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Tritium is a betta emitter radionuclide. Being an isotope of hydrogen, it is easily transferred to different environmental compartments, and to human and non-human biota. Considering that tritium levels are expected to rise in the upcoming decades with the development of nuclear facilities producing tritium using fission processes, investigating the potential toxicity of tritium to human and non-human biota is necessary. Tritiated thymidine, an organic form of tritium, has been used in this study to assess its toxicity on fish embryo development. Zebrafish embryos (3.5 hpf; hours post fertilization) have been exposed to tritiated thymidine at three different activity concentrations (7.5; 40; 110 kBq/mL) for four days. These experiments highlighted that zebrafish development was affected by the exposure to organic tritium, with smaller larvae at 3 dpf after exposure to the two lowest dose rates (22 and 170 µGy/h), a delayed hatching after exposure to the two highest dose rates (170 and 470 µGy/h), an increase in the spontaneous tail movement (1 dpf) and a decrease in the heartbeat (3 dpf) after exposure to the highest dose rate. The results also highlighted an increase in ROS production in larvae exposed to the intermediate dose rate. A dysregulation of many genes, involved in apoptosis, DNA repair or oxidative stress, was also found after 1 day of exposure to the lowest tritium dose rate. Our results thus suggest that exposure to tritiated thymidine from a dose rate as low as 22 µGy/h can lead to sublethal effects, with an effect on the development, dysregulation of many genes and increase of the ROS production. This paper provides valuable information on toxic effects arising from the exposure of fish to an organic form of tritium, which was the main objective of this study., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Recent laboratory studies focusing on multigenerational approach demonstrated drastic phenotypic effects after chronic fish irradiation exposure. No irradiation effect at phenotypic scale was observed for F0 (reproductive performances) while early mortality and malformations were observed in F1 offspring whether they were irradiated or not. The objective was to study molecular mechanisms likely to be involved in these phenotypic effects induced by parental irradiation. Thus, F0 adult zebrafish were irradiated for ten days until reproduction and maternal involvement in offspring development was assessed. Levels of maternal provided cortisol and vitellogenin, needed for embryo development, were not impacted by irradiation. However, maternal transcriptome highlighted irradiation effect on processes involved in oocyte development, as well as on essential maternal factors needed for offspring development. Therefore, this study highlighted the importance of parental exposure on offspring fate and of the importance of multigenerational exposure in risk assessment., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Embryonic development is particularly vulnerable to stress and DNA damage, as mutations can accumulate through cell proliferation in a wide number of cells and organs. However, the biological effects of chronic exposure to ionising radiation (IR) at low and moderate dose rates (< 6 mGy/h) remain largely controversial, raising concerns for environmental protection. The present study focuses on the molecular effects of IR (0.005 to 50 mGy/h) on zebrafish embryos at the gastrula stage (6 hpf), at both the transcriptomics and epigenetics levels. Our results show that exposure to IR modifies the expression of genes involved in mitochondrial activity from 0.5 to 50 mGy/h. In addition, important developmental pathways, namely, the Notch, retinoic acid, BMP and Wnt signalling pathways, were altered at 5 and 50 mGy/h. Transcriptional changes of genes involved in the morphogenesis of the ectoderm and mesoderm were detected at all dose rates, but were prominent from 0.5 to 50 mGy/h. At the epigenetic level, exposure to IR induced a hypomethylation of DNA in the promoter of genes that colocalised with both H3K27me3 and H3Kme4 histone marks and correlated with changes in transcriptional activity. Finally, pathway enrichment analysis demonstrated that the DNA methylation changes occurred in the promoter of important developmental genes, including morphogenesis of the ectoderm and mesoderm. Together, these results show that the transcriptional program regulating morphogenesis in gastrulating embryos was modified at dose rates greater than or equal to 0.5 mGy/h, which might predict potential neurogenesis and somitogenesis defects observed at similar dose rates later in development.

Promoting the regeneration or survival of retinal ganglion cells (RGCs) is one focus of regenerative medicine. Homeobox Barhl transcription factors might be instrumental in these processes. In mammals, only barhl2 is expressed in the retina and is required for both subtype identity acquisition of amacrine cells and for the survival of RGCs downstream of Atoh7, a transcription factor necessary for RGC genesis. The underlying mechanisms of this dual role of Barhl2 in mammals have remained elusive. Whole genome duplication in the teleost lineage generated the barhl1a and barhl2 paralogues. In the Zebrafish retina, Barhl2 functions as a determinant of subsets of amacrine cells lineally related to RGCs independently of Atoh7. In contrast, barhl1a expression depends on Atoh7 but its expression dynamics and function have not been studied. Here we describe for the first time a Barhl1a reporter line in vivo showing that barhl1a turns on exclusively in subsets of RGCs and their post-mitotic precursors. We also show transient expression of barhl1a:GFP in diencephalic neurons extending their axonal projections as part of the post-optic commissure, at the time of optic chiasm formation. This work sets the ground for future studies on RGC subtype identity, axonal projections and genetic specification of Barhl1a-positive RGCs and commissural neurons.

Multigenerational studies have become of great interest in ecotoxicology since the consequence of parental exposure to contaminants on offspring generations was established in situ or in laboratory conditions. The present study mainly examined the chronic effects of external Cs-137 gamma irradiation exposure at 4 dose rates (control, 0.5, 5, and 50 mGy h -1 ) on adult zebrafish (F0) exposed for 10 d and their progeny (F1) exposed or unexposed for 4 to 5 d. The main endpoints investigated included parental reproductive performance, embryo-larval survival, DNA alterations, and reactive oxygen species (ROS) production in F0 and F1. No effects on reproductive success, fecundity, or egg fertilization rate were observed. However, drastic effects were observed on F1 exposed to 50 mGy h -1 , resulting in a mortality rate of 100%. The drastic effects were also observed when the progeny was not irradiated. It was demonstrated that the sensitivity of the embryos was mainly attributable to parental irradiation. Moreover, these drastic effects induced by adult irradiation disappeared over time when 10 d-irradiated adults were placed in a nonirradiated condition. Alterations in larval DNA were observed for the 3 dose rates, and an increase of ROS production was also shown for the 2 lowest dose rates. The present study improves our understanding of the consequences of parental exposure conditions to the progeny. Furthermore, it provides an incentive to take transmitted generational effects into account in ecological risk assessments. Environ Toxicol Chem 2019;38:2556-2567. © 2019 SETAC., (© 2019 SETAC.)

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

HSF1 is a transcription factor that plays a key role in circadian resetting by temperature. We have used zebrafish embryos to decipher the roles of zHsf1, heat and light on zper2 transcription in vivo. Our results show that heat shock (HS) stimulated zper2 expression in the dark but has no cumulative effect combined with light. After light exposition, zper2 expression was 2.7 fold increased threefold in the hsf1-morphants in comparison to control embryos. Our results show that zHsf1 plays a positive role in HS-driven expression of zper2 in the dark but seems to act as an attenuator in the presence light.

Uranium is an actinide naturally found in the environment. Anthropogenic activities lead to the release of increasing amounts of uranium and depleted uranium (DU) in the environment, posing potential risks to aquatic organisms due to radiological and chemical toxicity of this radionucleide. Although environmental contaminations with high levels of uranium have already been observed, chronic exposures of non-human species to levels close to the environmental quality standards remain scarcely characterized. The present study focused on the identification of the molecular pathways impacted by a chronic exposure of zebrafish to 20 μg/L of DU during 10 days. The transcriptomic effects were evaluated by the use of the mRNAseq analysis in three organs of adult zebrafish, the brain the testis and the ovaries, and two developmental stages of the adult fish progeny, two-cells embryo and four-days larvae. The results highlight generic effects on the cell adhesion process, but also specific transcriptomic responses depending on the organ or the developmental stage investigated. The analysis of the transgenerational effects of DU-exposure on the four-day zebrafish larvae demonstrate an induction of genes involved in oxidative response (cat, mpx, sod1 and sod2), a decrease of expression of the two hatching enzymes (he1a and he1b), the deregulation of the expression of gene coding for the ATPase complex and the induction of cellular stress. Electron microscopy analysis of skeletal muscles on the four-days larvae highlights significant histological impacts on the ultrastructure of both the mitochondria and the myofibres. In addition, the comparison with the transcriptomic data obtained for the acetylcholine esterase mutant reveals the induction of protein-chaperons in the skeletal muscles of the progeny of fish chronically exposed to DU, pointing towards long lasting effects of this chemical in the muscles. The results presented in this study support the hypothesis that a chronic parental exposure to an environmentally relevant concentration of DU could impair the progeny development with significant effects observed both at the molecular level and on the histological ultrastructure of organs. This study provides a comprehensive transcriptomic dataset useful for ecotoxicological studies on other fish species at the molecular level. It also provides a key DU responsive gene, egr1, which may be a candidate biomarker for monitoring aquatic pollution by heavy metals.

This study investigated the effects of adult zebrafish exposure to a nominal concentration of 20μgL -1 of depleted uranium (DU) for six days upon DNA methylation, gene expression and the appearance of histopathological damage in their progeny. In the embryos at the 2-8 cell stage, the parental exposure induced significant DU accumulation, with levels seven times higher than those measured in the control embryos, but in larvae 96h post-fertilisation (hpf), uranium concentration had already returned to a level identical to that of the control larvae. A significant two-fold increase in the global level of DNA methylation was observed in embryos as early as the prim5 (24 hpf) stage and was still maintained at the 96 hpf stage despite the fact that DU had already been depurated at the later stage. RNA sequencing analysis indicated an impact of parental exposure upon the total RNAs transmitted from the mother to eggs, and the up-regulated genes were those associated with post-traductional protein modification and trafficking and cellular signalling pathways, whereas the down-regulated genes concerned the translational process, cell cycle regulation and several cell signalling pathways. Alterations of photoreceptor cells and the axon-axon junctions between photoreceptors were observed in the eyes of adult fish exposed for 10days to DU. Actin and myosin filament disorganisation was observed in the skeletal muscles of 96 hpf larvae, at a stage when the maternally transmitted DU had already been excreted. These data reveal the extreme sensitivity of zebrafish embryos to DU transmitted through the oocyte by exposed females., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The zebrafish has become a model to study adult vertebrate neurogenesis. In particular, the adult telencephalon has been an intensely studied structure in the zebrafish brain. Differential expression of transcriptional regulators (TRs) is a key feature of development and tissue homeostasis. Here we report an expression map of 1,202 TR genes in the telencephalon of adult zebrafish. Our results are summarized in a database with search and clustering functions to identify genes expressed in particular regions of the telencephalon. We classified 562 genes into 13 distinct patterns, including genes expressed in the proliferative zone. The remaining 640 genes displayed unique and complex patterns of expression and could thus not be grouped into distinct classes. The neurogenic ventricular regions express overlapping but distinct sets of TR genes, suggesting regional differences in the neurogenic niches in the telencephalon. In summary, the small telencephalon of the zebrafish shows a remarkable complexity in TR gene expression. The adult zebrafish telencephalon has become a model to study neurogenesis. We established the expression pattern of more than 1200 transcription regulators (TR) in the adult telencephalon. The neurogenic regions express overlapping but distinct sets of TR genes suggesting regional differences in the neurogenic potential., (© 2015 Wiley Periodicals, Inc.)

The teleost brain has the remarkable ability to generate new neurons and to repair injuries during adult life stages. Maintaining life-long neurogenesis requires careful management of neural stem cell pools. In a genome-wide expression screen for transcription regulators, the id1 gene, encoding a negative regulator of E-proteins, was found to be upregulated in response to injury. id1 expression was mapped to quiescent type I neural stem cells in the adult telencephalic stem cell niche. Gain and loss of id1 function in vivo demonstrated that Id1 promotes stem cell quiescence. The increased id1 expression observed in neural stem cells in response to injury appeared independent of inflammatory signals, suggesting multiple antagonistic pathways in the regulation of reactive neurogenesis. Together, we propose that Id1 acts to maintain the neural stem cell pool by counteracting neurogenesis-promoting signals., (© 2014 AlphaMed Press.)

The cornea is a central component of the camera eye of vertebrates and even slight corneal disturbances severely affect vision. The transcription factor PAX6 is required for normal eye development, namely the proper separation of the lens from the developing cornea and the formation of the iris and anterior chamber. Human PAX6 mutations are associated with severe ocular disorders such as aniridia, Peters anomaly and chronic limbal stem cell insufficiency. To develop the zebrafish as a model for corneal disease, we first performed transcriptome and in situ expression analysis to identify marker genes to characterise the cornea in normal and pathological conditions. We show that, at 7 days post fertilisation (dpf), the zebrafish cornea expresses the majority of marker genes (67/84 tested genes) found also expressed in the cornea of juvenile and adult stages. We also characterised homozygous pax6b mutants. Mutant embryos have a thick cornea, iris hypoplasia, a shallow anterior chamber and a small lens. Ultrastructure analysis revealed a disrupted corneal endothelium. pax6b mutants show loss of corneal epithelial gene expression including regulatory genes (sox3, tfap2a, foxc1a and pitx2). In contrast, several genes (pitx2, ctnnb2, dcn and fabp7a) were ectopically expressed in the malformed corneal endothelium. Lack of pax6b function leads to severe disturbance of the corneal gene regulatory programme.

Neutrophils are the most abundant polymorphonuclear leukocytes, presenting the first line of defence against infection or tissue damage. To characterize the molecular changes on the protein level in neutrophils during sterile inflammation we established the chemically-induced inflammation (ChIn) assay in adult zebrafish and investigated the proteome dynamics within neutrophils of adult zebrafish upon inflammation. Through label-free proteomics we identified 48 proteins that were differentially regulated during inflammation. Gene ontology analysis revealed that these proteins were associated with cell cycle, nitric oxide signalling, regulation of cytoskeleton rearrangement and intermediate filaments as well as immune-related processes such as antigen presentation, leucocyte chemotaxis and IL-6 signalling. Comparison of protein expression dynamics with transcript expression dynamics suggests the existence of regulatory mechanisms confined to the protein level for some genes. This is the first proteome analysis of adult zebrafish neutrophils upon chemically-induced inflammation providing a valuable reference for future studies using zebrafish inflammation models., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

A core promoter is a stretch of DNA surrounding the transcription start site (TSS) that integrates regulatory inputs and recruits general transcription factors to initiate transcription. The nature and causative relationship of the DNA sequence and chromatin signals that govern the selection of most TSSs by RNA polymerase II remain unresolved. Maternal to zygotic transition represents the most marked change of the transcriptome repertoire in the vertebrate life cycle. Early embryonic development in zebrafish is characterized by a series of transcriptionally silent cell cycles regulated by inherited maternal gene products: zygotic genome activation commences at the tenth cell cycle, marking the mid-blastula transition. This transition provides a unique opportunity to study the rules of TSS selection and the hierarchy of events linking transcription initiation with key chromatin modifications. We analysed TSS usage during zebrafish early embryonic development at high resolution using cap analysis of gene expression, and determined the positions of H3K4me3-marked promoter-associated nucleosomes. Here we show that the transition from the maternal to zygotic transcriptome is characterized by a switch between two fundamentally different modes of defining transcription initiation, which drive the dynamic change of TSS usage and promoter shape. A maternal-specific TSS selection, which requires an A/T-rich (W-box) motif, is replaced with a zygotic TSS selection grammar characterized by broader patterns of dinucleotide enrichments, precisely aligned with the first downstream (+1) nucleosome. The developmental dynamics of the H3K4me3-marked nucleosomes reveal their DNA-sequence-associated positioning at promoters before zygotic transcription and subsequent transcription-independent adjustment to the final position downstream of the zygotic TSS. The two TSS-defining grammars coexist, often physically overlapping, in core promoters of constitutively expressed genes to enable their expression in the two regulatory environments. The dissection of overlapping core promoter determinants represents a framework for future studies of promoter structure and function across different regulatory contexts.

The precise spatial and temporal control of gene expression is a key process in the development, maintenance and regeneration of the vertebrate body. A substantial proportion of vertebrate genomes encode genes that control the transcription of the genetic information into mRNA. The zebrafish is particularly well suited to investigate gene regulatory networks underlying the control of gene expression during development due to the external development of its transparent embryos and the increasingly sophisticated tools for genetic manipulation available for this model system. We review here recent data on the analysis of cis-regulatory modules, transcriptional regulators and their integration into gene regulatory networks in the zebrafish, using the developing spinal cord as example.

Spatiotemporal control of gene expression is central to animal development. Core promoters represent a previously unanticipated regulatory level by interacting with cis-regulatory elements and transcription initiation in different physiological and developmental contexts. Here, we provide a first and comprehensive description of the core promoter repertoire and its dynamic use during the development of a vertebrate embryo. By using cap analysis of gene expression (CAGE), we mapped transcription initiation events at single nucleotide resolution across 12 stages of zebrafish development. These CAGE-based transcriptome maps reveal genome-wide rules of core promoter usage, structure, and dynamics, key to understanding the control of gene regulation during vertebrate ontogeny. They revealed the existence of multiple classes of pervasive intra- and intergenic post-transcriptionally processed RNA products and their developmental dynamics. Among these RNAs, we report splice donor site-associated intronic RNA (sRNA) to be specific to genes of the splicing machinery. For the identification of conserved features, we compared the zebrafish data sets to the first CAGE promoter map of Tetraodon and the existing human CAGE data. We show that a number of features, such as promoter type, newly discovered promoter properties such as a specialized purine-rich initiator motif, as well as sRNAs and the genes in which they are detected, are conserved in mammalian and Tetraodon CAGE-defined promoter maps. The zebrafish developmental promoterome represents a powerful resource for studying developmental gene regulation and revealing promoter features shared across vertebrates.

Transcription is the primary step in the retrieval of genetic information. A substantial proportion of the protein repertoire of each organism consists of transcriptional regulators (TRs). It is believed that the differential expression and combinatorial action of these TRs is essential for vertebrate development and body homeostasis. We mined the zebrafish genome exhaustively for genes encoding TRs and determined their expression in the zebrafish embryo by sequencing to saturation and in situ hybridisation. At the evolutionary conserved phylotypic stage, 75% of the 3302 TR genes encoded in the genome are already expressed. The number of expressed TR genes increases only marginally in subsequent stages and is maintained during adulthood suggesting important roles of the TR genes in body homeostasis. Fewer than half of the TR genes (45%, n=1711 genes) are expressed in a tissue-restricted manner in the embryo. Transcripts of 207 genes were detected in a single tissue in the 24h embryo, potentially acting as regulators of specific processes. Other TR genes were expressed in multiple tissues. However, with the exception of certain territories in the nervous system, we did not find significant synexpression suggesting that most tissue-restricted TRs act in a freely combinatorial fashion. Our data indicate that elaboration of body pattern and function from the phylotypic stage onward relies mostly on redeployment of TRs and post-transcriptional processes., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Methyl mercury (MeHg) is a neurotoxicant with adverse effects on the development of the nervous system from fish to man. Despite a detailed understanding of the molecular mechanisms by which MeHg affects cellular homeostasis, it is still not clear how MeHg causes developmental neurotoxicity. We performed here a genome-wide transcriptional analysis of MeHg-exposed zebrafish embryos and combined this with a whole-mount in situ expression analysis of 88 MeHg-affected genes. The majority of the analyzed genes showed tissue- and region-restricted responses in various organs and tissues. The genes were linked to gene ontology terms like oxidative stress, transport and cell protection. Areas even within the central nervous system (CNS) are affected differently resulting in distinct cellular stress responses. Our study revealed an unexpected heterogeneity in gene responses to MeHg exposure in different tissues and neuronal subregions, even though the known molecular action of MeHg would predict a similar burden of exposed cells. The overall structure of the developing brain of MeHg-exposed embryos appeared normal, suggesting that the mechanism leading to differentiation of the CNS is not overtly affected by exposure to MeHg. We propose that MeHg disturbs the function of the CNS by disturbing the cellular homeostasis. As these cellular stress responses comprise genes that are also involved in normal neuronal activity and learning, MeHg may affect the developing CNS in a subtle manner that manifests itself in behavioral deficits.

Most organisms possess circadian clocks that are able to anticipate the day/night cycle and are reset or "entrained" by the ambient light. In the zebrafish, many organs and even cultured cell lines are directly light responsive, allowing for direct entrainment of the clock by light. Here, we have characterized light induced gene transcription in the zebrafish at several organizational levels. Larvae, heart organ cultures and cell cultures were exposed to 1- or 3-hour light pulses, and changes in gene expression were compared with controls kept in the dark. We identified 117 light regulated genes, with the majority being induced and some repressed by light. Cluster analysis groups the genes into five major classes that show regulation at all levels of organization or in different subset combinations. The regulated genes cover a variety of functions, and the analysis of gene ontology categories reveals an enrichment of genes involved in circadian rhythms, stress response and DNA repair, consistent with the exposure to visible wavelengths of light priming cells for UV-induced damage repair. Promoter analysis of the induced genes shows an enrichment of various short sequence motifs, including E- and D-box enhancers that have previously been implicated in light regulation of the zebrafish period2 gene. Heterologous reporter constructs with sequences matching these motifs reveal light regulation of D-box elements in both cells and larvae. Morpholino-mediated knock-down studies of two homologues of the D-box binding factor Tef indicate that these are differentially involved in the cell autonomous light induction in a gene-specific manner. These findings suggest that the mechanisms involved in period2 regulation might represent a more general pathway leading to light induced gene expression.

Zebrafish is an attractive model to investigate regeneration of the nervous system. Despite major progress in our understanding of the underlying processes, the transcriptomic changes are largely unknown. We carried out a computational analysis of the transcriptome of the regenerating telencephalon integrating changes in the expression of mRNAs, their splice variants and investigated the putative role of regulatory RNAs in the modulation of these transcriptional changes. Profound changes in the expression of genes and their splice variants engaged in many distinct processes were observed. Differential transcription and splicing are important processes in response to injury of the telencephalon. As exemplified by the coordinated regulation of the cholesterol synthesizing enzymes and transporters, the genome responded to injury of the telencephalon in a multi-tiered manner with distinct and interwoven changes in expression of enzymes, transporters and their regulatory molecules. This coordinated genomic response involved a decrease of the mRNA of the key transcription factor SREBF2, induction of microRNAs (miR-182 , miR-155 , miR-146 , miR-31) targeting cholesterol genes, shifts in abundance of splice variants as well as regulation of long non-coding RNAs. Cholesterol metabolism appears to be switched from synthesis to relocation of cholesterol. Based on our in silico analyses, this switch involves complementary and synergistic inputs by different regulatory principles. Our studies suggest that adaptation of cholesterol metabolism is a key process involved in regeneration of the injured zebrafish brain. [ABSTRACT FROM AUTHOR]

Background: Alterations in neurotransmitter phenotypes of specific neurons can cause imbalances in excitation and inhibition in the central nervous system (CNS), leading to diseases. Therefore, the correct specification and maintenance of neurotransmitter phenotypes is vital. As with other neuronal properties, neurotransmitter phenotypes are often specified and maintained by particular transcription factors. However, the specific molecular mechanisms and transcription factors that regulate neurotransmitter phenotypes remain largely unknown. Methods: In this paper we use single mutant, double mutant and transgenic zebrafish embryos to elucidate the functions of Lmx1ba and Lmx1bb in the regulation of spinal cord interneuron neurotransmitter phenotypes. Results: We demonstrate that lmx1ba and lmx1bb are both expressed in zebrafish spinal cord and that lmx1bb is expressed by both V0v cells and dI5 cells. Our functional analyses demonstrate that these transcription factors are not required for neurotransmitter fate specification at early stages of development, but that in embryos with at least two lmx1ba and/or lmx1bb mutant alleles there is a reduced number of excitatory (glutamatergic) spinal interneurons at later stages of development. In contrast, there is no change in the numbers of V0v or dI5 cells. These data suggest that lmx1b-expressing spinal neurons still form normally, but at least a subset of them lose, or do not form, their normal excitatory fates. As the reduction in glutamatergic cells is only seen at later stages of development, Lmx1b is probably required either for the maintenance of glutamatergic fates or to specify glutamatergic phenotypes of a subset of later forming neurons. Using double labeling experiments, we also show that at least some of the cells that lose their normal glutamatergic phenotype are V0v cells. Finally, we also establish that Evx1 and Evx2, two transcription factors that are required for V0v cells to acquire their excitatory neurotransmitter phenotype, are also required for lmx1ba and lmx1bb expression in these cells, suggesting that Lmx1ba and Lmx1bb act downstream of Evx1 and Evx2 in V0v cells. Conclusions: Lmx1ba and Lmx1bb function at least partially redundantly in the spinal cord and three functional lmx1b alleles are required in zebrafish for correct numbers of excitatory spinal interneurons at later developmental stages. Taken together, our data significantly enhance our understanding of how spinal cord neurotransmitter fates are regulated. [ABSTRACT FROM AUTHOR]

The teleost brain has the remarkable ability to generate new neurons and to repair injuries during adult life stages. Maintaining life-long neurogenesis requires careful management of neural stem cell pools. In a genome-wide expression screen for transcription regulators, the id1 gene, encoding a negative regulator of E-proteins, was found to be upregulated in response to injury. id1 expression was mapped to quiescent type I neural stem cells in the adult telencephalic stem cell niche. Gain and loss of id1 function in vivo demonstrated that Id1 promotes stem cell quiescence. The increased id1 expression observed in neural stem cells in response to injury appeared independent of inflammatory signals, suggesting multiple antagonistic pathways in the regulation of reactive neurogenesis. Together, we propose that Id1 acts to maintain the neural stem cell pool by counteracting neurogenesis-promoting signals. S tem C ells 2015;33:892-903 [ABSTRACT FROM AUTHOR]