neural network was used as a method for retinal fundus identification. The steps performed prior to identification were pre-processing (green channel, contrast limited adapative histogram qualization (CLAHE), morphological close, background exclusion, thresholding and connected component analysis), feature extraction using zoning. The results show that the proposed method is able to identify retinal fundus with an accuracy of 95% with maximum epoch of 1500.

Evaluating human cancer cell metastasis in zebrafish

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

2013-01-01

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

Molecular cloning and developmental expression of Tlx (Hox11) genes in zebrafish (Danio rerio).

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Langenau, D M; Palomero, T; Kanki, J P; Ferrando, A A; Zhou, Y; Zon, L I; Look, A T

2002-09-01

Tlx (Hox11) genes are orphan homeobox genes that play critical roles in the regulation of early developmental processes in vertebrates. Here, we report the identification and expression patterns of three members of the zebrafish Tlx family. These genes share similar, but not identical, expression patterns with other vertebrate Tlx-1 and Tlx-3 genes. Tlx-1 is expressed early in the developing hindbrain and pharyngeal arches, and later in the putative splenic primordium. However, unlike its orthologues, zebrafish Tlx-1 is not expressed in the cranial sensory ganglia or spinal cord. Two homologues of Tlx-3 were identified: Tlx-3a and Tlx-3b, which are both expressed in discrete regions of the developing nervous system, including the cranial sensory ganglia and Rohon-Beard neurons. However, only Tlx-3a is expressed in the statoacoustic cranial ganglia, enteric neurons and non-neural tissues such as the fin bud and pharyngeal arches and Tlx-3b is only expressed in the dorsal root ganglia. Copyright 2002 Elsevier Science Ireland Ltd.

Normal anatomy and histology of the adult zebrafish.

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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.

Characterization of genetically targeted neuron types in the zebrafish optic tectum

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Estuardo eRobles

2011-02-01

Full Text Available The optically transparent larval zebrafish is ideally suited for in vivo analyses of neural circuitry controlling visually guided behaviors. However, there is a lack of information regarding specific cell types in the major retinorecipient brain region of the fish, the optic tectum. Here we report the characterization of three previously unidentified tectal cell types that are specifically labeled by dlx5/6 enhancer elements. In vivo laser scanning microscopy in conjunction with ex vivo array tomography revealed that these neurons differ in their morphologies, synaptic connectivity, and neurotransmitter phenotypes. The first type is an excitatory bistratified periventricular interneuron (bsPVIN that forms a dendritic arbor in the retinorecipient stratum fibrosum griseum et superficiale (SFGS and an axonal arbor in the stratum griseum centrale (SGC. The second type, a GABAergic nonstratified periventricular interneuron (nsPVIN, extends a bushy arbor containing both dendrites and axons into the SGC and the deepest sublayers of the SFGS. The third type is a GABAergic periventricular projection neuron (PVPN that extends a dendritic arbor into the SGC and a long axon to the torus semicircularis, medulla oblongata, and anterior hindbrain. Interestingly, the same axons form en passant synapses within the deepest neuropil layer of the tectum, the stratum album centrale. This approach revealed several novel aspects of tectal circuitry, including: (1 a glutamatergic mode of transmission from the superficial, retinorecipient neuropil layers to the deeper, output layers, (2 the presence of interneurons with mixed dendrite/axon arbors likely involved in local processing, and (3 a heretofore unknown GABAergic tectofugal projection to midbrain and hindbrain. These observations establish a framework for studying the morphological and functional differentiation of neural circuits in the zebrafish visual system.

Evaluation of visible implant elastomer tags in zebrafish (Danio rerio

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Claudia Hohn

2013-11-01

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

The neurotoxic effects of N-methyl-N-nitrosourea on the electrophysiological property and visual signal transmission of rat's retina.

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Tao, Ye; Chen, Tao; Liu, Bei; Yang, Guo Qing; Peng, Guanghua; Zhang, Hua; Huang, Yi Fei

2015-07-01

The neurotoxic effects of N-methyl-N-nitrosourea (MNU) on the inner retinal neurons and related visual signal circuits have not been described in any animal models or human, despite ample morphological evidences about the MNU induced photoreceptor (PR) degeneration. With the helping of MEA (multielectrode array) recording system, we gained the opportunity to systemically explore the neural activities and visual signal pathways of MNU administrated rats. Our MEA research identified remarkable alterations in the electrophysiological properties and firstly provided instructive information about the neurotoxicity of MNU that affects the signal transmission in the inner retina. Moreover, the spatial electrophysiological functions of retina were monitored and found that the focal PRs had different vulnerabilities to the MNU. The MNU-induced PR dysfunction exhibited a distinct spatial- and time-dependent progression. In contrast, the spiking activities of both central and peripheral RGCs altered synchronously in response to the MNU administration. Pharmacological tests suggested that gap junctions played a pivotal role in this homogeneous response of RGCs. SNR analysis of MNU treated retina suggested that the signaling efficiency and fidelity of inner retinal circuits have been ruined by this toxicant, although the microstructure of the inner retina seemed relatively consolidated. The present study provided an appropriate example of MEA investigations on the toxicant induced pathological models and the effects of the pharmacological compounds on neuron activities. The positional MEA information would enrich our knowledge about the pathology of MNU induced RP models, and eventually be instrumental for elucidating the underlying mechanism of human RP. Copyright © 2015 Elsevier Inc. All rights reserved.

Neural crest contributions to the lamprey head

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McCauley, David W.; Bronner-Fraser, Marianne

2003-01-01

The neural crest is a vertebrate-specific cell population that contributes to the facial skeleton and other derivatives. We have performed focal DiI injection into the cranial neural tube of the developing lamprey in order to follow the migratory pathways of discrete groups of cells from origin to destination and to compare neural crest migratory pathways in a basal vertebrate to those of gnathostomes. The results show that the general pathways of cranial neural crest migration are conserved throughout the vertebrates, with cells migrating in streams analogous to the mandibular and hyoid streams. Caudal branchial neural crest cells migrate ventrally as a sheet of cells from the hindbrain and super-pharyngeal region of the neural tube and form a cylinder surrounding a core of mesoderm in each pharyngeal arch, similar to that seen in zebrafish and axolotl. In addition to these similarities, we also uncovered important differences. Migration into the presumptive caudal branchial arches of the lamprey involves both rostral and caudal movements of neural crest cells that have not been described in gnathostomes, suggesting that barriers that constrain rostrocaudal movement of cranial neural crest cells may have arisen after the agnathan/gnathostome split. Accordingly, neural crest cells from a single axial level contributed to multiple arches and there was extensive mixing between populations. There was no apparent filling of neural crest derivatives in a ventral-to-dorsal order, as has been observed in higher vertebrates, nor did we find evidence of a neural crest contribution to cranial sensory ganglia. These results suggest that migratory constraints and additional neural crest derivatives arose later in gnathostome evolution.

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

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

2014-08-06

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

Topography of ganglion cell production in the cat's retina

International Nuclear Information System (INIS)

Walsh, C.; Polley, E.H.

1985-01-01

The ganglion cells of the cat's retina form several classes distinguishable in terms of soma size, axon diameter, dendritic morphology, physiological properties, and central connections. Labeling with [ 3 H]thymidine shows that the ganglion cells which survive in the adult are produced as several temporally shifted, overlapping waves: medium-sized cells are produced before large cells, whereas the smallest ganglion cells are produced throughout the period of ganglion cell generation. Large cells and medium-sized cells show the same distinctive pattern of production, forming rough spirals around the area centralis. The oldest cells tend to lie superior and nasal to the area centralis, whereas cells in the inferior nasal retina and inferior temporal retina are, in general, progressively younger. Within each retinal quadrant, cells nearer the area centralis tend to be older than cells in the periphery, but there is substantial overlap. The retinal raphe divides the superior temporal quadrant into two zones with different patterns of cell addition. Superior temporal retina near the vertical meridian adds cells only slightly later than superior nasal retina, whereas superior temporal retina near the horizontal meridian adds cells very late, contemporaneously with inferior temporal retina. The broader wave of production of smaller ganglion cells seems to follow this same spiral pattern at its beginning and end. The presence of the area centralis as a nodal point about which ganglion cell production in the retinal quadrants pivots suggests that the area centralis is already an important retinal landmark even at the earliest stages of retinal development

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.

A dominant negative zebrafish Ahr2 partially protects developing zebrafish from dioxin toxicity.

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Kevin A Lanham

Full Text Available The toxicity by 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD is thought to be caused by activation of the aryl hydrocarbon receptor (AHR. However, our understanding of how AHR activation by TCDD leads to toxic effects is poor. Ideally we would like to manipulate AHR activity in specific tissues and at specific times. One route to this is expressing dominant negative AHRs (dnAHRs. This work describes the construction and characterization of dominant negative forms of the zebrafish Ahr2 in which the C-terminal transactivation domain was either removed, or replaced with the inhibitory domain from the Drosophila engrailed repressor protein. One of these dnAhr2s was selected for expression from the ubiquitously active e2fα promoter in transgenic zebrafish. We found that these transgenic zebrafish expressing dnAhr2 had reduced TCDD induction of the Ahr2 target gene cyp1a, as measured by 7-ethoxyresorufin-O-deethylase activity. Furthermore, the cardiotoxicity produced by TCDD, pericardial edema, heart malformation, and reduced blood flow, were all mitigated in the zebrafish expressing the dnAhr2. These results provide in vivo proof-of-principle results demonstrating the effectiveness of dnAHRs in manipulating AHR activity in vivo, and demonstrating that this approach can be a means for blocking TCDD toxicity.

The Difference between Anxiolytic and Anxiogenic Effects Induced by Acute and Chronic Alcohol Exposure and Changes in Associative Learning and Memory Based on Color Preference and the Cause of Parkinson-Like Behaviors in Zebrafish.

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Xiang Li

Full Text Available We describe an interdisciplinary comparison of the effects of acute and chronic alcohol exposure in terms of their disturbance of light, dark and color preferences and the occurrence of Parkinson-like behavior in zebrafish through computer visual tracking, data mining, and behavioral and physiological analyses. We found that zebrafish in anxiolytic and anxious states, which are induced by acute and chronic repeated alcohol exposure, respectively, display distinct emotional reactions in light/dark preference tests as well as distinct learning and memory abilities in color-enhanced conditional place preference (CPP tests. Additionally, compared with the chronic alcohol (1.0% treatment, acute alcohol exposure had a significant, dose-dependent effect on anxiety, learning and memory (color preference as well as locomotive activities. Acute exposure doses (0.5%, 1.0%, and 1.5% generated an "inverted V" dose-dependent pattern in all of the behavioral parameters, with 1.0% having the greatest effect, while the chronic treatment had a moderate effect. Furthermore, by measuring locomotive activity, learning and memory performance, the number of dopaminergic neurons, tyrosine hydroxylase expression, and the change in the photoreceptors in the retina, we found that acute and chronic alcohol exposure induced varying degrees of Parkinson-like symptoms in zebrafish. Taken together, these results illuminated the behavioral and physiological mechanisms underlying the changes associated with learning and memory and the cause of potential Parkinson-like behaviors in zebrafish due to acute and chronic alcohol exposure.

The Difference between Anxiolytic and Anxiogenic Effects Induced by Acute and Chronic Alcohol Exposure and Changes in Associative Learning and Memory Based on Color Preference and the Cause of Parkinson-Like Behaviors in Zebrafish.

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Li, Xiang; Li, Xu; Li, Yi-Xiang; Zhang, Yuan; Chen, Di; Sun, Ming-Zhu; Zhao, Xin; Chen, Dong-Yan; Feng, Xi-Zeng

2015-01-01

We describe an interdisciplinary comparison of the effects of acute and chronic alcohol exposure in terms of their disturbance of light, dark and color preferences and the occurrence of Parkinson-like behavior in zebrafish through computer visual tracking, data mining, and behavioral and physiological analyses. We found that zebrafish in anxiolytic and anxious states, which are induced by acute and chronic repeated alcohol exposure, respectively, display distinct emotional reactions in light/dark preference tests as well as distinct learning and memory abilities in color-enhanced conditional place preference (CPP) tests. Additionally, compared with the chronic alcohol (1.0%) treatment, acute alcohol exposure had a significant, dose-dependent effect on anxiety, learning and memory (color preference) as well as locomotive activities. Acute exposure doses (0.5%, 1.0%, and 1.5%) generated an "inverted V" dose-dependent pattern in all of the behavioral parameters, with 1.0% having the greatest effect, while the chronic treatment had a moderate effect. Furthermore, by measuring locomotive activity, learning and memory performance, the number of dopaminergic neurons, tyrosine hydroxylase expression, and the change in the photoreceptors in the retina, we found that acute and chronic alcohol exposure induced varying degrees of Parkinson-like symptoms in zebrafish. Taken together, these results illuminated the behavioral and physiological mechanisms underlying the changes associated with learning and memory and the cause of potential Parkinson-like behaviors in zebrafish due to acute and chronic alcohol exposure.

Hypoxia-induced retinal neovascularization in zebrafish embryos: a potential model of retinopathy of prematurity.

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Wu, Yu-Ching; Chang, Chao-Yuan; Kao, Alex; Hsi, Brian; Lee, Shwu-Huey; Chen, Yau-Hung; Wang, I-Jong

2015-01-01

Retinopathy of prematurity, formerly known as a retrolental fibroplasia, is a leading cause of infantile blindness worldwide. Retinopathy of prematurity is caused by the failure of central retinal vessels to reach the retinal periphery, creating a nonperfused peripheral retina, resulting in retinal hypoxia, neovascularization, vitreous hemorrhage, vitreoretinal fibrosis, and loss of vision. We established a potential retinopathy of prematurity model by using a green fluorescent vascular endothelium zebrafish transgenic line treated with cobalt chloride (a hypoxia-inducing agent), followed by GS4012 (a vascular endothelial growth factor inducer) at 24 hours postfertilization, and observed that the number of vascular branches and sprouts significantly increased in the central retinal vascular trunks 2-4 days after treatment. We created an angiography method by using tetramethylrhodamine dextran, which exhibited severe vascular leakage through the vessel wall into the surrounding retinal tissues. The quantification of mRNA extracted from the heads of the larvae by using real-time quantitative polymerase chain reaction revealed a twofold increase in vegfaa and vegfr2 expression compared with the control group, indicating increased vascular endothelial growth factor signaling in the hypoxic condition. In addition, we demonstrated that the hypoxic insult could be effectively rescued by several antivascular endothelial growth factor agents such as SU5416, bevacizumab, and ranibizumab. In conclusion, we provide a simple, highly reproducible, and clinically relevant retinopathy of prematurity model based on zebrafish embryos; this model may serve as a useful platform for clarifying the mechanisms of human retinopathy of prematurity and its progression.

Helping the Retina Regenerate

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... the retina News Brief 03/30/17 A new report gives recommendations for regenerating retinal ganglion cells (RGCs), crucial neurons in the back of the eye that carry visual information to the brain. Authored ...

Triclosan Lacks (Anti-Estrogenic Effects in Zebrafish Cells but Modulates Estrogen Response in Zebrafish Embryos

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Hélène Serra

2018-04-01

Full Text Available Triclosan (TCS, an antimicrobial agent widely found in the aquatic environment, is suspected to act as an endocrine disrupting compound, however mechanistic information is lacking in regards to aquatic species. This study assessed the ability of TCS to interfere with estrogen receptor (ER transcriptional activity, in zebrafish-specific in vitro and in vivo reporter gene assays. We report that TCS exhibits a lack of either agonistic or antagonistic effects on a panel of ER-expressing zebrafish (ZELH-zfERα and -zfERβ and human (MELN cell lines. At the organism level, TCS at concentrations of up to 0.3 µM had no effect on ER-regulated brain aromatase gene expression in transgenic cyp19a1b-GFP zebrafish embryos. At a concentration of 1 µM, TCS interfered with the E2 response in an ambivalent manner by potentializing a low E2 response (0.625 nM, but decreasing a high E2 response (10 nM. Altogether, our study suggests that while modulation of ER-regulated genes by TCS may occur in zebrafish, it does so irrespective of a direct binding and activation of zfERs.

Meduloepitelioma teratóide da retina: relato de caso Teratoid medulloepithelioma of the retina: case report

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Ramon Coral Ghanem

2004-06-01

Full Text Available O meduloepitelioma é um tumor intra-ocular congênito originário do epitélio medular primitivo que, por sua vez, é responsável pela formação do epitélio não pigmentado do corpo ciliar. Ocorre geralmente na infância, de forma unilateral, acometendo o corpo ciliar. O objetivo deste trabalho é documentar um caso raro de meduloepitelioma teratóide originário da retina. Paciente de nove anos, feminina, apresentava baixa acuidade visual (AV, estrabismo e leucocoria no olho esquerdo (OE. A AV era de 1,0 no olho direito e movimentos de mão no OE. Foi observada tumoração retrocristaliniana branco-acinzentada no OE, aparentemente subretiniana, vascularizada, de grande extensão, com alterações císticas na sua superfície. Foram realizadas tomografia de crânio e órbitas e ecografia ocular. A paciente foi submetida à enucleação com suspeita clínica de retinoblastoma. Pelo aspecto histopatológico foi feito o diagnóstico de meduloepitelioma teratóide benigno originário da retina. Na maioria dos casos apresentados na literatura o meduloepitelioma tem origem a partir do epitélio não pigmentado do corpo ciliar. No nosso caso, a neoplasia parece ter tido origem a partir da retina, já que os cortes revelaram epitélio do corpo ciliar preservado e não foi reconhecida a estrutura normal da retina. Embora o tumor apresentado neste relato tenha sido classificado como benigno, o fato de ser lesão de grandes proporções e de crescimento aparentemente recente, justifica a conduta cirúrgica empregada. O tratamento do meduloepitelioma deve objetivar a intervenção cirúrgica precoce, na tentativa de se evitar a disseminação extra-ocular.Medulloepithelioma is a congenital intraocular tumor that usually arises from the primitive medullary epithelium that is destined to form the nonpigmented ciliary epithelium of the ciliary body. It occurs most frequently in early childhood and is unilateral. This report documents a rare case of

A Zebrafish Heart Failure Model for Assessing Therapeutic Agents.

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

2018-03-20

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

Biosecurity and Health Monitoring at the Zebrafish International Resource Center.

Science.gov (United States)

Murray, Katrina N; Varga, Zoltán M; Kent, Michael L

2016-07-01

The Zebrafish International Resource Center (ZIRC) is a repository and distribution center for mutant, transgenic, and wild-type zebrafish. In recent years annual imports of new zebrafish lines to ZIRC have increased tremendously. In addition, after 15 years of research, we have identified some of the most virulent pathogens affecting zebrafish that should be avoided in large production facilities, such as ZIRC. Therefore, while importing a high volume of new lines we prioritize safeguarding the health of our in-house fish colony. Here, we describe the biosecurity and health-monitoring program implemented at ZIRC. This strategy was designed to prevent introduction of new zebrafish pathogens, minimize pathogens already present in the facility, and ensure a healthy zebrafish colony for in-house uses and shipment to customers.

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

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

2013-01-01

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

Why has Nature Chosen Lutein and Zeaxanthin to Protect the Retina?

OpenAIRE

Widomska, Justyna; Subczynski, Witold K

2014-01-01

Age-related macular degeneration (AMD) is associated with a low level of macular carotenoids in the eye retina. Only two carotenoids, namely lutein and zeaxanthin are selectively accumulated in the human eye retina from blood plasma where more than twenty other carotenoids are available. The third carotenoid which is found in the human retina, meso-zeaxanthin is formed directly in the retina from lutein. All these carotenoids, named also macular xanthophylls, play key roles in eye health and ...

Rhythmic ganglion cell activity in bleached and blind adult mouse retinas.

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Menzler, Jacob; Channappa, Lakshmi; Zeck, Guenther

2014-01-01

In retinitis pigmentosa--a degenerative disease which often leads to incurable blindness--the loss of photoreceptors deprives the retina from a continuous excitatory input, the so-called dark current. In rodent models of this disease this deprivation leads to oscillatory electrical activity in the remaining circuitry, which is reflected in the rhythmic spiking of retinal ganglion cells (RGCs). It remained unclear, however, if the rhythmic RGC activity is attributed to circuit alterations occurring during photoreceptor degeneration or if rhythmic activity is an intrinsic property of healthy retinal circuitry which is masked by the photoreceptor's dark current. Here we tested these hypotheses by inducing and analysing oscillatory activity in adult healthy (C57/Bl6) and blind mouse retinas (rd10 and rd1). Rhythmic RGC activity in healthy retinas was detected upon partial photoreceptor bleaching using an extracellular high-density multi-transistor-array. The mean fundamental spiking frequency in bleached retinas was 4.3 Hz; close to the RGC rhythm detected in blind rd10 mouse retinas (6.5 Hz). Crosscorrelation analysis of neighbouring wild-type and rd10 RGCs (separation distance rhythmic RGC spiking in these retinas is driven by a network of presynaptic neurons. The inhibition of glutamatergic ganglion cell input or the inhibition of gap junctional coupling abolished the rhythmic pattern. In rd10 and rd1 retinas the presynaptic network leads to local field potentials, whereas in bleached retinas additional pharmacological disinhibition is required to achieve detectable field potentials. Our results demonstrate that photoreceptor bleaching unmasks oscillatory activity in healthy retinas which shares many features with the functional phenotype detected in rd10 retinas. The quantitative physiological differences advance the understanding of the degeneration process and may guide future rescue strategies.

Synaptic Mechanisms Generating Orientation Selectivity in the ON Pathway of the Rabbit Retina.

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Venkataramani, Sowmya; Taylor, W Rowland

2016-03-16

Neurons that signal the orientation of edges within the visual field have been widely studied in primary visual cortex. Much less is known about the mechanisms of orientation selectivity that arise earlier in the visual stream. Here we examine the synaptic and morphological properties of a subtype of orientation-selective ganglion cell in the rabbit retina. The receptive field has an excitatory ON center, flanked by excitatory OFF regions, a structure similar to simple cell receptive fields in primary visual cortex. Examination of the light-evoked postsynaptic currents in these ON-type orientation-selective ganglion cells (ON-OSGCs) reveals that synaptic input is mediated almost exclusively through the ON pathway. Orientation selectivity is generated by larger excitation for preferred relative to orthogonal stimuli, and conversely larger inhibition for orthogonal relative to preferred stimuli. Excitatory orientation selectivity arises in part from the morphology of the dendritic arbors. Blocking GABAA receptors reduces orientation selectivity of the inhibitory synaptic inputs and the spiking responses. Negative contrast stimuli in the flanking regions produce orientation-selective excitation in part by disinhibition of a tonic NMDA receptor-mediated input arising from ON bipolar cells. Comparison with earlier studies of OFF-type OSGCs indicates that diverse synaptic circuits have evolved in the retina to detect the orientation of edges in the visual input. A core goal for visual neuroscientists is to understand how neural circuits at each stage of the visual system extract and encode features from the visual scene. This study documents a novel type of orientation-selective ganglion cell in the retina and shows that the receptive field structure is remarkably similar to that of simple cells in primary visual cortex. However, the data indicate that, unlike in the cortex, orientation selectivity in the retina depends on the activity of inhibitory interneurons. The

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

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Alessandra M. Welker

2016-02-01

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

Zebrafish con/disp1 reveals multiple spatiotemporal requirements for Hedgehog-signaling in craniofacial development

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Schwend Tyler

2009-11-01

Full Text Available Abstract Background The vertebrate head skeleton is derived largely from cranial neural crest cells (CNCC. Genetic studies in zebrafish and mice have established that the Hedgehog (Hh-signaling pathway plays a critical role in craniofacial development, partly due to the pathway's role in CNCC development. Disruption of the Hh-signaling pathway in humans can lead to the spectral disorder of Holoprosencephaly (HPE, which is often characterized by a variety of craniofacial defects including midline facial clefting and cyclopia 12. Previous work has uncovered a role for Hh-signaling in zebrafish dorsal neurocranium patterning and chondrogenesis, however Hh-signaling mutants have not been described with respect to the ventral pharyngeal arch (PA skeleton. Lipid-modified Hh-ligands require the transmembrane-spanning receptor Dispatched 1 (Disp1 for proper secretion from Hh-synthesizing cells to the extracellular field where they act on target cells. Here we study chameleon mutants, lacking a functional disp1(con/disp1. Results con/disp1 mutants display reduced and dysmorphic mandibular and hyoid arch cartilages and lack all ceratobranchial cartilage elements. CNCC specification and migration into the PA primorida occurs normally in con/disp1 mutants, however disp1 is necessary for post-migratory CNCC patterning and differentiation. We show that disp1 is required for post-migratory CNCC to become properly patterned within the first arch, while the gene is dispensable for CNCC condensation and patterning in more posterior arches. Upon residing in well-formed pharyngeal epithelium, neural crest condensations in the posterior PA fail to maintain expression of two transcription factors essential for chondrogenesis, sox9a and dlx2a, yet continue to robustly express other neural crest markers. Histology reveals that posterior arch residing-CNCC differentiate into fibrous-connective tissue, rather than becoming chondrocytes. Treatments with Cyclopamine, to

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

Science.gov (United States)

Postlethwait, J H

2006-01-01

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

[Degenerative lesions of the peripheral retina].

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Conart, J-B; Baron, D; Berrod, J-P

2014-01-01

Degenerative lesions of the peripheral retina are present from teenage years onwards and increase with age. These abnormabilities are frequent, some of them being benign while others predispose to retinal tears and detachment. In the latter case, the lesions are rhegmatogenous and may justify prophylactic treatment by laser photocoagulation. We distinguish congenital lesions of the peripheral retina and intraretinal, chorioretinal and vitreoretinal degenerations. The holes and tears observed in 2% of the population consist of round atrophic holes, "horseshoe" tears, oral dialyses and giant tears. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Zebrafish models in neuropsychopharmacology and CNS drug discovery.

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Khan, Kanza M; Collier, Adam D; Meshalkina, Darya A; Kysil, Elana V; Khatsko, Sergey L; Kolesnikova, Tatyana; Morzherin, Yury Yu; Warnick, Jason E; Kalueff, Allan V; Echevarria, David J

2017-07-01

Despite the high prevalence of neuropsychiatric disorders, their aetiology and molecular mechanisms remain poorly understood. The zebrafish (Danio rerio) is increasingly utilized as a powerful animal model in neuropharmacology research and in vivo drug screening. Collectively, this makes zebrafish a useful tool for drug discovery and the identification of disordered molecular pathways. Here, we discuss zebrafish models of selected human neuropsychiatric disorders and drug-induced phenotypes. As well as covering a broad range of brain disorders (from anxiety and psychoses to neurodegeneration), we also summarize recent developments in zebrafish genetics and small molecule screening, which markedly enhance the disease modelling and the discovery of novel drug targets. © 2017 The British Pharmacological Society.

Detection of vitellogenin incorporation into zebrafish oocytes by FITC fluorescence

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Yokoi Hayato

2011-04-01

Full Text Available Abstract Background Large volumes of lymph can be collected from the eye-sacs of bubble-eye goldfish. We attempted to induce vitellogenin (Vtg in the eye-sac lymph of bubble-eye goldfish and develop a method for visualizing Vtg incorporation by zebrafish oocytes using FITC-labeling. Methods Estrogen efficiently induced Vtg in the eye-sac lymph of goldfish. After FITC-labeled Vtg was prepared, it was injected into mature female zebrafish. Results Incorporation of FITC-labeled Vtg by zebrafish oocytes was detected in in vivo and in vitro experiments. The embryos obtained from zebrafish females injected with FITC-labeled Vtg emitted FITC fluorescence from the yolk sac and developed normally. Conclusion This method for achieving Vtg incorporation by zebrafish oocytes could be useful in experiments related to the development and endocrinology of zebrafish oocytes.

Definition of the zebrafish genome using flow cytometry and cytogenetic mapping

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Zhou Yi

2007-06-01

Full Text Available Abstract Background The zebrafish (Danio rerio is an important vertebrate model organism system for biomedical research. The syntenic conservation between the zebrafish and human genome allows one to investigate the function of human genes using the zebrafish model. To facilitate analysis of the zebrafish genome, genetic maps have been constructed and sequence annotation of a reference zebrafish genome is ongoing. However, the duplicative nature of teleost genomes, including the zebrafish, complicates accurate assembly and annotation of a representative genome sequence. Cytogenetic approaches provide "anchors" that can be integrated with accumulating genomic data. Results Here, we cytogenetically define the zebrafish genome by first estimating the size of each linkage group (LG chromosome using flow cytometry, followed by the cytogenetic mapping of 575 bacterial artificial chromosome (BAC clones onto metaphase chromosomes. Of the 575 BAC clones, 544 clones localized to apparently unique chromosomal locations. 93.8% of these clones were assigned to a specific LG chromosome location using fluorescence in situ hybridization (FISH and compared to the LG chromosome assignment reported in the zebrafish genome databases. Thirty-one BAC clones localized to multiple chromosomal locations in several different hybridization patterns. From these data, a refined second generation probe panel for each LG chromosome was also constructed. Conclusion The chromosomal mapping of the 575 large-insert DNA clones allows for these clones to be integrated into existing zebrafish mapping data. An accurately annotated zebrafish reference genome serves as a valuable resource for investigating the molecular basis of human diseases using zebrafish mutant models.

Neurotransmitter properties of the newborn human retina

International Nuclear Information System (INIS)

Hollyfield, J.G.; Frederick, J.M.; Rayborn, M.E.

1983-01-01

Human retinal tissue from a newborn was examined autoradiographically for the presence of high-affinity uptake and localization of the following putative neurotransmitters: dopamine, glycine, GABA, aspartate, and glutamate. In addition, the dopamine content of this newborn retina was measured by high pressure liquid chromatography. Our study reveals that specific uptake mechanisms for 3 H-glycine, 3 H-dopamine, and 3 H-GABA are present at birth. However, the number and distribution of cells labeled with each of these 3 H-transmitters are not identical to those observed in adult human retinas. Furthermore, the amount of endogenous dopamine in the newborn retina is approximately 1/20 the adult level. Photoreceptor-specific uptake of 3 H-glutamate and 3 H-aspartate are not observed. These findings indicate that, while some neurotransmitter-specific properties are present at birth, significant maturation of neurotransmitter systems occurs postnatally

Biosecurity and Health Monitoring at the Zebrafish International Resource Center

OpenAIRE

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

2016-01-01

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

A digital retina-like low-level vision processor.

Science.gov (United States)

Mertoguno, S; Bourbakis, N G

2003-01-01

This correspondence presents the basic design and the simulation of a low level multilayer vision processor that emulates to some degree the functional behavior of a human retina. This retina-like multilayer processor is the lower part of an autonomous self-organized vision system, called Kydon, that could be used on visually impaired people with a damaged visual cerebral cortex. The Kydon vision system, however, is not presented in this paper. The retina-like processor consists of four major layers, where each of them is an array processor based on hexagonal, autonomous processing elements that perform a certain set of low level vision tasks, such as smoothing and light adaptation, edge detection, segmentation, line recognition and region-graph generation. At each layer, the array processor is a 2D array of k/spl times/m hexagonal identical autonomous cells that simultaneously execute certain low level vision tasks. Thus, the hardware design and the simulation at the transistor level of the processing elements (PEs) of the retina-like processor and its simulated functionality with illustrative examples are provided in this paper.

The mammalian retina as a clock

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Tosini, Gianluca; Fukuhara, Chiaki

2002-01-01

Many physiological, cellular, and biochemical parameters in the retina of vertebrates show daily rhythms that, in many cases, also persist under constant conditions. This demonstrates that they are driven by a circadian pacemaker. The presence of an autonomous circadian clock in the retina of vertebrates was first demonstrated in Xenopus laevis and then, several years later, in mammals. In X. laevis and in chicken, the retinal circadian pacemaker has been localized in the photoreceptor layer, whereas in mammals, such information is not yet available. Recent advances in molecular techniques have led to the identification of a group of genes that are believed to constitute the molecular core of the circadian clock. These genes are expressed in the retina, although with a slightly different 24-h profile from that observed in the central circadian pacemaker. This result suggests that some difference (at the molecular level) may exist between the retinal clock and the clock located in the suprachiasmatic nuclei of hypothalamus. The present review will focus on the current knowledge of the retinal rhythmicity and the mechanisms responsible for its control.

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

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Adam C Roberts

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

Navigation by environmental geometry: the use of zebrafish as a model.

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Lee, Sang Ah; Vallortigara, Giorgio; Flore, Michele; Spelke, Elizabeth S; Sovrano, Valeria A

2013-10-01

Sensitivity to environmental shape in spatial navigation has been found, at both behavioural and neural levels, in virtually every species tested, starting early in development. Moreover, evidence that genetic deletions can cause selective deficits in such navigation behaviours suggests a genetic basis to navigation by environmental geometry. Nevertheless, the geometric computations underlying navigation have not been specified in any species. The present study teases apart the geometric components within the traditionally used rectangular enclosure and finds that zebrafish selectively represent distance and directional relationships between extended boundary surfaces. Similar behavioural results in geometric navigation tasks with human children provide prima facie evidence for similar underlying cognitive computations and open new doors for probing the genetic foundations that give rise to these computations.

Characterization of behavioral and endocrine effects of LSD on zebrafish.

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Grossman, Leah; Utterback, Eli; Stewart, Adam; Gaikwad, Siddharth; Chung, Kyung Min; Suciu, Christopher; Wong, Keith; Elegante, Marco; Elkhayat, Salem; Tan, Julia; Gilder, Thomas; Wu, Nadine; Dileo, John; Cachat, Jonathan; Kalueff, Allan V

2010-12-25

Lysergic acid diethylamide (LSD) is a potent hallucinogenic drug that strongly affects animal and human behavior. Although adult zebrafish (Danio rerio) are emerging as a promising neurobehavioral model, the effects of LSD on zebrafish have not been investigated previously. Several behavioral paradigms (the novel tank, observation cylinder, light-dark box, open field, T-maze, social preference and shoaling tests), as well as modern video-tracking tools and whole-body cortisol assay were used to characterize the effects of acute LSD in zebrafish. While lower doses (5-100 microg/L) did not affect zebrafish behavior, 250 microg/L LSD increased top dwelling and reduced freezing in the novel tank and observation cylinder tests, also affecting spatiotemporal patterns of activity (as assessed by 3D reconstruction of zebrafish traces and ethograms). LSD evoked mild thigmotaxis in the open field test, increased light behavior in the light-dark test, reduced the number of arm entries and freezing in the T-maze and social preference test, without affecting social preference. In contrast, LSD affected zebrafish shoaling (increasing the inter-fish distance in a group), and elevated whole-body cortisol levels. Overall, our findings show sensitivity of zebrafish to LSD action, and support the use of zebrafish models to study hallucinogenic drugs of abuse. Copyright (c) 2010 Elsevier B.V. All rights reserved.

Characterization of the Zebrafish Homolog of Zipper Interacting Protein Kinase

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Brandon W. Carr

2014-06-01

Full Text Available Zipper-interacting protein kinase (ZIPK is a conserved vertebrate-specific regulator of actomyosin contractility in smooth muscle and non-muscle cells. Murine ZIPK has undergone an unusual divergence in sequence and regulation compared to other ZIPK orthologs. In humans, subcellular localization is controlled by phosphorylation of threonines 299 and 300. In contrast, ZIPK subcellular localization in mouse and rat is controlled by interaction with PAR-4. We carried out a comparative biochemical characterization of the regulation of the zebrafish ortholog of ZIPK. Like the human orthologs zebrafish ZIPK undergoes nucleocytoplasmic-shuttling and is abundant in the cytoplasm, unlike the primarily nuclear rat ZIPK. Rat ZIPK, but not human or zebrafish ZIPK, interacts with zebrafish PAR-4. Mutation of the conserved residues required for activation of the mammalian orthologs abrogated activity of the zebrafish ZIPK. In contrast to the human ortholog, mutation of threonine 299 and 300 in the zebrafish ZIPK has no effect on the activity or subcellular localization. Thus, we found that zebrafish ZIPK functions in a manner most similar to the human ZIPK and quite distinct from murine orthologs, yet the regulation of subcellular localization is not conserved.

A key role for poly(ADP-ribose polymerase 3 in ectodermal specification and neural crest development.

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Michèle Rouleau

2011-01-01

Full Text Available The PARP family member poly(ADP-ribose polymerase 3 (PARP3 is structurally related to the well characterized PARP1 that orchestrates cellular responses to DNA strand breaks and cell death by the synthesis of poly(ADP-ribose. In contrast to PARP1 and PARP2, the functions of PARP3 are undefined. Here, we reveal critical functions for PARP3 during vertebrate development.We have used several in vitro and in vivo approaches to examine the possible functions of PARP3 as a transcriptional regulator, a function suggested from its previously reported association with several Polycomb group (PcG proteins. We demonstrate that PARP3 gene occupancy in the human neuroblastoma cell line SK-N-SH occurs preferentially with developmental genes regulating cell fate specification, tissue patterning, craniofacial development and neurogenesis. Addressing the significance of this association during zebrafish development, we show that morpholino oligonucleotide-directed inhibition of parp3 expression in zebrafish impairs the expression of the neural crest cell specifier sox9a and of dlx3b/dlx4b, the formation of cranial sensory placodes, inner ears and pectoral fins. It delays pigmentation and severely impedes the development of the median fin fold and tail bud.Our findings demonstrate that Parp3 is crucial in the early stages of zebrafish development, possibly by exerting its transcriptional regulatory functions as early as during the specification of the neural plate border.

HCV IRES-mediated core expression in zebrafish.

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Ye Zhao

Full Text Available The lack of small animal models for hepatitis C virus has impeded the discovery and development of anti-HCV drugs. HCV-IRES plays an important role in HCV gene expression, and is an attractive target for antiviral therapy. In this study, we report a zebrafish model with a biscistron expression construct that can co-transcribe GFP and HCV-core genes by human hepatic lipase promoter and zebrafish liver fatty acid binding protein enhancer. HCV core translation was designed mediated by HCV-IRES sequence and gfp was by a canonical cap-dependent mechanism. Results of fluorescence image and in situ hybridization indicate that expression of HCV core and GFP is liver-specific; RT-PCR and Western blotting show that both core and gfp expression are elevated in a time-dependent manner for both transcription and translation. It means that the HCV-IRES exerted its role in this zebrafish model. Furthermore, the liver-pathological impact associated with HCV-infection was detected by examination of gene markers and some of them were elevated, such as adiponectin receptor, heparanase, TGF-β, PDGF-α, etc. The model was used to evaluate three clinical drugs, ribavirin, IFNα-2b and vitamin B12. The results show that vitamin B12 inhibited core expression in mRNA and protein levels in dose-dependent manner, but failed to impact gfp expression. Also VB12 down-regulated some gene transcriptions involved in fat liver, liver fibrosis and HCV-associated pathological process in the larvae. It reveals that HCV-IRES responds to vitamin B12 sensitively in the zebrafish model. Ribavirin did not disturb core expression, hinting that HCV-IRES is not a target site of ribavirin. IFNα-2b was not active, which maybe resulted from its degradation in vivo for the long time. These findings demonstrate the feasibility of the zebrafish model for screening of anti-HCV drugs targeting to HCV-IRES. The zebrafish system provides a novel evidence of using zebrafish as a HCV model organism.

Analysis of Lethality and Malformations During Zebrafish (Danio rerio) Development.

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Raghunath, Azhwar; Perumal, Ekambaram

2018-01-01

The versatility offered by zebrafish (Danio rerio) makes it a powerful and an attractive vertebrate model in developmental toxicity and teratogenicity assays. Apart from the newly introduced chemicals as drugs, xenobiotics also induce abnormal developmental abnormalities and congenital malformations in living organisms. Over the recent decades, zebrafish embryo/larva has emerged as a potential tool to test teratogenicity potential of these chemicals. Zebrafish responds to compounds as mammals do as they share similarities in their development, metabolism, physiology, and signaling pathways with that of mammals. The methodology used by the different scientists varies enormously in the zebrafish embryotoxicity test. In this chapter, we present methods to assess lethality and malformations during zebrafish development. We propose two major malformations scoring systems: binomial and relative morphological scoring systems to assess the malformations in zebrafish embryos/larvae. Based on the scoring of the malformations, the test compound can be classified as a teratogen or a nonteratogen and its teratogenic potential is evaluated.

Expression of nitric oxide synthase during the development of RCS rat retinas.

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Sharma, R K; Warfvinge, K; Ehinger, B

2001-01-01

Nitric oxide (NO) has been reported to be both neurodestructive and neuroprotective in the central nervous system and could possibly play an important role in neurodegenerative disorders. On the assumption that NO synthesis may influence degenerative processes in the retina, we have examined the development and distribution of nitric-oxide-synthase(NOS)-immunoreactive cells in developing Royal College of Surgeons (RCS) rat retinas, which is an animal model for retinal degeneration. An antibody against constitutive neuronal NOS was used for immunocytochemistry on RCS rat retinas from postnatal (PN) days 3, 7, 10, 14, 35, 70 and 281 and compared with that in the normal rats of PN days 3, 7, 10, 14, 54 and adults. Immunoreactive cells were not seen in PN 3 retinas but were distinctly seen in the PN 7 retina along with a plexus in the inner plexiform layer. In both groups (normal and RCS rats) a distinct sublayering of the plexus in the inner plexiform layer could be seen at PN 10, which became more distinct at PN 14. The immunoreactive cells were detected also in the oldest retina examined, which was PN 281 in the case of RCS rats. In both groups, certain amacrine cells, certain bipolar cells and certain horizontal cells were found to be immunoreactive. In conclusion, the developmental timetable of the NOS immunoreactivity was identical in the normal and the RCS rat retinas. The NOS-immunoreactive cells persisted in the RCS retinas even when the retina had degenerated extensively. Abnormalities with the inducible isoforms of NOS cannot be ruled out from this study. We conclude that the chronological and qualitative development of the constitutive neuronal NOS immunoreactivity is normal in RCS rat retinas. Copyright 2001 S. Karger AG, Basel

A zebrafish model of inflammatory lymphangiogenesis

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Kazuhide S. Okuda

2015-10-01

Full Text Available Inflammatory bowel disease (IBD is a disabling chronic inflammatory disease of the gastrointestinal tract. IBD patients have increased intestinal lymphatic vessel density and recent studies have shown that this may contribute to the resolution of IBD. However, the molecular mechanisms involved in IBD-associated lymphangiogenesis are still unclear. In this study, we established a novel inflammatory lymphangiogenesis model in zebrafish larvae involving colitogenic challenge stimulated by exposure to 2,4,6-trinitrobenzenesulfonic acid (TNBS or dextran sodium sulphate (DSS. Treatment with either TNBS or DSS resulted in vascular endothelial growth factor receptor (Vegfr-dependent lymphangiogenesis in the zebrafish intestine. Reduction of intestinal inflammation by the administration of the IBD therapeutic, 5-aminosalicylic acid, reduced intestinal lymphatic expansion. Zebrafish macrophages express vascular growth factors vegfaa, vegfc and vegfd and chemical ablation of these cells inhibits intestinal lymphatic expansion, suggesting that the recruitment of macrophages to the intestine upon colitogenic challenge is required for intestinal inflammatory lymphangiogenesis. Importantly, this study highlights the potential of zebrafish as an inflammatory lymphangiogenesis model that can be used to investigate the role and mechanism of lymphangiogenesis in inflammatory diseases such as IBD.

Toward automated selective retina treatment (SRT): an optical microbubble detection technique

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Seifert, Eric; Park, Young-Gun; Theisen-Kunde, Dirk; Roh, Young-Jung; Brinkmann, Ralf

2018-02-01

Selective retina therapy (SRT) is an ophthalmological laser technique, targeting the retinal pigment epithelium (RPE) with repetitive microsecond laser pulses, while causing no thermal damage to the neural retina, the photoreceptors as well as the choroid. The RPE cells get damaged mechanically by microbubbles originating, at the intracellular melanosomes. Beneficial effects of SRT on Central Serous Retinopathy (CSR) and Diabetic Macula Edema (DME) have already been shown. Variations in the transmission of the anterior eye media and pigmentation variation of RPE yield in intra- and inter- individual thresholds of the pulse energy required for selective RPE damage. Those selective RPE lesions are not visible. Thus, dosimetry-systems, designed to detect microbubbles as an indicator for RPE cell damage, are demanded elements to facilitate SRT application. Therefore, a technique based on the evaluation of backscattered treatment light has been developed. Data of 127 spots, acquired during 10 clinical treatments of CSR patients, were assigned to a RPE cell damage class, validated by fluorescence angiography (FLA). An algorithm has been designed to match the FLA based information. A sensitivity of 0.9 with a specificity close to 1 is achieved. The data can be processed within microseconds. Thus, the process can be implemented in existing SRT lasers with an automatic pulse wise increasing energy and an automatic irradiation ceasing ability to enable automated treatment close above threshold to prevent adverse effects caused by too high pulse energy. Alternatively, a guidance procedure, informing the treating clinician about the adequacy of the actual settings, is possible.

Developmental and Architectural Principles of the Lateral-line Neural Map

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Hernan eLopez-Schier

2013-03-01

Full Text Available The transmission and central representation of sensory cues through the accurate construction of neural maps is essential for animals to react to environmental stimuli. Structural diversity of sensorineural maps along a continuum between discrete- and continuous-map architectures can influence behavior. The mechanosensory lateral line of fishes and amphibians, for example, detects complex hydrodynamics occurring around the animal body. It. It triggers innate fast escape reactions but also modulates complex navigation behaviors that require constant knowledge about the environment. The aim of this article is to summarize recent work in the zebrafish that has shed light on the development and structure of the lateralis neural map, which is helping to understand how individual sensory modalities generate appropriate behavioral responses to the sensory context.

Real-time simulation of the retina allowing visualization of each processing stage

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Teeters, Jeffrey L.; Werblin, Frank S.

1991-08-01

The retina computes to let us see, but can we see the retina compute? Until now, the answer has been no, because the unconscious nature of the processing hides it from our view. Here the authors describe a method of seeing computations performed throughout the retina. This is achieved by using neurophysiological data to construct a model of the retina, and using a special-purpose image processing computer (PIPE) to implement the model in real time. Processing in the model is organized into stages corresponding to computations performed by each retinal cell type. The final stage is the transient (change detecting) ganglion cell. A CCD camera forms the input image, and the activity of a selected retinal cell type is the output which is displayed on a TV monitor. By changing the retina cell driving the monitor, the progressive transformations of the image by the retina can be observed. These simulations demonstrate the ubiquitous presence of temporal and spatial variations in the patterns of activity generated by the retina which are fed into the brain. The dynamical aspects make these patterns very different from those generated by the common DOG (Difference of Gaussian) model of receptive field. Because the retina is so successful in biological vision systems, the processing described here may be useful in machine vision.

Silver nanoparticles induce endoplasmatic reticulum stress response in zebrafish

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Christen, Verena [University of Applied Sciences and Arts Northwestern Switzerland, School of Life Sciences, Gründenstrasse 40, CH-4132 Muttenz (Switzerland); Capelle, Martinus [Crucell, P.O. Box 2048, NL-2301 Leiden (Netherlands); Fent, Karl, E-mail: karl.fent@fhnw.ch [University of Applied Sciences and Arts Northwestern Switzerland, School of Life Sciences, Gründenstrasse 40, CH-4132 Muttenz (Switzerland); Swiss Federal Institute of Technology Zürich, Department of Environmental Systems Science, CH-8092 Zürich (Switzerland)

2013-10-15

Silver nanoparticles (AgNPs) find increasing applications, and therefore humans and the environment are increasingly exposed to them. However, potential toxicological implications are not sufficiently known. Here we investigate effects of AgNPs (average size 120 nm) on zebrafish in vitro and in vivo, and compare them to human hepatoma cells (Huh7). AgNPs are incorporated in zebrafish liver cells (ZFL) and Huh7, and in zebrafish embryos. In ZFL cells AgNPs lead to induction of reactive oxygen species (ROS), endoplasmatic reticulum (ER) stress response, and TNF-α. Transcriptional alterations also occur in pro-apoptotic genes p53 and Bax. The transcriptional profile differed in ZFL and Huh7 cells. In ZFL cells, the ER stress marker BiP is induced, concomitant with the ER stress marker ATF-6 and spliced XBP-1 after 6 h and 24 h exposure to 0.5 g/L and 0.05 g/L AgNPs, respectively. This indicates the induction of different pathways of the ER stress response. Moreover, AgNPs induce TNF-α. In zebrafish embryos exposed to 0.01, 0.1, 1 and 5 mg/L AgNPs hatching was affected and morphological defects occurred at high concentrations. ER stress related gene transcripts BiP and Synv are significantly up-regulated after 24 h at 0.1 and 5 mg/L AgNPs. Furthermore, transcriptional alterations occurred in the pro-apoptotic genes Noxa and p21. The ER stress response was strong in ZFL cells and occurred in zebrafish embryos as well. Our data demonstrate for the first time that AgNPs lead to induction of ER stress in zebrafish. The induction of ER stress can have several consequences including the activation of apoptotic and inflammatory pathways. - Highlights: • Effects of silver nanoparticles (120 nm AgNPs) are investigated in zebrafish. • AgNPs induce all ER stress reponses in vitro in zebrafish liver cells. • AgNPs induce weak ER stress in zebrafish embryos. • AgNPs induce oxidative stress and transcripts of pro-apoptosis genes.

Silver nanoparticles induce endoplasmatic reticulum stress response in zebrafish

International Nuclear Information System (INIS)

Christen, Verena; Capelle, Martinus; Fent, Karl

2013-01-01

Silver nanoparticles (AgNPs) find increasing applications, and therefore humans and the environment are increasingly exposed to them. However, potential toxicological implications are not sufficiently known. Here we investigate effects of AgNPs (average size 120 nm) on zebrafish in vitro and in vivo, and compare them to human hepatoma cells (Huh7). AgNPs are incorporated in zebrafish liver cells (ZFL) and Huh7, and in zebrafish embryos. In ZFL cells AgNPs lead to induction of reactive oxygen species (ROS), endoplasmatic reticulum (ER) stress response, and TNF-α. Transcriptional alterations also occur in pro-apoptotic genes p53 and Bax. The transcriptional profile differed in ZFL and Huh7 cells. In ZFL cells, the ER stress marker BiP is induced, concomitant with the ER stress marker ATF-6 and spliced XBP-1 after 6 h and 24 h exposure to 0.5 g/L and 0.05 g/L AgNPs, respectively. This indicates the induction of different pathways of the ER stress response. Moreover, AgNPs induce TNF-α. In zebrafish embryos exposed to 0.01, 0.1, 1 and 5 mg/L AgNPs hatching was affected and morphological defects occurred at high concentrations. ER stress related gene transcripts BiP and Synv are significantly up-regulated after 24 h at 0.1 and 5 mg/L AgNPs. Furthermore, transcriptional alterations occurred in the pro-apoptotic genes Noxa and p21. The ER stress response was strong in ZFL cells and occurred in zebrafish embryos as well. Our data demonstrate for the first time that AgNPs lead to induction of ER stress in zebrafish. The induction of ER stress can have several consequences including the activation of apoptotic and inflammatory pathways. - Highlights: • Effects of silver nanoparticles (120 nm AgNPs) are investigated in zebrafish. • AgNPs induce all ER stress reponses in vitro in zebrafish liver cells. • AgNPs induce weak ER stress in zebrafish embryos. • AgNPs induce oxidative stress and transcripts of pro-apoptosis genes

Functional Architecture of the Retina: Development and Disease

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Hoon, Mrinalini; Okawa, Haruhisa; Santina, Luca Della; Wong, Rachel O.L.

2014-01-01

Structure and function are highly correlated in the vertebrate retina, a sensory tissue that is organized into cell layers with microcircuits working in parallel and together to encode visual information. All vertebrate retinas share a fundamental plan, comprising five major neuronal cell classes with cell body distributions and connectivity arranged in stereotypic patterns. Conserved features in retinal design have enabled detailed analysis and comparisons of structure, connectivity and function across species. Each species, however, can adopt structural and/or functional retinal specializations, implementing variations to the basic design in order to satisfy unique requirements in visual function. Recent advances in molecular tools, imaging and electrophysiological approaches have greatly facilitated identification of the cellular and molecular mechanisms that establish the fundamental organization of the retina and the specializations of its microcircuits during development. Here, we review advances in our understanding of how these mechanisms act to shape structure and function at the single cell level, to coordinate the assembly of cell populations, and to define their specific circuitry. We also highlight how structure is rearranged and function is disrupted in disease, and discuss current approaches to re-establish the intricate functional architecture of the retina. PMID:24984227

Culturable gut microbiota diversity in zebrafish.

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Cantas, Leon; Sørby, Jan Roger Torp; Aleström, Peter; Sørum, Henning

2012-03-01

The zebrafish (Danio rerio) is an increasingly used laboratory animal model in basic biology and biomedicine, novel drug development, and toxicology. The wide use has increased the demand for optimized husbandry protocols to ensure animal health care and welfare. The knowledge about the correlation between culturable zebrafish intestinal microbiota and health in relation to environmental factors and management procedures is very limited. A semi-quantitative level of growth of individual types of bacteria was determined and associated with sampling points. A total of 72 TAB line zebrafish from four laboratories (Labs A-D) in the Zebrafish Network Norway were used. Diagnostic was based on traditional bacterial culture methods and biochemical characterization using commercial kits, followed by 16S rDNA gene sequencing from pure subcultures. Also selected Gram-negative isolates were analyzed for antibiotic susceptibility to 8 different antibiotics. A total of 13 morphologically different bacterial species were the most prevalent: Aeromonas hydrophila, Aeromonas sobria, Vibrio parahaemolyticus, Photobacterium damselae, Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas luteola, Comamonas testosteroni, Ochrobactrum anthropi, Staphylococcus cohnii, Staphylococcus epidermidis, Staphylococcus capitis, and Staphylococcus warneri. Only Lab B had significantly higher levels of total bacterial growth (OR=2.03), whereas numbers from Lab C (OR=1.01) and Lab D (OR=1.12) were found to be similar to the baseline Lab A. Sexually immature individuals had a significantly higher level of harvested total bacterial growth than mature fish (OR=0.82), no statistically significant differences were found between male and female fish (OR=1.01), and the posterior intestinal segment demonstrated a higher degree of culturable bacteria than the anterior segment (OR=4.1). Multiple antibiotic (>3) resistance was observed in 17% of the strains. We propose that a rapid conventional

Toxicity assessment of zebrafish following exposure to CdTe QDs

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Zhang, Wei, E-mail: wzhang@ecust.edu.cn [State Environmental Protection Key Laboratory of Environmental Risk Assessment and Control on Chemical Process, East China University of Science and Technology, Shanghai 200237 (China); Shanghai Key Laboratory of Functional Materials Chemistry, East China University of Science and Technology, Shanghai 200237 (China); School of Resource and Environmental Engineering, East China University of Science and Technology, Shanghai 200237 (China); Lin, Kuangfei, E-mail: kflin@ecust.edu.cn [State Environmental Protection Key Laboratory of Environmental Risk Assessment and Control on Chemical Process, East China University of Science and Technology, Shanghai 200237 (China); Shanghai Key Laboratory of Functional Materials Chemistry, East China University of Science and Technology, Shanghai 200237 (China); School of Resource and Environmental Engineering, East China University of Science and Technology, Shanghai 200237 (China); Miao, Youna [State Environmental Protection Key Laboratory of Environmental Risk Assessment and Control on Chemical Process, East China University of Science and Technology, Shanghai 200237 (China); Shanghai Key Laboratory of Functional Materials Chemistry, East China University of Science and Technology, Shanghai 200237 (China); School of Resource and Environmental Engineering, East China University of Science and Technology, Shanghai 200237 (China); Dong, Qiaoxiang; Huang, Changjiang; Wang, Huili [Zhejiang Provincial Key Lab for Technology and Application of Model Organisms, Wenzhou Medical College, Wenzhou 325035 (China); Guo, Meijin [State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237 (China); Cui, Xinhong [Shanghai Institute of Landscape Gardening, Shanghai 200233 (China)

2012-04-30

Highlights: Black-Right-Pointing-Pointer The LC{sub 50} of TGA-CdTe for zebrafish at 120 hpf was 185.9 nM. Black-Right-Pointing-Pointer Zebrafish exposed to TGA-CdTe resulted in lower hatch rate and more malformation. Black-Right-Pointing-Pointer Body length and heart beat of zebrafish declined after exposure to TGA-CdTe. Black-Right-Pointing-Pointer Larvae exposure to TGA-CdTe elicited a higher basal swimming rate. Black-Right-Pointing-Pointer Abnormal vascular of FLI-1 transgenic zebrafish larvae exposed to TGA-CdTe occurred. - Abstract: CdTe quantum dots (QDs) are nanocrystals of unique composition and properties that have found many new commercial applications; therefore, their potential toxicity to aquatic organisms has become a hot research topic. The lab study was performed to determine the developmental and behavioral toxicities to zebrafish under continuous exposure to low concentrations of CdTe QDs (1-400 nM) coated with thioglycolic acid (TGA). The results show: (1) the 120 h LC{sub 50} of 185.9 nM, (2) the lower hatch rate and body length, more malformations, and less heart beat and swimming speed of the exposed zebrafish, (3) the brief burst and a higher basal swimming rate of the exposed zebrafish larvae during a rapid transition from light-to-dark, and (4) the vascular hyperplasia, vascular bifurcation, vascular crossing and turbulence of the exposed FLI-1 transgenic zebrafish larvae.

Modeling laser damage to the retina

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Clark, Clifton D.

This dissertation presents recent progress in several areas related to modeling laser damage to the retina. In Chapter 3, we consider the consequences of using the Arrhenius damage model to predict the damage thresholds of multiple pulse, or repetitive pulse, exposures. We have identified a few fundamental trends associated with the multiple pulse damage predictions made by the Arrhenius model. These trends differ from what would be expected by non-thermal mechanisms, and could prove useful in differentiating thermal and non-thermal damage. Chapter 4 presents a new rate equation damage model hypothesized to describe photochemical damage. The model adds a temperature dependent term to the simple rate equation implied by the principle of reciprocity that is characteristic of photochemical damage thresholds. A recent damage threshold study, conducted in-vitro, has revealed a very sharp transition between thermal and photochemical damage threshold trends. For the wavelength used in the experiment (413 nm), thermal damage thresholds were observed at exposure levels that were twice the expected photochemical damage threshold, based on the traditional understanding of photochemical damage. Our model accounts for this observed trend by introducing a temperature dependent quenching, or repair, rate to the photochemical damage rate. For long exposures that give a very small temperature rise, the model reduces to the principle of reciprocity. Near the transition region between thermal and photochemical damage, the model allows the damage threshold to be set by thermal mechanisms, even at exposure above the reciprocity exposure. In Chapter 5, we describe a retina damage model that includes thermal lensing in the eye by coupling beam propagation and heat transfer models together. Thermal lensing has recently been suggested as a contributing factor to the large increase in measured retinal damage thresholds in the near infrared. The transmission of the vitreous decreases

Illumination-invariant face recognition with a contrast sensitive silicon retina

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Buhmann, J.M. [Rheinische Friedrich-Wilhelms-Univ., Bonn (Germany). Inst. fuer Informatik II; Lades, M. [Bochum Univ. (Germany). Inst. fuer Neuroinformatik; Eeckman, F. [Lawrence Livermore National Lab., CA (United States)

1993-11-29

Changes in lighting conditions strongly effect the performance and reliability of computer vision systems. We report face recognition results under drastically changing lighting conditions for a computer vision system which concurrently uses a contrast sensitive silicon retina and a conventional, gain controlled CCD camera. For both input devices the face recognition system employs an elastic matching algorithm with wavelet based features to classify unknown faces. To assess the effect of analog on-chip preprocessing by the silicon retina the CCD images have been digitally preprocessed with a bandpass filter to adjust the power spectrum. The silicon retina with its ability to adjust sensitivity increases the recognition rate up to 50 percent. These comparative experiments demonstrate that preprocessing with an analog VLSI silicon retina generates image data enriched with object-constant features.

Zebrafish syntenic relationship to human/mouse genomes revealed by radiation hybrid mapping

International Nuclear Information System (INIS)

Samonte, Irene E.

2007-01-01

Zebrafish (Danio rerio) is an excellent model system for vertebrate developmental analysis and a new model for human disorders. In this study, however, zebrafish was used to determine its syntenic relationship to human/mouse genomes using the zebrafish-hamster radiation hybrid panel. The focus was on genes residing on chromosomes 6 and 17 of human and mouse, respectively, and some other genes of either immunologic or evolutionary importance. Gene sequences of interest and zebrafish expressed sequence tags deposited in the GenBank were used in identifying zebrafish homologs. Polymerase chain reaction (PCR) amplification, cloning and subcloning, sequencing, and phylogenetic analysis were done to confirm the homology of the candidate genes in zebrafish. The promising markers were then tested in the 94 zebrafish-hamster radiation hybrid panel cell lines and submitted for logarithm of the odds (LOD) score analysis to position genes on the zebrafish map. A total of 19 loci were successfully mapped to zebrafish linkage groups 1, 14, 15, 19, and 20. Four of these loci were positioned in linkage group 20, whereas, 3 more loci were added in linkage group 19, thus increasing to 34 loci the number of human genes syntenic to the group. With the sequencing of the zebrafish genome, about 20 more MHC genes were reported linked on the same group. (Author)

Maturação funcional da retina em bebês prematuros

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Adriana Berezovsky

2007-06-01

Full Text Available A retina humana ainda não está totalmente desenvolvida no nascimento. Só após o nascimento é que ocorrem mudanças anatômicas como o aumento na densidade de cones centrais e o alongamento do segmento externo dos fotorreceptores. As mudanças funcionais que ocorrem na retina com a maturação no primeiro ano de vida podem ser avaliadas pela técnica do eletrorretinograma de campo total, que representa a atividade somada da retina distal em resposta à luz. Abordaremos aspectos da maturação funcional da retina avaliada pelo eletrorretinograma em bebês prematuros.

Myomaker mediates fusion of fast myocytes in zebrafish embryos

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Landemaine, Aurélie; Rescan, Pierre-Yves; Gabillard, Jean-Charles, E-mail: Jean-charles.gabillard@rennes.inra.fr

2014-09-05

Highlights: • Myomaker is transiently expressed in fast myocytes during embryonic myogenesis. • Myomaker is essential for fast myocyte fusion in zebrafish. • The function of myomaker is conserved among Teleostomi. - Abstract: Myomaker (also called Tmem8c), a new membrane activator of myocyte fusion was recently discovered in mice. Using whole mount in situ hybridization on zebrafish embryos at different stages of embryonic development, we show that myomaker is transiently expressed in fast myocytes forming the bulk of zebrafish myotome. Zebrafish embryos injected with morpholino targeted against myomaker were alive after yolk resorption and appeared morphologically normal, but they were unable to swim, even under effect of a tactile stimulation. Confocal observations showed a marked phenotype characterized by the persistence of mononucleated muscle cells in the fast myotome at developmental stages where these cells normally fuse to form multinucleated myotubes. This indicates that myomaker is essential for myocyte fusion in zebrafish. Thus, there is an evolutionary conservation of myomaker expression and function among Teleostomi.

Connecting the Retina to the Brain

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Lynda Erskine

2014-12-01

Full Text Available The visual system is beautifully crafted to transmit information of the external world to visual processing and cognitive centers in the brain. For visual information to be relayed to the brain, a series of axon pathfinding events must take place to ensure that the axons of retinal ganglion cells, the only neuronal cell type in the retina that sends axons out of the retina, find their way out of the eye to connect with targets in the brain. In the past few decades, the power of molecular and genetic tools, including the generation of genetically manipulated mouse lines, have multiplied our knowledge about the molecular mechanisms involved in the sculpting of the visual system. Here, we review major advances in our understanding of the mechanisms controlling the differentiation of RGCs, guidance of their axons from the retina to the primary visual centers, and the refinement processes essential for the establishment of topographic maps and eye-specific axon segregation. Human disorders, such as albinism and achiasmia, that impair RGC axon growth and guidance and, thus, the establishment of a fully functioning visual system will also be discussed.

Transcriptome analysis of zebrafish embryogenesis using microarrays.

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Sinnakaruppan Mathavan

2005-08-01

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

The Zebrafish Anillin-eGFP Reporter Marks Late Dividing Retinal Precursors and Stem Cells Entering Neuronal Lineages.

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Meret Cepero Malo

Full Text Available Monitoring cycling behaviours of stem and somatic cells in the living animal is a powerful tool to better understand tissue development and homeostasis. The tg(anillin:anillin-eGFP transgenic line carries the full-length zebrafish F-actin binding protein Anillin fused to eGFP from a bacterial artificial chromosome (BAC containing Anillin cis-regulatory sequences. Here we report the suitability of the Anillin-eGFP reporter as a direct indicator of cycling cells in the late embryonic and post-embryonic retina. We show that combining the anillin:anillin-eGFP with other transgenes such as ptf1a:dsRed and atoh7:gap-RFP allows obtaining spatial and temporal resolution of the mitotic potentials of specific retinal cell populations. This is exemplified by the analysis of the origin of the previously reported apically and non-apically dividing late committed precursors of the photoreceptor and horizontal cell layers.

Stimulus-dependent maximum entropy models of neural population codes.

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Einat Granot-Atedgi

Full Text Available Neural populations encode information about their stimulus in a collective fashion, by joint activity patterns of spiking and silence. A full account of this mapping from stimulus to neural activity is given by the conditional probability distribution over neural codewords given the sensory input. For large populations, direct sampling of these distributions is impossible, and so we must rely on constructing appropriate models. We show here that in a population of 100 retinal ganglion cells in the salamander retina responding to temporal white-noise stimuli, dependencies between cells play an important encoding role. We introduce the stimulus-dependent maximum entropy (SDME model-a minimal extension of the canonical linear-nonlinear model of a single neuron, to a pairwise-coupled neural population. We find that the SDME model gives a more accurate account of single cell responses and in particular significantly outperforms uncoupled models in reproducing the distributions of population codewords emitted in response to a stimulus. We show how the SDME model, in conjunction with static maximum entropy models of population vocabulary, can be used to estimate information-theoretic quantities like average surprise and information transmission in a neural population.

The zebrafish world of colors and shapes: preference and discrimination.

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Oliveira, Jessica; Silveira, Mayara; Chacon, Diana; Luchiari, Ana

2015-04-01

Natural environment imposes many challenges to animals, which have to use cognitive abilities to cope with and exploit it to enhance their fitness. Since zebrafish is a well-established model for cognitive studies and high-throughput screening for drugs and diseases that affect cognition, we tested their ability for ambient color preference and 3D objects discrimination to establish a protocol for memory evaluation. For the color preference test, zebrafish were observed in a multiple-chamber tank with different environmental color options. Zebrafish showed preference for blue and green, and avoided yellow and red. For the 3D objects discrimination, zebrafish were allowed to explore two equal objects and then observed in a one-trial test in which a new color, size, or shape of the object was presented. Zebrafish showed discrimination for color, shape, and color+shape combined, but not size. These results imply that zebrafish seem to use some categorical system to discriminate items, and distracters affect their ability for discrimination. The type of variables available (color and shape) may favor zebrafish objects perception and facilitate discrimination processing. We suggest that this easy and simple memory test could serve as a useful screening tool for cognitive dysfunction and neurotoxicological studies.

Microcystin-LR exposure induces developmental neurotoxicity in zebrafish embryo

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Wu, Qin; Yan, Wei; Liu, Chunsheng; Li, Li; Yu, Liqin; Zhao, Sujuan; Li, Guangyu

2016-01-01

Microcystin-LR (MCLR) is a commonly acting potent hepatotoxin and has been pointed out of potentially causing developmental neurotoxicity, but the exact mechanism is little known. In this study, zebrafish embryos were exposed to 0, 0.8, 1.6 or 3.2 mg/L MCLR for 120 h. MCLR exposure through submersion caused serious hatching delay and body length decrease. The content of MCLR in zebrafish larvae was analyzed and the results demonstrated that MCLR can accumulate in zebrafish larvae. The locomotor speed of zebrafish larvae was decreased. Furthermore, the dopamine and acetylcholine (ACh) content were detected to be significantly decreased in MCLR exposure groups. And the acetylcholinesterase (AChE) activity was significantly increased after exposure to 1.6 and 3.2 mg/L MCLR. The transcription pattern of manf, chrnα7 and ache gene was consistent with the change of the dopamine content, ACh content and AChE activity. Gene expression involved in the development of neurons was also measured. α1-tubulin and shha gene expression were down-regulated, whereas mbp and gap43 gene expression were observed to be significantly up-regulated upon exposure to MCLR. The above results indicated that MCLR-induced developmental toxicity might attribute to the disorder of cholinergic system, dopaminergic signaling, and the development of neurons. - Highlights: • MCLR accumulation induces developmental neurotoxicity in zebrafish embryo. • The decrease of dopamine levels might be associated with the MCLR-induced developmental neurotoxicity in zebrafish larvae. • The alternation of cholinergic system might contribute to the change of neurobehavior in zebrafish larvae exposure with MCLR. - MCLR accumulation induces developmental neurotoxicity by affecting cholinergic system, dopaminergic signaling, and the development of neurons in zebrafish embryo.

Increased radial glia quiescence, decreased reactivation upon injury and unaltered neuroblast behavior underlie decreased neurogenesis in the aging zebrafish telencephalon.

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Edelmann, Kathrin; Glashauser, Lena; Sprungala, Susanne; Hesl, Birgit; Fritschle, Maike; Ninkovic, Jovica; Godinho, Leanne; Chapouton, Prisca

2013-09-01

The zebrafish has recently become a source of new data on the mechanisms of neural stem cell (NSC) maintenance and ongoing neurogenesis in adult brains. In this vertebrate, neurogenesis occurs at high levels in all ventricular regions of the brain, and brain injuries recover successfully, owing to the recruitment of radial glia, which function as NSCs. This new vertebrate model of adult neurogenesis is thus advancing our knowledge of the molecular cues in use for the activation of NSCs and fate of their progeny. Because the regenerative potential of somatic stem cells generally weakens with increasing age, it is important to assess the extent to which zebrafish NSC potential decreases or remains unaltered with age. We found that neurogenesis in the ventricular zone, in the olfactory bulb, and in a newly identified parenchymal zone of the telencephalon indeed declines as the fish ages and that oligodendrogenesis also declines. In the ventricular zone, the radial glial cell population remains largely unaltered morphologically but enters less frequently into the cell cycle and hence produces fewer neuroblasts. The neuroblasts themselves do not change their behavior with age and produce the same number of postmitotic neurons. Thus, decreased neurogenesis in the physiologically aging zebrafish brain is correlated with an increasing quiescence of radial glia. After injuries, radial glia in aged brains are reactivated, and the percentage of cell cycle entry is increased in the radial glia population. However, this reaction is far less pronounced than in younger animals, pointing to irreversible changes in aging zebrafish radial glia. Copyright © 2013 Wiley Periodicals, Inc.

Coincidence detection of single-photon responses in the inner retina at the sensitivity limit of vision.

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Ala-Laurila, Petri; Rieke, Fred

2014-12-15

Vision in starlight relies on our ability to detect single absorbed photons. Indeed, the sensitivity of dark-adapted vision approaches limits set by the quantal nature of light. This sensitivity requires neural mechanisms that selectively transmit quantal responses and suppress noise. Such mechanisms face an inevitable tradeoff because signal and noise cannot be perfectly separated, and rejecting noise also means rejecting signal. We report measurements of single-photon responses in the output signals of the primate retina. We find that visual signals arising from a few absorbed photons are read out fundamentally differently by primate On and Off parasol ganglion cells, key retinal output neurons. Off parasol cells respond linearly to near-threshold flashes, retaining sensitivity to each absorbed photon but maintaining a high level of noise. On parasol cells respond nonlinearly due to thresholding of their excitatory synaptic inputs. This nonlinearity reduces neural noise but also limits information about single-photon absorptions. The long-standing idea that information about each photon absorption is available for behavior at the sensitivity limit of vision is not universally true across retinal outputs. More generally, our work shows how a neural circuit balances the competing needs for sensitivity and noise rejection. Copyright © 2014 Elsevier Ltd. All rights reserved.

Zebrafish Axenic Larvae Colonization with Human Intestinal Microbiota.

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Arias-Jayo, Nerea; Alonso-Saez, Laura; Ramirez-Garcia, Andoni; Pardo, Miguel A

2018-04-01

The human intestine hosts a vast and complex microbial community that is vital for maintaining several functions related with host health. The processes that determine the gut microbiome composition are poorly understood, being the interaction between species, the external environment, and the relationship with the host the most feasible. Animal models offer the opportunity to understand the interactions between the host and the microbiota. There are different gnotobiotic mice or rat models colonized with the human microbiota, however, to our knowledge, there are no reports on the colonization of germ-free zebrafish with a complex human intestinal microbiota. In the present study, we have successfully colonized 5 days postfertilization germ-free zebrafish larvae with the human intestinal microbiota previously extracted from a donor and analyzed by high-throughput sequencing the composition of the transferred microbial communities that established inside the zebrafish gut. Thus, we describe for first time which human bacteria phylotypes are able to colonize the zebrafish digestive tract. Species with relevant interest because of their linkage to dysbiosis in different human diseases, such as Akkermansia muciniphila, Eubacterium rectale, Faecalibacterium prausnitzii, Prevotella spp., or Roseburia spp. have been successfully transferred inside the zebrafish digestive tract.

Effects of 2,2',4,4'-tetrabromodiphenyl ether on neurobehavior and memory change and bcl-2, c-fos, grin1b and lingo1b gene expression in male zebrafish (Danio rerio).

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Zheng, Shukai; Liu, Caixia; Huang, Yanhong; Bao, Mian; Huang, Yuanni; Wu, Kusheng

2017-10-15

Polybrominated diphenyl ethers (PBDEs) are persistent organic pollutants in various environmental matrices and organisms and pose a threat to neural systems of organisms. However, though quite a few studies have explored the effect of PBDEs on neural behaviors such as learning and memory abilities in animals, their mechanisms are less known. We used the zebrafish model to evaluate neurotoxicity of PBDEs and observe changes in behavior and related gene expression. In behavioral testing, 50 zebrafish were divided into five groups treated with different concentrations of BDE-47. T-maze exploration was used for learning and memory testing, which was recorded by camera every 7days. After 21days, all fish were killed, and the gene expression of c-fos, bcl-2, lingo1b and grin1b in brain tissue was analyzed by RT-qPCR. The behavioral changes (latency to leave the start zone, reach the reward zone, and stay in the reward zone; accuracy in choosing the right maze arm, accumulation of freezing bouts, etc.) were related to BDE-47 concentration and had a time-effect relation with increasing exposure days, especially with 500μg/L BDE-47. BDE-47 elevated brain bcl-2, grin1b and lingo1b expression. The expression of c-fos showed an increase with 50 and 100μg/L BDE-47 exposure. The PBDE BDE-47 had a negative impact on the neurobehaviors of zebrafish and affected the expression of c-fos, bcl-2, lingo1b and grin1b in zebrafish brain tissue. Copyright © 2017 Elsevier Inc. All rights reserved.

Effect of Monocular Deprivation on Rabbit Neural Retinal Cell Densities

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Mwachaka, Philip Maseghe; Saidi, Hassan; Odula, Paul Ochieng; Mandela, Pamela Idenya

2015-01-01

Purpose: To describe the effect of monocular deprivation on densities of neural retinal cells in rabbits. Methods: Thirty rabbits, comprised of 18 subject and 12 control animals, were included and monocular deprivation was achieved through unilateral lid suturing in all subject animals. The rabbits were observed for three weeks. At the end of each week, 6 experimental and 3 control animals were euthanized, their retinas was harvested and processed for light microscopy. Photomicrographs of ...

Zebrafish Database: Customizable, Free, and Open-Source Solution for Facility Management.

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Yakulov, Toma Antonov; Walz, Gerd

2015-12-01

Zebrafish Database is a web-based customizable database solution, which can be easily adapted to serve both single laboratories and facilities housing thousands of zebrafish lines. The database allows the users to keep track of details regarding the various genomic features, zebrafish lines, zebrafish batches, and their respective locations. Advanced search and reporting options are available. Unique features are the ability to upload files and images that are associated with the respective records and an integrated calendar component that supports multiple calendars and categories. Built on the basis of the Joomla content management system, the Zebrafish Database is easily extendable without the need for advanced programming skills.

Xenotransplantation of human neural progenitor cells to the subretinal space of nonimmunosuppressed pigs

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Warfvinge, Karin; Schwartz, Philip H; Kiilgaard, Jens Folke

2011-01-01

To investigate the feasibility of transplanting human neural progenitor cells (hNPCs) to the retina of nonimmunosuppressed pigs, cultured hNPCs were injected into the subretinal space of 5 adult pigs after laser burns were applied to promote donor cell integration. Postoperatively, the retinal ve...... that modulation of host immunity is likely necessary for prolonged xenograft survival in this model....

Method for somatic cell nuclear transfer in zebrafish.

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Siripattarapravat, Kannika; Cibelli, Jose B

2011-01-01

Somatic cell nuclear transfer (SCNT) has been a well-known technique for decades and widely applied to generate identical animals, including ones with genetic alterations. The system has been demonstrated successfully in zebrafish. The elaborated requirements of SCNT, however, limit reproducibility of the established model to a few groups in zebrafish research community. In this chapter, we meticulously outline each step of the published protocol as well as preparations of equipments and reagents used in zebrafish SCNT. All describable detailed-tips are elaborated in texts and figures. Copyright © 2011 Elsevier Inc. All rights reserved.

Concentration Sensing by the Moving Nucleus in Cell Fate Determination: A Computational Analysis.

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Varun Aggarwal

Full Text Available During development of the vertebrate neuroepithelium, the nucleus in neural progenitor cells (NPCs moves from the apex toward the base and returns to the apex (called interkinetic nuclear migration at which point the cell divides. The fate of the resulting daughter cells is thought to depend on the sampling by the moving nucleus of a spatial concentration profile of the cytoplasmic Notch intracellular domain (NICD. However, the nucleus executes complex stochastic motions including random waiting and back and forth motions, which can expose the nucleus to randomly varying levels of cytoplasmic NICD. How nuclear position can determine daughter cell fate despite the stochastic nature of nuclear migration is not clear. Here we derived a mathematical model for reaction, diffusion, and nuclear accumulation of NICD in NPCs during interkinetic nuclear migration (INM. Using experimentally measured trajectory-dependent probabilities of nuclear turning, nuclear waiting times and average nuclear speeds in NPCs in the developing zebrafish retina, we performed stochastic simulations to compute the nuclear trajectory-dependent probabilities of NPC differentiation. Comparison with experimentally measured nuclear NICD concentrations and trajectory-dependent probabilities of differentiation allowed estimation of the NICD cytoplasmic gradient. Spatially polarized production of NICD, rapid NICD cytoplasmic consumption and the time-averaging effect of nuclear import/export kinetics are sufficient to explain the experimentally observed differentiation probabilities. Our computational studies lend quantitative support to the feasibility of the nuclear concentration-sensing mechanism for NPC fate determination in zebrafish retina.

Zebrafish Models of Human Leukemia: Technological Advances and Mechanistic Insights.

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Harrison, Nicholas R; Laroche, Fabrice J F; Gutierrez, Alejandro; Feng, Hui

2016-01-01

Insights concerning leukemic pathophysiology have been acquired in various animal models and further efforts to understand the mechanisms underlying leukemic treatment resistance and disease relapse promise to improve therapeutic strategies. The zebrafish (Danio rerio) is a vertebrate organism with a conserved hematopoietic program and unique experimental strengths suiting it for the investigation of human leukemia. Recent technological advances in zebrafish research including efficient transgenesis, precise genome editing, and straightforward transplantation techniques have led to the generation of a number of leukemia models. The transparency of the zebrafish when coupled with improved lineage-tracing and imaging techniques has revealed exquisite details of leukemic initiation, progression, and regression. With these advantages, the zebrafish represents a unique experimental system for leukemic research and additionally, advances in zebrafish-based high-throughput drug screening promise to hasten the discovery of novel leukemia therapeutics. To date, investigators have accumulated knowledge of the genetic underpinnings critical to leukemic transformation and treatment resistance and without doubt, zebrafish are rapidly expanding our understanding of disease mechanisms and helping to shape therapeutic strategies for improved outcomes in leukemic patients.

Benign familial fleck retina: multimodal imaging including optical coherence tomography angiography.

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Garcia, Jose Mauricio Botto de Barros; Isaac, David Leonardo Cruvinel; Sardeiro, Tainara; Aquino, Érika; Avila, Marcos

2017-01-01

This report presents multimodal imaging of a 27-year-old woman diagnosed with benign familial fleck retina (OMIM 228980), an uncommon disorder. Fundus photographs revealed retinal flecks that affected her post-equatorial retina but spared the macular area. Fundus autofluorescence and infrared imaging demonstrated a symmetrical pattern of yellow-white fleck lesions that affected both eyes. Her full-field electroretinogram and electrooculogram were normal. An optical coherence tomography B-scan was performed for both eyes, revealing increased thickness of the retinal pigmented epithelium leading to multiple small pigmented epithelium detachments. The outer retina remained intact in both eyes. Spectral-domain optical coherence tomography angiography with split-spectrum amplitude decorrelation algorithm and 3 × 3 mm structural en face optical coherence tomography did not show macular lesions. Benign familial fleck retina belongs to a heterogenous group of so-called flecked retina syndromes, and should be considered in patients with yellowish-white retinal lesions without involvement of the macula.

Benign familial fleck retina: multimodal imaging including optical coherence tomography angiography

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Jose Mauricio Botto de Barros Garcia

Full Text Available ABSTRACT This report presents multimodal imaging of a 27-year-old woman diagnosed with benign familial fleck retina (OMIM 228980, an uncommon disorder. Fundus photographs revealed retinal flecks that affected her post-equatorial retina but spared the macular area. Fundus autofluorescence and infrared imaging demonstrated a symmetrical pattern of yellow-white fleck lesions that affected both eyes. Her full-field electroretinogram and electrooculogram were normal. An optical coherence tomography B-scan was performed for both eyes, revealing increased thickness of the retinal pigmented epithelium leading to multiple small pigmented epithelium detachments. The outer retina remained intact in both eyes. Spectral-domain optical coherence tomography angiography with split-spectrum amplitude decorrelation algorithm and 3 × 3 mm structural en face optical coherence tomography did not show macular lesions. Benign familial fleck retina belongs to a heterogenous group of so-called flecked retina syndromes, and should be considered in patients with yellowish-white retinal lesions without involvement of the macula.

Brain-wide neuronal dynamics during motor adaptation in zebrafish.

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

2012-05-09

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

GLUT2-mediated glucose uptake and availability are required for embryonic brain development in zebrafish.

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Marín-Juez, Rubén; Rovira, Mireia; Crespo, Diego; van der Vaart, Michiel; Spaink, Herman P; Planas, Josep V

2015-01-01

Glucose transporter 2 (GLUT2; gene name SLC2A2) has a key role in the regulation of glucose dynamics in organs central to metabolism. Although GLUT2 has been studied in the context of its participation in peripheral and central glucose sensing, its role in the brain is not well understood. To decipher the role of GLUT2 in brain development, we knocked down slc2a2 (glut2), the functional ortholog of human GLUT2, in zebrafish. Abrogation of glut2 led to defective brain organogenesis, reduced glucose uptake and increased programmed cell death in the brain. Coinciding with the observed localization of glut2 expression in the zebrafish hindbrain, glut2 deficiency affected the development of neural progenitor cells expressing the proneural genes atoh1b and ptf1a but not those expressing neurod. Specificity of the morphant phenotype was demonstrated by the restoration of brain organogenesis, whole-embryo glucose uptake, brain apoptosis, and expression of proneural markers in rescue experiments. These results indicate that glut2 has an essential role during brain development by facilitating the uptake and availability of glucose and support the involvement of glut2 in brain glucose sensing.

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

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Moayad Saad

2018-02-01

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

A review of monoaminergic neuropsychopharmacology in zebrafish.

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Maximino, Caio; Herculano, Anderson Manoel

2010-12-01

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

Hypoxia-induced retinopathy model in adult zebrafish

DEFF Research Database (Denmark)

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

2010-01-01

Hypoxia-induced vascular responses, including angiogenesis, vascular remodeling and vascular leakage, significantly contribute to the onset, development and progression of retinopathy. However, until recently there were no appropriate animal disease models recapitulating adult retinopathy available....... 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......-induced retinopathy model in adult zebrafish. This model provides a unique opportunity to study kinetically the development of retinopathy in adult animals using noninvasive protocols and to assess therapeutic efficacy of orally active antiangiogenic drugs....

Genetics of lattice degeneration of the retina.

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Murakami, F; Ohba, N

1982-01-01

First-degree relatives of proband patients with lattice degeneration of the retina revealed a significantly higher prevalence of the disease than the prevalence in the general population: the former had the disease about three times as frequently as the latter. The observed data were analyzed in terms of their accordance with recognized genetic models. The inheritance pattern did not fit well to a monogenic mode of inheritance, and it was hypothesized that a polygenic or multifactorial mode of inheritance is the most likely for lattice degeneration of the retina.

Functional inhibition of UQCRB suppresses angiogenesis in zebrafish

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Cho, Yoon Sun; Jung, Hye Jin [Chemical Genomics National Research Laboratory, Department of Biotechnology, Translational Research Center for Protein Function Control, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of); Seok, Seung Hyeok [Department of Microbiology and Immunology, Institute for Experimental Animals, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Payumo, Alexander Y.; Chen, James K. [Department of Chemical and Systems Biology, Stanford University School of Medicine, Stanford, CA 94305 (United States); Kwon, Ho Jeong, E-mail: kwonhj@yonsei.ac.kr [Chemical Genomics National Research Laboratory, Department of Biotechnology, Translational Research Center for Protein Function Control, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of)

2013-04-19

Highlights: ► This is the first functional characterization of UQCRB in vivo model. ► Angiogenesis is inhibited with UQCRB loss of function in zebrafish. ► UQCRB is introduced as a prognostic marker for mitochondria- and angiogenesis-related diseases. -- Abstract: As a subunit of mitochondrial complex III, UQCRB plays an important role in complex III stability, electron transport, and cellular oxygen sensing. Herein, we report UQCRB function regarding angiogenesis in vivo with the zebrafish (Danio rerio). UQCRB knockdown inhibited angiogenesis in zebrafish leading to the suppression of VEGF expression. Moreover, the UQCRB-targeting small molecule terpestacin also inhibited angiogenesis and VEGF levels in zebrafish, supporting the role of UQCRB in angiogenesis. Collectively, UQCRB loss of function by either genetic and pharmacological means inhibited angiogenesis, indicating that UQCRB plays a key role in this process and can be a prognostic marker of angiogenesis- and mitochondria-related diseases.

Phenotype classification of zebrafish embryos by supervised learning.

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Nathalie Jeanray

Full Text Available Zebrafish is increasingly used to assess biological properties of chemical substances and thus is becoming a specific tool for toxicological and pharmacological studies. The effects of chemical substances on embryo survival and development are generally evaluated manually through microscopic observation by an expert and documented by several typical photographs. Here, we present a methodology to automatically classify brightfield images of wildtype zebrafish embryos according to their defects by using an image analysis approach based on supervised machine learning. We show that, compared to manual classification, automatic classification results in 90 to 100% agreement with consensus voting of biological experts in nine out of eleven considered defects in 3 days old zebrafish larvae. Automation of the analysis and classification of zebrafish embryo pictures reduces the workload and time required for the biological expert and increases the reproducibility and objectivity of this classification.

Functional inhibition of UQCRB suppresses angiogenesis in zebrafish

International Nuclear Information System (INIS)

Cho, Yoon Sun; Jung, Hye Jin; Seok, Seung Hyeok; Payumo, Alexander Y.; Chen, James K.; Kwon, Ho Jeong

2013-01-01

Highlights: ► This is the first functional characterization of UQCRB in vivo model. ► Angiogenesis is inhibited with UQCRB loss of function in zebrafish. ► UQCRB is introduced as a prognostic marker for mitochondria- and angiogenesis-related diseases. -- Abstract: As a subunit of mitochondrial complex III, UQCRB plays an important role in complex III stability, electron transport, and cellular oxygen sensing. Herein, we report UQCRB function regarding angiogenesis in vivo with the zebrafish (Danio rerio). UQCRB knockdown inhibited angiogenesis in zebrafish leading to the suppression of VEGF expression. Moreover, the UQCRB-targeting small molecule terpestacin also inhibited angiogenesis and VEGF levels in zebrafish, supporting the role of UQCRB in angiogenesis. Collectively, UQCRB loss of function by either genetic and pharmacological means inhibited angiogenesis, indicating that UQCRB plays a key role in this process and can be a prognostic marker of angiogenesis- and mitochondria-related diseases

Rip3 knockdown rescues photoreceptor cell death in blind pde6c zebrafish.

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Viringipurampeer, I A; Shan, X; Gregory-Evans, K; Zhang, J P; Mohammadi, Z; Gregory-Evans, C Y

2014-05-01

Achromatopsia is a progressive autosomal recessive retinal disease characterized by early loss of cone photoreceptors and later rod photoreceptor loss. In most cases, mutations have been identified in CNGA3, CNGB3, GNAT2, PDE6C or PDE6H genes. Owing to this genetic heterogeneity, mutation-independent therapeutic schemes aimed at preventing cone cell death are very attractive treatment strategies. In pde6c(w59) mutant zebrafish, cone photoreceptors expressed high levels of receptor-interacting protein kinase 1 (RIP1) and receptor-interacting protein kinase 3 (RIP3) kinases, key regulators of necroptotic cell death. In contrast, rod photoreceptor cells were alternatively immunopositive for caspase-3 indicating activation of caspase-dependent apoptosis in these cells. Morpholino gene knockdown of rip3 in pde6c(w59) embryos rescued the dying cone photoreceptors by inhibiting the formation of reactive oxygen species and by inhibiting second-order neuron remodelling in the inner retina. In rip3 morphant larvae, visual function was restored in the cones by upregulation of the rod phosphodiesterase genes (pde6a and pde6b), compensating for the lack of cone pde6c suggesting that cones are able to adapt to their local environment. Furthermore, we demonstrated through pharmacological inhibition of RIP1 and RIP3 activity that cone cell death was also delayed. Collectively, these results demonstrate that the underlying mechanism of cone cell death in the pde6c(w59) mutant retina is through necroptosis, whereas rod photoreceptor bystander death occurs through a caspase-dependent mechanism. This suggests that targeting the RIP kinase signalling pathway could be an effective therapeutic intervention in retinal degeneration patients. As bystander cell death is an important feature of many retinal diseases, combinatorial approaches targeting different cell death pathways may evolve as an important general principle in treatment.

Retinoic acid temporally orchestrates colonization of the gut by vagal neural crest cells.

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Uribe, Rosa A; Hong, Stephanie S; Bronner, Marianne E

2018-01-01

The enteric nervous system arises from neural crest cells that migrate as chains into and along the primitive gut, subsequently differentiating into enteric neurons and glia. Little is known about the mechanisms governing neural crest migration en route to and along the gut in vivo. Here, we report that Retinoic Acid (RA) temporally controls zebrafish enteric neural crest cell chain migration. In vivo imaging reveals that RA loss severely compromises the integrity and migration of the chain of neural crest cells during the window of time window when they are moving along the foregut. After loss of RA, enteric progenitors accumulate in the foregut and differentiate into enteric neurons, but subsequently undergo apoptosis resulting in a striking neuronal deficit. Moreover, ectopic expression of the transcription factor meis3 and/or the receptor ret, partially rescues enteric neuron colonization after RA attenuation. Collectively, our findings suggest that retinoic acid plays a critical temporal role in promoting enteric neural crest chain migration and neuronal survival upstream of Meis3 and RET in vivo. Copyright © 2017 Elsevier Inc. All rights reserved.

Cardiac Ca2+ signalling in zebrafish: Translation of findings to man.

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van Opbergen, Chantal J M; van der Voorn, Stephanie M; Vos, Marc A; de Boer, Teun P; van Veen, Toon A B

2018-05-07

Sudden cardiac death is a leading cause of death worldwide, mainly caused by highly disturbed electrical activation patterns in the heart. Currently, murine models are the most popular model to study underlying molecular mechanisms of inherited or acquired cardiac electrical abnormalities, although the numerous electrophysiological discrepancies between mouse and human raise the question whether mice are the optimal model to study cardiac rhythm disorders. Recently it has been uncovered that the zebrafish cardiac electrophysiology seems surprisingly similar to the human heart, mainly because the zebrafish AP contains a clear plateau phase and ECG characteristics show alignment with the human ECG. Although, before using zebrafish as a model to study cardiac arrhythmogenesis, however, it is very important to gain a better insight into the electrophysiological characteristics of the zebrafish heart. In this review we outline the electrophysiological machinery of the zebrafish cardiomyocytes, with a special focus on the intracellular Ca 2+ dynamics and excitation-contraction coupling. We debate the potential of zebrafish as a model to study human cardiovascular diseases and postulate steps to employ zebrafish into a more 'humanized' model. Copyright © 2018 Elsevier Ltd. All rights reserved.

Activation of Sterol Regulatory Element Binding Factors by Fenofibrate and Gemfibrozil Stimulate Myelination in Zebrafish

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Yuhei Nishimura

2016-07-01

Full Text Available Oligodendrocytes are major myelin-producing cells and play essential roles in the function of a healthy nervous system. However, they are also one of the most vulnerable neural cell types in the central nervous system (CNS, and myelin abnormalities in the CNS are found in a wide variety of neurological disorders, including multiple sclerosis, adrenoleukodystrophy, and schizophrenia. There is an urgent need to identify small molecular weight compounds that can stimulate myelination. In this study, we performed comparative transcriptome analysis to identify pharmacodynamic effects common to miconazole and clobetasol, which have been shown to stimulate myelination by mouse oligodendrocyte progenitor cells (OPCs. Of the genes differentially expressed in both miconazole- and clobetasol-treated mouse OPCs compared with untreated cells, we identified differentially expressed genes (DEGs common to both drug treatments. Gene ontology analysis revealed that these DEGs are significantly associated with the sterol biosynthetic pathway, and further bioinformatics analysis suggested that sterol regulatory element binding factors (SREBFs might be key upstream regulators of the DEGs. In silico screening of a public database for chemicals associated with SREBF activation identified fenofibrate, a peroxisome proliferator-activated receptor α (PPARα agonist, as a drug that increases the expression of known SREBF targets, raising the possibility that fenofibrate may also stimulate myelination. To test this, we performed in vivo imaging of zebrafish expressing a fluorescent reporter protein under the control of the myelin basic protein (mbp promoter. Treatment of zebrafish with fenofibrate significantly increased expression of the fluorescent reporter compared with untreated zebrafish. This increase was attenuated by co-treatment with fatostatin, a specific inhibitor of SREBFs, confirming that the fenofibrate effect was mediated via SREBFs. Furthermore, incubation

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

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

2012-07-13

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

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

International Nuclear Information System (INIS)

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

2012-01-01

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

Direct Visualization of DNA Replication Dynamics in Zebrafish Cells.

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Kuriya, Kenji; Higashiyama, Eriko; Avşar-Ban, Eriko; Tamaru, Yutaka; Ogata, Shin; Takebayashi, Shin-ichiro; Ogata, Masato; Okumura, Katsuzumi

2015-12-01

Spatiotemporal regulation of DNA replication in the S-phase nucleus has been extensively studied in mammalian cells because it is tightly coupled with the regulation of other nuclear processes such as transcription. However, little is known about the replication dynamics in nonmammalian cells. Here, we analyzed the DNA replication processes of zebrafish (Danio rerio) cells through the direct visualization of replicating DNA in the nucleus and on DNA fiber molecules isolated from the nucleus. We found that zebrafish chromosomal DNA at the nuclear interior was replicated first, followed by replication of DNA at the nuclear periphery, which is reminiscent of the spatiotemporal regulation of mammalian DNA replication. However, the relative duration of interior DNA replication in zebrafish cells was longer compared to mammalian cells, possibly reflecting zebrafish-specific genomic organization. The rate of replication fork progression and ori-to-ori distance measured by the DNA combing technique were ∼ 1.4 kb/min and 100 kb, respectively, which are comparable to those in mammalian cells. To our knowledge, this is a first report that measures replication dynamics in zebrafish cells.

Damage and functional recovery of the mouse retina after exposure to genotoxic agents

International Nuclear Information System (INIS)

Vinogradova, Yu.V.; Tronov, V.A.; Lyakhova, K.N.; Ostrovskij, M.A.

2013-01-01

As is known, the mature retina is characterized by high radiation resistance. We showed earlier that ionizing radiation at a dose of ≥25 Gy and the chemical genotoxic agent methylnitrosourea (MNU) in a concentration of ≥60 mg/kg induce acute retinal degeneration, combined with proapoptotic protein expression. The process has a high genotoxic threshold, below which no degeneration signs were traced. The aim of this work was to study the damaging effect of ionizing radiation and MNU on the functional activity of the retina and its ability to recover after exposure to these genotoxicants. The functional activity of the mouse retina was evaluated with electroretinograms (ERG). In parallel, morphological changes in the retina were controlled, and the TUNEL detection of the death of its cell elements was performed. It has been shown that gamma rays or accelerated proton irradiation below 15 Gy cause no structural or functional changes in the mouse retina, which confirms the mature retina's high radiation resistance. Irradiation with a higher dose of 25 Gy leads to photoreceptor layer destruction. This goes along with an increase in the number of the TUNEL-positive photoreceptors, among which are cells with fragmented nuclei, which are typical of apoptosis. MNU in a concentration of 70 mg/kg caused the irreversible loss of the retina's physiological activity, and the morphological degeneration of photoreceptors and their mass death. In a concentration of 35 mg/kg, however, MNU had no cytotoxic effect on the retina. Moreover, this dose caused a reversible ERG amplitude decrease. Also, adaptive response was observed in the retina, which became apparent after two consecutive MNU injections - first, at a dose of 17 mg/kg; then, at a cytotoxic dose of 70 mg/kg. These results point to the possibility of the neurohormesis effect, which was described concerning the retina's exposure to ionizing radiation and some chemicals.

Radiation therapy in and about the retina, optic nerve, and anterior visual pathway: psychophysical assessment

International Nuclear Information System (INIS)

Fitzgerald, C.R.; Enoch, J.M.; Temme, L.A.

1981-01-01

Visual changes may develop in patients receiving radiation therapy for malignant neoplasms in and about the optic nerve and anterior visual pathway. Cases have been studied using a series of psychophysical tests, including kinetic perimetry, increment threshold determinations, Flashing Repeat Static Test, and sustained- and transient-like functions. A characteristic time-dependent reduction in sensitivity has been identified in these patients. This finding, in addition to the presence of nerve fiber bundle defects, appears to place the pathologic changes in the axon of the ganglion cell posterior to the lamina cribrosa. Any change in the sustained- and transient-like functions, the organization of which appears to be in the neural retina, was seen only if a concomitant radiation retinopathy was identified

Radiation therapy in and about the retina, optic nerve, and anterior visual pathway. Psychophysical assessment

International Nuclear Information System (INIS)

Fitzgerald, C.R.; Enoch, J.M.; Temme, L.A.

1981-01-01

Visual changes may develop in patients receiving radiation therapy for malignant neoplasms in and about the optic nerve and anterior visual pathway. Cases have been studied using a series of psychophysical tests, including kinetic perimetry, increment threshold determinations, Flashing Repeat Static Test, and sustained- and transient-like functions. A characteristic time-dependent reduction in sensitivity has been identified in these patients. This finding, in addition to the presence of nerve fiber bundle defects, appears to place the pathologic changes in the axon of the ganglion cell posterior to the lamina cribrosa. Any change in the sustained- and transient-like functions, the organization of which appears to be in the neural retina, was seen only if a concomitant radiation retinopathy was identified

Whole-body and multispectral photoacoustic imaging of adult zebrafish

Science.gov (United States)

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.

Heart-specific expression of laminopathic mutations in transgenic zebrafish.

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Verma, Ajay D; Parnaik, Veena K

2017-07-01

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

Distribution of tubulin, kinesin, and dynein in light- and dark-adapted octopus retinas.

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Martinez, J M; Elfarissi, H; De Velasco, B; Ochoa, G H; Miller, A M; Clark, Y M; Matsumoto, B; Robles, L J

2000-01-01

Cephalopod retinas exhibit several responses to light and dark adaptation, including rhabdom size changes, photopigment movements, and pigment granule migration. Light- and dark-directed rearrangements of microfilament and microtubule cytoskeletal transport pathways could drive these changes. Recently, we localized actin-binding proteins in light-/dark-adapted octopus rhabdoms and suggested that actin cytoskeletal rearrangements bring about the formation and degradation of rhabdomere microvilli subsets. To determine if the microtubule cytoskeleton and associated motor proteins control the other light/dark changes, we used immunoblotting and immunocytochemical procedures to map the distribution of tubulin, kinesin, and dynein in dorsal and ventral halves of light- and dark-adapted octopus retinas. Immunoblots detected alpha- and beta-tubulin, dynein intermediate chain, and kinesin heavy chain in extracts of whole retinas. Epifluorescence and confocal microscopy showed that the tubulin proteins were distributed throughout the retina with more immunoreactivity in retinas exposed to light. Kinesin localization was heavy in the pigment layer of light- and dark-adapted ventral retinas but was less prominent in the dorsal region. Dynein distribution also varied in dorsal and ventral retinas with more immunoreactivity in light- and dark-adapted ventral retinas and confocal microscopy emphasized the granular nature of this labeling. We suggest that light may regulate the distribution of microtubule cytoskeletal proteins in the octopus retina and that position, dorsal versus ventral, also influences the distribution of motor proteins. The microtubule cytoskeleton is most likely involved in pigment granule migration in the light and dark and with the movement of transport vesicles from the photoreceptor inner segments to the rhabdoms.

Ionic channels underlying the ventricular action potential in zebrafish embryo.

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Alday, Aintzane; Alonso, Hiart; Gallego, Monica; Urrutia, Janire; Letamendia, Ainhoa; Callol, Carles; Casis, Oscar

2014-06-01

Over the last years zebrafish has become a popular model in the study of cardiac physiology, pathology and pharmacology. Recently, the application of the 3Rs regulation and the characteristics of the embryo have reduced the use of adult zebrafish use in many studies. However, the zebrafish embryo cardiac physiology is poorly characterized since most works have used indirect techniques and direct recordings of cardiac action potential and ionic currents are scarce. In order to optimize the zebrafish embryo model, we used electrophysiological, pharmacological and immunofluorescence tools to identify the characteristics and the ionic channels involved in the ventricular action potentials of zebrafish embryos. The application of Na(+) or T-type Ca(+2) channel blockers eliminated the cardiac electrical activity, indicating that the action potential upstroke depends on Na(+) and T-type Ca(+2) currents. The plateau phase depends on L-type Ca(+2) channels since it is abolished by specific blockade. The direct channel blockade indicates that the action potential repolarization and diastolic potential depends on ERG K(+) channels. The presence in the embryonic heart of the Nav1.5, Cav1.2, Cav3.2 and ERG channels was also confirmed by immunofluorescence, while the absence of effect of specific blockers and immunostaining indicate that two K(+) repolarizing currents present in human heart, Ito and IKs, are absent in the embryonic zebrafish heart. Our results describe the ionic channels present and its role in the zebrafish embryo heart and support the use of zebrafish embryos to study human diseases and their use for drug testing. Copyright © 2014 Elsevier Ltd. All rights reserved.

High magnetic field induced otolith fusion in the zebrafish larvae.

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Pais-Roldán, Patricia; Singh, Ajeet Pratap; Schulz, Hildegard; Yu, Xin

2016-04-11

Magnetoreception in animals illustrates the interaction of biological systems with the geomagnetic field (geoMF). However, there are few studies that identified the impact of high magnetic field (MF) exposure from Magnetic Resonance Imaging (MRI) scanners (>100,000 times of geoMF) on specific biological targets. Here, we investigated the effects of a 14 Tesla MRI scanner on zebrafish larvae. All zebrafish larvae aligned parallel to the B0 field, i.e. the static MF, in the MRI scanner. The two otoliths (ear stones) in the otic vesicles of zebrafish larvae older than 24 hours post fertilization (hpf) fused together after the high MF exposure as short as 2 hours, yielding a single-otolith phenotype with aberrant swimming behavior. The otolith fusion was blocked in zebrafish larvae under anesthesia or embedded in agarose. Hair cells may play an important role on the MF-induced otolith fusion. This work provided direct evidence to show that high MF interacts with the otic vesicle of zebrafish larvae and causes otolith fusion in an "all-or-none" manner. The MF-induced otolith fusion may facilitate the searching for MF sensors using genetically amenable vertebrate animal models, such as zebrafish.

Neutrophil Reverse Migration Becomes Transparent with Zebrafish

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Taylor W. Starnes

2012-01-01

Full Text Available The precise control of neutrophil-mediated inflammation is critical for both host defense and the prevention of immunopathology. In vivo imaging studies in zebrafish, and more recently in mice, have made the novel observation that neutrophils leave a site of inflammation through a process called neutrophil reverse migration. The application of advanced imaging techniques to the genetically tractable, optically transparent zebrafish larvae was critical for these advances. Still, the mechanisms underlying neutrophil reverse migration and its effects on the resolution or priming of immune responses remain unclear. Here, we review the current knowledge of neutrophil reverse migration, its potential roles in host immunity, and the live imaging tools that make zebrafish a valuable model for increasing our knowledge of neutrophil behavior in vivo.

Acute Exposure to Fluoxetine Alters Aggressive Behavior of Zebrafish and Expression of Genes Involved in Serotonergic System Regulation

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Michail Pavlidis

2017-04-01

Full Text Available Zebrafish, Danio rerio, is an emerging model organism in stress and neurobehavioral studies. In nature, the species forms shoals, yet when kept in pairs it exhibits an agonistic and anxiety-like behavior that leads to the establishment of dominant-subordinate relationships. Fluoxetine, a selective serotonin reuptake inhibitor, is used as an anxiolytic tool to alter aggressive behavior in several vertebrates and as an antidepressant drug in humans. Pairs of male zebrafish were held overnight to develop dominant—subordinate behavior, either treated or non-treated for 2 h with fluoxetine (5 mg L−1, and allowed to interact once more for 1 h. Behavior was recorded both prior and after fluoxetine administration. At the end of the experiment, trunk and brain samples were also taken for cortisol determination and mRNA expression studies, respectively. Fluoxetine treatment significantly affected zebrafish behavior and the expression levels of several genes, by decreasing offensive aggression in dominants and by eliminating freezing in the subordinates. There was no statistically significant difference in whole-trunk cortisol concentrations between dominant and subordinate fish, while fluoxetine treatment resulted in higher (P = 0.004 cortisol concentrations in both groups. There were statistically significant differences between dominant and subordinate fish in brain mRNA expression levels of genes involved in stress axis (gr, mr, neural activity (bdnf, c-fos, and the serotonergic system (htr2b, slc6a4b. The significant decrease in the offensive and defensive aggression following fluoxetine treatment was concomitant with a reversed pattern in c-fos expression levels. Overall, an acute administration of a selective serotonin reuptake inhibitor alters aggressive behavior in male zebrafish in association with changes in the neuroendocrine mediators of coping styles.

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

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Ramsay, J.M.; Watral, Virginia G.; 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.

[Lattice degeneration of the peripheral retina: ultrastructural study].

Science.gov (United States)

Bec, P; Malecaze, F; Arne, J L; Mathis, A

1985-01-01

The ultrastructural study of a case of snail track degeneration shows the presence of lipid inclusions in both the glial and the macrophage cells in every layer of the retina, and the existence of intraretinal fibers different from collagen fibers appearing to be glial filaments similar to those found in astrocytic gliomes and to the Rosenthal fibers observed in senile nervous cells. Other features were thinning of the retina and absence of blood vessels in the retina. There are no abnormalities of the vitreo-retinal juncture. All the lesions are in agreement with those observed by Daicker [Ophthalmologica, Basel 165: 360-365, 1972; Klin. Mbl. Augenheilk. 172: 581-583, 1978] with some differences, however. They are different from those found in lattice degeneration. They show that snail track degeneration is a specific form of peripheral retinal degeneration which is quite different from lattice degeneration and must not be considered similar.

Impact of MCT1 Haploinsufficiency on the Mouse Retina

KAUST Repository

Peachey, Neal S.

2018-05-02

The monocarboxylate transporter 1 (MCT1) is highly expressed in the outer retina, suggesting that it plays a critical role in photoreceptors. We examined MCT1+/− heterozygotes, which express half of the normal complement of MCT1. The MCT1+/− retina developed normally and retained normal function, indicating that MCT1 is expressed at sufficient levels to support outer retinal metabolism.

Impact of MCT1 Haploinsufficiency on the Mouse Retina

KAUST Repository

Peachey, Neal S.; Yu, Minzhong; Han, John Y. S.; Lengacher, Sylvain; Magistretti, Pierre J.; Pellerin, Luc; Philp, Nancy J.

2018-01-01

The monocarboxylate transporter 1 (MCT1) is highly expressed in the outer retina, suggesting that it plays a critical role in photoreceptors. We examined MCT1+/− heterozygotes, which express half of the normal complement of MCT1. The MCT1+/− retina developed normally and retained normal function, indicating that MCT1 is expressed at sufficient levels to support outer retinal metabolism.

Impact of bronchopulmonary dysplasia on brain and retina

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Annie Wing Hoi Poon

2016-04-01

Full Text Available Many premature newborns develop bronchopulmonary dysplasia (BPD, a chronic lung disease resulting from prolonged mechanical ventilation and hyperoxia. BPD survivors typically suffer long-term injuries not only to the lungs, but also to the brain and retina. However, currently it is not clear whether the brain and retinal injuries in these newborns are related only to their prematurity, or also to BPD. We investigated whether the hyperoxia known to cause histologic changes in the lungs similar to BPD in an animal model also causes brain and retinal injuries. Sprague Dawley rat pups were exposed to hyperoxia (95% O2, ‘BPD’ group or room air (21% O2, ‘control’ group from postnatal day 4–14 (P4–14; the rat pups were housed in room air between P14 and P28. At P28, they were sacrificed, and their lungs, brain, and eyes were extracted. Hematoxylin and eosin staining was performed on lung and brain sections; retinas were stained with Toluidine Blue. Hyperoxia exposure resulted in an increased mean linear intercept in the lungs (P<0.0001. This increase was associated with a decrease in some brain structures [especially the whole-brain surface (P=0.02], as well as a decrease in the thickness of the retinal layers [especially the total retina (P=0.0008], compared to the room air control group. In addition, a significant negative relationship was observed between the lung structures and the brain (r=−0.49, P=0.02 and retina (r=−0.70, P=0.0008 structures. In conclusion, hyperoxia exposure impaired lung, brain, and retina structures. More severe lung injuries correlated with more severe brain and retinal injuries. This result suggests that the same animal model of chronic neonatal hyperoxia can be used to simultaneously study lung, brain and retinal injuries related to hyperoxia.

Zebrafish in Toxicology and Environmental Health.

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Bambino, Kathryn; Chu, Jaime

2017-01-01

As manufacturing processes and development of new synthetic compounds increase to keep pace with the expanding global demand, environmental health, and the effects of toxicant exposure are emerging as critical public health concerns. Additionally, chemicals that naturally occur in the environment, such as metals, have profound effects on human and animal health. Many of these compounds are in the news: lead, arsenic, and endocrine disruptors such as bisphenol A have all been widely publicized as causing disease or damage to humans and wildlife in recent years. Despite the widespread appreciation that environmental toxins can be harmful, there is limited understanding of how many toxins cause disease. Zebrafish are at the forefront of toxicology research; this system has been widely used as a tool to detect toxins in water samples and to investigate the mechanisms of action of environmental toxins and their related diseases. The benefits of zebrafish for studying vertebrate development are equally useful for studying teratogens. Here, we review how zebrafish are being used both to detect the presence of some toxins as well as to identify how environmental exposures affect human health and disease. We focus on areas where zebrafish have been most effectively used in ecotoxicology and in environmental health, including investigation of exposures to endocrine disruptors, industrial waste byproducts, and arsenic. © 2017 Elsevier Inc. All rights reserved.

Axonal regeneration in zebrafish spinal cord

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Hui, Subhra Prakash

2018-01-01

Abstract In the present review we discuss two interrelated events—axonal damage and repair—known to occur after spinal cord injury (SCI) in the zebrafish. Adult zebrafish are capable of regenerating axonal tracts and can restore full functionality after SCI. Unlike fish, axon regeneration in the adult mammalian central nervous system is extremely limited. As a consequence of an injury there is very little repair of disengaged axons and therefore functional deficit persists after SCI in adult mammals. In contrast, peripheral nervous system axons readily regenerate following injury and hence allow functional recovery both in mammals and fish. A better mechanistic understanding of these three scenarios could provide a more comprehensive insight into the success or failure of axonal regeneration after SCI. This review summarizes the present understanding of the cellular and molecular basis of axonal regeneration, in both the peripheral nervous system and the central nervous system, and large scale gene expression analysis is used to focus on different events during regeneration. The discovery and identification of genes involved in zebrafish spinal cord regeneration and subsequent functional experimentation will provide more insight into the endogenous mechanism of myelination and remyelination. Furthermore, precise knowledge of the mechanism underlying the extraordinary axonal regeneration process in zebrafish will also allow us to unravel the potential therapeutic strategies to be implemented for enhancing regrowth and remyelination of axons in mammals. PMID:29721326

Effects of alpha particles on zebrafish embryos

International Nuclear Information System (INIS)

Yum, E.H.W.; Choi, V.W.Y.; Yu, K.N.; Li, V.W.T.; Cheng, S.H.

2008-01-01

Full text: Ionizing radiation such as X-ray and alpha particles can damage cellular macromolecules, which can lead to DNA single- and double-strand breaks. In the present work, we studied the effects of alpha particles on dechorionated zebrafish embryos. Thin polyallyldiglycol carbonate (PADC) films with a thickness of 16 μm were prepared from commercially available PADC films (with thickness of 100 μm) by chemical etching and used as support substrates for holding zebrafish embryos for alpha-particle irradiation. These films recorded alpha-particle hit positions, quantified the number and energy of alpha particles actually incident on the embryo cells, and thus enabled the calculation of the dose absorbed by the embryo cells. Irradiation was made at 1.25 hours post fertilization (hpf) with various absorbed dose. TdT-mediated dUTP Nick-End Labeling (TUNEL) assay was performed on the embryos at different time stages after irradiation. Marked apoptosis was detected only in embryos at earlier time stages. The results showed that DNA double-strand break during zebrafish embryogenesis can be induced by alpha-particle irradiation, which suggests that zebrafish is a potential model for assessing the effects of alpha-particle radiation

Defective glycinergic synaptic transmission in zebrafish motility mutants

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Hiromi Hirata

2010-01-01

Full Text Available Glycine is a major inhibitory neurotransmitter in the spinal cord and brainstem. Recently, in vivo analysis of glycinergic synaptic transmission has been pursued in zebrafish using molecular genetics. An ENU mutagenesis screen identified two behavioral mutants that are defective in glycinergic synaptic transmission. Zebrafish bandoneon (beo mutants have a defect in glrbb, one of the duplicated glycine receptor (GlyR β subunit genes. These mutants exhibit a loss of glycinergic synaptic transmission due to a lack of synaptic aggregation of GlyRs. Due to the consequent loss of reciprocal inhibition of motor circuits between the two sides of the spinal cord, motor neurons activate simultaneously on both sides resulting in bilateral contraction of axial muscles of beo mutants, eliciting the so-called ‘accordion’ phenotype. Similar defects in GlyR subunit genes have been observed in several mammals and are the basis for human hyperekplexia/startle disease. By contrast, zebrafish shocked (sho mutants have a defect in slc6a9, encoding GlyT1, a glycine transporter that is expressed by astroglial cells surrounding the glycinergic synapse in the hindbrain and spinal cord. GlyT1 mediates rapid uptake of glycine from the synaptic cleft, terminating synaptic transmission. In zebrafish sho mutants, there appears to be elevated extracellular glycine resulting in persistent inhibition of postsynaptic neurons and subsequent reduced motility, causing the ‘twitch once’ phenotype. We review current knowledge regarding zebrafish ‘accordion’ and ‘twitch once’ mutants, including beo and sho, and report the identification of a new α2 subunit that revises the phylogeny of zebrafish GlyRs.

Defective Glycinergic Synaptic Transmission in Zebrafish Motility Mutants

Science.gov (United States)

Hirata, Hiromi; Carta, Eloisa; Yamanaka, Iori; Harvey, Robert J.; Kuwada, John Y.

2009-01-01

Glycine is a major inhibitory neurotransmitter in the spinal cord and brainstem. Recently, in vivo analysis of glycinergic synaptic transmission has been pursued in zebrafish using molecular genetics. An ENU mutagenesis screen identified two behavioral mutants that are defective in glycinergic synaptic transmission. Zebrafish bandoneon (beo) mutants have a defect in glrbb, one of the duplicated glycine receptor (GlyR) β subunit genes. These mutants exhibit a loss of glycinergic synaptic transmission due to a lack of synaptic aggregation of GlyRs. Due to the consequent loss of reciprocal inhibition of motor circuits between the two sides of the spinal cord, motor neurons activate simultaneously on both sides resulting in bilateral contraction of axial muscles of beo mutants, eliciting the so-called ‘accordion’ phenotype. Similar defects in GlyR subunit genes have been observed in several mammals and are the basis for human hyperekplexia/startle disease. By contrast, zebrafish shocked (sho) mutants have a defect in slc6a9, encoding GlyT1, a glycine transporter that is expressed by astroglial cells surrounding the glycinergic synapse in the hindbrain and spinal cord. GlyT1 mediates rapid uptake of glycine from the synaptic cleft, terminating synaptic transmission. In zebrafish sho mutants, there appears to be elevated extracellular glycine resulting in persistent inhibition of postsynaptic neurons and subsequent reduced motility, causing the ‘twitch-once’ phenotype. We review current knowledge regarding zebrafish ‘accordion’ and ‘twitch-once’ mutants, including beo and sho, and report the identification of a new α2 subunit that revises the phylogeny of zebrafish GlyRs. PMID:20161699

BMP signaling modulates hepcidin expression in zebrafish embryos independent of hemojuvelin.

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Yann Gibert

2011-01-01

Full Text Available Hemojuvelin (Hjv, a member of the repulsive-guidance molecule (RGM family, upregulates transcription of the iron regulatory hormone hepcidin by activating the bone morphogenetic protein (BMP signaling pathway in mammalian cells. Mammalian models have identified furin, neogenin, and matriptase-2 as modifiers of Hjv's function. Using the zebrafish model, we evaluated the effects of hjv and its interacting proteins on hepcidin expression during embryonic development. We found that hjv is strongly expressed in the notochord and somites of the zebrafish embryo and that morpholino knockdown of hjv impaired the development of these structures. Knockdown of hjv or other hjv-related genes, including zebrafish orthologs of furin or neogenin, however, failed to decrease hepcidin expression relative to liver size. In contrast, overexpression of bmp2b or knockdown of matriptase-2 enhanced the intensity and extent of hepcidin expression in zebrafish embryos, but this occurred in an hjv-independent manner. Furthermore, we demonstrated that zebrafish hjv can activate the human hepcidin promoter and enhance BMP responsive gene expression in vitro, but is expressed at low levels in the zebrafish embryonic liver. Taken together, these data support an alternative mechanism for hepcidin regulation during zebrafish embryonic development, which is independent of hjv.

The HDAC Inhibitor TSA Ameliorates a Zebrafish Model of Duchenne Muscular Dystrophy.

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Johnson, Nathan M; Farr, Gist H; Maves, Lisa

2013-09-17

Zebrafish are an excellent model for Duchenne muscular dystrophy. In particular, zebrafish provide a system for rapid, easy, and low-cost screening of small molecules that can ameliorate muscle damage in dystrophic larvae. Here we identify an optimal anti-sense morpholino cocktail that robustly knocks down zebrafish Dystrophin (dmd-MO). We use two approaches, muscle birefringence and muscle actin expression, to quantify muscle damage and show that the dmd-MO dystrophic phenotype closely resembles the zebrafish dmd mutant phenotype. We then show that the histone deacetylase (HDAC) inhibitor TSA, which has been shown to ameliorate the mdx mouse Duchenne model, can rescue muscle fiber damage in both dmd-MO and dmd mutant larvae. Our study identifies optimal morpholino and phenotypic scoring approaches for dystrophic zebrafish, further enhancing the zebrafish dmd model for rapid and cost-effective small molecule screening.

Zebrafish Expression Ontology of Gene Sets (ZEOGS): A Tool to Analyze Enrichment of Zebrafish Anatomical Terms in Large Gene Sets

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Marsico, Annalisa

2013-01-01

Abstract The zebrafish (Danio rerio) is an established model organism for developmental and biomedical research. It is frequently used for high-throughput functional genomics experiments, such as genome-wide gene expression measurements, to systematically analyze molecular mechanisms. However, the use of whole embryos or larvae in such experiments leads to a loss of the spatial information. To address this problem, we have developed a tool called Zebrafish Expression Ontology of Gene Sets (ZEOGS) to assess the enrichment of anatomical terms in large gene sets. ZEOGS uses gene expression pattern data from several sources: first, in situ hybridization experiments from the Zebrafish Model Organism Database (ZFIN); second, it uses the Zebrafish Anatomical Ontology, a controlled vocabulary that describes connected anatomical structures; and third, the available connections between expression patterns and anatomical terms contained in ZFIN. Upon input of a gene set, ZEOGS determines which anatomical structures are overrepresented in the input gene set. ZEOGS allows one for the first time to look at groups of genes and to describe them in terms of shared anatomical structures. To establish ZEOGS, we first tested it on random gene selections and on two public microarray datasets with known tissue-specific gene expression changes. These tests showed that ZEOGS could reliably identify the tissues affected, whereas only very few enriched terms to none were found in the random gene sets. Next we applied ZEOGS to microarray datasets of 24 and 72 h postfertilization zebrafish embryos treated with beclomethasone, a potent glucocorticoid. This analysis resulted in the identification of several anatomical terms related to glucocorticoid-responsive tissues, some of which were stage-specific. Our studies highlight the ability of ZEOGS to extract spatial information from datasets derived from whole embryos, indicating that ZEOGS could be a useful tool to automatically analyze gene

Zebrafish Expression Ontology of Gene Sets (ZEOGS): a tool to analyze enrichment of zebrafish anatomical terms in large gene sets.

Science.gov (United States)

Prykhozhij, Sergey V; Marsico, Annalisa; Meijsing, Sebastiaan H

2013-09-01

The zebrafish (Danio rerio) is an established model organism for developmental and biomedical research. It is frequently used for high-throughput functional genomics experiments, such as genome-wide gene expression measurements, to systematically analyze molecular mechanisms. However, the use of whole embryos or larvae in such experiments leads to a loss of the spatial information. To address this problem, we have developed a tool called Zebrafish Expression Ontology of Gene Sets (ZEOGS) to assess the enrichment of anatomical terms in large gene sets. ZEOGS uses gene expression pattern data from several sources: first, in situ hybridization experiments from the Zebrafish Model Organism Database (ZFIN); second, it uses the Zebrafish Anatomical Ontology, a controlled vocabulary that describes connected anatomical structures; and third, the available connections between expression patterns and anatomical terms contained in ZFIN. Upon input of a gene set, ZEOGS determines which anatomical structures are overrepresented in the input gene set. ZEOGS allows one for the first time to look at groups of genes and to describe them in terms of shared anatomical structures. To establish ZEOGS, we first tested it on random gene selections and on two public microarray datasets with known tissue-specific gene expression changes. These tests showed that ZEOGS could reliably identify the tissues affected, whereas only very few enriched terms to none were found in the random gene sets. Next we applied ZEOGS to microarray datasets of 24 and 72 h postfertilization zebrafish embryos treated with beclomethasone, a potent glucocorticoid. This analysis resulted in the identification of several anatomical terms related to glucocorticoid-responsive tissues, some of which were stage-specific. Our studies highlight the ability of ZEOGS to extract spatial information from datasets derived from whole embryos, indicating that ZEOGS could be a useful tool to automatically analyze gene expression

Quaternary and tertiary aldoxime antidotes for organophosphate exposure in a zebrafish model system

Energy Technology Data Exchange (ETDEWEB)

Schmidt, Hayden R. [Department of Biology, Whittier College, Whittier, CA 90608 (United States); Radić, Zoran; Taylor, Palmer [Department of Pharmacology, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California at San Diego, La Jolla, CA 92093-0650 (United States); Fradinger, Erica A., E-mail: efrading@whittier.edu [Department of Biology, Whittier College, Whittier, CA 90608 (United States)

2015-04-15

The zebrafish is rapidly becoming an important model system for screening of new therapeutics. Here we evaluated the zebrafish as a potential pharmacological model for screening novel oxime antidotes to organophosphate (OP)-inhibited acetylcholinesterase (AChE). The k{sub i} values determined for chlorpyrifos oxon (CPO) and dichlorvos (DDVP) showed that CPO was a more potent inhibitor of both human and zebrafish AChE, but overall zebrafish AChE was less sensitive to OP inhibition. In contrast, aldoxime antidotes, the quaternary ammonium 2-PAM and tertiary amine RS-194B, showed generally similar overall reactivation kinetics, k{sub r}, in both zebrafish and human AChE. However, differences between the K{sub ox} and k{sub 2} constants suggest that zebrafish AChE associates more tightly with oximes, but has a slower maximal reactivation rate than human AChE. Homology modeling suggests that these kinetic differences result from divergences in the amino acids lining the entrance to the active site gorge. Although 2-PAM had the more favorable in vitro reactivation kinetics, RS-194B was more effective antidote in vivo. In intact zebrafish embryos, antidotal treatment with RS-194B rescued embryos from OP toxicity, whereas 2-PAM had no effect. Dechorionation of the embryos prior to antidotal treatment allowed both 2-PAM and RS-194B to rescue zebrafish embryos from OP toxicity. Interestingly, RS-194B and 2-PAM alone increased cholinergic motor activity in dechorionated embryos possibly due to the reversible inhibition kinetics, K{sub i} and αK{sub i}, of the oximes. Together these results demonstrate that the zebrafish at various developmental stages provides an excellent model for investigating membrane penetrant antidotes to OP exposure. - Highlights: • Zebrafish AChE shares significant structural similarities with human AChE. • OP-inhibited zebrafish and human AChE exhibit similar reactivation kinetics. • The zebrafish chorion is permeable to BBB penetrant and not

Cadmium inhibits neurogenesis in zebrafish embryonic brain development

Energy Technology Data Exchange (ETDEWEB)

Chow, Elly Suk Hen [Division of Biology, California Institute of Technology, 1200 California Boulevard, Pasadena, CA 91125 (United States); Hui, Michelle Nga Yu; Lin Chunchi [Department of Biology and Chemistry, City University of Hong Kong, 83 Tat Chee Avenue, Kowloon, Hong Kong (China); Cheng Shukhan [Department of Biology and Chemistry, City University of Hong Kong, 83 Tat Chee Avenue, Kowloon, Hong Kong (China)], E-mail: bhcheng@cityu.edu.hk

2008-05-01

Cadmium is a non-essential heavy metal found abundantly in the environment. Children of women exposed to cadmium during pregnancy display lower motor and perceptual abilities. High cadmium body burden in children is also related to impaired intelligence and lowered school achievement. However, little is known about the molecular and cellular basis of developmental neurotoxicity in the sensitive early life stages of animals. In this study, we explore neurological deficits caused by cadmium during early embryonic stages in zebrafish by examining regionalization of the neural tube, pattern formation and cell fate determination, commitment of proneural genes and induction of neurogenesis. We show that cadmium-treated embryos developed a smaller head with unclear boundaries between the brain subdivisions, particularly in the mid-hindbrain region. Embryos display normal anterior to posterior regionalization; however, the commitment of neural progenitor cells was affected by cadmium. We observe prominent reductions in the expression of several proneuronal genes including ngn1 in cell clusters, zash1a in the developing optic tectum, and zash1b in the telencephalon and tectum. Cadmium-treated embryos also have fewer differentiated neurons and glia in the facial sensory ganglia as indicated by decreased zn-12 expression. Also, a lower transcription level of neurogenic genes, ngn1 and neuroD, is observed in neurons. Our data suggest that cadmium-induced neurotoxicity can be caused by impaired neurogenesis, resulting in markedly reduced neuronal differentiation and axonogenesis.

Cadmium inhibits neurogenesis in zebrafish embryonic brain development

International Nuclear Information System (INIS)

Chow, Elly Suk Hen; Hui, Michelle Nga Yu; Lin Chunchi; Cheng Shukhan

2008-01-01

Cadmium is a non-essential heavy metal found abundantly in the environment. Children of women exposed to cadmium during pregnancy display lower motor and perceptual abilities. High cadmium body burden in children is also related to impaired intelligence and lowered school achievement. However, little is known about the molecular and cellular basis of developmental neurotoxicity in the sensitive early life stages of animals. In this study, we explore neurological deficits caused by cadmium during early embryonic stages in zebrafish by examining regionalization of the neural tube, pattern formation and cell fate determination, commitment of proneural genes and induction of neurogenesis. We show that cadmium-treated embryos developed a smaller head with unclear boundaries between the brain subdivisions, particularly in the mid-hindbrain region. Embryos display normal anterior to posterior regionalization; however, the commitment of neural progenitor cells was affected by cadmium. We observe prominent reductions in the expression of several proneuronal genes including ngn1 in cell clusters, zash1a in the developing optic tectum, and zash1b in the telencephalon and tectum. Cadmium-treated embryos also have fewer differentiated neurons and glia in the facial sensory ganglia as indicated by decreased zn-12 expression. Also, a lower transcription level of neurogenic genes, ngn1 and neuroD, is observed in neurons. Our data suggest that cadmium-induced neurotoxicity can be caused by impaired neurogenesis, resulting in markedly reduced neuronal differentiation and axonogenesis

Defects of the Glycinergic Synapse in Zebrafish

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Ogino, Kazutoyo; Hirata, Hiromi

2016-01-01

Glycine mediates fast inhibitory synaptic transmission. Physiological importance of the glycinergic synapse is well established in the brainstem and the spinal cord. In humans, the loss of glycinergic function in the spinal cord and brainstem leads to hyperekplexia, which is characterized by an excess startle reflex to sudden acoustic or tactile stimulation. In addition, glycinergic synapses in this region are also involved in the regulation of respiration and locomotion, and in the nociceptive processing. The importance of the glycinergic synapse is conserved across vertebrate species. A teleost fish, the zebrafish, offers several advantages as a vertebrate model for research of glycinergic synapse. Mutagenesis screens in zebrafish have isolated two motor defective mutants that have pathogenic mutations in glycinergic synaptic transmission: bandoneon (beo) and shocked (sho). Beo mutants have a loss-of-function mutation of glycine receptor (GlyR) β-subunit b, alternatively, sho mutant is a glycinergic transporter 1 (GlyT1) defective mutant. These mutants are useful animal models for understanding of glycinergic synaptic transmission and for identification of novel therapeutic agents for human diseases arising from defect in glycinergic transmission, such as hyperekplexia or glycine encephalopathy. Recent advances in techniques for genome editing and for imaging and manipulating of a molecule or a physiological process make zebrafish more attractive model. In this review, we describe the glycinergic defective zebrafish mutants and the technical advances in both forward and reverse genetic approaches as well as in vivo visualization and manipulation approaches for the study of the glycinergic synapse in zebrafish. PMID:27445686

Mapping of brain lipid binding protein (Blbp) in the brain of adult zebrafish, co-expression with aromatase B and links with proliferation.

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Diotel, Nicolas; Vaillant, Colette; Kah, Olivier; Pellegrini, Elisabeth

2016-01-01

Adult fish exhibit a strong neurogenic capacity due to the persistence of radial glial cells. In zebrafish, radial glial cells display well-established markers such as the estrogen-synthesizing enzyme (AroB) and the brain lipid binding protein (Blbp), which is known to strongly bind omega-3 polyunsaturated fatty acids such as docosahexaenoic acid (DHA). While Blpb is mainly described in the telencephalon of adult zebrafish, its expression in the remaining regions of the brain is poorly documented. The present study was designed to further investigate Blbp expression in the brain, its co-expression with AroB, and its link with radial glial cells proliferation in zebrafish. We generated a complete and detailed mapping of Blbp expression in the whole brain and show its complete co-expression with AroB, except in some tectal and hypothalamic regions. By performing PCNA and Blbp immunohistochemistry on cyp19a1b-GFP (AroB-GFP) fish, we also demonstrated preferential Blbp expression in proliferative radial glial cells in almost all regions studied. To our knowledge, this is the first complete and detailed mapping of Blbp-expressing cells showing strong association between Blbp and radial glial cell proliferation in the adult brain of fish. Given that zebrafish is now recognized models for studying neurogenesis and brain repair, our data provide detailed characterization of Blbp in the entire brain and open up a broad field of research investigating the role of omega-3 polyunsaturated fatty acids in neural stem cell activity in fish. Copyright © 2015 Elsevier B.V. All rights reserved.

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.

Zebrafish Models of Prader-Willi Syndrome: Fast Track to Pharmacotherapeutics

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Emma D. Spikol

2016-03-01

Full Text Available Prader-Willi syndrome (PWS is a rare genetic neurodevelopmental disorder characterized by an insatiable appetite, leading to chronic overeating and obesity. Additional features include short stature, intellectual disability, behavioral problems and incomplete sexual development. Although significant progress has been made in understanding the genetic basis of PWS, the mechanisms underlying the pathogenesis of the disorder remain poorly understood. Treatment for PWS consists mainly of palliative therapies; curative therapies are sorely needed. Zebrafish, Danio rerio, represent a promising way forward for elucidating physiological problems such as obesity and identifying new pharmacotherapeutic options for PWS. Over the last decade, an increased appreciation for the highly conserved biology among vertebrates and the ability to perform high-throughput drug screening has seen an explosion in the use of zebrafish for disease modeling and drug discovery. Here, we review recent advances in developing zebrafish models of human disease. Aspects of zebrafish genetics and physiology that are relevant to PWS will be discussed, and the advantages and disadvantages of zebrafish models will be contrasted with current animal models for this syndrome. Finally, we will present a paradigm for drug screening in zebrafish that is potentially the fastest route for identifying and delivering curative pharmacotherapies to PWS patients.

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.

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

Science.gov (United States)

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

2016-10-07

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

Mutagenesis and phenotyping resources in zebrafish for studying development and human disease

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Varshney, Gaurav Kumar

2014-01-01

The zebrafish (Danio rerio) is an important model organism for studying development and human disease. The zebrafish has an excellent reference genome and the functions of hundreds of genes have been tested using both forward and reverse genetic approaches. Recent years have seen an increasing number of large-scale mutagenesis projects and the number of mutants or gene knockouts in zebrafish has increased rapidly, including for the first time conditional knockout technologies. In addition, targeted mutagenesis techniques such as zinc finger nucleases, transcription activator-like effector nucleases and clustered regularly interspaced short sequences (CRISPR) or CRISPR-associated (Cas), have all been shown to effectively target zebrafish genes as well as the first reported germline homologous recombination, further expanding the utility and power of zebrafish genetics. Given this explosion of mutagenesis resources, it is now possible to perform systematic, high-throughput phenotype analysis of all zebrafish gene knockouts. PMID:24162064

Adenosine as a signaling molecule in the retina: biochemical and developmental aspects

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ROBERTO PAES-DE-CARVALHO

2002-09-01

Full Text Available The nucleoside adenosine plays an important role as a neurotransmitter or neuromodulator in the central nervous system, including the retina. In the present paper we review compelling evidence showing that adenosine is a signaling molecule in the developing retina. In the chick retina, adenosine transporters are present since early stages of development before the appearance of adenosine A1 receptors modulating dopamine-dependent adenylate cyclase activity or A2 receptors that directly activate the enzyme. Experiments using retinal cell cultures revealed that adenosine is taken up by specific cell populations that when stimulated by depolarization or neurotransmitters such as dopamine or glutamate, release the nucleoside through calcium-dependent transporter-mediated mechanisms. The presence of adenosine in the extracellular medium and the long-term activation of adenosine receptors is able to regulate the survival of retinal neurons and blocks glutamate excitoxicity. Thus, adenosine besides working as a neurotransmitter or neuromodulator in the mature retina, is considered as an important signaling molecule during retinal development having important functions such as regulation of neuronal survival and differentiation.O nucleosídeo adenosina apresenta um importante papel como neurotransmissor ou neuromodulador no sistema nervoso central, inclusive na retina. Neste artigo apresentamos uma revisão das evidências que mostram que a adenosina é uma molécula sinalizadora na retina em desenvolvimento. Na retina de pinto, transportadores de adenosina estão presentes desde estágios precoces do desenvolvimento, antes do aparecimento dos receptores A1 que modulam a atividade adenilato ciclase dependente de dopamina ou dos receptores A2 que ativam diretamente a enzima. Experimentos usando culturas de células de retina revelaram que a adenosina é captada por populações celulares específicas que, quando estimuladas por despolarização ou por

Identification of endogenous flurophores in the layered retina

Science.gov (United States)

Xu, Gaixia; Chen, Danni; Sun, Yiwen; Qu, Junle; Lin, Ziyang; Ding, Zhihua; Niu, Hanben

2007-05-01

In this paper, we measured and analyzed the characteristic of endogenous fluorophores in porcine layered retina by using advanced fluorescence spectroscopy and microscopy imaging technology. It was found that there were obvious contrasts corresponding to the different layers of retina, which may be important for fundus disease diagnosis. The retinal pigment epithelium cells exhibited strong autofluorescence with as emission peak of 600+/-10nm when excited with 860-nm light. The emission peak of photoreceptors was at 652+/-5nm, and the emission peak of retinal vessels layer was weak and at 640~700nm, when excited with 488-nm light. Autofluorescence images of three layers of retina were obtained using the same setup. We concluded that the main endogenous fluorophore in PRE was lipofuscin and that in retinal vessels was porphyrin. What's more, the FMHW (full width at half. maximum) of retinal fluorescence spectrum was broad, which suggested that there wasn't only one endogenous fluorophores of tissues excited.

Evaluation of color preference in zebrafish for learning and memory.

Science.gov (United States)

Avdesh, Avdesh; Martin-Iverson, Mathew T; Mondal, Alinda; Chen, Mengqi; Askraba, Sreten; Morgan, Newman; Lardelli, Michael; Groth, David M; Verdile, Giuseppe; Martins, Ralph N

2012-01-01

There is growing interest in using zebrafish (Danio rerio) as a model of neurodegenerative disorders such as Alzheimer's disease. A zebrafish model of tauopathies has recently been developed and characterized in terms of presence of the pathological hallmarks (i.e., neurofibrillary tangles and cell death). However, it is also necessary to validate these models for function by assessing learning and memory. The majority of tools to assess memory and learning in animal models involve visual stimuli, including color preference. The color preference of zebrafish has received little attention. To validate zebrafish as a model for color-associated-learning and memory, it is necessary to evaluate its natural preferences or any pre-existing biases towards specific colors. In the present study, we have used four different colors (red, yellow, green, and blue) to test natural color preferences of the zebrafish using two procedures: Place preference and T-maze. Results from both experiments indicate a strong aversion toward blue color relative to all other colors (red, yellow, and green) when tested in combinations. No preferences or biases were found among reds, yellows, and greens in the place preference procedure. However, red and green were equally preferred and both were preferred over yellow by zebrafish in the T-maze procedure. The results from the present study show a strong aversion towards blue color compared to red, green, and yellow, with yellow being less preferred relative to red and green. The findings from this study may underpin any further designing of color-based learning and memory paradigms or experiments involving aversion, anxiety, or fear in the zebrafish.

Standard anatomical and visual space for the mouse retina: computational reconstruction and transformation of flattened retinae with the Retistruct package.

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David C Sterratt

Full Text Available The concept of topographic mapping is central to the understanding of the visual system at many levels, from the developmental to the computational. It is important to be able to relate different coordinate systems, e.g. maps of the visual field and maps of the retina. Retinal maps are frequently based on flat-mount preparations. These use dissection and relaxing cuts to render the quasi-spherical retina into a 2D preparation. The variable nature of relaxing cuts and associated tears limits quantitative cross-animal comparisons. We present an algorithm, "Retistruct," that reconstructs retinal flat-mounts by mapping them into a standard, spherical retinal space. This is achieved by: stitching the marked-up cuts of the flat-mount outline; dividing the stitched outline into a mesh whose vertices then are mapped onto a curtailed sphere; and finally moving the vertices so as to minimise a physically-inspired deformation energy function. Our validation studies indicate that the algorithm can estimate the position of a point on the intact adult retina to within 8° of arc (3.6% of nasotemporal axis. The coordinates in reconstructed retinae can be transformed to visuotopic coordinates. Retistruct is used to investigate the organisation of the adult mouse visual system. We orient the retina relative to the nictitating membrane and compare this to eye muscle insertions. To align the retinotopic and visuotopic coordinate systems in the mouse, we utilised the geometry of binocular vision. In standard retinal space, the composite decussation line for the uncrossed retinal projection is located 64° away from the retinal pole. Projecting anatomically defined uncrossed retinal projections into visual space gives binocular congruence if the optical axis of the mouse eye is oriented at 64° azimuth and 22° elevation, in concordance with previous results. Moreover, using these coordinates, the dorsoventral boundary for S-opsin expressing cones closely matches

Localization and Developmental Expression Patterns of CSPG in the RCS Rat Retina

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Li-Feng Chen

2011-05-01

Full Text Available Purpose: Investigate changes in chondroitin sulfate proteoglycan (CSPG distribution in Royal College of Surgeons (RCS rat retinae. Could CSPGs distribution act as a physical barrier to transplanted cell migration in degenerating retinae? Methods: CSPG expression was examined in RCS and Long-Evans rat retinae from birth to postnatal day 150 (PND150 using immunofluorescence and western-blots. Results: Both groups showed a rapid rise in CSPG expression on PND14, which peaked on PND21 before declining to lower levels by PND35. CSPG expression had risen again by PND90 and remained elevated for the duration of the study (PND150. However, from PND21, CSPG expression was significantly higher (p ≤ 0.05, n = 5 in Long-Evans rat retinae. CSPG-positive cells were localized in the ganglion cell layer (GCL and the photoreceptor outer segment debris zone (DZ; CSPG expression in the DZ was the main contributor to the higher expression in older animals for both groups. Conclusions: Increased expression of CSPGs in the DZ may act as a physical barrier following retinal cellular transplantation. CSPGs in the GCL is probably related to dendritic changes. CSPG accumulation in the older retinae suggests that aging influences the microenvironment in the retina, which may affect the efficacy of cell transplantation.

An autonomous circadian clock in the inner mouse retina regulated by dopamine and GABA.

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Guo-Xiang Ruan

2008-10-01

Full Text Available The influence of the mammalian retinal circadian clock on retinal physiology and function is widely recognized, yet the cellular elements and neural regulation of retinal circadian pacemaking remain unclear due to the challenge of long-term culture of adult mammalian retina and the lack of an ideal experimental measure of the retinal circadian clock. In the current study, we developed a protocol for long-term culture of intact mouse retinas, which allows retinal circadian rhythms to be monitored in real time as luminescence rhythms from a PERIOD2::LUCIFERASE (PER2::LUC clock gene reporter. With this in vitro assay, we studied the characteristics and location within the retina of circadian PER2::LUC rhythms, the influence of major retinal neurotransmitters, and the resetting of the retinal circadian clock by light. Retinal PER2::LUC rhythms were routinely measured from whole-mount retinal explants for 10 d and for up to 30 d. Imaging of vertical retinal slices demonstrated that the rhythmic luminescence signals were concentrated in the inner nuclear layer. Interruption of cell communication via the major neurotransmitter systems of photoreceptors and ganglion cells (melatonin and glutamate and the inner nuclear layer (dopamine, acetylcholine, GABA, glycine, and glutamate did not disrupt generation of retinal circadian PER2::LUC rhythms, nor did interruption of intercellular communication through sodium-dependent action potentials or connexin 36 (cx36-containing gap junctions, indicating that PER2::LUC rhythms generation in the inner nuclear layer is likely cell autonomous. However, dopamine, acting through D1 receptors, and GABA, acting through membrane hyperpolarization and casein kinase, set the phase and amplitude of retinal PER2::LUC rhythms, respectively. Light pulses reset the phase of the in vitro retinal oscillator and dopamine D1 receptor antagonists attenuated these phase shifts. Thus, dopamine and GABA act at the molecular level of PER

Bcl-2 expression during the development and degeneration of RCS rat retinae.

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Sharma, R K

2001-12-14

In various hereditary retinal degenerations, including that in Royal College of Surgeons (RCS) rats, the photoreceptors ultimately die by apoptosis. Bcl-2 is one of the genes, which regulates apoptosis and is thought to promote survival of cells. This study has investigated the developmental expression of Bcl-2 in RCS rat, which is a well-studied animal model for hereditary retinal degeneration. An antibody against Bcl-2 was used for its immunohistochemical localization in dystrophic RCS rat retinae from postnatal (PN) days 4, 7, 13, 35, 45, 70, 202 and 14 months. Results were compared with Bcl-2 localization in congenic non-dystrophic rats from PN 4, 7, 13, 44, 202 and 14 months. Bcl-2 immunoreactivity in non-dystrophic retinae was already present in PN 4 retinae in the nerve fiber layer (presumably in the endfeet of immature Müller cells) and in the proximal parts of certain radially aligned neuroepithelial cells/immature Müller cell radial processes. With increasing age the immunoreactivity in relatively more mature Müller cell radial processes spread distally towards the outer retina and between PN 13 and 44 it reached the adult distribution. No cell bodies in the ganglion cell layer were found to be immunoreactive. Expression of Bcl-2 immunoreactivity in dystrophic RCS rat retinae closely resembled that of non-dystrophic retinae. No immunoreactivity was seen in photoreceptors or retinal pigment epithelium in dystrophic or non-dystrophic retinae. In conclusion, Bcl-2 expression is not altered, either in terms of its chronology or the cell type expressing it, during retinal degeneration in RCS rats.

Evaluation of MWNT toxic effects on daphnia and zebrafish embryos

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Olasagasti, Maider; Rainieri, Sandra [AZTI-TECNALIA, Parque Tecnologico de Bizkaia 609, 48160 Derio (Spain)], E-mail: srainieri@azti.es; Alvarez, Noelia; Vera, Carolina [INASMET-TECNALIA, Mikeletegi pasealekua, 2, Parque Tecnologico, 20009 San Sebastian (Spain)

2009-05-01

Organisms of daphnia (Daphnia magna) and zebrafish (Danio rerio) embryos were exposed to a range of different concentrations of COOH-functionalized MWCNT suspended in an aqueous solution of Tween 20. Immobilization of daphnia and growth retardation, inhibition and malformation of zebrafish embryos were the endpoints tested after 24 and 48 hours. Immobilization of daphnia could be observed from 3 to 16 ppm and an increasing mortality of zebrafish embryo was detected at all the concentration tested. To identify more subtle toxic effects, we took advantage of the extensive information available on the zebrafish genome and monitored by RT-PCR the expression patterns of different zebrafish genes that could act as toxicity bio-markers. At some of the concentrations tested, changes in the expression profiles of the genes examined were detected. Our results suggest that MWCNT could potentially represent a risk to human health and environment, therefore a wider range of concentrations and further testing of this molecules should be carried out to define possible limitations in their use.

Pharmacological analyses of learning and memory in zebrafish (Danio rerio).

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Bailey, Jordan M; Oliveri, Anthony N; Levin, Edward D

2015-12-01

Over the last decade, zebrafish (Danio rerio) have become valuable as a complementary model in behavioral pharmacology, opening a new avenue for understanding the relationships between drug action and behavior. This species offers a useful intermediate approach bridging the gap between in vitro studies and traditional mammalian models. Zebrafish offer great advantages of economy compared to their rodent counterparts, their complex brains and behavioral repertoire offer great translational potential relative to in vitro models. The development and validation of a variety of tests to measure behavior, including cognition, in zebrafish have set the stage for the use of this animal for behavioral pharmacology studies. This has led to research into the basic mechanisms of cognitive function as well as screening for potential cognition-improving drug therapies, among other lines of research. As with all models, zebrafish have limitations, which span pharmacokinetic challenges to difficulties quantifying behavior. The use, efficacy and limitations associated with a zebrafish model of cognitive function are discussed in this review, within the context of behavioral pharmacology. Copyright © 2015 Elsevier Inc. All rights reserved.

Protein changes in the retina following experimental retinal detachment in rabbits

DEFF Research Database (Denmark)

Mandal, Nakul; Lewis, Geoffrey P.; Fisher, Steven K.

2011-01-01

Retinal detachment leads to the widespread cellular remodeling of the retina. The purpose of this study was to identify protein changes that accompany these cellular alterations by comparing the proteomic profiles of sham and experimentally detached rabbit retina. Elucidation of the proteins most...

Genetic and immunohistochemical analysis of HSPA5 in mouse and human retinas.

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Chintalapudi, Sumana R; Wang, XiaoFei; Li, Huiling; Lau, Yin H Chan; Williams, Robert W; Jablonski, Monica M

2016-01-01

Photoreceptor degenerative diseases are among the leading causes of vision loss. Although the causative genetic mutations are often known, mechanisms leading to photoreceptor degeneration remain poorly defined. We have previously demonstrated that the photoreceptor membrane-associated protein XAP-1 antigen is a product of the HSPA5 gene. In this study, we used systems genetic methods, statistical modeling, and immunostaining to identify and analyze candidate genes that modulate Hspa5 expression in the retina. Quantitative trait locus (QTL) mapping was used to map the genomic region that regulates Hspa5 in the cross between C57BL/6J X DBA/2J mice (BXD) genetic reference panel. The stepwise refinement of candidate genes was based on expression QTL mapping, gene expression correlation analyses (direct and partial), and analysis of regional sequence variants. The subcellular localization of candidate proteins and HSPA5 in mouse and human retinas was evaluated by immunohistochemistry. Differences in the localization of extracellular HSPA5 were assessed between healthy human donor and atrophic age-related macular degeneration (AMD) donor eyes. In the eyes of healthy mice, extracellular HSPA5 was confined to the area around the cone photoreceptor outer segments. Mapping variation in Hspa5 mRNA expression levels in the retina revealed a statistically significant trans -acting expression QTL (eQTL) on Chromosome 2 (Chr 2) and a suggestive locus on Chr 15. Sulf2 on Chr 2 was the strongest candidate gene based on partial correlation analysis, Pearson correlation with Hspa5 , expression levels in the retina, a missense variant in exon 14, and its reported function in the extracellular matrix and interphotoreceptor matrix. SULF2 is localized to the rod and cone photoreceptors in both human and mouse retinas. In human retinas with no pathology, extracellular HSPA5 was localized around many cones within the macular area. In contrast, fewer HSPA5-immunopositive cones were

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...

Variations in dysfunction of sister chromatid cohesion in esco2 mutant zebrafish reflect the phenotypic diversity of Roberts syndrome

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Stefanie M. Percival

2015-08-01

Full Text Available Mutations in ESCO2, one of two establishment of cohesion factors necessary for proper sister chromatid cohesion (SCC, cause a spectrum of developmental defects in the autosomal-recessive disorder Roberts syndrome (RBS, warranting in vivo analysis of the consequence of cohesion dysfunction. Through a genetic screen in zebrafish targeting embryonic-lethal mutants that have increased genomic instability, we have identified an esco2 mutant zebrafish. Utilizing the natural transparency of zebrafish embryos, we have developed a novel technique to observe chromosome dynamics within a single cell during mitosis in a live vertebrate embryo. Within esco2 mutant embryos, we observed premature chromatid separation, a unique chromosome scattering, prolonged mitotic delay, and genomic instability in the form of anaphase bridges and micronuclei formation. Cytogenetic studies indicated complete chromatid separation and high levels of aneuploidy within mutant embryos. Amongst aneuploid spreads, we predominantly observed decreases in chromosome number, suggesting that either cells with micronuclei or micronuclei themselves are eliminated. We also demonstrated that the genomic instability leads to p53-dependent neural tube apoptosis. Surprisingly, although many cells required Esco2 to establish cohesion, 10-20% of cells had only weakened cohesion in the absence of Esco2, suggesting that compensatory cohesion mechanisms exist in these cells that undergo a normal mitotic division. These studies provide a unique in vivo vertebrate view of the mitotic defects and consequences of cohesion establishment loss, and they provide a compensation-based model to explain the RBS phenotypes.

TOXICITY EVALUATION OF NEW ENGINEERED NANOMATERIALS IN ZEBRAFISH

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Maria Violetta Brundo

2016-04-01

Full Text Available The effect of the nanoparticles on the marine organisms, depends on their size, chemical composition, surface structure, solubility and shape.In order to take advantage from their activity, preserving the surrounding environment from a possible pollution, we are trying to trap the nanoparticles into new nanomaterials. The nanomaterials tested were synthesized proposing a ground-breaking approach by an upside-down vision of the Au/TiO2nano-system to avoid the release of nanoparticles. The system was synthesized by wrapping Au nanoparticles with a thin layer of TiO2. The non-toxicity of the nano-system was established by testing the effect of the material on zebrafish larvae. Danio rerio o zebrafish was considered a excellent model for the environmental biomonitoring of aquatic environments and the Zebrafish Embryo Toxicity Test is considered an alternative method of animal test. For this reason zebrafish larvae were exposed to different concentrations of nanoparticles of TiO2 and Au and new nanomaterials. As biomarkers of exposure, we evaluated the expression of metallothioneins by immunohistochemistry analysis and western blotting analysis also. The results obtained by toxicity test showed that neither mortality as well as sublethal effects were induced by the different nanomaterials and nanoparticles tested. Only zebrafish larvae exposed to free Au nanoparticles showed a different response to anti-MT antibody. In fact, the immunolocalization analysis highlighted an increase of the metallothioneins synthesis.

EVALUATION OF INHOMOGENEITIES IN HISTOLOGICAL STRUCTURES (CARTILAGE, RETINA

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Lutz Muche

2011-05-01

Full Text Available This paper investigates histological tissues by means of image analysis and spatial statistics. For the quantification of cell frequencies and accumulations two statistical characteristics, intensity function and cluster density, are suggested. The samples are histological sections of human articular cartilage and human retina considered in view of changes during the ageing process. The articular cartilage is characterized by continuous changes of both functions, the cell intensity as well as the clusterization. In contrast, the retina is a trilaminar structure formed in the early embryonic stage without changes by ageing.

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

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

2018-05-01

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

Sulforaphane protects rodent retinas against ischemia-reperfusion injury through the activation of the Nrf2/HO-1 antioxidant pathway.

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Hong Pan

Full Text Available Retinal ischemia-reperfusion (I/R injury induces oxidative stress, leukocyte infiltration, and neuronal cell death. Sulforaphane (SF, which can be obtained in cruciferous vegetables such as broccoli, exerts protective effects in response to oxidative stress in various tissues. These effects can be initiated through nuclear factor E2-related factor 2 (Nrf2-mediated induction of heme oxygenase-1 (HO-1. This investigation was designed to elucidate the neural protective mechanisms of SF in the retinal I/R rat model. Animals were intraperitoneally (i.p. injected with SF (12.5 mg/kg or vehicle (corn oil once a day for 7 consecutive days. Then, retinal I/R was made by elevating the intraocular pressure (IOP to 130 mmHg for 1 h. To determine if HO-1 was involved in the Nrf2 antioxidant pathway, rats were subjected to protoporphyrin IX zinc (II (ZnPP, 30 mg/kg, i.p. treatments at 24 h before retinal ischemia. The neuroprotective effects of SF were assessed by determining the morphology of the retina, counting the infiltrating inflammatory cells and the surviving retinal ganglion cells (RGCs and amacrine cells, and measuring apoptosis in the retinal layers. The expression of Nrf2 and HO-1 was studied by immunofluorescence analysis and western blotting. I/R induced a marked increase of ROS generation, caused pronounced inflammation, increased the apoptosis of RGCs and amacrine cells and caused the thinning of the inner retinal layer (IRL, and these effects were diminished or abolished by SF pretreatment. Meanwhile, SF pretreatment significantly elevated the nuclear accumulation of Nrf2 and the level of HO-1 expression in the I/R retinas; however, ZnPP reversed the protective effects of SF on I/R retinas. Together, we offer direct evidence that SF had protective effects on I/R retinas, which could be attributed, at least in part, to the activation of the Nrf2/HO-1 antioxidant pathway.

Optic nerve signals in a neuromorphic chip I: Outer and inner retina models.

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Zaghloul, Kareem A; Boahen, Kwabena

2004-04-01

We present a novel model for the mammalian retina and analyze its behavior. Our outer retina model performs bandpass spatiotemporal filtering. It is comprised of two reciprocally connected resistive grids that model the cone and horizontal cell syncytia. We show analytically that its sensitivity is proportional to the space-constant-ratio of the two grids while its half-max response is set by the local average intensity. Thus, this outer retina model realizes luminance adaptation. Our inner retina model performs high-pass temporal filtering. It features slow negative feedback whose strength is modulated by a locally computed measure of temporal contrast, modeling two kinds of amacrine cells, one narrow-field, the other wide-field. We show analytically that, when the input is spectrally pure, the corner-frequency tracks the input frequency. But when the input is broadband, the corner frequency is proportional to contrast. Thus, this inner retina model realizes temporal frequency adaptation as well as contrast gain control. We present CMOS circuit designs for our retina model in this paper as well. Experimental measurements from the fabricated chip, and validation of our analytical results, are presented in the companion paper [Zaghloul and Boahen (2004)].

Thyroid Hormone Signaling in the Mouse Retina.

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Patrick Arbogast

Full Text Available Thyroid hormone is a crucial regulator of gene expression in the developing and adult retina. Here we sought to map sites of thyroid hormone signaling at the cellular level using the transgenic FINDT3 reporter mouse model in which neurons express β-galactosidase (β-gal under the control of a hybrid Gal4-TRα receptor when triiodothyronine (T3 and cofactors of thyroid receptor signaling are present. In the adult retina, nearly all neurons of the ganglion cell layer (GCL, ganglion cells and displaced amacrine cells showed strong β-gal labeling. In the inner nuclear layer (INL, a minority of glycineric and GABAergic amacrine cells showed β-gal labeling, whereas the majority of amacrine cells were unlabeled. At the level of amacrine types, β-gal labeling was found in a large proportion of the glycinergic AII amacrines, but only in a small proportion of the cholinergic/GABAergic 'starburst' amacrines. At postnatal day 10, there also was a high density of strongly β-gal-labeled neurons in the GCL, but only few amacrine cells were labeled in the INL. There was no labeling of bipolar cells, horizontal cells and Müller glia cells at both stages. Most surprisingly, the photoreceptor somata in the outer nuclear layer also showed no β-gal label, although thyroid hormone is known to control cone opsin expression. This is the first record of thyroid hormone signaling in the inner retina of an adult mammal. We hypothesize that T3 levels in photoreceptors are below the detection threshold of the reporter system. The topographical distribution of β-gal-positive cells in the GCL follows the overall neuron distribution in that layer, with more T3-signaling cells in the ventral than the dorsal half-retina.

A bioenergetic model for zebrafish Danio rerio (Hamilton)

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Chizinski, C.J.; Sharma, Bibek; Pope, K.L.; Patino, R.

2008-01-01

A bioenergetics model was developed from observed consumption, respiration and growth rates for zebrafish Danio rerio across a range (18-32?? C) of water temperatures, and evaluated with a 50 day laboratory trial at 28?? C. No significant bias in variable estimates was found during the validation trial; namely, predicted zebrafish mass generally agreed with observed mass. ?? 2008 The Authors.

Neural Responses to Injury: Prevention, Protection and Repair; Volume 7: Role Growth Factors and Cell Signaling in the Response of Brain and Retina to Injury

National Research Council Canada - National Science Library

Bazan, Nicolas

1996-01-01

...: Prevention, Protection, and Repair, Subproject: Role of Growth Factors and Cell Signaling in the Response of Brain and Retina to Injury, are as follows: Species Rat(Albino Wistar), Number Allowed...

Zebrafish models of cardiovascular diseases and their applications in herbal medicine research.

Science.gov (United States)

Seto, Sai-Wang; Kiat, Hosen; Lee, Simon M Y; Bensoussan, Alan; Sun, Yu-Ting; Hoi, Maggie P M; Chang, Dennis

2015-12-05

The zebrafish (Danio rerio) has recently become a powerful animal model for cardiovascular research and drug discovery due to its ease of maintenance, genetic manipulability and ability for high-throughput screening. Recent advances in imaging techniques and generation of transgenic zebrafish have greatly facilitated in vivo analysis of cellular events of cardiovascular development and pathogenesis. More importantly, recent studies have demonstrated the functional similarity of drug metabolism systems between zebrafish and humans, highlighting the clinical relevance of employing zebrafish in identifying lead compounds in Chinese herbal medicine with potential beneficial cardiovascular effects. This paper seeks to summarise the scope of zebrafish models employed in cardiovascular studies and the application of these research models in Chinese herbal medicine to date. Crown Copyright © 2015. Published by Elsevier B.V. All rights reserved.

In-silico experiments of zebrafish behaviour: modeling swimming in three dimensions

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Mwaffo, Violet; Butail, Sachit; Porfiri, Maurizio

2017-01-01

Zebrafish is fast becoming a species of choice in biomedical research for the investigation of functional and dysfunctional processes coupled with their genetic and pharmacological modulation. As with mammals, experimentation with zebrafish constitutes a complicated ethical issue that calls for the exploration of alternative testing methods to reduce the number of subjects, refine experimental designs, and replace live animals. Inspired by the demonstrated advantages of computational studies in other life science domains, we establish an authentic data-driven modelling framework to simulate zebrafish swimming in three dimensions. The model encapsulates burst-and-coast swimming style, speed modulation, and wall interaction, laying the foundations for in-silico experiments of zebrafish behaviour. Through computational studies, we demonstrate the ability of the model to replicate common ethological observables such as speed and spatial preference, and anticipate experimental observations on the correlation between tank dimensions on zebrafish behaviour. Reaching to other experimental paradigms, our framework is expected to contribute to a reduction in animal use and suffering.

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

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Ricardo J Figueroa

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

Modeling and Simulation of Microelectrode-Retina Interactions

Energy Technology Data Exchange (ETDEWEB)

Beckerman, M

2002-11-30

The goal of the retinal prosthesis project is the development of an implantable microelectrode array that can be used to supply visually-driven electrical input to cells in the retina, bypassing nonfunctional rod and cone cells, thereby restoring vision to blind individuals. This goal will be achieved through the study of the fundamentals of electrical engineering, vision research, and biomedical engineering with the aim of acquiring the knowledge needed to engineer a high-density microelectrode-tissue hybrid sensor that will restore vision to millions of blind persons. The modeling and simulation task within this project is intended to address the question how best to stimulate, and communicate with, cells in the retina using implanted microelectrodes.

The Hypocretin/Orexin Neuronal Networks in Zebrafish.

Science.gov (United States)

Elbaz, Idan; Levitas-Djerbi, Talia; Appelbaum, Lior

2017-01-01

The hypothalamic Hypocretin/Orexin (Hcrt) neurons secrete two Hcrt neuropeptides. These neurons and peptides play a major role in the regulation of feeding, sleep wake cycle, reward-seeking, addiction, and stress. Loss of Hcrt neurons causes the sleep disorder narcolepsy. The zebrafish has become an attractive model to study the Hcrt neuronal network because it is a transparent vertebrate that enables simple genetic manipulation, imaging of the structure and function of neuronal circuits in live animals, and high-throughput monitoring of behavioral performance during both day and night. The zebrafish Hcrt network comprises ~16-60 neurons, which similar to mammals, are located in the hypothalamus and widely innervate the brain and spinal cord, and regulate various fundamental behaviors such as feeding, sleep, and wakefulness. Here we review how the zebrafish contributes to the study of the Hcrt neuronal system molecularly, anatomically, physiologically, and pathologically.

Expression of sall4 in taste buds of zebrafish.

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Jackson, Robyn; Braubach, Oliver R; Bilkey, Jessica; Zhang, Jing; Akimenko, Marie-Andrée; Fine, Alan; Croll, Roger P; Jonz, Michael G

2013-07-01

We characterized the expression of sall4, a gene encoding a zinc finger transcription factor involved in the maintenance of embryonic stem cells, in taste buds of zebrafish (Danio rerio). Using an enhancer trap line (ET5), we detected enhanced green fluorescent protein (EGFP) in developing and adult transgenic zebrafish in regions containing taste buds: the lips, branchial arches, and the nasal and maxillary barbels. Localization of EGFP to taste cells of the branchial arches and lips was confirmed by co-immunolabeling with antibodies against calretinin and serotonin, and a zebrafish-derived neuronal marker (zn-12). Transgenic insertion of the ET construct into the zebrafish genome was evaluated and mapped to chromosome 23 in proximity (i.e. 23 kb) to the sall4 gene. In situ hybridization and expression analysis between 24 and 96 h post-fertilization (hpf) demonstrated that transgenic egfp expression in ET5 zebrafish was correlated with the spatial and temporal pattern of expression of sall4 in the wild-type. Expression was first observed in the central nervous system and branchial arches at 24 hpf. At 48 hpf, sall4 and egfp expression was observed in taste bud primordia surrounding the mouth and branchial arches. At 72 and 96 hpf, expression was detected in the upper and lower lips and branchial arches. Double fluorescence in situ hybridization at 3 and 10 dpf confirmed colocalization of sall4 and egfp in the lips and branchial arches. These studies reveal sall4 expression in chemosensory cells and implicate this transcription factor in the development and renewal of taste epithelia in zebrafish. Copyright © 2013 Wiley Periodicals, Inc.

Zebrafish as a Model for the Study of Human Myeloid Malignancies

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Jeng-Wei Lu

2015-01-01

Full Text Available Myeloid malignancies are heterogeneous disorders characterized by uncontrolled proliferation or/and blockage of differentiation of myeloid progenitor cells. Although a substantial number of gene alterations have been identified, the mechanism by which these abnormalities interact has yet to be elucidated. Over the past decades, zebrafish have become an important model organism, especially in biomedical research. Several zebrafish models have been developed to recapitulate the characteristics of specific myeloid malignancies that provide novel insight into the pathogenesis of these diseases and allow the evaluation of novel small molecule drugs. This report will focus on illustrative examples of applications of zebrafish models, including transgenesis, zebrafish xenograft models, and cell transplantation approaches, to the study of human myeloid malignancies.

Intrinsic and extrinsic innervation of the heart in zebrafish (Danio rerio).

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Stoyek, Matthew R; Croll, Roger P; Smith, Frank M

2015-08-01

In the vertebrate heart the intracardiac nervous system is the final common pathway for autonomic control of cardiac output, but the neuroanatomy of this system is not well understood. In this study we investigated the innervation of the heart in a model vertebrate, the zebrafish. We used antibodies against acetylated tubulin, human neuronal protein C/D, choline acetyltransferase, tyrosine hydroxylase, neuronal nitric oxide synthase, and vasoactive intestinal polypeptide to visualize neural elements and their neurotransmitter content. Most neurons were located at the venous pole in a plexus around the sinoatrial valve; mean total number of cells was 197 ± 23, and 92% were choline acetyltransferase positive, implying a cholinergic role. The plexus contained cholinergic, adrenergic, and nitrergic axons and vasoactive intestinal polypeptide-positive terminals, some innervating somata. Putative pacemaker cells near the plexus showed immunoreactivity for hyperpolarization-activated cyclic nucleotide-gated channel 4 (HCN4) and the transcription factor Islet-1 (Isl1). The neurotracer neurobiotin showed that extrinsic axons from the left and right vagosympathetic trunks innervated the sinoatrial plexus proximal to their entry into the heart; some extrinsic axons from each trunk also projected into the medial dorsal plexus region. Extrinsic axons also innervated the atrial and ventricular walls. An extracardiac nerve trunk innervated the bulbus arteriosus and entered the arterial pole of the heart to innervate the proximal ventricle. We have shown that the intracardiac nervous system in the zebrafish is anatomically and neurochemically complex, providing a substrate for autonomic control of cardiac effectors in all chambers. © 2015 Wiley Periodicals, Inc.

Distribution of photon absorption rates across the rat retina.

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Williams, T P; Webbers, J P; Giordano, L; Henderson, R P

1998-04-15

1. An investigation into the distribution of light intensity across the rat retina was carried out on excised, intact rat eyes exposed to Ganzfeld illumination from a helium-neon laser (543 nm). 2. Some of the light entering the eyes exits through the sclera where its intensity can be monitored with an optical 'pick-up' that samples the intensity coming from a small region of external sclera and underlying retina. The spatial resolution of the pick-up is such that it samples light that has passed through ca 2 % of the rods in the rat eye. 3. Some of the laser light is absorbed by the rod pigment, rhodopsin, which gradually bleaches. Bleaching in the retina, in turn, causes an exponential increase in intensity emanating from the sclera. By monitoring this intensity increase, we are able to measure two important parameters in a single bleaching run: the local rhodopsin concentration and the local intensity falling on the rods. 4. With an ocular transmission photometer, we have measured both the local intensity and the local rhodopsin concentration across wide regions of rat retina. Both pigmented and albino rats were studied. 5. The distributions of rhodopsin and intensity were both nearly uniform; consequently, the product, (rhodopsin concentration) x (intensity), was similarly nearly equal across the retina. This means that the initial rate of photon absorption is about the same at all retinal locations. 6. Interpreted in terms of photostasis (the regulation of daily photon catch), this means that the rate of photon absorption is about the same in each rod, viz. 14 400 photons absorbed per rod per second. Since this rate of absorption is sufficient to saturate the rod, one possible purpose of photostasis is to maintain the rod system in a saturated state during daylight hours.

Cyp1a reporter zebrafish reveals target tissues for dioxin

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Kim, Kun-Hee [Department of Biomedical Sciences, Chonnam National University Medical School, Gwangju (Korea, Republic of); Department of Microbiology, Chonnam National University Medical School, Gwangju (Korea, Republic of); Park, Hye-Jeong [Department of Biomedical Sciences, Chonnam National University Medical School, Gwangju (Korea, Republic of); Kim, Jin Hee [Department of Biomedical Sciences, Chonnam National University Medical School, Gwangju (Korea, Republic of); Department of Microbiology, Chonnam National University Medical School, Gwangju (Korea, Republic of); Kim, Suhyun [Graduate School of Medicine, Korea University, Ansan (Korea, Republic of); Williams, Darren R. [New Drug Targets Laboratory, School of Life Sciences, Gwangju Institute of Science and Technology, Gwangju (Korea, Republic of); Kim, Myeong-Kyu [Department of Neurology, Chonnam National University Medical School, Gwangju (Korea, Republic of); Jung, Young Do [Department of Biochemistry, Chonnam National University Medical School, Gwangju (Korea, Republic of); Teraoka, Hiroki [School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu (Japan); Park, Hae-Chul [Graduate School of Medicine, Korea University, Ansan (Korea, Republic of); Choy, Hyon E., E-mail: hyonchoy@chonnam.ac.kr [Department of Microbiology, Chonnam National University Medical School, Gwangju (Korea, Republic of); Shin, Boo Ahn, E-mail: bashin@chonnam.ac.kr [Department of Microbiology, Chonnam National University Medical School, Gwangju (Korea, Republic of); Choi, Seok-Yong, E-mail: zebrafish@chonnam.ac.kr [Department of Biomedical Sciences, Chonnam National University Medical School, Gwangju (Korea, Republic of); School of Biological Sciences and Technology, Chonnam National University, Gwangju (Korea, Republic of)

2013-06-15

Highlights: •2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most toxic anthropogenic substance ever identified. •Transgenic cyp1a reporter zebrafish reveals target tissues for TCDD. •The retinal bipolar cells, otic vesicle, lateral line, pancreas, cloaca and pectoral fin bud are novel targets in zebrafish for TCDD. •Our findings will further understanding of human health risks by TCDD. -- Abstract: 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the unintentional byproduct of various industrial processes, is classified as human carcinogen and could disrupt reproductive, developmental and endocrine systems. Induction of cyp1a1 is used as an indicator of TCDD exposure. We sought to determine tissues that are vulnerable to TCDD toxicity using a transgenic zebrafish (Danio rerio) model. We inserted a nuclear enhanced green fluorescent protein gene (EGFP) into the start codon of a zebrafish cyp1a gene in a fosmid clone using DNA recombineering. The resulting recombineered fosmid was then used to generate cyp1a reporter zebrafish, embryos of which were exposed to TCDD. Expression pattern of EGFP in the reporter zebrafish mirrored that of endogenous cyp1a mRNA. In addition, exposure of the embryos to TCDD at as low as 10 pM for 72 h, which does not elicit morphological abnormalities of embryos, markedly increased GFP expression. Furthermore, the reporter embryos responded to other AhR ligands as well. Exposure of the embryos to TCDD revealed previously reported (the cardiovascular system, liver, pancreas, kidney, swim bladder and skin) and unreported target tissues (retinal bipolar cells, otic vesicle, lateral line, cloaca and pectoral fin bud) for TCDD. Transgenic cyp1a reporter zebrafish we have developed can further understanding of ecotoxicological relevance and human health risks by TCDD. In addition, they could be used to identify agonists of AhR and antidotes to TCDD toxicity.

Influence of carbon nanotube length on toxicity to zebrafish embryos

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Cheng J

2012-07-01

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

The Adhesion Molecule-Characteristic HNK-1 Carbohydrate Contributes to Functional Recovery After Spinal Cord Injury in Adult Zebrafish.

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Ma, Liping; Shen, Hui-Fan; Shen, Yan-Qin; Schachner, Melitta

2017-07-01

The human natural killer cell antigen-1 (HNK-1) is functionally important in development, synaptic activity, and regeneration after injury in the nervous system of several mammalian species. It contains a sulfated glucuronic acid which is carried by neural adhesion molecules and expressed in nonmammalian species, including zebrafish, which, as opposed to mammals, spontaneously regenerate after injury in the adult. To evaluate HNK-1's role in recovery of function after spinal cord injury (SCI) of adult zebrafish, we assessed the effects of the two HNK-1 synthesizing enzymes, glucuronyl transferase and HNK-1 sulfotransferase. Expression of these two enzymes was increased at the messenger RNA (mRNA) level 11 days after injury in the brainstem nuclei that are capable of regrowth of severed axons, namely, the nucleus of medial longitudinal fascicle and intermediate reticular formation, but not at earlier time points after SCI. mRNA levels of glucuronyl transferase and sulfotransferase were increased in neurons, not only of these nuclei but also in the spinal cord caudal to the injury site at 11 days. Mauthner neurons which are not capable of regeneration did not show increased levels of enzyme mRNAs after injury. Reducing protein levels of the enzymes by application of anti-sense morpholinos resulted in reduction of locomotor recovery for glucuronyl transferase, but not for HNK-1 sulfotransferase. The combined results indicate that HNK-1 is upregulated in expression only in those neurons that are intrinsically capable of regeneration and contributes to regeneration after spinal cord injury in adult zebrafish in the absence of its sulfate moiety.

Molecular genetics of pituitary development in zebrafish.

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Pogoda, Hans-Martin; Hammerschmidt, Matthias

2007-08-01

The pituitary gland of vertebrates consists of two major parts, the neurohypophysis (NH) and the adenohypophysis (AH). As a central part of the hypothalamo-hypophyseal system (HHS), it constitutes a functional link between the nervous and the endocrine system to regulate basic body functions, such as growth, metabolism and reproduction. The development of the AH has been intensively studied in mouse, serving as a model for organogenesis and differential cell specification. However, given that the AH is a relatively recent evolutionary advance of the chordate phylum, it is also interesting to understand its development in lower chordate systems. In recent years, the zebrafish has emerged as a powerful lower vertebrate system for developmental studies, being amenable for large-scale genetic approaches, embryological manipulations, and in vivo imaging. Here, we present an overview of current knowledge of the mechanisms and genetic control of pituitary formation during zebrafish development. First, we describe the components of the zebrafish HHS, and the different pituitary cell types and hormones, followed by a description of the different steps of normal pituitary development. The central part of the review deals with the genes found to be essential for zebrafish AH development, accompanied by a description of the corresponding mutant phenotypes. Finally, we discuss future directions, with particular focus on evolutionary aspects, and some novel functional aspects with growing medical and social relevance.

A two-scale model for correlation between B cell VDJ usage in zebrafish

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Pan, Keyao; Deem, Michael

2011-03-01

The zebrafish (Danio rerio) is one of the model animals for study of immunology. The dynamics of the adaptive immune system in zebrafish is similar to that in higher animals. In this work, we built a two-scale model to simulate the dynamics of B cells in primary and secondary immune reactions in zebrafish and to explain the reported correlation between VDJ usage of B cell repertoires in distinct zebrafish. The first scale of the model consists of a generalized NK model to simulate the B cell maturation process in the 10-day primary immune response. The second scale uses a delay ordinary differential equation system to model the immune responses in the 6-month lifespan of zebrafish. The generalized NK model shows that mature B cells specific to one antigen mostly possess a single VDJ recombination. The probability that mature B cells in two zebrafish have the same VDJ recombination increases with the B cell population size or the B cell selection intensity and decreases with the B cell hypermutation rate. The ODE model shows a distribution of correlation in the VDJ usage of the B cell repertoires in two six-month-old zebrafish that is highly similar to that from experiment. This work presents a simple theory to explain the experimentally observed correlation in VDJ usage of distinct zebrafish B cell repertoires after an immune response.

Induced Pluripotent Stem Cell Therapies for Degenerative Disease of the Outer Retina: Disease Modeling and Cell Replacement.

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Di Foggia, Valentina; Makwana, Priyanka; Ali, Robin R; Sowden, Jane C

2016-06-01

Stem cell therapies are being explored as potential treatments for retinal disease. How to replace neurons in a degenerated retina presents a continued challenge for the regenerative medicine field that, if achieved, could restore sight. The major issues are: (i) the source and availability of donor cells for transplantation; (ii) the differentiation of stem cells into the required retinal cells; and (iii) the delivery, integration, functionality, and survival of new cells in the host neural network. This review considers the use of induced pluripotent stem cells (iPSC), currently under intense investigation, as a platform for cell transplantation therapy. Moreover, patient-specific iPSC are being developed for autologous cell transplantation and as a tool for modeling specific retinal diseases, testing gene therapies, and drug screening.

Characterization of Pax3 and Sox10 transgenic Xenopus laevis embryos as tools to study neural crest development.

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Alkobtawi, Mansour; Ray, Heather; Barriga, Elias H; Moreno, Mauricio; Kerney, Ryan; Monsoro-Burq, Anne-Helene; Saint-Jeannet, Jean-Pierre; Mayor, Roberto

2018-03-06

The neural crest is a multipotent population of cells that originates a variety of cell types. Many animal models are used to study neural crest induction, migration and differentiation, with amphibians and birds being the most widely used systems. A major technological advance to study neural crest development in mouse, chick and zebrafish has been the generation of transgenic animals in which neural crest specific enhancers/promoters drive the expression of either fluorescent proteins for use as lineage tracers, or modified genes for use in functional studies. Unfortunately, no such transgenic animals currently exist for the amphibians Xenopus laevis and tropicalis, key model systems for studying neural crest development. Here we describe the generation and characterization of two transgenic Xenopus laevis lines, Pax3-GFP and Sox10-GFP, in which GFP is expressed in the pre-migratory and migratory neural crest, respectively. We show that Pax3-GFP could be a powerful tool to study neural crest induction, whereas Sox10-GFP could be used in the study of neural crest migration in living embryos. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

ZebrafishMiner: an open source software for interactive evaluation of domain-specific fluorescence in zebrafish

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Reischl Markus

2017-09-01

Full Text Available High-throughput microscopy makes it possible to observe the morphology of zebrafish on large scale to quantify genetic, toxic or drug effects. The image acquisition is done by automated microscopy, images are evaluated automatically by image processing pipelines, tailored specifically to the requirements of the scientific question. The transfer of such algorithms to other projects, however, is complex due to missing guidelines and lack of mathematical or programming knowledge. In this work, we implement an image processing pipeline for automatic fluorescence quantification in user-defined domains of zebrafish embryos and larvae of different age. The pipeline is capable of detecting embryos and larvae in image stacks and quantifying domain activity. To make this protocol available to the community, we developed an open source software package called „ZebrafishMiner“ which guides the user through all steps of the processing pipeline and makes the algorithms available and easy to handle. We implemented all routines in an MATLAB-based graphical user interface (GUI that gives the user control over all image processing parameters. The software is shipped with a manual of 30 pages and three tutorial datasets, which guide the user through the manual step by step. It can be downloaded at https://sourceforge.net/projects/scixminer/.

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

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

2009-01-01

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

Genetic and immunohistochemical analysis of HSPA5 in mouse and human retinas

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Chintalapudi, Sumana R.; Wang, XiaoFei; Li, Huiling; Lau, Yin H. Chan; Williams, Robert W.; Jablonski, Monica M.

2016-01-01

Purpose Photoreceptor degenerative diseases are among the leading causes of vision loss. Although the causative genetic mutations are often known, mechanisms leading to photoreceptor degeneration remain poorly defined. We have previously demonstrated that the photoreceptor membrane-associated protein XAP-1 antigen is a product of the HSPA5 gene. In this study, we used systems genetic methods, statistical modeling, and immunostaining to identify and analyze candidate genes that modulate Hspa5 expression in the retina. Methods Quantitative trait locus (QTL) mapping was used to map the genomic region that regulates Hspa5 in the cross between C57BL/6J X DBA/2J mice (BXD) genetic reference panel. The stepwise refinement of candidate genes was based on expression QTL mapping, gene expression correlation analyses (direct and partial), and analysis of regional sequence variants. The subcellular localization of candidate proteins and HSPA5 in mouse and human retinas was evaluated by immunohistochemistry. Differences in the localization of extracellular HSPA5 were assessed between healthy human donor and atrophic age-related macular degeneration (AMD) donor eyes. Results In the eyes of healthy mice, extracellular HSPA5 was confined to the area around the cone photoreceptor outer segments. Mapping variation in Hspa5 mRNA expression levels in the retina revealed a statistically significant trans-acting expression QTL (eQTL) on Chromosome 2 (Chr 2) and a suggestive locus on Chr 15. Sulf2 on Chr 2 was the strongest candidate gene based on partial correlation analysis, Pearson correlation with Hspa5, expression levels in the retina, a missense variant in exon 14, and its reported function in the extracellular matrix and interphotoreceptor matrix. SULF2 is localized to the rod and cone photoreceptors in both human and mouse retinas. In human retinas with no pathology, extracellular HSPA5 was localized around many cones within the macular area. In contrast, fewer HSPA5

Using local chromatin structure to improve CRISPR/Cas9 efficiency in zebrafish.

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Chen, Yunru; Zeng, Shiyang; Hu, Ruikun; Wang, Xiangxiu; Huang, Weilai; Liu, Jiangfang; Wang, Luying; Liu, Guifen; Cao, Ying; Zhang, Yong

2017-01-01

Although the CRISPR/Cas9 has been successfully applied in zebrafish, considerable variations in efficiency have been observed for different gRNAs. The workload and cost of zebrafish mutant screening is largely dependent on the mutation rate of injected embryos; therefore, selecting more effective gRNAs is especially important for zebrafish mutant construction. Besides the sequence features, local chromatin structures may have effects on CRISPR/Cas9 efficiency, which remain largely unexplored. In the only related study in zebrafish, nucleosome organization was not found to have an effect on CRISPR/Cas9 efficiency, which is inconsistent with recent studies in vitro and in mammalian cell lines. To understand the effects of local chromatin structure on CRISPR/Cas9 efficiency in zebrafish, we first determined that CRISPR/Cas9 introduced genome editing mainly before the dome stage. Based on this observation, we reanalyzed our published nucleosome organization profiles and generated chromatin accessibility profiles in the 256-cell and dome stages using ATAC-seq technology. Our study demonstrated that chromatin accessibility showed positive correlation with CRISPR/Cas9 efficiency, but we did not observe a clear correlation between nucleosome organization and CRISPR/Cas9 efficiency. We constructed an online database for zebrafish gRNA selection based on local chromatin structure features that could prove beneficial to zebrafish homozygous mutant construction via CRISPR/Cas9.

Nutritional manipulation of primate retinas, III: Effects of lutein or zeaxanthin supplementation on adipose tissue and retina of xanthophyll-free monkeys.

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Johnson, Elizabeth J; Neuringer, Martha; Russell, Robert M; Schalch, Wolfgang; Snodderly, D Max

2005-02-01

Macular pigment (MP) is composed of the xanthophylls lutein (L) and zeaxanthin (Z) and may help to prevent age-related macular degeneration or retard its progression. In this study the effects of L or Z supplementation on carotenoid levels was examined in serum, adipose tissue, and retina in rhesus monkeys with no previous intake of xanthophylls. From birth to 7 to 16 years of age, 18 rhesus monkeys were fed semipurified diets containing all essential nutrients but no xanthophylls. Six were supplemented with pure L and 6 with pure Z at 3.9 micromol/kg per day for 24 to 101 weeks. At baseline and at 4- to 12-week intervals, carotenoids in adipose tissue were measured by HPLC. At study completion, carotenoids in serum and retina (central 4 mm, 8-mm annulus, and the periphery) were determined. Results were compared with data from control monkeys fed a standard laboratory diet. Monkeys fed xanthophyll-free diets had no L or Z in serum or tissues. After L or Z supplementation, serum and adipose tissue concentrations significantly increased in the supplemented groups. Both L and 3R,3'S-Z (RSZ or meso-Z, not present in the diet) were incorporated into retinas of monkeys supplemented with L, with RSZ present only in the macula (central 4 mm). All-trans Z, but no RSZ, accumulated in retinas of monkeys supplemented with Z. L is the precursor of RSZ, a major component of macular pigment. Xanthophyll-free monkeys can accumulate retinal xanthophylls and provide a valuable model for examining their uptake and conversion.

Frequency of Toxoplasma gondii in the retina in eye banks in Brazil.

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Costa, Deise F; Nascimento, Heloisa; Sutili, Aline; Nobrega, Fernando A J; Fowler, Flavio; Nobrega, Mario Junqueira; Garrido, Cristina; de Oliveira Dias, Janaina; Adán, Consuelo B D; Rizzo, Luiz Vicente; Silveira, Claudio; Belfort, Rubens; Commodaro, Alessandra G

2017-07-01

Ocular toxoplasmosis is the main cause of posterior uveitis worldwide frequently leading to vision loss. In Brazil, the seroprevalence of Toxoplasma gondii infection ranges from 50 to 80% depending of the region studied. The frequency of toxoplasmic retinal scar may reach 18% of the adults in the South of Brazil. Our goal was to determine the frequency of T. gondii DNA in retinas from eye banks from different regions in Brazil. A total of 162 eyes were obtained from eye banks in Manaus (n = 60), Sao Paulo (n = 60), Chapeco (n = 26), and Joinville (n = 16). The retinas were macroscopically analyzed and collected for DNA extraction. Real-time PCR (qPCR) was performed using the T. gondii B1 marker. By qPCR, a higher frequency of T. gondii DNA in the retinas from the eye bank of Joinville (25%) was found when compared to Manaus (5%). The retinas from Sao Paulo and Chapeco were qPCR negative. Clinical examination determined the retina lesions to be compatible with toxoplasmosis in the following frequencies: Joinville (62.5%), Manaus (10%), Sao Paulo (6.7%), and Chapeco (15.4%).

Some indications of structural damage in retina by heavy ion radiation

International Nuclear Information System (INIS)

Nelson, A.C.; Hayes, T.L.; Tobias, C.A.; Yang, T.C.

1981-01-01

At the Lawrence Berkeley Laboratory Bevalac Facility, iron nuclei were accelerated to an energy of 600 MeV/amu. The beam of iron thus obtained was used to irradiate living biological specimens in order to study possible microscopic tissue damage with the aid of SEM. The experiments involved total head irradiation of live rats which were subsequently returned to their cages to remain for 1 day and 30 days before further examination. After the 1 day and 30 day waits, both eyes were enucleated and placed in chemical fixative followed by ethanol dehydration and critical point drying. Retinas were carefully removed from the eye cups and loaded separately on aluminum stubs which were sputter coated. SEM of the 1 day and 30 day retinas revealed lesions which were not found at all in control retinas. The 1 day and 30 day retinas manifest regions where outer rod segments were missing or rearranged. A single energetic iron nucleus may be capable of generating a retinal lesion which becomes enlarged as biological processes intervene during the 1 day and 30 day waits. Being composed of highly specialized nerve cells, retinas cannot regenerate following irradiation which severely damages the rod cells. Thus one would expect the observed radiation induced retinal lesions to correspond to permanent tissue damage and possible loss of visual acuity in the intact animal

A theory of how active behavior stabilises neural activity: Neural gain modulation by closed-loop environmental feedback.

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Christopher L Buckley

2018-01-01

Full Text Available During active behaviours like running, swimming, whisking or sniffing, motor actions shape sensory input and sensory percepts guide future motor commands. Ongoing cycles of sensory and motor processing constitute a closed-loop feedback system which is central to motor control and, it has been argued, for perceptual processes. This closed-loop feedback is mediated by brainwide neural circuits but how the presence of feedback signals impacts on the dynamics and function of neurons is not well understood. Here we present a simple theory suggesting that closed-loop feedback between the brain/body/environment can modulate neural gain and, consequently, change endogenous neural fluctuations and responses to sensory input. We support this theory with modeling and data analysis in two vertebrate systems. First, in a model of rodent whisking we show that negative feedback mediated by whisking vibrissa can suppress coherent neural fluctuations and neural responses to sensory input in the barrel cortex. We argue this suppression provides an appealing account of a brain state transition (a marked change in global brain activity coincident with the onset of whisking in rodents. Moreover, this mechanism suggests a novel signal detection mechanism that selectively accentuates active, rather than passive, whisker touch signals. This mechanism is consistent with a predictive coding strategy that is sensitive to the consequences of motor actions rather than the difference between the predicted and actual sensory input. We further support the theory by re-analysing previously published two-photon data recorded in zebrafish larvae performing closed-loop optomotor behaviour in a virtual swim simulator. We show, as predicted by this theory, that the degree to which each cell contributes in linking sensory and motor signals well explains how much its neural fluctuations are suppressed by closed-loop optomotor behaviour. More generally we argue that our results

A theory of how active behavior stabilises neural activity: Neural gain modulation by closed-loop environmental feedback.

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Buckley, Christopher L; Toyoizumi, Taro

2018-01-01

During active behaviours like running, swimming, whisking or sniffing, motor actions shape sensory input and sensory percepts guide future motor commands. Ongoing cycles of sensory and motor processing constitute a closed-loop feedback system which is central to motor control and, it has been argued, for perceptual processes. This closed-loop feedback is mediated by brainwide neural circuits but how the presence of feedback signals impacts on the dynamics and function of neurons is not well understood. Here we present a simple theory suggesting that closed-loop feedback between the brain/body/environment can modulate neural gain and, consequently, change endogenous neural fluctuations and responses to sensory input. We support this theory with modeling and data analysis in two vertebrate systems. First, in a model of rodent whisking we show that negative feedback mediated by whisking vibrissa can suppress coherent neural fluctuations and neural responses to sensory input in the barrel cortex. We argue this suppression provides an appealing account of a brain state transition (a marked change in global brain activity) coincident with the onset of whisking in rodents. Moreover, this mechanism suggests a novel signal detection mechanism that selectively accentuates active, rather than passive, whisker touch signals. This mechanism is consistent with a predictive coding strategy that is sensitive to the consequences of motor actions rather than the difference between the predicted and actual sensory input. We further support the theory by re-analysing previously published two-photon data recorded in zebrafish larvae performing closed-loop optomotor behaviour in a virtual swim simulator. We show, as predicted by this theory, that the degree to which each cell contributes in linking sensory and motor signals well explains how much its neural fluctuations are suppressed by closed-loop optomotor behaviour. More generally we argue that our results demonstrate the dependence

Tributyltin and Zebrafish: Swimming in Dangerous Water

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Clemilson Berto-Júnior

2018-04-01

Full Text Available Zebrafish has been established as a reliable biological model with important insertion in academy (morphologic, biochemical, and pathophysiological studies and pharmaceutical industry (toxicology and drug development due to its molecular complexity and similar systems biology that recapitulate those from other organisms. Considering the toxicological aspects, many efforts using zebrafish models are being done in order to elucidate the effects of endocrine disruptors, and some of them are focused on tributyltin (TBT and its mechanism of action. TBT is an antifouling agent applied in ship’s hull that is constantly released into the water and absorbed by marine organisms, leading to bioaccumulation and biomagnification effects. Thus, several findings of malformations and changes in the normal biochemical and physiologic aspects of these marine animals have been related to TBT contamination. In the present review, we have compiled the most significant studies related to TBT effects in zebrafish, also taking into consideration the effects found in other study models.

Tributyltin and Zebrafish: Swimming in Dangerous Water

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Berto-Júnior, Clemilson; de Carvalho, Denise Pires; Soares, Paula; Miranda-Alves, Leandro

2018-01-01

Zebrafish has been established as a reliable biological model with important insertion in academy (morphologic, biochemical, and pathophysiological studies) and pharmaceutical industry (toxicology and drug development) due to its molecular complexity and similar systems biology that recapitulate those from other organisms. Considering the toxicological aspects, many efforts using zebrafish models are being done in order to elucidate the effects of endocrine disruptors, and some of them are focused on tributyltin (TBT) and its mechanism of action. TBT is an antifouling agent applied in ship’s hull that is constantly released into the water and absorbed by marine organisms, leading to bioaccumulation and biomagnification effects. Thus, several findings of malformations and changes in the normal biochemical and physiologic aspects of these marine animals have been related to TBT contamination. In the present review, we have compiled the most significant studies related to TBT effects in zebrafish, also taking into consideration the effects found in other study models. PMID:29692757

Disease modeling in genetic kidney diseases: zebrafish.

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Schenk, Heiko; Müller-Deile, Janina; Kinast, Mark; Schiffer, Mario

2017-07-01

Growing numbers of translational genomics studies are based on the highly efficient and versatile zebrafish (Danio rerio) vertebrate model. The increasing types of zebrafish models have improved our understanding of inherited kidney diseases, since they not only display pathophysiological changes but also give us the opportunity to develop and test novel treatment options in a high-throughput manner. New paradigms in inherited kidney diseases have been developed on the basis of the distinct genome conservation of approximately 70 % between zebrafish and humans in terms of existing gene orthologs. Several options are available to determine the functional role of a specific gene or gene sets. Permanent genome editing can be induced via complete gene knockout by using the CRISPR/Cas-system, among others, or via transient modification by using various morpholino techniques. Cross-species rescues succeeding knockdown techniques are employed to determine the functional significance of a target gene or a specific mutation. This article summarizes the current techniques and discusses their perspectives.

Use of zebrafish to study Shigella infection

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Duggan, Gina M.

2018-01-01

ABSTRACT Shigella is a leading cause of dysentery worldwide, responsible for up to 165 million cases of shigellosis each year. Shigella is also recognised as an exceptional model pathogen to study key issues in cell biology and innate immunity. Several infection models have been useful to explore Shigella biology; however, we still lack information regarding the events taking place during the Shigella infection process in vivo. Here, we discuss a selection of mechanistic insights recently gained from studying Shigella infection of zebrafish (Danio rerio), with a focus on cytoskeleton rearrangements and cellular immunity. We also discuss how infection of zebrafish can be used to investigate new concepts underlying infection control, including emergency granulopoiesis and the use of predatory bacteria to combat antimicrobial resistance. Collectively, these insights illustrate how Shigella infection of zebrafish can provide fundamental advances in our understanding of bacterial pathogenesis and vertebrate host defence. This information should also provide vital clues for the discovery of new therapeutic strategies against infectious disease in humans. PMID:29590642

Use of zebrafish to study Shigella infection

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Gina M. Duggan

2018-02-01

Full Text Available Shigella is a leading cause of dysentery worldwide, responsible for up to 165 million cases of shigellosis each year. Shigella is also recognised as an exceptional model pathogen to study key issues in cell biology and innate immunity. Several infection models have been useful to explore Shigella biology; however, we still lack information regarding the events taking place during the Shigella infection process in vivo. Here, we discuss a selection of mechanistic insights recently gained from studying Shigella infection of zebrafish (Danio rerio, with a focus on cytoskeleton rearrangements and cellular immunity. We also discuss how infection of zebrafish can be used to investigate new concepts underlying infection control, including emergency granulopoiesis and the use of predatory bacteria to combat antimicrobial resistance. Collectively, these insights illustrate how Shigella infection of zebrafish can provide fundamental advances in our understanding of bacterial pathogenesis and vertebrate host defence. This information should also provide vital clues for the discovery of new therapeutic strategies against infectious disease in humans.

Cloning and expression of a zebrafish SCN1B ortholog and identification of a species-specific splice variant

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Slat Emily A

2007-07-01

Full Text Available Abstract Background Voltage-gated Na+ channel β1 (Scn1b subunits are multi-functional proteins that play roles in current modulation, channel cell surface expression, cell adhesion, cell migration, and neurite outgrowth. We have shown previously that β1 modulates electrical excitability in vivo using a mouse model. Scn1b null mice exhibit spontaneous seizures and ataxia, slowed action potential conduction, decreased numbers of nodes of Ranvier in myelinated axons, alterations in nodal architecture, and differences in Na+ channel α subunit localization. The early death of these mice at postnatal day 19, however, make them a challenging model system to study. As a first step toward development of an alternative model to investigate the physiological roles of β1 subunits in vivo we cloned two β1-like subunit cDNAs from D. rerio. Results Two β1-like subunit mRNAs from zebrafish, scn1ba_tv1 and scn1ba_tv2, arise from alternative splicing of scn1ba. The deduced amino acid sequences of Scn1ba_tv1 and Scn1ba_tv2 are identical except for their C-terminal domains. The C-terminus of Scn1ba_tv1 contains a tyrosine residue similar to that found to be critical for ankyrin association and Na+ channel modulation in mammalian β1. In contrast, Scn1ba_tv2 contains a unique, species-specific C-terminal domain that does not contain a tyrosine. Immunohistochemical analysis shows that, while the expression patterns of Scn1ba_tv1 and Scn1ba_tv2 overlap in some areas of the brain, retina, spinal cord, and skeletal muscle, only Scn1ba_tv1 is expressed in optic nerve where its staining pattern suggests nodal expression. Both scn1ba splice forms modulate Na+ currents expressed by zebrafish scn8aa, resulting in shifts in channel gating mode, increased current amplitude, negative shifts in the voltage dependence of current activation and inactivation, and increases in the rate of recovery from inactivation, similar to the function of mammalian β1 subunits. In

Glyphosate and Roundup® alter morphology and behavior in zebrafish.

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

2017-12-01

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

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.

Defective Glycinergic Synaptic Transmission in Zebrafish Motility Mutants

OpenAIRE

Hirata, Hiromi; Carta, Eloisa; Yamanaka, Iori; Harvey, Robert J.; Kuwada, John Y.

2010-01-01

Glycine is a major inhibitory neurotransmitter in the spinal cord and brainstem. Recently, in vivo analysis of glycinergic synaptic transmission has been pursued in zebrafish using molecular genetics. An ENU mutagenesis screen identified two behavioral mutants that are defective in glycinergic synaptic transmission. Zebrafish bandoneon (beo) mutants have a defect in glrbb, one of the duplicated glycine receptor (GlyR) β subunit genes. These mutants exhibit a loss of glycinergic synaptic ...

Insulin-like activity in the retina

International Nuclear Information System (INIS)

Das, A.

1986-01-01

A number of studies have recently demonstrated that insulin or a homologous peptide may be synthesized outside the pancreas also. The present study was designed to investigate whether insulin-like activity exists in the retina, and if it exists, whether it is due to local synthesis of insulin or a similar peptide in the retina. To determine whether the insulin-like immunoreactivity in retinal glial cells is due to binding and uptake or local synthesis of insulin, a combined approach of immunocytochemistry and in situ DNA-RNA hybridization techniques was used on cultured rat retinal glial cells. Insulin-like immunoreactivity was demonstrated in the cytoplasma of these cells. In situ hybridization studies using labeled rat insulin cDNA indicated that these cells contain the mRNA necessary for de novo synthesis of insulin or a closely homologous peptide. Since human retinal cells have, as yet, not been conveniently grown in culture, an ocular tumor cell line, human Y79 retinoblastoma was used as a model to extend these investigations. The presence of insulin-like immunoreactivity as well as insulin-specific mRNA was demonstrated in this cell line. Light microscopic autoradiography following incubation of isolated rat retinal cells with 125 I-insulin showed the presence of insulin binding sites on the photoreceptors and amarcine cells. On the basis of these observations that rat retina glial cells, including Muller cells are sites of synthesis of insulin or a similar peptide, a model for the pathogenesis of dabetic retinopathy is proposed

Metastatic behaviour of primary human tumours in a zebrafish xenotransplantation model

International Nuclear Information System (INIS)

Marques, Ines J; Bagowski, Christoph P; Weiss, Frank Ulrich; Vlecken, Danielle H; Nitsche, Claudia; Bakkers, Jeroen; Lagendijk, Anne K; Partecke, Lars Ivo; Heidecke, Claus-Dieter; Lerch, Markus M

2009-01-01

Aberrant regulation of cell migration drives progression of many diseases, including cancer cell invasion and metastasis formation. Analysis of tumour invasion and metastasis in living organisms to date is cumbersome and involves difficult and time consuming investigative techniques. For primary human tumours we establish here a simple, fast, sensitive and cost-effective in vivo model to analyse tumour invasion and metastatic behaviour. We fluorescently labelled small explants from gastrointestinal human tumours and investigated their metastatic behaviour after transplantation into zebrafish embryos and larvae. The transparency of the zebrafish embryos allows to follow invasion, migration and micrometastasis formation in real-time. High resolution imaging was achieved through laser scanning confocal microscopy of live zebrafish. In the transparent zebrafish embryos invasion, circulation of tumour cells in blood vessels, migration and micrometastasis formation can be followed in real-time. Xenografts of primary human tumours showed invasiveness and micrometastasis formation within 24 hours after transplantation, which was absent when non-tumour tissue was implanted. Furthermore, primary human tumour cells, when organotopically implanted in the zebrafish liver, demonstrated invasiveness and metastatic behaviour, whereas primary control cells remained in the liver. Pancreatic tumour cells showed no metastatic behaviour when injected into cloche mutant embryos, which lack a functional vasculature. Our results show that the zebrafish is a useful in vivo animal model for rapid analysis of invasion and metastatic behaviour of primary human tumour specimen

Functional and Cellular Responses to Laser Injury in the Rat Snake Retina

National Research Council Canada - National Science Library

Glickman, Randolph D; Elliott, III, W. R; Kumar, Neeru

2007-01-01

.... This animal is of interest for vision research because its eye has an all-cone retina. A linear array of 5 thermal lesions was placed in the retina of anesthetized animals, near the area centralis, using a Nd:VO4 laser (532 nm...

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

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Claudia Hohn

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

Comparison of Antemortem and Environmental Samples for Zebrafish Health Monitoring and Quarantine

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Crim, Marcus J; Lawrence, Christian; Livingston, Robert S; Rakitin, Andrei; Hurley, Shane J; Riley, Lela K

2017-01-01

Molecular diagnostic assays offer both exquisite sensitivity and the ability to test a wide variety of sample types. Various types of environmental sample, such as detritus and concentrated water, might provide a useful adjunct to sentinels in routine zebrafish health monitoring. Similarly, antemortem sampling would be advantageous for expediting zebrafish quarantine, without euthanasia of valuable fish. We evaluated the detection of Mycobacterium chelonae, M. fortuitum, M. peregrinum, Pseudocapillaria tomentosa, and Pseudoloma neurophilia in zebrafish, detritus, pooled feces, and filter membranes after filtration of 1000-, 500-, and 150-mL water samples by real-time PCR analysis. Sensitivity varied according to sample type and pathogen, and environmental sampling was significantly more sensitive than zebrafish sampling for detecting Mycobacterium spp. but not for Pseudocapillaria neurophilia or Pseudoloma tomentosa. The results of these experiments provide strong evidence of the utility of multiple sample types for detecting pathogens according to each pathogen's life cycle and ecological niche within zebrafish systems. In a separate experiment, zebrafish subclinically infected with M. chelonae, M. marinum, Pleistophora hyphessobryconis, Pseudocapillaria tomentosa, or Pseudoloma neurophilia were pair-spawned and individually tested with subsets of embryos from each clutch that received no rinse, a fluidizing rinse, or were surface-disinfected with sodium hypochlorite. Frequently, one or both parents were subclinically infected with pathogen(s) that were not detected in any embryo subset. Therefore, negative results from embryo samples may not reflect the health status of the parent zebrafish. PMID:28724491

Imaging Subcellular Structures in the Living Zebrafish Embryo.

Science.gov (United States)

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

2016-04-02

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

Preserving information in neural transmission.

Science.gov (United States)

Sincich, Lawrence C; Horton, Jonathan C; Sharpee, Tatyana O

2009-05-13

Along most neural pathways, the spike trains transmitted from one neuron to the next are altered. In the process, neurons can either achieve a more efficient stimulus representation, or extract some biologically important stimulus parameter, or succeed at both. We recorded the inputs from single retinal ganglion cells and the outputs from connected lateral geniculate neurons in the macaque to examine how visual signals are relayed from retina to cortex. We found that geniculate neurons re-encoded multiple temporal stimulus features to yield output spikes that carried more information about stimuli than was available in each input spike. The coding transformation of some relay neurons occurred with no decrement in information rate, despite output spike rates that averaged half the input spike rates. This preservation of transmitted information was achieved by the short-term summation of inputs that geniculate neurons require to spike. A reduced model of the retinal and geniculate visual responses, based on two stimulus features and their associated nonlinearities, could account for >85% of the total information available in the spike trains and the preserved information transmission. These results apply to neurons operating on a single time-varying input, suggesting that synaptic temporal integration can alter the temporal receptive field properties to create a more efficient representation of visual signals in the thalamus than the retina.

Retina-like sensor image coordinates transformation and display

Science.gov (United States)

Cao, Fengmei; Cao, Nan; Bai, Tingzhu; Song, Shengyu

2015-03-01

For a new kind of retina-like senor camera, the image acquisition, coordinates transformation and interpolation need to be realized. Both of the coordinates transformation and interpolation are computed in polar coordinate due to the sensor's particular pixels distribution. The image interpolation is based on sub-pixel interpolation and its relative weights are got in polar coordinates. The hardware platform is composed of retina-like senor camera, image grabber and PC. Combined the MIL and OpenCV library, the software program is composed in VC++ on VS 2010. Experience results show that the system can realizes the real-time image acquisition, coordinate transformation and interpolation.

A comparison of some organizational characteristics of the mouse central retina and the human macula.

Science.gov (United States)

Volland, Stefanie; Esteve-Rudd, Julian; Hoo, Juyea; Yee, Claudine; Williams, David S

2015-01-01

Mouse models have greatly assisted our understanding of retinal degenerations. However, the mouse retina does not have a macula, leading to the question of whether the mouse is a relevant model for macular degeneration. In the present study, a quantitative comparison between the organization of the central mouse retina and the human macula was made, focusing on some structural characteristics that have been suggested to be important in predisposing the macula to stresses leading to degeneration: photoreceptor density, phagocytic load on the RPE, and the relative thinness of Bruch's membrane. Light and electron microscopy measurements from retinas of two strains of mice, together with published data on human retinas, were used for calculations and subsequent comparisons. As in the human retina, the central region of the mouse retina possesses a higher photoreceptor cell density and a thinner Bruch's membrane than in the periphery; however, the magnitudes of these periphery to center gradients are larger in the human. Of potentially greater relevance is the actual photoreceptor cell density, which is much greater in the mouse central retina than in the human macula, underlying a higher phagocytic load for the mouse RPE. Moreover, at eccentricities that correspond to the peripheral half of the human macula, the rod to cone ratio is similar between mouse and human. Hence, with respect to photoreceptor density and phagocytic load of the RPE, the central mouse retina models at least the more peripheral part of the macula, where macular degeneration is often first evident.

Large-scale multielectrode recording and stimulation of neural activity

International Nuclear Information System (INIS)

Sher, A.; Chichilnisky, E.J.; Dabrowski, W.; Grillo, A.A.; Grivich, M.; Gunning, D.; Hottowy, P.; Kachiguine, S.; Litke, A.M.; Mathieson, K.; Petrusca, D.

2007-01-01

Large circuits of neurons are employed by the brain to encode and process information. How this encoding and processing is carried out is one of the central questions in neuroscience. Since individual neurons communicate with each other through electrical signals (action potentials), the recording of neural activity with arrays of extracellular electrodes is uniquely suited for the investigation of this question. Such recordings provide the combination of the best spatial (individual neurons) and temporal (individual action-potentials) resolutions compared to other large-scale imaging methods. Electrical stimulation of neural activity in turn has two very important applications: it enhances our understanding of neural circuits by allowing active interactions with them, and it is a basis for a large variety of neural prosthetic devices. Until recently, the state-of-the-art in neural activity recording systems consisted of several dozen electrodes with inter-electrode spacing ranging from tens to hundreds of microns. Using silicon microstrip detector expertise acquired in the field of high-energy physics, we created a unique neural activity readout and stimulation framework that consists of high-density electrode arrays, multi-channel custom-designed integrated circuits, a data acquisition system, and data-processing software. Using this framework we developed a number of neural readout and stimulation systems: (1) a 512-electrode system for recording the simultaneous activity of as many as hundreds of neurons, (2) a 61-electrode system for electrical stimulation and readout of neural activity in retinas and brain-tissue slices, and (3) a system with telemetry capabilities for recording neural activity in the intact brain of awake, naturally behaving animals. We will report on these systems, their various applications to the field of neurobiology, and novel scientific results obtained with some of them. We will also outline future directions

Cold Shock Proteins Are Expressed in the Retina Following Exposure to Low Temperatures.

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Ignacio M Larrayoz

Full Text Available Hypothermia has been proposed as a therapeutic intervention for some retinal conditions, including ischemic insults. Cold exposure elevates expression of cold-shock proteins (CSP, including RNA-binding motif protein 3 (RBM3 and cold inducible RNA-binding protein (CIRP, but their presence in mammalian retina is so far unknown. Here we show the effects of hypothermia on the expression of these CSPs in retina-derived cell lines and in the retina of newborn and adult rats. Two cell lines of retinal origin, R28 and mRPE, were exposed to 32°C for different time periods and CSP expression was measured by qRT-PCR and Western blotting. Neonatal and adult Sprague-Dawley rats were exposed to a cold environment (8°C and expression of CSPs in their retinas was studied by Western blotting, multiple inmunofluorescence, and confocal microscopy. RBM3 expression was upregulated by cold in both R28 and mRPE cells in a time-dependent fashion. On the other hand, CIRP was upregulated in R28 cells but not in mRPE. In vivo, expression of CSPs was negligible in the retina of newborn and adult rats kept at room temperature (24°C. Exposure to a cold environment elicited a strong expression of both proteins, especially in retinal pigment epithelium cells, photoreceptors, bipolar, amacrine and horizontal cells, Müller cells, and ganglion cells. In conclusion, CSP expression rapidly rises in the mammalian retina following exposure to hypothermia in a cell type-specific pattern. This observation may be at the basis of the molecular mechanism by which hypothermia exerts its therapeutic effects in the retina.

Cross-Modal Learning between Visual and Vibration Signals in Zebrafish Danio Rerio

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Mu-Yun Wang

2011-10-01

Full Text Available Animals are always integrating environmental information from multiple sensory modalities, but the mechanisms underneath are highly underexploited. Crossmodal interactions in animal perception have been found in several species including human, mice and flies. Here we subjected zebrafish as model because its genetic effects on brain and sense organ development are well studied, but the attentional processes are mainly unexplored. Zebrafish show impressive behaviour capabilities with relatively small or “simple” brains which make their nervous system relatively more accessible to experimentation than large-brained mammals. When conditioned with both vision and vibration signals, zebrafish were able to make higher correct choices than only one sensation. After multimodal training, zebrafish were also able to transfer the memory to unimodal conditioning when only given vision or vibration signals. This study provided basic findings for how animals process multimodal sensory from the environment, and showed crossmodal interactions in zebrafish for the first time.

Two-Photon-Based Photoactivation in Live Zebrafish Embryos

OpenAIRE

Russek-Blum, Niva; Nabel-Rosen, Helit; Levkowitz, Gil

2010-01-01

Photoactivation of target compounds in a living organism has proven a valuable approach to investigate various biological processes such as embryonic development, cellular signaling and adult physiology. In this respect, the use of multi-photon microscopy enables quantitative photoactivation of a given light responsive agent in deep tissues at a single cell resolution. As zebrafish embryos are optically transparent, their development can be monitored in vivo. These traits make the zebrafish a...

Autofluorescence from the outer retina and subretinal space: hypothesis and review.

Science.gov (United States)

Spaide, Richard

2008-01-01

To review the pathophysiologic principles underlying increased autofluorescence from the outer retina and subretinal space using selected diseases as examples. The ocular imaging information and histopathologic features, when known, were integrated for diseases causing increased autofluorescence from the outer retina and subretinal space. Inferences were taken from this information and used to create a classification scheme. These diseases are principally those that cause separation of the outer retina from the retinal pigment epithelium, thereby preventing proper phagocytosis of photoreceptor outer segments. The separation can arise from increased exudation into the subretinal space or inadequate removal of fluid from the subretinal space. Lack of normal outer segment processing initially leads to increased accumulation of outer segments on the outer retina and subretinal space. Over time, this material is visible as an increasingly thick coating on the outer retina, is yellow, and is autofluorescent. Over time, atrophy develops with thinning of the deposited material and decreasing autofluorescence. The accumulated material is ultimately capable of inducing damage to the retinal pigment epithelium. Diseases causing accumulation of the material include central serous chorioretinopathy, vitelliform macular dystrophy, acute exudative polymorphous vitelliform maculopathy, choroidal tumors, and vitreomacular traction syndrome. The physical separation of the retinal outer segments from the retinal pigment epithelium hinders proper phagocytosis of the outer segments. Accumulation of the shed but not phagocytized outer segments plays a role in disease manifestations for a number of macular diseases.

Directed Differentiation of Zebrafish Pluripotent Embryonic Cells to Functional Cardiomyocytes

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Yao Xiao

2016-09-01

Full Text Available A cardiomyocyte differentiation in vitro system from zebrafish embryos remains to be established. Here, we have determined pluripotency window of zebrafish embryos by analyzing their gene-expression patterns of pluripotency factors together with markers of three germ layers, and have found that zebrafish undergoes a very narrow period of pluripotency maintenance from zygotic genome activation to a brief moment after oblong stage. Based on the pluripotency and a combination of appropriate conditions, we established a rapid and efficient method for cardiomyocyte generation in vitro from primary embryonic cells. The induced cardiomyocytes differentiated into functional and specific cardiomyocyte subtypes. Notably, these in vitro generated cardiomyocytes exhibited typical contractile kinetics and electrophysiological features. The system provides a new paradigm of cardiomyocyte differentiation from primary embryonic cells in zebrafish. The technology provides a new platform for the study of heart development and regeneration, in addition to drug discovery, disease modeling, and assessment of cardiotoxic agents.

Elucidating the mechanism of action of tributyltin (TBT) in zebrafish.

Science.gov (United States)

McGinnis, Courtney L; Crivello, Joseph F

2011-05-01

Tributyltin (TBT), an antifouling agent, has been implicated in the masculinization of fish species worldwide, but the masculinizing mechanism is not fully understood. We have examined the actions of TBT as an endocrine disruptor in zebrafish (Danio rerio). In HeLa cells transiently co-transfected with plasmid constructs containing the zebrafish estrogen receptors (zfERα, zfERβ(1) and zfERβ(2)) and the zebrafish estrogen response element (zfERE-tk-luc), ethinyl estradiol (EE2) induced luciferase activity 4 to 6-fold and was inhibited by TBT. In HeLa cells transiently co-transfected with the zebrafish androgen receptor (zfAR) and the murine androgen receptor response element (ARE-slp-luc), testosterone induced luciferase activity was not inhibited by TBT. In HeLa cells co-transfected with zfERα, zfERβ(1) and zfERβ(2) and a plasmid containing zebrafish aromatase (zfCyp19b-luc), TBT inhibited luciferase activity. In zebrafish exposed to 1mg/kg and 5mg/kg TBT in vivo, there was a increase in liver sulfotransferase and a decrease acyl-CoA testosterone acyltransferase activity. Real-time PCR analysis of sexual differentiation markers in fish exposed to TBT in vivo revealed a tissue-specific response. In brain there was increased production of Sox9, Dax1, and SF1 mRNA, an androgenizing effect, while in the liver there was increased production of Dax1, Cyp19a and zfERβ(1) mRNA but decreased production of Sox9 mRNA, a feminizing effect. In the gonads there was increased production of zfERα and zfCyp19a mRNA, again a feminizing effect. TBT has an overall masculinizing effect but the masculinizing effect is tempered by a feminizing effect on gene transcription in certain tissues. These results are discussed in the context of TBT as an endocrine disruptor in zebrafish. Copyright © 2011 Elsevier B.V. All rights reserved.

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

Science.gov (United States)

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

2018-06-01

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

The hypoxia factor Hif-1α controls neural crest chemotaxis and epithelial to mesenchymal transition

Science.gov (United States)

Barriga, Elias H.; Maxwell, Patrick H.

2013-01-01

One of the most important mechanisms that promotes metastasis is the stabilization of Hif-1 (hypoxia-inducible transcription factor 1). We decided to test whether Hif-1α also was required for early embryonic development. We focused our attention on the development of the neural crest, a highly migratory embryonic cell population whose behavior has been likened to cancer metastasis. Inhibition of Hif-1α by antisense morpholinos in Xenopus laevis or zebrafish embryos led to complete inhibition of neural crest migration. We show that Hif-1α controls the expression of Twist, which in turn represses E-cadherin during epithelial to mesenchymal transition (EMT) of neural crest cells. Thus, Hif-1α allows cells to initiate migration by promoting the release of cell–cell adhesions. Additionally, Hif-1α controls chemotaxis toward the chemokine SDF-1 by regulating expression of its receptor Cxcr4. Our results point to Hif-1α as a novel and key regulator that integrates EMT and chemotaxis during migration of neural crest cells. PMID:23712262

The zebrafish reference genome sequence and its relationship to the human genome.

Science.gov (United States)

Howe, Kerstin; Clark, Matthew D; Torroja, Carlos F; Torrance, James; Berthelot, Camille; Muffato, Matthieu; Collins, John E; Humphray, Sean; McLaren, Karen; Matthews, Lucy; McLaren, Stuart; Sealy, Ian; Caccamo, Mario; Churcher, Carol; Scott, Carol; Barrett, Jeffrey C; Koch, Romke; Rauch, Gerd-Jörg; White, Simon; Chow, William; Kilian, Britt; Quintais, Leonor T; Guerra-Assunção, José A; Zhou, Yi; Gu, Yong; Yen, Jennifer; Vogel, Jan-Hinnerk; Eyre, Tina; Redmond, Seth; Banerjee, Ruby; Chi, Jianxiang; Fu, Beiyuan; Langley, Elizabeth; Maguire, Sean F; Laird, Gavin K; Lloyd, David; Kenyon, Emma; Donaldson, Sarah; Sehra, Harminder; Almeida-King, Jeff; Loveland, Jane; Trevanion, Stephen; Jones, Matt; Quail, Mike; Willey, Dave; Hunt, Adrienne; Burton, John; Sims, Sarah; McLay, Kirsten; Plumb, Bob; Davis, Joy; Clee, Chris; Oliver, Karen; Clark, Richard; Riddle, Clare; Elliot, David; Eliott, David; Threadgold, Glen; Harden, Glenn; Ware, Darren; Begum, Sharmin; Mortimore, Beverley; Mortimer, Beverly; Kerry, Giselle; Heath, Paul; Phillimore, Benjamin; Tracey, Alan; Corby, Nicole; Dunn, Matthew; Johnson, Christopher; Wood, Jonathan; Clark, Susan; Pelan, Sarah; Griffiths, Guy; Smith, Michelle; Glithero, Rebecca; Howden, Philip; Barker, Nicholas; Lloyd, Christine; Stevens, Christopher; Harley, Joanna; Holt, Karen; Panagiotidis, Georgios; Lovell, Jamieson; Beasley, Helen; Henderson, Carl; Gordon, Daria; Auger, Katherine; Wright, Deborah; Collins, Joanna; Raisen, Claire; Dyer, Lauren; Leung, Kenric; Robertson, Lauren; Ambridge, Kirsty; Leongamornlert, Daniel; McGuire, Sarah; Gilderthorp, Ruth; Griffiths, Coline; Manthravadi, Deepa; Nichol, Sarah; Barker, Gary; Whitehead, Siobhan; Kay, Michael; Brown, Jacqueline; Murnane, Clare; Gray, Emma; Humphries, Matthew; Sycamore, Neil; Barker, Darren; Saunders, David; Wallis, Justene; Babbage, Anne; Hammond, Sian; Mashreghi-Mohammadi, Maryam; Barr, Lucy; Martin, Sancha; Wray, Paul; Ellington, Andrew; Matthews, Nicholas; Ellwood, Matthew; Woodmansey, Rebecca; Clark, Graham; Cooper, James D; Cooper, James; Tromans, Anthony; Grafham, Darren; Skuce, Carl; Pandian, Richard; Andrews, Robert; Harrison, Elliot; Kimberley, Andrew; Garnett, Jane; Fosker, Nigel; Hall, Rebekah; Garner, Patrick; Kelly, Daniel; Bird, Christine; Palmer, Sophie; Gehring, Ines; Berger, Andrea; Dooley, Christopher M; Ersan-Ürün, Zübeyde; Eser, Cigdem; Geiger, Horst; Geisler, Maria; Karotki, Lena; Kirn, Anette; Konantz, Judith; Konantz, Martina; Oberländer, Martina; Rudolph-Geiger, Silke; Teucke, Mathias; Lanz, Christa; Raddatz, Günter; Osoegawa, Kazutoyo; Zhu, Baoli; Rapp, Amanda; Widaa, Sara; Langford, Cordelia; Yang, Fengtang; Schuster, Stephan C; Carter, Nigel P; Harrow, Jennifer; Ning, Zemin; Herrero, Javier; Searle, Steve M J; Enright, Anton; Geisler, Robert; Plasterk, Ronald H A; Lee, Charles; Westerfield, Monte; de Jong, Pieter J; Zon, Leonard I; Postlethwait, John H; Nüsslein-Volhard, Christiane; Hubbard, Tim J P; Roest Crollius, Hugues; Rogers, Jane; Stemple, Derek L

2013-04-25

Zebrafish have become a popular organism for the study of vertebrate gene function. The virtually transparent embryos of this species, and the ability to accelerate genetic studies by gene knockdown or overexpression, have led to the widespread use of zebrafish in the detailed investigation of vertebrate gene function and increasingly, the study of human genetic disease. However, for effective modelling of human genetic disease it is important to understand the extent to which zebrafish genes and gene structures are related to orthologous human genes. To examine this, we generated a high-quality sequence assembly of the zebrafish genome, made up of an overlapping set of completely sequenced large-insert clones that were ordered and oriented using a high-resolution high-density meiotic map. Detailed automatic and manual annotation provides evidence of more than 26,000 protein-coding genes, the largest gene set of any vertebrate so far sequenced. Comparison to the human reference genome shows that approximately 70% of human genes have at least one obvious zebrafish orthologue. In addition, the high quality of this genome assembly provides a clearer understanding of key genomic features such as a unique repeat content, a scarcity of pseudogenes, an enrichment of zebrafish-specific genes on chromosome 4 and chromosomal regions that influence sex determination.

Complement anaphylatoxin C3a is a potent inducer of embryonic chick retina regeneration

Science.gov (United States)

Haynes, Tracy; Luz-Madrigal, Agustin; Reis, Edimara S.; Echeverri Ruiz, Nancy P.; Grajales-Esquivel, Erika; Tzekou, Apostolia; Tsonis, Panagiotis A.; Lambris, John D.; Del Rio-Tsonis, Katia

2013-01-01

Identifying the initiation signals for tissue regeneration in vertebrates is one of the major challenges in regenerative biology. Much of the research thus far has indicated that certain growth factors have key roles. Here we show that complement fragment C3a is sufficient to induce complete regeneration of the embryonic chick retina from stem/progenitor cells present in the eye, independent of fibroblast growth factor receptor signaling. Instead, C3a induces retina regeneration via STAT3 activation, which in turn activates the injury- and inflammation-responsive factors, IL-6, IL-8 and TNF-α. This activation sets forth regulation of Wnt2b, Six3 and Sox2, genes associated with retina stem and progenitor cells. Thus, our results establish a mechanism for retina regeneration based on injury and inflammation signals. Furthermore, our results indicate a unique function for complement anaphylatoxins that implicate these molecules in the induction and complete regeneration of the retina, opening new avenues of experimentation in the field. PMID:23942241

The essential role of endogenous ghrelin in growth hormone expression during zebrafish adenohypophysis development.

Science.gov (United States)

Li, Xi; He, Jiangyan; Hu, Wei; Yin, Zhan

2009-06-01

Ghrelin, a multifunctional hormone, including potent GH stimulation activity, has been suggested to be important during embryonic development. Expression of ghrelin has been confirmed in the zebrafish pancreas during embryonic stages. Interfering with ghrelin function using two specific antisense morpholino oligonucleotides causes defects during zebrafish embryonic development. In ghrelin morphants the expression of GH was abolished in zebrafish somatotropes, whereas the expression patterns of the other key molecules involved in hypothalamic-pituitary development and distinct pituitary hormones genes remain largely intact at the appropriate time during zebrafish adenohypophysis development. Effective rescue of the ghrelin morphants with exogenous ghrelin mRNA showed that the correct gene had been targeted. Moreover, by analyzing the efficiencies of the ghrelin morphants rescue experiments with various forms of exogenous mutant ghrelin mRNAs, we also demonstrated the essentiality of the form acyl-ghrelin on GH stimulation during zebrafish adenohypophysis development. Our in vivo experiments, for the first time, also provided evidence of the existence of functional obestatin in the C-terminal part of zebrafish proghrelin peptides. Our research here has demonstrated that zebrafish is a unique model for functional studies of endogenous ghrelin, especially during embryonic development.

Developmental effects of aerosols and coal burning particles in zebrafish embryos

International Nuclear Information System (INIS)

Olivares, Alba; Drooge, Barend L. van; Casado, Marta; Prats, Eva; Serra, Montserrat; Ven, Leo T. van der; Kamstra, Jorke H.; Hamers, Timo; Hermsen, Sanne; Grimalt, Joan O.; Piña, Benjamin

2013-01-01

Embryo toxicity of particles generated by combustion processes is of special concern for human health. A significant part of these toxic effects is linked to the binding of some pollutants (like polycyclic aromatic hydrocarbons or PAHs) to the Aryl hydrocarbon Receptor (AhR) and the activation of target genes, like the cytochrome P4501A. This activity was analyzed for ambient air and coal-combustion particle extracts in zebrafish embryos (the cyp1aDarT assay) and in two single-cell bioassays: the yeast-based YCM-RYA and the DR-luc (rat cells) assay. Observed AhR ligand activity of samples generally correlated to the predicted toxic effect according to their PAH composition, except for one of the coal combustion samples with an anomalously high activity in the cyp1aDarT assay. This sample induced deformities in zebrafish embryos. We concluded that the combination of morphological and molecular assays may detect embryonic toxic effects that cannot be predicted from chemical analyses or single-cell bioassays. -- Highlights: ► Samples from air particulated matter and coal waste gob showed embryo toxicity in zebrafish. ► PAHs composition of samples does not adequately predict the toxic effects in zebrafish. ► Active coal waste gob samples show maximal AhR-ligand activity and induce deformations in zebrafish embryos. -- Aerosols and coal burning particles showed a strong developmental toxicity in zebrafish, in a degree that cannot be directly predicted from chemical analyses or single-cell bioassays

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

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

2018-01-01

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

A multi-scale model for correlation in B cell VDJ usage of zebrafish

International Nuclear Information System (INIS)

Pan, Keyao; Deem, Michael W

2011-01-01

The zebrafish (Danio rerio) is one of the model animals used for the study of immunology because the dynamics in the adaptive immune system of zebrafish are similar to that in higher animals. In this work, we built a multi-scale model to simulate the dynamics of B cells in the primary and secondary immune responses of zebrafish. We use this model to explain the reported correlation between VDJ usage of B cell repertoires in individual zebrafish. We use a delay ordinary differential equation (ODE) system to model the immune responses in the 6-month lifespan of a zebrafish. This mean field theory gives the number of high-affinity B cells as a function of time during an infection. The sequences of those B cells are then taken from a distribution calculated by a 'microscopic' random energy model. This generalized NK model shows that mature B cells specific to one antigen largely possess a single VDJ recombination. The model allows first-principle calculation of the probability, p, that two zebrafish responding to the same antigen will select the same VDJ recombination. This probability p increases with the B cell population size and the B cell selection intensity. The probability p decreases with the B cell hypermutation rate. The multi-scale model predicts correlations in the immune system of the zebrafish that are highly similar to that from experiment

A multi-scale model for correlation in B cell VDJ usage of zebrafish

Science.gov (United States)

Pan, Keyao; Deem, Michael W.

2011-10-01

The zebrafish (Danio rerio) is one of the model animals used for the study of immunology because the dynamics in the adaptive immune system of zebrafish are similar to that in higher animals. In this work, we built a multi-scale model to simulate the dynamics of B cells in the primary and secondary immune responses of zebrafish. We use this model to explain the reported correlation between VDJ usage of B cell repertoires in individual zebrafish. We use a delay ordinary differential equation (ODE) system to model the immune responses in the 6-month lifespan of a zebrafish. This mean field theory gives the number of high-affinity B cells as a function of time during an infection. The sequences of those B cells are then taken from a distribution calculated by a 'microscopic' random energy model. This generalized NK model shows that mature B cells specific to one antigen largely possess a single VDJ recombination. The model allows first-principle calculation of the probability, p, that two zebrafish responding to the same antigen will select the same VDJ recombination. This probability p increases with the B cell population size and the B cell selection intensity. The probability p decreases with the B cell hypermutation rate. The multi-scale model predicts correlations in the immune system of the zebrafish that are highly similar to that from experiment.

Endocrine disruption of courtship behaviour and reproduction in zebrafish (Danio rerio)

DEFF Research Database (Denmark)

Broch-Lips, Mia Gina Gruwier

2011-01-01

of the reversibility of hormonally induced shifts in sex ratio of zebrafish. In the first part of this study zebrafish were exposed to three different environmentally relevant concentrations of the synthetic oestrogen17α-ethinylestradiol (EE2) from egg stage to sexual maturity. Secondary sexual characteristics...... as fertilizing the spawned eggs. It was further demonstrated that the exposure to TB led to irreversible masculinisation of zebrafish which is in contrast with the partial reversibility of oestrogen induced sex change. During my investigations leading to this thesis it became apparent that sexual behaviour...... courtship behaviour have only been scarcely investigated. The aim of this project was to learn more about the effects of EDCS on the courtship behaviour and reproduction in zebrafish as well as investigating the reversibility of observed effects. I furthermore observed some interesting aspects...

Repairing quite swimmingly: advances in regenerative medicine using zebrafish.

Science.gov (United States)

Goessling, Wolfram; North, Trista E

2014-07-01

Regenerative medicine has the promise to alleviate morbidity and mortality caused by organ dysfunction, longstanding injury and trauma. Although regenerative approaches for a few diseases have been highly successful, some organs either do not regenerate well or have no current treatment approach to harness their intrinsic regenerative potential. In this Review, we describe the modeling of human disease and tissue repair in zebrafish, through the discovery of disease-causing genes using classical forward-genetic screens and by modulating clinically relevant phenotypes through chemical genetic screening approaches. Furthermore, we present an overview of those organ systems that regenerate well in zebrafish in contrast to mammalian tissue, as well as those organs in which the regenerative potential is conserved from fish to mammals, enabling drug discovery in preclinical disease-relevant models. We provide two examples from our own work in which the clinical translation of zebrafish findings is either imminent or has already proven successful. The promising results in multiple organs suggest that further insight into regenerative mechanisms and novel clinically relevant therapeutic approaches will emerge from zebrafish research in the future. © 2014. Published by The Company of Biologists Ltd.

Modeling tuberculous meningitis in zebrafish using Mycobacterium marinum

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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.

Training and professional profile of retinologists in Spain: Retina 2 project, Report 4.

Science.gov (United States)

Pastor, J Carlos; Fernández, Itziar; Rojas, Jimena; Coco, Rosa; Sanabria, Maria R; Rodríguez-de la Rúa, Enrique; Sánchez, Diego; Valverde, Carmen; Sala-Puigdollers, Anna

2011-01-01

Uniform postresidency systems to train medical specialists have not been developed in most European countries. Before developing a framework for such a system, we established the learning and professional profiles of Spanish ophthalmologists dedicated to medical retina and vitreoretina subspecialties. After identification of presumed subspecialists by experts from different autonomous regions, a self-administered questionnaire was mailed in 2006. A reminder was sent three weeks later. Postal mail was used. Nonresponder bias was determined. Of 492 possible retina subspecialists, 261 replied to the questionnaires. While about 86% received specific retinal training, standardized fellowship programs were uncommon for both medical retina and vitreoretina (around 10%). Of the responders, 24.5% performed only medical retina, 11.8% vitreoretina, and 63.6% both. Most (60.5%) practiced anterior segment surgery, and 78.7% declared skills in vitrectomy. We have developed a database of Spanish ophthalmologists dedicated to retinal pathologies and identified some characteristics of their professional profile. Although most of them have received specific retinal training, standardized mastership programs are still uncommon. These data will be useful in creating a standardized Retina Mastership, an important goal of the European Higher Education Area.

Biotransformation of ginsenosides F4 and Rg6 in zebrafish.

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

2017-03-28

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

Developmental toxicity of thyroid-active compounds in a zebrafish embryotoxicity test

NARCIS (Netherlands)

Jomaa, B.; Hermsen, S.A.B.; Kessels, M.Y.; Berg, van den J.H.J.; Peijenburg, A.C.M.; Aarts, J.M.M.J.G.; Piersma, A.H.; Rietjens, I.

2014-01-01

Zebrafish embryos were exposed to concentration ranges of selected thyroid-active model compounds in order to assess the applicability of zebrafish-based developmental scoring systems within an alternative testing strategy to detect the developmental toxicity of thyroid-active compounds. Model

Sex specific response in cholesterol level in zebrafish (Danio rerio) after long-term exposure of difenoconazole

International Nuclear Information System (INIS)

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

2015-01-01

Difenoconazole is a widely used triazole fungicide, its extensive application may potentially cause toxic effects on non-target organisms. To investigate the effect of difenoconazole on cholesterol content and related mechanism, adult zebrafish were exposed to environmental related dosage (0.1, 10 and 500 μg/L) difenoconazole. The body weight and hepatic total cholesterol (TCHO) level was tested at 7, 15 and 30 days post exposure (dpe). The expressions of eight cholesterol synthesis genes and one cholesterol metabolism gene were assessed via Quantitative PCR method. The significant decrease of TCHO level in male zebrafish liver was observed at 15 and 30 dpe, which was accompanied by apparent hepatic cholesterol-genesis genes expression decline. In comparison with males, female zebrafish showed different transcription modification of tested genes, and the cholesterol content maintain normal level during the whole exposure. - Highlights: • Difenoconazle could reduce TCHO level in male zebrafish liver. • Difenoconazole could inhibit sterol-genesis genes expression in male zebrafish. • Female zebrafish didn't show obvious change of TCHO level after exposure. • Difenoconazole could inhibit body weight of both male and female zebrafish. - Difenoconazle could reduce cholesterol level and sterol-genesis genes expression in male zebrafish. While female zebrafish showed no obvious cholesterol content change during exposure

Automated processing of zebrafish imaging data: a survey.

Science.gov (United States)

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

2013-09-01

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

Automated Processing of Zebrafish Imaging Data: A Survey

Science.gov (United States)

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

2013-01-01

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

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.

Short-term memory in zebrafish (Danio rerio).

Science.gov (United States)

Jia, Jason; Fernandes, Yohaan; Gerlai, Robert

2014-08-15

Learning and memory represent perhaps the most complex behavioral phenomena. Although their underlying mechanisms have been extensively analyzed, only a fraction of the potential molecular components have been identified. The zebrafish has been proposed as a screening tool with which mechanisms of complex brain functions may be systematically uncovered. However, as a relative newcomer in behavioral neuroscience, the zebrafish has not been well characterized for its cognitive and mnemonic features, thus learning and/or memory screens with adults have not been feasible. Here we study short-term memory of adult zebrafish. We show animated images of conspecifics (the stimulus) to the experimental subject during 1 min intervals on ten occasions separated by different (2, 4, 8 or 16 min long) inter-stimulus intervals (ISI), a between subject experimental design. We quantify the distance of the subject from the image presentation screen during each stimulus presentation interval, during each of the 1-min post-stimulus intervals immediately following the stimulus presentations and during each of the 1-min intervals furthest away from the last stimulus presentation interval and just before the next interval (pre-stimulus interval), respectively. Our results demonstrate significant retention of short-term memory even in the longest ISI group but suggest no acquisition of reference memory. Because in the employed paradigm both stimulus presentation and behavioral response quantification is computer automated, we argue that high-throughput screening for drugs or mutations that alter short-term memory performance of adult zebrafish is now becoming feasible. Copyright © 2014 Elsevier B.V. All rights reserved.

Fish from Head to Tail: The 9th European Zebrafish Meeting in Oslo.

Science.gov (United States)

Griffiths, Gareth; Müller, Ferenc; Ledin, Johan; Patton, E Elizabeth; Gjøen, Tor; Lobert, Viola Hélène; Winther-Larsen, Hanne Cecilie; Mullins, Mary; Joly, Jean-Stephane; Weltzien, Finn-Arne; Press, Charles McLean; Aleström, Peter

2016-04-01

The 9th European Zebrafish Meeting took place recently in Oslo (June 28-July 2, 2015). A total of 650 participants came to hear the latest research news focused on the zebrafish, Danio rerio, and to its distant evolutionary relative medaka, Oryzias latipes. The packed program included keynote and plenary talks, short oral presentations and poster sessions, workshops, and strategic discussions. The meeting was a great success and revealed dramatically how important the zebrafish in particular has become as a model system for topics, such as developmental biology, functional genomics, biomedicine, toxicology, and drug development. A new emphasis was given to its potential as a model for aquaculture, a topic of great economic interest to the host country Norway and for the future global food supply in general. Zebrafish husbandry as well as its use in teaching were also covered in separate workshops. As has become a tradition in these meetings, there was a well-attended Wellcome Trust Sanger Institute and ZFIN workshop focused on Zebrafish Genome Resources on the first day. The full EZM 2015 program with abstracts can be read and downloaded from the EZM 2015 Web site zebrafish2015.org .

Molecular regionalization of the developing amphioxus neural tube challenges major partitions of the vertebrate brain.

Science.gov (United States)

Albuixech-Crespo, Beatriz; López-Blanch, Laura; Burguera, Demian; Maeso, Ignacio; Sánchez-Arrones, Luisa; Moreno-Bravo, Juan Antonio; Somorjai, Ildiko; Pascual-Anaya, Juan; Puelles, Eduardo; Bovolenta, Paola; Garcia-Fernàndez, Jordi; Puelles, Luis; Irimia, Manuel; Ferran, José Luis

2017-04-01

All vertebrate brains develop following a common Bauplan defined by anteroposterior (AP) and dorsoventral (DV) subdivisions, characterized by largely conserved differential expression of gene markers. However, it is still unclear how this Bauplan originated during evolution. We studied the relative expression of 48 genes with key roles in vertebrate neural patterning in a representative amphioxus embryonic stage. Unlike nonchordates, amphioxus develops its central nervous system (CNS) from a neural plate that is homologous to that of vertebrates, allowing direct topological comparisons. The resulting genoarchitectonic model revealed that the amphioxus incipient neural tube is unexpectedly complex, consisting of several AP and DV molecular partitions. Strikingly, comparison with vertebrates indicates that the vertebrate thalamus, pretectum, and midbrain domains jointly correspond to a single amphioxus region, which we termed Di-Mesencephalic primordium (DiMes). This suggests that these domains have a common developmental and evolutionary origin, as supported by functional experiments manipulating secondary organizers in zebrafish and mice.

Social dominance modulates eavesdropping in zebrafish

Science.gov (United States)

Abril-de-Abreu, Rodrigo; Cruz, Ana S.; Oliveira, Rui F.

2015-01-01

Group living animals may eavesdrop on signalling interactions between conspecifics and integrate it with their own past social experience in order to optimize the use of relevant information from others. However, little is known about this interplay between public (eavesdropped) and private social information. To investigate it, we first manipulated the dominance status of bystander zebrafish. Next, we either allowed or prevented bystanders from observing a fight. Finally, we assessed their behaviour towards the winners and losers of the interaction, using a custom-made video-tracking system and directional analysis. We found that only dominant bystanders who had seen the fight revealed a significant increase in directional focus (a measure of attention) towards the losers of the fights. Furthermore, our results indicate that information about the fighters' acquired status was collected from the signalling interaction itself and not from post-interaction status cues, which implies the existence of individual recognition in zebrafish. Thus, we show for the first time that zebrafish, a highly social model organism, eavesdrop on conspecific agonistic interactions and that this process is modulated by the eavesdroppers' dominance status. We suggest that this type of integration of public and private information may be ubiquitous in social learning processes. PMID:26361550

Afferent connectivity of the zebrafish habenulae

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Katherine Jane Turner

2016-04-01

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

Non-invasive electrocardiogram detection of in vivo zebrafish embryos using electric potential sensors

Science.gov (United States)

Rendon-Morales, E.; Prance, R. J.; Prance, H.; Aviles-Espinosa, R.

2015-11-01

In this letter, we report the continuous detection of the cardiac electrical activity in embryonic zebrafish using a non-invasive approach. We present a portable and cost-effective platform based on the electric potential sensing technology, to monitor in vivo electrocardiogram activity from the zebrafish heart. This proof of principle demonstration shows how electrocardiogram measurements from the embryonic zebrafish may become accessible by using electric field detection. We present preliminary results using the prototype, which enables the acquisition of electrophysiological signals from in vivo 3 and 5 days-post-fertilization zebrafish embryos. The recorded waveforms show electrocardiogram traces including detailed features such as QRS complex, P and T waves.

Comparative Analyses of Zebrafish Anxiety-Like Behavior Using Conflict-Based Novelty Tests.

Science.gov (United States)

Kysil, Elana V; Meshalkina, Darya A; Frick, Erin E; Echevarria, David J; Rosemberg, Denis B; Maximino, Caio; Lima, Monica Gomes; Abreu, Murilo S; Giacomini, Ana C; Barcellos, Leonardo J G; Song, Cai; Kalueff, Allan V

2017-06-01

Modeling of stress and anxiety in adult zebrafish (Danio rerio) is increasingly utilized in neuroscience research and central nervous system (CNS) drug discovery. Representing the most commonly used zebrafish anxiety models, the novel tank test (NTT) focuses on zebrafish diving in response to potentially threatening stimuli, whereas the light-dark test (LDT) is based on fish scototaxis (innate preference for dark vs. bright areas). Here, we systematically evaluate the utility of these two tests, combining meta-analyses of published literature with comparative in vivo behavioral and whole-body endocrine (cortisol) testing. Overall, the NTT and LDT behaviors demonstrate a generally good cross-test correlation in vivo, whereas meta-analyses of published literature show that both tests have similar sensitivity to zebrafish anxiety-like states. Finally, NTT evokes higher levels of cortisol, likely representing a more stressful procedure than LDT. Collectively, our study reappraises NTT and LDT for studying anxiety-like states in zebrafish, and emphasizes their developing utility for neurobehavioral research. These findings can help optimize drug screening procedures by choosing more appropriate models for testing anxiolytic or anxiogenic drugs.

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

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Yuxiao Yao

2017-09-01

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

From schooling to shoaling: patterns of collective motion in zebrafish (Danio rerio.

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Noam Miller

Full Text Available Animal groups on the move can take different configurations. For example, groups of fish can either be 'shoals' or 'schools': shoals are simply aggregations of individuals; schools are shoals exhibiting polarized, synchronized motion. Here we demonstrate that polarization distributions of groups of zebrafish (Danio rerio are bimodal, showing two distinct modes of collective motion corresponding to the definitions of shoaling and schooling. Other features of the group's motion also vary consistently between the two modes: zebrafish schools are faster and less dense than zebrafish shoals. Habituation to an environment can also alter the proportion of time zebrafish groups spend schooling or shoaling. Models of collective motion suggest that the degree and stability of group polarization increases with the group's density. Examining zebrafish groups of different sizes from 5 to 50, we show that larger groups are less polarized than smaller groups. Decreased fearfulness in larger groups may function similarly to habituation, causing them to spend more time shoaling than schooling, contrary to most models' predictions.

Protective Role of Comfrey Leave Extracts on UV-induced Zebrafish Fin Damage.

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Cheng, Chien-Chung; Chou, Chi-Yuan; Chang, Yao-Chin; Wang, Hsuan-Wen; Wen, Chi-Chung; Chen, Yau-Hung

2014-07-01

In zebrafish, UV exposure leads to fin malformation phenotypes including fin reduction or absence. The present study evaluated UV-protective activities of comfrey leaves extracts in a zebrafish model by recording fin morphological changes. Chemopreventive effects of comfrey leave extracts were evaluated using Kaplan-Meier analysis and Cox proportional hazards regression. The results showed that (1) the mean times of return to normal fin in the UV+comfrey (50 and 100 ppm) groups were 3.43 and 2.86 days and were quicker compared with that in the UV only group (4.21 days); (2) zebrafish fins in the UV+comfrey (50 and 100 ppm) groups were 2.05 and 3.25 times more likely to return to normal than those in the UV only group; and (3) comfrey leave extracts had UV-absorbance abilities and significantly reduced ROS production in UV-exposed zebrafish embryos, which may attenuate UV-mediated apoptosis. In conclusion, comfrey leaves extracts may have the potential to be developed as UV-protective agents to protect zebrafish embryos from UV-induced damage.

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...

Zebrafish heart as a model to study the integrative autonomic control of pacemaker function

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Stoyek, Matthew R.; Quinn, T. Alexander; Croll, Roger P.

2016-01-01

The cardiac pacemaker sets the heart's primary rate, with pacemaker discharge controlled by the autonomic nervous system through intracardiac ganglia. A fundamental issue in understanding the relationship between neural activity and cardiac chronotropy is the identification of neuronal populations that control pacemaker cells. To date, most studies of neurocardiac control have been done in mammalian species, where neurons are embedded in and distributed throughout the heart, so they are largely inaccessible for whole-organ, integrative studies. Here, we establish the isolated, innervated zebrafish heart as a novel alternative model for studies of autonomic control of heart rate. Stimulation of individual cardiac vagosympathetic nerve trunks evoked bradycardia (parasympathetic activation) and tachycardia (sympathetic activation). Simultaneous stimulation of both vagosympathetic nerve trunks evoked a summative effect. Effects of nerve stimulation were mimicked by direct application of cholinergic and adrenergic agents. Optical mapping of electrical activity confirmed the sinoatrial region as the site of origin of normal pacemaker activity and identified a secondary pacemaker in the atrioventricular region. Strong vagosympathetic nerve stimulation resulted in a shift in the origin of initial excitation from the sinoatrial pacemaker to the atrioventricular pacemaker. Putative pacemaker cells in the sinoatrial and atrioventricular regions expressed adrenergic β2 and cholinergic muscarinic type 2 receptors. Collectively, we have demonstrated that the zebrafish heart contains the accepted hallmarks of vertebrate cardiac control, establishing this preparation as a viable model for studies of integrative physiological control of cardiac function by intracardiac neurons. PMID:27342878

Corticotropin-releasing factor critical for zebrafish camouflage behavior is regulated by light and sensitive to ethanol.

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Wagle, Mahendra; Mathur, Priya; Guo, Su

2011-01-05

The zebrafish camouflage response is an innate "hard-wired" behavior that offers an excellent opportunity to explore neural circuit assembly and function. Moreover, the camouflage response is sensitive to ethanol, making it a tractable system for understanding how ethanol influences neural circuit development and function. Here we report the identification of corticotropin-releasing factor (CRF) as a critical component of the camouflage response pathway. We further show that ethanol, having no direct effect on the visual sensory system or the melanocytes, acts downstream of retinal ganglion cells and requires the CRF-proopiomelanocortin pathway to exert its effect on camouflage. Treatment with ethanol, as well as alteration of light exposure that changes sensory input into the camouflage circuit, robustly modifies CRF expression in subsets of neurons. Activity of both adenylyl cyclase 5 and extracellular signal-regulated kinase (ERK) is required for such ethanol-induced or light-induced plasticity of crf expression. These results reveal an essential role of a peptidergic pathway in camouflage that is regulated by light and influenced by ethanol at concentrations relevant to abuse and anxiolysis, in a cAMP-dependent and ERK-dependent manner. We conclude that this ethanol-modulated camouflage response represents a novel and relevant system for molecular genetic dissection of a neural circuit that is regulated by light and sensitive to ethanol.

Image Signal Transfer Method in Artificial Retina using Laser

Energy Technology Data Exchange (ETDEWEB)

Yoon, I.Y.; Lee, B.H.; Kim, S.J. [Seoul National University, Seoul (Korea)

2002-05-01

Recently, the research on artificial retina for the blind is active. In this paper a new optical link method for the retinal prosthesis is proposed. Laser diode system was chosen to transfer image into the eye in this project and the new optical system was designed and evaluated. The use of laser diode array in artificial retina system makes system simple for lack of signal processing part inside of the eyeball. Designed optical system is enough to focus laser diode array on photodiode array in 20X20 application. (author). 11 refs., 7 figs., 2 tabs.

Modeling Myeloid Malignancies Using Zebrafish

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Kathryn S. Potts

2017-12-01

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

Developmental toxicity evaluation of three hexabromocyclododecane diastereoisomers on zebrafish embryos

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Du Miaomiao [Key Lab of Urban Environment and Health, Institute of Urban Environment, Chinese Academy of Sciences, Xiamen 361021 (China); Graduate University of Chinese Academy of Sciences, Beijing 100049 (China); Zhang Dandan; Yan Changzhou [Key Lab of Urban Environment and Health, Institute of Urban Environment, Chinese Academy of Sciences, Xiamen 361021 (China); Zhang Xian, E-mail: xzhang@iue.ac.cn [Key Lab of Urban Environment and Health, Institute of Urban Environment, Chinese Academy of Sciences, Xiamen 361021 (China)

2012-05-15

Structural dissimilarities of hexabromocyclododecane diastereoisomers could raise substantial differences in physicochemical, biological and toxicological properties. In order to fully assess the environmental safety and health risk of hexabromocyclododecanes (HBCDs), zebrafish embryos were used to evaluate the developmental toxicity of individual HBCD diastereoisomers ({alpha}-HBCD, {beta}-HBCD and {gamma}-HBCD). Four-hour post-fertilization (hpf) zebrafish embryos were exposed to different concentrations of HBCD diastereoisomers (0, 0.01, 0.1 and 1.0 mg/l) until 120 hpf. The results showed that exposure to HBCDs can affect the development of zebrafish embryos/larvae in a dose-dependent and diastereoselective manner. The diastereoisomers {alpha}-, {beta}- and {gamma}-HBCD at 0.01 mg/l had little effect on the development of zebrafish embryos except that exposure to 0.01 mg/l {gamma}-HBCD significantly delayed hatching (P < 0.05). At 0.1 mg/l, {alpha}-HBCD resulted in depressed heart rate of larvae (96 hpf) and delayed hatching, whereas {beta}- and {gamma}-HBCD both caused significant hatching delay and growth inhibition (P < 0.05). In addition, a remarkable and significant increase in mortality and malformation rate was noted at 0.1 mg/l {gamma}-HBCD exposure groups (P < 0.05). At 1.0 mg/l, {alpha}-, {beta}- and {gamma}-HBCD significantly affected all of the endpoints monitored (P < 0.05). Additionally, HBCD diastereoisomers could induce the generation of reactive oxygen species (ROS) and the activities of caspase-3 and caspase-9 in a dose-dependent manner. The results indicated that HBCD diastereoisomers could cause developmental toxicity to zebrafish embryos through inducing apoptosis by ROS formation. The overall results showed a good agreement confirming that the order of developmental toxicity of HBCD diastereoisomers in zebrafish is {gamma}-HBCD > {beta}-HBCD > {alpha}-HBCD.

The zebrafish reference genome sequence and its relationship to the human genome

Science.gov (United States)

Howe, Kerstin; Clark, Matthew D.; Torroja, Carlos F.; Torrance, James; Berthelot, Camille; Muffato, Matthieu; Collins, John E.; Humphray, Sean; McLaren, Karen; Matthews, Lucy; McLaren, Stuart; Sealy, Ian; Caccamo, Mario; Churcher, Carol; Scott, Carol; Barrett, Jeffrey C.; Koch, Romke; Rauch, Gerd-Jörg; White, Simon; Chow, William; Kilian, Britt; Quintais, Leonor T.; Guerra-Assunção, José A.; Zhou, Yi; Gu, Yong; Yen, Jennifer; Vogel, Jan-Hinnerk; Eyre, Tina; Redmond, Seth; Banerjee, Ruby; Chi, Jianxiang; Fu, Beiyuan; Langley, Elizabeth; Maguire, Sean F.; Laird, Gavin K.; Lloyd, David; Kenyon, Emma; Donaldson, Sarah; Sehra, Harminder; Almeida-King, Jeff; Loveland, Jane; Trevanion, Stephen; Jones, Matt; Quail, Mike; Willey, Dave; Hunt, Adrienne; Burton, John; Sims, Sarah; McLay, Kirsten; Plumb, Bob; Davis, Joy; Clee, Chris; Oliver, Karen; Clark, Richard; Riddle, Clare; Eliott, David; Threadgold, Glen; Harden, Glenn; Ware, Darren; Mortimer, Beverly; Kerry, Giselle; Heath, Paul; Phillimore, Benjamin; Tracey, Alan; Corby, Nicole; Dunn, Matthew; Johnson, Christopher; Wood, Jonathan; Clark, Susan; Pelan, Sarah; Griffiths, Guy; Smith, Michelle; Glithero, Rebecca; Howden, Philip; Barker, Nicholas; Stevens, Christopher; Harley, Joanna; Holt, Karen; Panagiotidis, Georgios; Lovell, Jamieson; Beasley, Helen; Henderson, Carl; Gordon, Daria; Auger, Katherine; Wright, Deborah; Collins, Joanna; Raisen, Claire; Dyer, Lauren; Leung, Kenric; Robertson, Lauren; Ambridge, Kirsty; Leongamornlert, Daniel; McGuire, Sarah; Gilderthorp, Ruth; Griffiths, Coline; Manthravadi, Deepa; Nichol, Sarah; Barker, Gary; Whitehead, Siobhan; Kay, Michael; Brown, Jacqueline; Murnane, Clare; Gray, Emma; Humphries, Matthew; Sycamore, Neil; Barker, Darren; Saunders, David; Wallis, Justene; Babbage, Anne; Hammond, Sian; Mashreghi-Mohammadi, Maryam; Barr, Lucy; Martin, Sancha; Wray, Paul; Ellington, Andrew; Matthews, Nicholas; Ellwood, Matthew; Woodmansey, Rebecca; Clark, Graham; Cooper, James; Tromans, Anthony; Grafham, Darren; Skuce, Carl; Pandian, Richard; Andrews, Robert; Harrison, Elliot; Kimberley, Andrew; Garnett, Jane; Fosker, Nigel; Hall, Rebekah; Garner, Patrick; Kelly, Daniel; Bird, Christine; Palmer, Sophie; Gehring, Ines; Berger, Andrea; Dooley, Christopher M.; Ersan-Ürün, Zübeyde; Eser, Cigdem; Geiger, Horst; Geisler, Maria; Karotki, Lena; Kirn, Anette; Konantz, Judith; Konantz, Martina; Oberländer, Martina; Rudolph-Geiger, Silke; Teucke, Mathias; Osoegawa, Kazutoyo; Zhu, Baoli; Rapp, Amanda; Widaa, Sara; Langford, Cordelia; Yang, Fengtang; Carter, Nigel P.; Harrow, Jennifer; Ning, Zemin; Herrero, Javier; Searle, Steve M. J.; Enright, Anton; Geisler, Robert; Plasterk, Ronald H. A.; Lee, Charles; Westerfield, Monte; de Jong, Pieter J.; Zon, Leonard I.; Postlethwait, John H.; Nüsslein-Volhard, Christiane; Hubbard, Tim J. P.; Crollius, Hugues Roest; Rogers, Jane; Stemple, Derek L.

2013-01-01

Zebrafish have become a popular organism for the study of vertebrate gene function1,2. The virtually transparent embryos of this species, and the ability to accelerate genetic studies by gene knockdown or overexpression, have led to the widespread use of zebrafish in the detailed investigation of vertebrate gene function and increasingly, the study of human genetic disease3–5. However, for effective modelling of human genetic disease it is important to understand the extent to which zebrafish genes and gene structures are related to orthologous human genes. To examine this, we generated a high-quality sequence assembly of the zebrafish genome, made up of an overlapping set of completely sequenced large-insert clones that were ordered and oriented using a high-resolution high-density meiotic map. Detailed automatic and manual annotation provides evidence of more than 26,000 protein-coding genes6, the largest gene set of any vertebrate so far sequenced. Comparison to the human reference genome shows that approximately 70% of human genes have at least one obvious zebrafish orthologue. In addition, the high quality of this genome assembly provides a clearer understanding of key genomic features such as a unique repeat content, a scarcity of pseudogenes, an enrichment of zebrafish-specific genes on chromosome 4 and chromosomal regions that influence sex determination. PMID:23594743

Comprehensive analysis of coding-lncRNA gene co-expression network uncovers conserved functional lncRNAs in zebrafish.

Science.gov (United States)

Chen, Wen; Zhang, Xuan; Li, Jing; Huang, Shulan; Xiang, Shuanglin; Hu, Xiang; Liu, Changning

2018-05-09

Zebrafish is a full-developed model system for studying development processes and human disease. Recent studies of deep sequencing had discovered a large number of long non-coding RNAs (lncRNAs) in zebrafish. However, only few of them had been functionally characterized. Therefore, how to take advantage of the mature zebrafish system to deeply investigate the lncRNAs' function and conservation is really intriguing. We systematically collected and analyzed a series of zebrafish RNA-seq data, then combined them with resources from known database and literatures. As a result, we obtained by far the most complete dataset of zebrafish lncRNAs, containing 13,604 lncRNA genes (21,128 transcripts) in total. Based on that, a co-expression network upon zebrafish coding and lncRNA genes was constructed and analyzed, and used to predict the Gene Ontology (GO) and the KEGG annotation of lncRNA. Meanwhile, we made a conservation analysis on zebrafish lncRNA, identifying 1828 conserved zebrafish lncRNA genes (1890 transcripts) that have their putative mammalian orthologs. We also found that zebrafish lncRNAs play important roles in regulation of the development and function of nervous system; these conserved lncRNAs present a significant sequential and functional conservation, with their mammalian counterparts. By integrative data analysis and construction of coding-lncRNA gene co-expression network, we gained the most comprehensive dataset of zebrafish lncRNAs up to present, as well as their systematic annotations and comprehensive analyses on function and conservation. Our study provides a reliable zebrafish-based platform to deeply explore lncRNA function and mechanism, as well as the lncRNA commonality between zebrafish and human.

Acute toxicity and gene responses induced by endosulfan in zebrafish (Danio rerio embryos

Directory of Open Access Journals (Sweden)

Young-Sun Moon

2016-10-01

Full Text Available Endosulfan has been listed as a persistent organic pollutant, and is frequently found in agricultural environments during monitoring processes owing to its heavy use and persistent characteristics. This study was conducted to understand the effects of endosulfan on the development of zebrafish (Danio rerio embryos by exposing them to a specific range of endosulfan concentrations. Exposing zebrafish embryos to endosulfan for 96 h yielded no acute toxicity until the concentration reached 1500 μg L−1, whereas malformed zebrafish larvae developed severely curved spines and shortened tails. About 50% of zebrafish larvae were malformed when exposed to 600 μg L−1 of endosulfan. Comparative gene expression using real-time quantitative polymerase chain reaction was assessed using endosulfan-exposed zebrafish embryos. CYP1A and CYP3A were significantly enhanced in response to endosulfan treatment. Two genes, acacb and fasn, encoding acetyl-CoA carboxylase b and fatty acid synthase proteins, respectively, were also up-regulated after treating zebrafish embryos with endosulfan. These genes are also involved in fatty acid biosynthesis. The genes encoding vitellogenin and Hsp70 increased in a concentration-dependent manner in embryos. Finally, biochemical studies showed that acetylcholinesterase activity was reduced, whereas glutathione S-transferase and carboxylesterase activities were enhanced in zebrafish embryos after endosulfan treatment. These biochemical and molecular biological differences might be used for tools to determine contamination of endosulfan in the aquatic environment.

Two-photon-based photoactivation in live zebrafish embryos.

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Russek-Blum, Niva; Nabel-Rosen, Helit; Levkowitz, Gil

2010-12-24

Photoactivation of target compounds in a living organism has proven a valuable approach to investigate various biological processes such as embryonic development, cellular signaling and adult physiology. In this respect, the use of multi-photon microscopy enables quantitative photoactivation of a given light responsive agent in deep tissues at a single cell resolution. As zebrafish embryos are optically transparent, their development can be monitored in vivo. These traits make the zebrafish a perfect model organism for controlling the activity of a variety of chemical agents and proteins by focused light. Here we describe the use of two-photon microscopy to induce the activation of chemically caged fluorescein, which in turn allows us to follow cell's destiny in live zebrafish embryos. We use embryos expressing a live genetic landmark (GFP) to locate and precisely target any cells of interest. This procedure can be similarly used for precise light induced activation of proteins, hormones, small molecules and other caged compounds.

Ibuprofen slows migration and inhibits bowel colonization by enteric nervous system precursors in zebrafish, chick and mouse

Science.gov (United States)

Schill, Ellen Merrick; Lake, Jonathan I.; Tusheva, Olga A.; Nagy, Nandor; Bery, Saya K.; Foster, Lynne; Avetisyan, Marina; Johnson, Stephen L.; Stenson, William F.; Goldstein, Allan M.; Heuckeroth, Robert O.

2016-01-01

Hirschsprung Disease (HSCR) is a potentially deadly birth defect characterized by the absence of the enteric nervous system (ENS) in distal bowel. Although HSCR has clear genetic causes, no HSCR-associated mutation is 100% penetrant, suggesting gene-gene and gene-environment interactions determine HSCR occurrence. To test the hypothesis that certain medicines might alter HSCR risk we treated zebrafish with medications commonly used during early human pregnancy and discovered that ibuprofen caused HSCR-like absence of enteric neurons in distal bowel. Using fetal CF-1 mouse gut slice cultures, we found that ibuprofen treated enteric neural crest-derived cells (ENCDC) had reduced migration, fewer lamellipodia and lower levels of active RAC1/CDC42. Additionally, inhibiting ROCK, a RHOA effector and known RAC1 antagonist, reversed ibuprofen effects on migrating mouse ENCDC in culture. Ibuprofen also inhibited colonization of Ret+/− mouse bowel by ENCDC in vivo and dramatically reduced bowel colonization by chick ENCDC in culture. Interestingly, ibuprofen did not affect ENCDC migration until after at least three hours of exposure. Furthermore, mice deficient in Ptgs1 (COX 1) and Ptgs2 (COX 2) had normal bowel colonization by ENCDC and normal ENCDC migration in vitro suggesting COX-independent effects. Consistent with selective and strain specific effects on ENCDC, ibuprofen did not affect migration of gut mesenchymal cells, NIH3T3, or WT C57BL/6 ENCDC, and did not affect dorsal root ganglion cell precursor migration in zebrafish. Thus, ibuprofen inhibits ENCDC migration in vitro and bowel colonization by ENCDC in vivo in zebrafish, mouse and chick, but there are cell type and strain specific responses. These data raise concern that ibuprofen may increase Hirschsprung disease risk in some genetically susceptible children. PMID:26586201

Dual cameras acquisition and display system of retina-like sensor camera and rectangular sensor camera

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Cao, Nan; Cao, Fengmei; Lin, Yabin; Bai, Tingzhu; Song, Shengyu

2015-04-01

For a new kind of retina-like senor camera and a traditional rectangular sensor camera, dual cameras acquisition and display system need to be built. We introduce the principle and the development of retina-like senor. Image coordinates transformation and interpolation based on sub-pixel interpolation need to be realized for our retina-like sensor's special pixels distribution. The hardware platform is composed of retina-like senor camera, rectangular sensor camera, image grabber and PC. Combined the MIL and OpenCV library, the software program is composed in VC++ on VS 2010. Experience results show that the system can realizes two cameras' acquisition and display.

Frequency spectrum might act as communication code between retina and visual cortex I.

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Yang, Xu; Gong, Bo; Lu, Jian-Wei

2015-01-01

To explore changes and possible communication relationship of local potential signals recorded simultaneously from retina and visual cortex I (V1). Fourteen C57BL/6J mice were measured with pattern electroretinogram (PERG) and pattern visually evoked potential (PVEP) and fast Fourier transform has been used to analyze the frequency components of those signals. The amplitude of PERG and PVEP was measured at about 36.7 µV and 112.5 µV respectively and the dominant frequency of PERG and PVEP, however, stay unchanged and both signals do not have second, or otherwise, harmonic generation. The results suggested that retina encodes visual information in the way of frequency spectrum and then transfers it to primary visual cortex. The primary visual cortex accepts and deciphers the input visual information coded from retina. Frequency spectrum may act as communication code between retina and V1.

Zebrafish embryos as models for embryotoxic and teratological effects of chemicals.

NARCIS (Netherlands)

Yang, Lixin; Ho, Nga Yu; Alshut, Rüdiger; Legradi, J.B.; Weiss, Carsten; Reischl, Markus; Mikut, Ralf; Liebel, Urban; Müller, Ferenc; Strähle, Uwe

2009-01-01

The experimental virtues of the zebrafish embryo such as small size, development outside of the mother, cheap maintenance of the adult made the zebrafish an excellent model for phenotypic genetic and more recently also chemical screens. The availability of a genome sequence and several thousand

Characterization of sleep in zebrafish and insomnia in hypocretin receptor mutants.

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Tohei Yokogawa

2007-10-01

Full Text Available Sleep is a fundamental biological process conserved across the animal kingdom. The study of how sleep regulatory networks are conserved is needed to better understand sleep across evolution. We present a detailed description of a sleep state in adult zebrafish characterized by reversible periods of immobility, increased arousal threshold, and place preference. Rest deprivation using gentle electrical stimulation is followed by a sleep rebound, indicating homeostatic regulation. In contrast to mammals and similarly to birds, light suppresses sleep in zebrafish, with no evidence for a sleep rebound. We also identify a null mutation in the sole receptor for the wake-promoting neuropeptide hypocretin (orexin in zebrafish. Fish lacking this receptor demonstrate short and fragmented sleep in the dark, in striking contrast to the excessive sleepiness and cataplexy of narcolepsy in mammals. Consistent with this observation, we find that the hypocretin receptor does not colocalize with known major wake-promoting monoaminergic and cholinergic cell groups in the zebrafish. Instead, it colocalizes with large populations of GABAergic neurons, including a subpopulation of Adra2a-positive GABAergic cells in the anterior hypothalamic area, neurons that could assume a sleep modulatory role. Our study validates the use of zebrafish for the study of sleep and indicates molecular diversity in sleep regulatory networks across vertebrates.

Chronic perfluorooctane sulfonate (PFOS) exposure induces hepatic steatosis in zebrafish

International Nuclear Information System (INIS)

Cheng, Jiangfei; Lv, Suping; Nie, Shangfei; Liu, Jing; Tong, Shoufang; Kang, Ning; Xiao, Yanyan; Dong, Qiaoxiang; Huang, Changjiang; Yang, Dongren

2016-01-01

Highlights: • PFOS chronic exposure induces sex-dependent hepatic steotosis in zebrafish. • PFOS interferes with β-oxidation, lipid synthesis, and lipid hepatic export process. • Zebrafish could be used as an alternative model for PFOS chronic toxicity screening. - Abstract: Perfluorooctane sulfonate (PFOS), one persistent organic pollutant, has been widely detected in the environment, wildlife and human. Currently few studies have documented the effects of chronic PFOS exposure on lipid metabolism, especially in aquatic organisms. The underlying mechanisms of hepatotoxicity induced by chronic PFOS exposure are still largely unknown. The present study defined the effects of chronic exposure to low level of PFOS on lipid metabolism using zebrafish as a model system. Our findings revealed a severe hepatic steatosis in the liver of males treated with 0.5 μM PFOS as evidenced by hepatosomatic index, histological assessment and liver lipid profiles. Quantitative PCR assay further indicated that PFOS significantly increase the transcriptional expression of nuclear receptors (nr1h3, rara, rxrgb, nr1l2) and the genes associated with fatty acid oxidation (acox1, acadm, cpt1a). In addition, chronic PFOS exposure significantly decreased liver ATP content and serum level of VLDL/LDL lipoprotein in males. Taken together, these findings suggest that chronic PFOS exposure induces hepatic steatosis in zebrafish via disturbing lipid biosynthesis, fatty acid β-oxidation and excretion of VLDL/LDL lipoprotein, and also demonstrate the validity of using zebrafish as an alternative model for PFOS chronic toxicity screening.

Chronic perfluorooctane sulfonate (PFOS) exposure induces hepatic steatosis in zebrafish

Energy Technology Data Exchange (ETDEWEB)

Cheng, Jiangfei; Lv, Suping; Nie, Shangfei; Liu, Jing; Tong, Shoufang; Kang, Ning; Xiao, Yanyan; Dong, Qiaoxiang [Zhejiang Provincial Key Laboratory for Technology and Application of Model Organisms (China); Institute of Environmental Safety and Human Health, Wenzhou Medical University, Wenzhou, 325035 (China); Huang, Changjiang, E-mail: cjhuang5711@163.com [Zhejiang Provincial Key Laboratory for Technology and Application of Model Organisms (China); Institute of Environmental Safety and Human Health, Wenzhou Medical University, Wenzhou, 325035 (China); Yang, Dongren, E-mail: yangdongren@yahoo.com [Zhejiang Provincial Key Laboratory for Technology and Application of Model Organisms (China); Institute of Environmental Safety and Human Health, Wenzhou Medical University, Wenzhou, 325035 (China)

2016-07-15

Highlights: • PFOS chronic exposure induces sex-dependent hepatic steotosis in zebrafish. • PFOS interferes with β-oxidation, lipid synthesis, and lipid hepatic export process. • Zebrafish could be used as an alternative model for PFOS chronic toxicity screening. - Abstract: Perfluorooctane sulfonate (PFOS), one persistent organic pollutant, has been widely detected in the environment, wildlife and human. Currently few studies have documented the effects of chronic PFOS exposure on lipid metabolism, especially in aquatic organisms. The underlying mechanisms of hepatotoxicity induced by chronic PFOS exposure are still largely unknown. The present study defined the effects of chronic exposure to low level of PFOS on lipid metabolism using zebrafish as a model system. Our findings revealed a severe hepatic steatosis in the liver of males treated with 0.5 μM PFOS as evidenced by hepatosomatic index, histological assessment and liver lipid profiles. Quantitative PCR assay further indicated that PFOS significantly increase the transcriptional expression of nuclear receptors (nr1h3, rara, rxrgb, nr1l2) and the genes associated with fatty acid oxidation (acox1, acadm, cpt1a). In addition, chronic PFOS exposure significantly decreased liver ATP content and serum level of VLDL/LDL lipoprotein in males. Taken together, these findings suggest that chronic PFOS exposure induces hepatic steatosis in zebrafish via disturbing lipid biosynthesis, fatty acid β-oxidation and excretion of VLDL/LDL lipoprotein, and also demonstrate the validity of using zebrafish as an alternative model for PFOS chronic toxicity screening.

Hypothalamic Projections to the Optic Tectum in Larval Zebrafish

Science.gov (United States)

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

2018-01-01

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

In Vivo Nanotoxicity Testing using the Zebrafish Embryo Assay.

Science.gov (United States)

Rizzo, Larissa Y; Golombek, Susanne K; Mertens, Marianne E; Pan, Yu; Laaf, Dominic; Broda, Janine; Jayapaul, Jabadurai; Möckel, Diana; Subr, Vladimir; Hennink, Wim E; Storm, Gert; Simon, Ulrich; Jahnen-Dechent, Willi; Kiessling, Fabian; Lammers, Twan

2013-06-10

Nanoparticles are increasingly used for biomedical purposes. Many different diagnostic and therapeutic applications are envisioned for nanoparticles, but there are often also serious concerns regarding their safety. Given the fact that numerous new nanomaterials are being developed every day, and that not much is known about the long-term toxicological impact of exposure to nanoparticles, there is an urgent need to establish efficient methods for nanotoxicity testing. The zebrafish (Danio rerio) embryo assay has recently emerged as an interesting 'intermediate' method for in vivo nanotoxicity screening, enabling (semi-) high-throughput analyses in a system significantly more complex than cultured cells, but at the same time also less 'invasive' and less expensive than large-scale biocompatibility studies in mice or rats. The zebrafish embryo assay is relatively well-established in the environmental sciences, but it has not yet gained wide notice in the nanomedicine field. Using prototypic polymeric drug carriers, gold-based nanodiagnostics and nanotherapeutics, and iron oxide-based nanodiagnostics, we here show that toxicity testing using zebrafish embryos is easy, efficient and informative, and faithfully reflects, yet significantly extends, cell-based toxicity testing. We therefore expect that the zebrafish embryo assay will become a popular future tool for in vivo nanotoxicity screening.

Melatonin mitigates neomycin-induced hair cell injury in zebrafish.

Science.gov (United States)

Oh, Kyoung Ho; Rah, Yoon Chan; Hwang, Kyu Ho; Lee, Seung Hoon; Kwon, Soon Young; Cha, Jae Hyung; Choi, June

2017-10-01

Ototoxicity due to medications, such as aminoglycosides, is irreversible, and free radicals in the inner ear are assumed to play a major role. Because melatonin has an antioxidant property, we hypothesize that it might mitigate hair cell injury by aminoglycosides. The objective of this study was to evaluate whether melatonin has an alleviative effect on neomycin-induced hair cell injury in zebrafish (Danio rerio). Various concentrations of melatonin were administered to 5-day post-fertilization zebrafish treated with 125 μM neomycin for 1 h. Surviving hair cells within four neuromasts were compared with that of a control group. Apoptosis was assessed via terminal deoxynucleotidyl transferase dUTP nick-end labeling assay. The changes of ultrastructure were confirmed using a scanning electron microscope. Melatonin alleviated neomycin-induced hair cell injury in neuromasts (neomycin + melatonin 100 μM: 13.88 ± 0.91 cells, neomycin only: 7.85 ± 0.90 cells; n = 10, p melatonin for 1 h in SEM findings. Melatonin is effective in alleviating aminoglycoside-induced hair cell injury in zebrafish. The results of this study demonstrated that melatonin has the potential to reduce apoptosis induced by aminoglycosides in zebrafish.

A Fully Automated High-Throughput Zebrafish Behavioral Ototoxicity Assay.

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Todd, Douglas W; Philip, Rohit C; Niihori, Maki; Ringle, Ryan A; Coyle, Kelsey R; Zehri, Sobia F; Zabala, Leanne; Mudery, Jordan A; Francis, Ross H; Rodriguez, Jeffrey J; Jacob, Abraham

2017-08-01

Zebrafish animal models lend themselves to behavioral assays that can facilitate rapid screening of ototoxic, otoprotective, and otoregenerative drugs. Structurally similar to human inner ear hair cells, the mechanosensory hair cells on their lateral line allow the zebrafish to sense water flow and orient head-to-current in a behavior called rheotaxis. This rheotaxis behavior deteriorates in a dose-dependent manner with increased exposure to the ototoxin cisplatin, thereby establishing itself as an excellent biomarker for anatomic damage to lateral line hair cells. Building on work by our group and others, we have built a new, fully automated high-throughput behavioral assay system that uses automated image analysis techniques to quantify rheotaxis behavior. This novel system consists of a custom-designed swimming apparatus and imaging system consisting of network-controlled Raspberry Pi microcomputers capturing infrared video. Automated analysis techniques detect individual zebrafish, compute their orientation, and quantify the rheotaxis behavior of a zebrafish test population, producing a powerful, high-throughput behavioral assay. Using our fully automated biological assay to test a standardized ototoxic dose of cisplatin against varying doses of compounds that protect or regenerate hair cells may facilitate rapid translation of candidate drugs into preclinical mammalian models of hearing loss.

Hypothalamic Projections to the Optic Tectum in Larval Zebrafish

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Lucy A. Heap

2018-01-01

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

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

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Brian eGriffiths

2012-10-01

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

Spatial pattern of cell geometry and cell-division orientation in zebrafish lens epithelium

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Toshiaki Mochizuki

2014-09-01

Full Text Available Cell proliferation is a key regulator of tissue morphogenesis. We examined cell proliferation and cell division in zebrafish lens epithelium by visualizing cell-cycle phases and nuclear positions, using fluorescent-labeled geminin and histone proteins. Proliferation was low in the anterior region of lens epithelium and higher in the marginal zone anterior to the equator, suggesting that the proliferation zone, called the germinative zone, is formed in zebrafish lens. Interestingly, cell-division orientation was biased longitudinally in the anterior region, shifted from longitudinal to circumferential along the anterior–posterior axis of lens sphere, and was biased circumferentially in the peripheral region. These data suggest that cell-division orientation is spatially regulated in zebrafish lens epithelium. The Hertwig rule indicates that cells tend to divide along their long axes. Orientation of long axes and cell division were biased similarly in zebrafish lens epithelium, suggesting that cell geometry correlates with cell-division orientation. A cell adhesion molecule, E-cadherin, is expressed in lens epithelium. In a zebrafish e-cadherin mutant, the long axes and cell-division orientation were shifted more longitudinally. These data suggest that E-cadherin is required for the spatial pattern of cell geometry and cell-division orientation in zebrafish lens epithelium.

Zebrafish as an In Vivo Model to Assess Epigenetic Effects of Ionizing Radiation

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Eva Yi Kong

2016-12-01

Full Text Available Exposure to ionizing radiations (IRs is ubiquitous in our environment and can be categorized into “targeted” effects and “non-targeted” effects. In addition to inducing deoxyribonucleic acid (DNA damage, IR exposure leads to epigenetic alterations that do not alter DNA sequence. Using an appropriate model to study the biological effects of radiation is crucial to better understand IR responses as well as to develop new strategies to alleviate exposure to IR. Zebrafish, Danio rerio, is a scientific model organism that has yielded scientific advances in several fields and recent studies show the usefulness of this vertebrate model in radiation biology. This review briefly describes both “targeted” and “non-targeted” effects, describes the findings in radiation biology using zebrafish as a model and highlights the potential of zebrafish to assess the epigenetic effects of IR, including DNA methylation, histone modifications and miRNA expression. Other in vivo models are included to compare observations made with zebrafish, or to illustrate the feasibility of in vivo models when the use of zebrafish was unavailable. Finally, tools to study epigenetic modifications in zebrafish, including changes in genome-wide DNA methylation, histone modifications and miRNA expression, are also described in this review.

LITTLE FISH, BIG DATA: ZEBRAFISH AS A MODEL FOR CARDIOVASCULAR AND METABOLIC DISEASE.

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Gut, Philipp; Reischauer, Sven; Stainier, Didier Y R; Arnaout, Rima

2017-07-01

The burden of cardiovascular and metabolic diseases worldwide is staggering. The emergence of systems approaches in biology promises new therapies, faster and cheaper diagnostics, and personalized medicine. However, a profound understanding of pathogenic mechanisms at the cellular and molecular levels remains a fundamental requirement for discovery and therapeutics. Animal models of human disease are cornerstones of drug discovery as they allow identification of novel pharmacological targets by linking gene function with pathogenesis. The zebrafish model has been used for decades to study development and pathophysiology. More than ever, the specific strengths of the zebrafish model make it a prime partner in an age of discovery transformed by big-data approaches to genomics and disease. Zebrafish share a largely conserved physiology and anatomy with mammals. They allow a wide range of genetic manipulations, including the latest genome engineering approaches. They can be bred and studied with remarkable speed, enabling a range of large-scale phenotypic screens. Finally, zebrafish demonstrate an impressive regenerative capacity scientists hope to unlock in humans. Here, we provide a comprehensive guide on applications of zebrafish to investigate cardiovascular and metabolic diseases. We delineate advantages and limitations of zebrafish models of human disease and summarize their most significant contributions to understanding disease progression to date. Copyright © 2017 the American Physiological Society.

The role of Sox6 in zebrafish muscle fiber type specification.

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

2015-01-01

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

Ethanol Exposure Causes Muscle Degeneration in Zebrafish

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Elizabeth C. Coffey

2018-03-01

Full Text Available Alcoholic myopathies are characterized by neuromusculoskeletal symptoms such as compromised movement and weakness. Although these symptoms have been attributed to neurological damage, EtOH may also target skeletal muscle. EtOH exposure during zebrafish primary muscle development or adulthood results in smaller muscle fibers. However, the effects of EtOH exposure on skeletal muscle during the growth period that follows primary muscle development are not well understood. We determined the effects of EtOH exposure on muscle during this phase of development. Strikingly, muscle fibers at this stage are acutely sensitive to EtOH treatment: EtOH induces muscle degeneration. The severity of EtOH-induced muscle damage varies but muscle becomes more refractory to EtOH as muscle develops. NF-kB induction in muscle indicates that EtOH triggers a pro-inflammatory response. EtOH-induced muscle damage is p53-independent. Uptake of Evans blue dye shows that EtOH treatment causes sarcolemmal instability before muscle fiber detachment. Dystrophin-null sapje mutant zebrafish also exhibit sarcolemmal instability. We tested whether Trichostatin A (TSA, which reduces muscle degeneration in sapje mutants, would affect EtOH-treated zebrafish. We found that TSA and EtOH are a lethal combination. EtOH does, however, exacerbate muscle degeneration in sapje mutants. EtOH also disrupts adhesion of muscle fibers to their extracellular matrix at the myotendinous junction: some detached muscle fibers retain beta-Dystroglycan indicating failure of muscle end attachments. Overexpression of Paxillin, which reduces muscle degeneration in zebrafish deficient for beta-Dystroglycan, is not sufficient to rescue degeneration. Taken together, our results suggest that EtOH exposure has pleiotropic deleterious effects on skeletal muscle.

Innovative Disease Model: Zebrafish as an In Vivo Platform for Intestinal Disorder and Tumors

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Jeng-Wei Lu

2017-09-01

Full Text Available Colorectal cancer (CRC is one of the world’s most common cancers and is the second leading cause of cancer deaths, causing more than 50,000 estimated deaths each year. Several risk factors are highly associated with CRC, including being overweight, eating a diet high in red meat and over-processed meat, having a history of inflammatory bowel disease, and smoking. Previous zebrafish studies have demonstrated that multiple oncogenes and tumor suppressor genes can be regulated through genetic or epigenetic alterations. Zebrafish research has also revealed that the activation of carcinogenesis-associated signal pathways plays an important role in CRC. The biology of cancer, intestinal disorders caused by carcinogens, and the morphological patterns of tumors have been found to be highly similar between zebrafish and humans. Therefore, the zebrafish has become an important animal model for translational medical research. Several zebrafish models have been developed to elucidate the characteristics of gastrointestinal diseases. This review article focuses on zebrafish models that have been used to study human intestinal disorders and tumors, including models involving mutant and transgenic fish. We also report on xenograft models and chemically-induced enterocolitis. This review demonstrates that excellent zebrafish models can provide novel insights into the pathogenesis of gastrointestinal diseases and help facilitate the evaluation of novel anti-tumor drugs.

Frequency spectrum might act as communication code between retina and visual cortex I

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Xu Yang

2015-12-01

Full Text Available AIM: To explore changes and possible communication relationship of local potential signals recorded simultaneously from retina and visual cortex I (V1. METHODS: Fourteen C57BL/6J mice were measured with pattern electroretinogram (PERG and pattern visually evoked potential (PVEP and fast Fourier transform has been used to analyze the frequency components of those signals. RESULTS: The amplitude of PERG and PVEP was measured at about 36.7 µV and 112.5 µV respectively and the dominant frequency of PERG and PVEP, however, stay unchanged and both signals do not have second, or otherwise, harmonic generation. CONCLUSION: The results suggested that retina encodes visual information in the way of frequency spectrum and then transfers it to primary visual cortex. The primary visual cortex accepts and deciphers the input visual information coded from retina. Frequency spectrum may act as communication code between retina and V1.

Aspartame-fed zebrafish exhibit acute deaths with swimming defects and saccharin-fed zebrafish have elevation of cholesteryl ester transfer protein activity in hypercholesterolemia.

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Kim, Jae-Yong; Seo, Juyi; Cho, Kyung-Hyun

2011-11-01

Although many artificial sweeteners (AS) have safety issues, the AS have been widely used in industry. To determine the physiologic effect of AS in the presence of hyperlipidemia, zebrafish were fed aspartame or saccharin with a high-cholesterol diet (HCD). After 12 days, 30% of zebrafish, which consumed aspartame and HCD, died with exhibiting swimming defects. The aspartame group had 65% survivability, while the control and saccharin groups had 100% survivability. Under HCD, the saccharin-fed groups had the highest increase in the serum cholesterol level (599 mg/dL). Aspartame-fed group showed a remarkable increase in serum glucose (up to 125 mg/dL), which was 58% greater than the increase in the HCD alone group. The saccharin and HCD groups had the highest cholesteryl ester transfer protein (CETP) activity (52% CE-transfer), while the HCD alone group had 42% CE-transfer. Histologic analysis revealed that the aspartame and HCD groups showed more infiltration of inflammatory cells in the brain and liver sections. Conclusively, under presence of hyperlipidemia, aspartame-fed zebrafish exhibited acute swimming defects with an increase in brain inflammation. Saccharin-fed zebrafish had an increased atherogenic serum lipid profile with elevation of CETP activity. Copyright © 2011 Elsevier Ltd. All rights reserved.