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Sample records for adult zebrafish brain

  1. Electrophysiological recording in the brain of intact adult zebrafish.

    PubMed

    Johnston, Lindsey; Ball, Rebecca E; Acuff, Seth; Gaudet, John; Sornborger, Andrew; Lauderdale, James D

    2013-01-01

    Previously, electrophysiological studies in adult zebrafish have been limited to slice preparations or to eye cup preparations and electrorentinogram recordings. This paper describes how an adult zebrafish can be immobilized, intubated, and used for in vivo electrophysiological experiments, allowing recording of neural activity. Immobilization of the adult requires a mechanism to deliver dissolved oxygen to the gills in lieu of buccal and opercular movement. With our technique, animals are immobilized and perfused with habitat water to fulfill this requirement. A craniotomy is performed under tricaine methanesulfonate (MS-222; tricaine) anesthesia to provide access to the brain. The primary electrode is then positioned within the craniotomy window to record extracellular brain activity. Through the use of a multitube perfusion system, a variety of pharmacological compounds can be administered to the adult fish and any alterations in the neural activity can be observed. The methodology not only allows for observations to be made regarding changes in neurological activity, but it also allows for comparisons to be made between larval and adult zebrafish. This gives researchers the ability to identify the alterations in neurological activity due to the introduction of various compounds at different life stages. PMID:24300281

  2. Regeneration, Plasticity, and Induced Molecular Programs in Adult Zebrafish Brain

    PubMed Central

    Cosacak, Mehmet Ilyas; Papadimitriou, Christos; Kizil, Caghan

    2015-01-01

    Regenerative capacity of the brain is a variable trait within animals. Aquatic vertebrates such as zebrafish have widespread ability to renew their brains upon damage, while mammals have—if not none—very limited overall regenerative competence. Underlying cause of such a disparity is not fully evident; however, one of the reasons could be activation of peculiar molecular programs, which might have specific roles after injury or damage, by the organisms that regenerate. If this hypothesis is correct, then there must be genes and pathways that (a) are expressed only after injury or damage in tissues, (b) are biologically and functionally relevant to restoration of neural tissue, and (c) are not detected in regenerating organisms. Presence of such programs might circumvent the initial detrimental effects of the damage and subsequently set up the stage for tissue redevelopment to take place by modulating the plasticity of the neural stem/progenitor cells. Additionally, if transferable, those “molecular mechanisms of regeneration” could open up new avenues for regenerative therapies of humans in clinical settings. This review focuses on the recent studies addressing injury/damage-induced molecular programs in zebrafish brain, underscoring the possibility of the presence of genes that could be used as biomarkers of neural plasticity and regeneration. PMID:26417601

  3. BDNF Expression in Larval and Adult Zebrafish Brain: Distribution and Cell Identification

    PubMed Central

    Cacialli, Pietro; Gueguen, Marie-Madeleine; Coumailleau, Pascal; D’Angelo, Livia; Kah, Olivier; Lucini, Carla; Pellegrini, Elisabeth

    2016-01-01

    Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, has emerged as an active mediator in many essential functions in the central nervous system of mammals. BDNF plays significant roles in neurogenesis, neuronal maturation and/or synaptic plasticity and is involved in cognitive functions such as learning and memory. Despite the vast literature present in mammals, studies devoted to BDNF in the brain of other animal models are scarse. Zebrafish is a teleost fish widely known for developmental genetic studies and is emerging as model for translational neuroscience research. In addition, its brain shows many sites of adult neurogenesis allowing higher regenerative properties after traumatic injuries. To add further knowledge on neurotrophic factors in vertebrate brain models, we decided to determine the distribution of bdnf mRNAs in the larval and adult zebrafish brain and to characterize the phenotype of cells expressing bdnf mRNAs by means of double staining studies. Our results showed that bdnf mRNAs were widely expressed in the brain of 7 days old larvae and throughout the whole brain of mature female and male zebrafish. In adults, bdnf mRNAs were mainly observed in the dorsal telencephalon, preoptic area, dorsal thalamus, posterior tuberculum, hypothalamus, synencephalon, optic tectum and medulla oblongata. By combining immunohistochemistry with in situ hybridization, we showed that bdnf mRNAs were never expressed by radial glial cells or proliferating cells. By contrast, bdnf transcripts were expressed in cells with neuronal phenotype in all brain regions investigated. Our results provide the first demonstration that the brain of zebrafish expresses bdnf mRNAs in neurons and open new fields of research on the role of the BDNF factor in brain mechanisms in normal and brain repairs situations. PMID:27336917

  4. Stab wound injury of the zebrafish adult telencephalon: a method to investigate vertebrate brain neurogenesis and regeneration.

    PubMed

    Schmidt, Rebecca; Beil, Tanja; Strähle, Uwe; Rastegar, Sepand

    2014-01-01

    Adult zebrafish have an amazing capacity to regenerate their central nervous system after injury. To investigate the cellular response and the molecular mechanisms involved in zebrafish adult central nervous system (CNS) regeneration and repair, we developed a zebrafish model of adult telencephalic injury. In this approach, we manually generate an injury by pushing an insulin syringe needle into the zebrafish adult telencephalon. At different post injury days, fish are sacrificed, their brains are dissected out and stained by immunohistochemistry and/or in situ hybridization (ISH) with appropriate markers to observe cell proliferation, gliogenesis, and neurogenesis. The contralateral unlesioned hemisphere serves as an internal control. This method combined for example with RNA deep sequencing can help to screen for new genes with a role in zebrafish adult telencephalon neurogenesis, regeneration, and repair. PMID:25146302

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

    PubMed

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

    2015-05-15

    Adult neural stem cells are the source for restoring injured brain tissue. We used repetitive imaging to follow single stem cells in the intact and injured adult zebrafish telencephalon in vivo and found that neurons are generated by both direct conversions of stem cells into postmitotic neurons and via intermediate progenitors amplifying the neuronal output. We observed an imbalance of direct conversion consuming the stem cells and asymmetric and symmetric self-renewing divisions, leading to depletion of stem cells over time. After brain injury, neuronal progenitors are recruited to the injury site. These progenitors are generated by symmetric divisions that deplete the pool of stem cells, a mode of neurogenesis absent in the intact telencephalon. Our analysis revealed changes in the behavior of stem cells underlying generation of additional neurons during regeneration. PMID:25977550

  6. Notch Receptor Expression in Neurogenic Regions of the Adult Zebrafish Brain

    PubMed Central

    de Oliveira-Carlos, Vanessa; Ganz, Julia; Hans, Stefan; Kaslin, Jan; Brand, Michael

    2013-01-01

    The adult zebrash brain has a remarkable constitutive neurogenic capacity. The regulation and maintenance of its adult neurogenic niches are poorly understood. In mammals, Notch signaling is involved in stem cell maintenance both in embryonic and adult CNS. To better understand how Notch signaling is involved in stem cell maintenance during adult neurogenesis in zebrafish we analysed Notch receptor expression in five neurogenic zones of the adult zebrafish brain. Combining proliferation and glial markers we identified several subsets of Notch receptor expressing cells. We found that 90 of proliferating radial glia express notch1a, notch1b and notch3. In contrast, the proliferating non-glial populations of the dorsal telencephalon and hypothalamus rarely express notch3 and about half express notch1a/1b. In the non-proliferating radial glia notch3 is the predominant receptor throughout the brain. In the ventral telencephalon and in the mitotic area of the optic tectum, where cells have neuroepithelial properties, notch1a/1b/3 are expressed in most proliferating cells. However, in the cerebellar niche, although progenitors also have neuroepithelial properties, only notch1a/1b are expressed in a high number of PCNA cells. In this region notch3 expression is mostly in Bergmann glia and at low levels in few PCNA cells. Additionally, we found that in the proliferation zone of the ventral telencephalon, Notch receptors display an apical high to basal low gradient of expression. Notch receptors are also expressed in subpopulations of oligodendrocytes, neurons and endothelial cells. We suggest that the partial regional heterogeneity observed for Notch expression in progenitor cells might be related to the cellular diversity present in each of these neurogenic niches. PMID:24039926

  7. Neurochemical measurements in the zebrafish brain

    PubMed Central

    Jones, Lauren J.; McCutcheon, James E.; Young, Andrew M. J.; Norton, William H. J.

    2015-01-01

    The zebrafish is an ideal model organism for behavioral genetics and neuroscience. The high conservation of genes and neurotransmitter pathways between zebrafish and other vertebrates permits the translation of research between species. Zebrafish behavior can be studied at both larval and adult stages and recent research has begun to establish zebrafish models for human disease. Fast scan cyclic voltammetry (FSCV) is an electrochemical technique that permits the detection of neurotransmitter release and reuptake. In this study we have used in vitro FSCV to measure the release of analytes in the adult zebrafish telencephalon. We compare different stimulation methods and present a characterization of neurochemical changes in the wild-type zebrafish brain. This study represents the first FSCV recordings in zebrafish, thus paving the way for neurochemical analysis of the fish brain. PMID:26441575

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

    PubMed Central

    Zou, Ming; De Koninck, Paul; Neve, Rachael L.; Friedrich, Rainer W.

    2014-01-01

    The zebrafish has various advantages as a model organism to analyze the structure and function of neural circuits but efficient viruses or other tools for fast gene transfer are lacking. We show that transgenes can be introduced directly into the adult zebrafish brain by herpes simplex type I viruses (HSV-1) or electroporation. We developed a new procedure to target electroporation to defined brain areas and identified promoters that produced strong long-term expression. The fast workflow of electroporation was exploited to express multiple channelrhodopsin-2 variants and genetically encoded calcium indicators in telencephalic neurons for measurements of neuronal activity and synaptic connectivity. The results demonstrate that HSV-1 and targeted electroporation are efficient tools for gene delivery into the zebrafish brain, similar to adeno-associated viruses and lentiviruses in other species. These methods fill an important gap in the spectrum of molecular tools for zebrafish and are likely to have a wide range of applications. PMID:24834028

  9. Zebrafish as an emerging model for studying complex brain disorders

    PubMed Central

    Kalueff, Allan V.; Stewart, Adam Michael; Gerlai, Robert

    2014-01-01

    The zebrafish (Danio rerio) is rapidly becoming a popular model organism in pharmacogenetics and neuropharmacology. Both larval and adult zebrafish are currently used to increase our understanding of brain function, dysfunction, and their genetic and pharmacological modulation. Here we review the developing utility of zebrafish in the analysis of complex brain disorders (including, for example, depression, autism, psychoses, drug abuse and cognitive disorders), also covering zebrafish applications towards the goal of modeling major human neuropsychiatric and drug-induced syndromes. We argue that zebrafish models of complex brain disorders and drug-induced conditions have become a rapidly emerging critical field in translational neuropharmacology research. PMID:24412421

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

    PubMed Central

    Dal Forno, Gonzalo Ogliari; Kist, Luiza Wilges; de Azevedo, Mariana Barbieri; Fritsch, Rachel Seemann; Pereira, Talita Carneiro Brandão; Britto, Roberta Socoowski; Guterres, Sílvia Stanisçuaski; Külkamp-Guerreiro, Irene Clemes; Bonan, Carla Denise; Monserrat, José María; Bogo, Maurício Reis

    2013-01-01

    Even though technologies involving nano/microparticles have great potential, it is crucial to determine possible toxicity of these technological products before extensive use. Fullerenes C60 are nanomaterials with unique physicochemical and biological properties that are important for the development of many technological applications. The aim of this study was to evaluate the consequences of nonphotoexcited fullerene C60 exposure in brain acetylcholinesterase expression and activity, antioxidant responses, and oxidative damage using adult zebrafish as an animal model. None of the doses tested (7.5, 15, and 30 mg/kg) altered AChE activity, antioxidant responses, and oxidative damage when zebrafish were exposed to nonphotoexcited C60 nano/microparticles during 6 and 12 hours. However, adult zebrafish exposed to the 30 mg/kg dose for 24 hours have shown enhanced AChE activity and augmented lipid peroxidation (TBARS assays) in brain. In addition, the up-regulation of brain AChE activity was neither related to the transcriptional control (RT-qPCR analysis) nor to the direct action of nonphotoexcited C60 nano/microparticles on the protein (in vitro results) but probably involved a posttranscriptional or posttranslational modulation of this enzymatic activity. Taken together these findings provided further evidence of toxic effects on brain after C60 exposure. PMID:23865059

  11. Regeneration of Zebrafish CNS: Adult Neurogenesis.

    PubMed

    Ghosh, Sukla; Hui, Subhra Prakash

    2016-01-01

    Regeneration in the animal kingdom is one of the most fascinating problems that have allowed scientists to address many issues of fundamental importance in basic biology. However, we came to know that the regenerative capability may vary across different species. Among vertebrates, fish and amphibians are capable of regenerating a variety of complex organs through epimorphosis. Zebrafish is an excellent animal model, which can repair several organs like damaged retina, severed spinal cord, injured brain and heart, and amputated fins. The focus of the present paper is on spinal cord regeneration in adult zebrafish. We intend to discuss our current understanding of the cellular and molecular mechanism(s) that allows formation of proliferating progenitors and controls neurogenesis, which involve changes in epigenetic and transcription programs. Unlike mammals, zebrafish retains radial glia, a nonneuronal cell type in their adult central nervous system. Injury induced proliferation involves radial glia which proliferate, transcribe embryonic genes, and can give rise to new neurons. Recent technological development of exquisite molecular tools in zebrafish, such as cell ablation, lineage analysis, and novel and substantial microarray, together with advancement in stem cell biology, allowed us to investigate how progenitor cells contribute to the generation of appropriate structures and various underlying mechanisms like reprogramming. PMID:27382491

  12. Regeneration of Zebrafish CNS: Adult Neurogenesis

    PubMed Central

    Ghosh, Sukla; Hui, Subhra Prakash

    2016-01-01

    Regeneration in the animal kingdom is one of the most fascinating problems that have allowed scientists to address many issues of fundamental importance in basic biology. However, we came to know that the regenerative capability may vary across different species. Among vertebrates, fish and amphibians are capable of regenerating a variety of complex organs through epimorphosis. Zebrafish is an excellent animal model, which can repair several organs like damaged retina, severed spinal cord, injured brain and heart, and amputated fins. The focus of the present paper is on spinal cord regeneration in adult zebrafish. We intend to discuss our current understanding of the cellular and molecular mechanism(s) that allows formation of proliferating progenitors and controls neurogenesis, which involve changes in epigenetic and transcription programs. Unlike mammals, zebrafish retains radial glia, a nonneuronal cell type in their adult central nervous system. Injury induced proliferation involves radial glia which proliferate, transcribe embryonic genes, and can give rise to new neurons. Recent technological development of exquisite molecular tools in zebrafish, such as cell ablation, lineage analysis, and novel and substantial microarray, together with advancement in stem cell biology, allowed us to investigate how progenitor cells contribute to the generation of appropriate structures and various underlying mechanisms like reprogramming. PMID:27382491

  13. Intraperitoneal Injection into Adult Zebrafish

    PubMed Central

    Kinkel, Mary D.; Eames, Stefani C.; Philipson, Louis H.; Prince, Victoria E.

    2010-01-01

    A convenient method for chemically treating zebrafish is to introduce the reagent into the tank water, where it will be taken up by the fish. However, this method makes it difficult to know how much reagent is absorbed or taken up per fish. Some experimental questions, particularly those related to metabolic studies, may be better addressed by delivering a defined quantity to each fish, based on weight. Here we present a method for intraperitoneal (IP) injection into adult zebrafish. Injection is into the abdominal cavity, posterior to the pelvic girdle. This procedure is adapted from veterinary methods used for larger fish. It is safe, as we have observed zero mortality. Additionally, we have seen bleeding at the injection site in only 5 out of 127 injections, and in each of those cases the bleeding was brief, lasting several seconds, and the quantity of blood lost was small. Success with this procedure requires gentle handling of the fish through several steps including fasting, weighing, anesthetizing, injection, and recovery. Precautions are required to minimize stress throughout the procedure. Our precautions include using a small injection volume and a 35G needle. We use Cortland salt solution as the vehicle, which is osmotically balanced for freshwater fish. Aeration of the gills is maintained during the injection procedure by first bringing the fish into a surgical plane of anesthesia, which allows slow operculum movements, and second, by holding the fish in a trough within a water-saturated sponge during the injection itself. We demonstrate the utility of IP injection by injecting glucose and monitoring the rise in blood glucose level and its subsequent return to normal. As stress is known to increase blood glucose in teleost fish, we compare blood glucose levels in vehicle-injected and non-injected adults and show that the procedure does not cause a significant rise in blood glucose. PMID:20834219

  14. Intraperitoneal injection into adult zebrafish.

    PubMed

    Kinkel, Mary D; Eames, Stefani C; Philipson, Louis H; Prince, Victoria E

    2010-01-01

    A convenient method for chemically treating zebrafish is to introduce the reagent into the tank water, where it will be taken up by the fish. However, this method makes it difficult to know how much reagent is absorbed or taken up per fish. Some experimental questions, particularly those related to metabolic studies, may be better addressed by delivering a defined quantity to each fish, based on weight. Here we present a method for intraperitoneal (IP) injection into adult zebrafish. Injection is into the abdominal cavity, posterior to the pelvic girdle. This procedure is adapted from veterinary methods used for larger fish. It is safe, as we have observed zero mortality. Additionally, we have seen bleeding at the injection site in only 5 out of 127 injections, and in each of those cases the bleeding was brief, lasting several seconds, and the quantity of blood lost was small. Success with this procedure requires gentle handling of the fish through several steps including fasting, weighing, anesthetizing, injection, and recovery. Precautions are required to minimize stress throughout the procedure. Our precautions include using a small injection volume and a 35G needle. We use Cortland salt solution as the vehicle, which is osmotically balanced for freshwater fish. Aeration of the gills is maintained during the injection procedure by first bringing the fish into a surgical plane of anesthesia, which allows slow operculum movements, and second, by holding the fish in a trough within a water-saturated sponge during the injection itself. We demonstrate the utility of IP injection by injecting glucose and monitoring the rise in blood glucose level and its subsequent return to normal. As stress is known to increase blood glucose in teleost fish, we compare blood glucose levels in vehicle-injected and non-injected adults and show that the procedure does not cause a significant rise in blood glucose. PMID:20834219

  15. Macrophages modulate adult zebrafish tail fin regeneration.

    PubMed

    Petrie, Timothy A; Strand, Nicholas S; Yang, Chao-Tsung; Tsung-Yang, Chao; Rabinowitz, Jeremy S; Moon, Randall T

    2014-07-01

    Neutrophils and macrophages, as key mediators of inflammation, have defined functionally important roles in mammalian tissue repair. Although recent evidence suggests that similar cells exist in zebrafish and also migrate to sites of injury in larvae, whether these cells are functionally important for wound healing or regeneration in adult zebrafish is unknown. To begin to address these questions, we first tracked neutrophils (lyzC(+), mpo(+)) and macrophages (mpeg1(+)) in adult zebrafish following amputation of the tail fin, and detailed a migratory timecourse that revealed conserved elements of the inflammatory cell response with mammals. Next, we used transgenic zebrafish in which we could selectively ablate macrophages, which allowed us to investigate whether macrophages were required for tail fin regeneration. We identified stage-dependent functional roles of macrophages in mediating fin tissue outgrowth and bony ray patterning, in part through modulating levels of blastema proliferation. Moreover, we also sought to detail molecular regulators of inflammation in adult zebrafish and identified Wnt/β-catenin as a signaling pathway that regulates the injury microenvironment, inflammatory cell migration and macrophage phenotype. These results provide a cellular and molecular link between components of the inflammation response and regeneration in adult zebrafish. PMID:24961798

  16. Thirty-Second Net Stressor Task in Adult Zebrafish

    PubMed Central

    Tran, Steven; Gerlai, Robert

    2016-01-01

    Zebrafish have become a popular animal model for behavioral neuroscience (Gerlai, 2014). Recent studies have demonstrated that brief experimental handling prior to euthanizing animals can subsequently alter biological measures quantified post-mortem (e.g. cortisol levels) (Ramsay et al., 2009; Tran et al., 2014). Here we provide a detailed protocol for a simple 30-sec net stressor task for adult zebrafish that increases whole-body cortisol levels without altering the levels of whole-brain dopamine, 3, 4-dihydroxyphenylacetic acid, serotonin, and 5-hydroxyindoleacetic acid (Tran et al., 2014).

  17. Brain tumor - primary - adults

    MedlinePlus

    ... Vestibular schwannoma (acoustic neuroma) - adults; Meningioma - adults; Cancer - brain tumor (adults) ... Primary brain tumors include any tumor that starts in the brain. Primary brain tumors can start from brain cells, ...

  18. Optimized cell transplantation using adult rag2 mutant zebrafish

    PubMed Central

    Tang, Qin; Abdelfattah, Nouran S.; Blackburn, Jessica S.; Moore, John C.; Martinez, Sarah A.; Moore, Finola E.; Lobbardi, Riadh; Tenente, Inês M.; Ignatius, Myron S.; Berman, Jason N.; Liwski, Robert S.; Houvras, Yariv; Langenau, David M.

    2014-01-01

    Cell transplantation into adult zebrafish has lagged behind mouse due to the lack of immune compromised models. Here, we have created homozygous rag2E450fs mutant zebrafish that have reduced numbers of functional T and B cells but are viable and fecund. Mutant fish engraft zebrafish muscle, blood stem cells, and cancers. rag2E450fs mutant zebrafish are the first immune compromised zebrafish model that permits robust, long-term engraftment of multiple tissues and cancer. PMID:25042784

  19. Methods for studying the zebrafish brain: past, present and future.

    PubMed

    Wyatt, Cameron; Bartoszek, Ewelina M; Yaksi, Emre

    2015-07-01

    The zebrafish (Danio rerio) is one of the most promising new model organisms. The increasing popularity of this amazing small vertebrate is evident from the exponentially growing numbers of research articles, funded projects and new discoveries associated with the use of zebrafish for studying development, brain function, human diseases and screening for new drugs. Thanks to the development of novel technologies, the range of zebrafish research is constantly expanding with new tools synergistically enhancing traditional techniques. In this review we will highlight the past and present techniques which have made, and continue to make, zebrafish an attractive model organism for various fields of biology, with a specific focus on neuroscience. PMID:25900095

  20. TCDD Inhibits Heart Regeneration in Adult Zebrafish

    PubMed Central

    Hofsteen, Peter; Mehta, Vatsal; Heideman, Warren

    2013-01-01

    Normal adult zebrafish can completely regenerate lost myocardium following partial amputation of the ventricle apex. We report that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) significantly impairs this regeneration. Adult male zebrafish were injected with vehicle (control) or TCDD (70ng/g, ip) 1 day prior to partial amputation of the ventricle apex. Gross observation and histological analysis of the amputated heart at 21 days postamputation revealed that TCDD-exposed fish had not progressed beyond the initial clot formation stage, whereas the vehicle control fish showed substantial recovery and almost complete resolution of the formed clot. In contrast, hearts that were not surgically wounded showed no signs of TCDD toxicity. Striking features in the TCDD-exposed hearts were the absence of the normal sheath of new tissue enveloping the wound and the absence of intense cell proliferation at the site of the wound. In addition, the patterns of collagen deposition at the wound site were different between the TCDD and vehicle groups. Because the receptor for TCDD is the aryl hydrocarbon receptor ligand-activated transcriptional regulator, we examined the effects of TCDD exposure on gene expression in the ventricle using DNA microarrays. Samples were collected just prior to amputation and at 6h and 7 days postamputation. TCDD-pretreated hearts had dysregulated expression of genes involved in heart function, tissue regeneration, cell growth, and extracellular matrix. Because embryonic, but not adult, hearts are major targets for TCDD-induced cardiotoxicity, we speculate that the need for embryonic-like cells in regeneration is connected with the effects of TCDD in inhibiting the response to wounding. PMID:23204111

  1. Adult Zebrafish model of streptococcal infection

    PubMed Central

    Phelps, Hilary A.; Runft, Donna L.

    2009-01-01

    Streptococcal pathogens cause a wide array of clinical syndromes in humans, including invasive systemic infections resulting in high mortality rates. Many of these pathogens are human specific, and therefore difficult to analyze in vivo using typical animal models, as these models rarely replicate what is observed in human infections. This unit describes the use of the zebrafish (Danio rerio) as an animal model for streptococcal infection to analyze multiple disease states. This model closely mimics the necrotizing fasciitis/myositis pathology observed in humans from a Streptococcus pyogenes infection. The use of a zoonotic pathogen, Streptococcus iniae, which replicates systemic infections caused by many streptococcal pathogens, including dissemination to the brain, is also described. Included protocols describe both intraperitoneal and intramuscular infections, as well as methods for histological and quantitative measurements of infection. PMID:19412913

  2. Comprehensive expression map of transcription regulators in the adult zebrafish telencephalon reveals distinct neurogenic niches.

    PubMed

    Diotel, Nicolas; Rodriguez Viales, Rebecca; Armant, Olivier; März, Martin; Ferg, Marco; Rastegar, Sepand; Strähle, Uwe

    2015-06-01

    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. PMID:25556858

  3. An assay for lateral line regeneration in adult zebrafish.

    PubMed

    Pisano, Gina C; Mason, Samantha M; Dhliwayo, Nyembezi; Intine, Robert V; Sarras, Michael P

    2014-01-01

    Due to the clinical importance of hearing and balance disorders in man, model organisms such as the zebrafish have been used to study lateral line development and regeneration. The zebrafish is particularly attractive for such studies because of its rapid development time and its high regenerative capacity. To date, zebrafish studies of lateral line regeneration have mainly utilized fish of the embryonic and larval stages because of the lower number of neuromasts at these stages. This has made quantitative analysis of lateral line regeneration/and or development easier in the earlier developmental stages. Because many zebrafish models of neurological and non-neurological diseases are studied in the adult fish and not in the embryo/larvae, we focused on developing a quantitative lateral line regenerative assay in adult zebrafish so that an assay was available that could be applied to current adult zebrafish disease models. Building on previous studies by Van Trump et al. that described procedures for ablation of hair cells in adult Mexican blind cave fish and zebrafish (Danio rerio), our assay was designed to allow quantitative comparison between control and experimental groups. This was accomplished by developing a regenerative neuromast standard curve based on the percent of neuromast reappearance over a 24 hr time period following gentamicin-induced necrosis of hair cells in a defined region of the lateral line. The assay was also designed to allow extension of the analysis to the individual hair cell level when a higher level of resolution is required. PMID:24747778

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

    PubMed

    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. PMID:26778924

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

    PubMed Central

    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

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

    PubMed Central

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

    2015-01-01

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

  7. Pharmacological Modulation of Hemodynamics in Adult Zebrafish In Vivo

    PubMed Central

    Brönnimann, Daniel; Dellenbach, Christian; Saveljic, Igor; Rieger, Michael; Rohr, Stephan; Filipovic, Nenad; Djonov, Valentin

    2016-01-01

    Introduction 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. Materials and Methods 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. Results 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). Discussion 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

  8. Proteome Mapping of Adult Zebrafish Marrow Neutrophils Reveals Partial Cross Species Conservation to Human Peripheral Neutrophils

    PubMed Central

    Singh, Sachin Kumar; Sethi, Sachin; Aravamudhan, Sriram; Krüger, Marcus; Grabher, Clemens

    2013-01-01

    Neutrophil granulocytes are pivotal cells within the first line of host defense of the innate immune system. In this study, we have used a gel-based LC-MS/MS approach to explore the proteome of primary marrow neutrophils from adult zebrafish. The identified proteins originated from all major cellular compartments. Gene ontology analysis revealed significant association of proteins with different immune-related network and pathway maps. 75% of proteins identified in neutrophils were identified in neutrophils only when compared to neutrophil-free brain tissue. Moreover, cross-species comparison with human peripheral blood neutrophils showed partial conservation of immune-related proteins between human and zebrafish. This study provides the first zebrafish neutrophil proteome and may serve as a valuable resource for an understanding of neutrophil biology and innate immunity. PMID:24019943

  9. Trim69 regulates zebrafish brain development by ap-1 pathway

    PubMed Central

    Han, Ruiqin; Wang, Renxian; Zhao, Qing; Han, Yongqing; Zong, Shudong; Miao, Shiying; Song, Wei; Wang, Linfang

    2016-01-01

    Proteins belonging to the TRIM family have been implicated in a variety of cellular processes such as apoptosis, differentiation, neurogenesis, muscular physiology and innate immune responses. Trim69, previously identified as a novel gene cloned from a human testis cDNA library, has a homologous gene in zebrafish and this study focused on investigating the function of trim69 in zebrafish neurogenesis. Trim69 was found to be expressed in zebrafish embryo brain at the early stages. Knockdown of trim69 led to deformed brain development, obvious signs of apoptosis present in the head, and decreased expression of neuronal differentiation and stem cell markers. This phenotype was rescued upon co-injection of human mRNA together along with the trim69 knockdown. Results of this study also showed an interaction between TRIM69 and c-Jun in human cells, and upon TRIM69 knock down c-Jun expression subsequently increased, whereas the over-expression of TRIM69 led to the down-regulation of c-Jun. Additionally, knockdown both c-Jun and trim69 can rescue the deformed brain, evident cellular apoptosis in the head and decreased expression of neuronal differentiation and stem cell markers. Overall, our results support a role for trim69 in the development of the zebrafish brain through ap-1 pathway. PMID:27050765

  10. Characterization of multiciliated ependymal cells that emerge in the neurogenic niche of the aged zebrafish brain.

    PubMed

    Ogino, Takashi; Sawada, Masato; Takase, Hiroshi; Nakai, Chiemi; Herranz-Pérez, Vicente; Cebrián-Silla, Arantxa; Kaneko, Naoko; García-Verdugo, José Manuel; Sawamoto, Kazunobu

    2016-10-15

    In mammals, ventricular walls of the developing brain maintain a neurogenic niche, in which radial glial cells act as neural stem cells (NSCs) and generate new neurons in the embryo. In the adult brain, the neurogenic niche is maintained in the ventricular-subventricular zone (V-SVZ) of the lateral wall of lateral ventricles and the hippocampal dentate gyrus. In the neonatal V-SVZ, radial glial cells transform into astrocytic postnatal NSCs and multiciliated ependymal cells. On the other hand, in zebrafish, radial glial cells continue to cover the surface of the adult telencephalic ventricle and maintain a higher neurogenic potential in the adult brain. However, the cell composition of the neurogenic niche of the aged zebrafish brain has not been investigated. Here we show that multiciliated ependymal cells emerge in the neurogenic niche of the aged zebrafish telencephalon. These multiciliated cells appear predominantly in the dorsal part of the ventral telencephalic ventricular zone, which also contains clusters of migrating new neurons. Scanning electron microscopy and live imaging analyses indicated that these multiple cilia beat coordinately and generate constant fluid flow within the ventral telencephalic ventricle. Analysis of the cell composition by transmission electron microscopy revealed that the neurogenic niche in the aged zebrafish contains different types of cells, with ultrastructures similar to those of ependymal cells, transit-amplifying cells, and migrating new neurons in postnatal mice. These data suggest that the transformation capacity of radial glial cells is conserved but that its timing is different between fish and mice. J. Comp. Neurol. 524:2982-2992, 2016. © 2016 Wiley Periodicals, Inc. PMID:26991819

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

    PubMed

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

    2015-01-01

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

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

    PubMed Central

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

    2015-01-01

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

  13. Unpredictable Chronic Stress Alters Adenosine Metabolism in Zebrafish Brain.

    PubMed

    Zimmermann, F F; Altenhofen, S; Kist, L W; Leite, C E; Bogo, M R; Cognato, G P; Bonan, C D

    2016-05-01

    Stress is considered a risk factor for several human disorders. Despite the broad knowledge of stress responses in mammals, data on the relationship between unpredictable chronic stress (UCS) and its effects on purinergic signaling are limited. ATP hydrolysis by ectonucleotidases is an important source of adenosine, and adenosine deaminase (ADA) contributes to the control of the nucleoside concentrations. Considering that some stress models could affect signaling systems, the objective of this study was to investigate whether UCS alters ectonucleotidase and ADA pathway in zebrafish brain. Additionally, we analyzed ATP metabolism as well as ada1, ada2.1, ada2.2, adaL, and adaasi gene expression in zebrafish brain. Our results have demonstrated that UCS did not alter ectonucleotidase and soluble ADA activities. However, ecto-ADA activity was significantly decreased (26.8%) in brain membranes of animals exposed to UCS when compared to the control group. Quantitative reverse transcription PCR (RT-PCR) analysis did not show significant changes on ADA gene expression after the UCS exposure. The brain ATP metabolism showed a marked increase in adenosine levels (ADO) in animals exposed to UCS. These data suggest an increase on extracellular adenosine levels in zebrafish brain. Since this nucleoside has neuromodulatory and anxiolytic effects, changes in adenosine levels could play a role in counteracting the stress, which could be related to a compensatory mechanism in order to restore the homeostasis. PMID:26081145

  14. Reactive gliosis in the adult zebrafish retina.

    PubMed

    Thomas, Jennifer L; Ranski, Alexandra H; Morgan, Gregory W; Thummel, Ryan

    2016-02-01

    In contrast to mammals, zebrafish posses the remarkable ability to regenerate retinal neurons. Damage to the zebrafish retina induces Müller glia to act as stem cells, generating retinal progenitors for regeneration. In contrast, injury in the mammalian retina results in Müller glial reactive gliosis, a characteristic gliotic response that is normally detrimental to vision. Understanding the signaling pathways that determine how Müller glia respond to injury is a critical step toward promoting regeneration in the mammalian retina. Here we report that zebrafish Müller glia exhibit signs of reactive gliosis even under normal regenerative conditions and that cell cycle inhibition increases this response. Persistently reactive Müller glia increase their neuroprotective functions, temporarily saving photoreceptors from a cytotoxic light lesion. However, the absence of a sustained proliferation response results in a significant inhibition of retinal regeneration. Interestingly, when cell cycle inhibition is released, a partial recovery of regeneration is observed. Together, these data demonstrate that zebrafish Müller glia possess both gliotic and regenerative potential. PMID:26492821

  15. Absence of rapid eye movements during sleep in adult zebrafish.

    PubMed

    Árnason, B B; Þorsteinsson, H; Karlsson, K Æ

    2015-09-15

    Sleep is not a uniform phenomenon, but is organized in alternating, fundamentally different states, rapid eye movement sleep and non-rapid eye movement sleep. Zebrafish (Danio rerio) have recently emerged as an excellent model for sleep research. Zebrafish are well characterized in terms of development, neurobiology and genetics. Moreover, there are many experimental tools not easily applied in mammalian models that can be readily applied to zebrafish, making them a valuable additional animal model for sleep research. Sleep in zebrafish is defined behaviorally and exhibits the hallmarks of mammalian sleep (e.g. sleep homeostasis and pressure). To our knowledge no attempts have been made to discern if sleep in zebrafish entails alternations of REM-NREM sleep cycles which are critical for further development of the model. In the current experiment we quantify two key REM sleep components, rapid eye movements and respiratory rates, across sleep-wake cycles. We find no sleep-related rapid eye movements. During sleep respiratory rates, however, are reduced and become less regular, further establishing that the behavioral definition used truly captures a change in the fish's physiology. We thus fail to find evidence for REM-NREM sleep cycles in zebrafish but demonstrate a physiological change that occurs concomitantly with the previously defined behavioral state of sleep. We do not rule out that other phasic REM components (e.g. atonia, cardiac arrhythmias, myoclonic twitches or desynchronized EEG) are coherently expressed during sleep but we conclude that adult zebrafish do not have REM-sleep-related rapid eye movements. PMID:26003945

  16. Stable multilineage xenogeneic replacement of definitive hematopoiesis in adult zebrafish.

    PubMed

    Hess, Isabell; Boehm, Thomas

    2016-01-01

    Bony fishes are the most numerous and phenotypically diverse group of vertebrates inhabiting our planet, making them an ideal target for identifying general principles of tissue development and function. However, lack of suitable experimental platforms prevents the exploitation of this rich source of natural phenotypic variation. Here, we use a zebrafish strain lacking definitive hematopoiesis for interspecific analysis of hematopoietic cell development. Without conditioning prior to transplantation, hematopoietic progenitor cells from goldfish stably engraft in adult zebrafish homozygous for the c-myb(I181N) mutation. However, in competitive repopulation experiments, zebrafish hematopoietic cells exhibit an advantage over their goldfish counterparts, possibly owing to subtle species-specific functional differences in hematopoietic microenvironments resulting from over 100 million years of independent evolution. Thus, our unique animal model provides an unprecedented opportunity to genetically and functionally disentangle universal and species-specific contributions of the microenvironment to hematopoietic progenitor cell maintenance and development. PMID:26777855

  17. Stable multilineage xenogeneic replacement of definitive hematopoiesis in adult zebrafish

    PubMed Central

    Hess, Isabell; Boehm, Thomas

    2016-01-01

    Bony fishes are the most numerous and phenotypically diverse group of vertebrates inhabiting our planet, making them an ideal target for identifying general principles of tissue development and function. However, lack of suitable experimental platforms prevents the exploitation of this rich source of natural phenotypic variation. Here, we use a zebrafish strain lacking definitive hematopoiesis for interspecific analysis of hematopoietic cell development. Without conditioning prior to transplantation, hematopoietic progenitor cells from goldfish stably engraft in adult zebrafish homozygous for the c-mybI181N mutation. However, in competitive repopulation experiments, zebrafish hematopoietic cells exhibit an advantage over their goldfish counterparts, possibly owing to subtle species-specific functional differences in hematopoietic microenvironments resulting from over 100 million years of independent evolution. Thus, our unique animal model provides an unprecedented opportunity to genetically and functionally disentangle universal and species-specific contributions of the microenvironment to hematopoietic progenitor cell maintenance and development. PMID:26777855

  18. Atlas of Cellular Dynamics during Zebrafish Adult Kidney Regeneration

    PubMed Central

    McCampbell, Kristen K.; Springer, Kristin N.; Wingert, Rebecca A.

    2015-01-01

    The zebrafish is a useful animal model to study the signaling pathways that orchestrate kidney regeneration, as its renal nephrons are simple, yet they maintain the biological complexity inherent to that of higher vertebrate organisms including mammals. Recent studies have suggested that administration of the aminoglycoside antibiotic gentamicin in zebrafish mimics human acute kidney injury (AKI) through the induction of nephron damage, but the timing and details of critical phenotypic events associated with the regeneration process, particularly in existing nephrons, have not been characterized. Here, we mapped the temporal progression of cellular and molecular changes that occur during renal epithelial regeneration of the proximal tubule in the adult zebrafish using a platform of histological and expression analysis techniques. This work establishes the timing of renal cell death after gentamicin injury, identifies proliferative compartments within the kidney, and documents gene expression changes associated with the regenerative response of proliferating cells. These data provide an important descriptive atlas that documents the series of events that ensue after damage in the zebrafish kidney, thus availing a valuable resource for the scientific community that can facilitate the implementation of zebrafish research to delineate the mechanisms that control renal regeneration. PMID:26089919

  19. Seizures induced by pentylenetetrazole in the adult zebrafish: a detailed behavioral characterization.

    PubMed

    Mussulini, Ben Hur M; Leite, Carlos E; Zenki, Kamila C; Moro, Luana; Baggio, Suelen; Rico, Eduardo P; Rosemberg, Denis B; Dias, Renato D; Souza, Tadeu M; Calcagnotto, Maria E; Campos, Maria M; Battastini, Ana M; de Oliveira, Diogo L

    2013-01-01

    Pentylenetetrazole (PTZ) is a common convulsant agent used in animal models to investigate the mechanisms of seizures. Although adult zebrafish have been recently used to study epileptic seizures, a thorough characterization of the PTZ-induced seizures in this animal model is missing. The goal of this study was to perform a detailed temporal behavior profile characterization of PTZ-induced seizure in adult zebrafish. The behavioral profile during 20 min of PTZ immersion (5, 7.5, 10, and 15 mM) was characterized by stages defined as scores: (0) short swim, (1) increased swimming activity and high frequency of opercular movement, (2) erratic movements, (3) circular movements, (4) clonic seizure-like behavior, (5) fall to the bottom of the tank and tonic seizure-like behavior, (6) death. Animals exposed to distinct PTZ concentrations presented different seizure profiles, intensities and latencies to reach all scores. Only animals immersed into 15 mM PTZ showed an increased time to return to the normal behavior (score 0), after exposure. Total mortality rate at 10 and 15 mM were 33% and 50%, respectively. Considering all behavioral parameters, 5, 7.5, 10, and 15 mM PTZ, induced seizures with low, intermediate, and high severity, respectively. Pretreatment with diazepam (DZP) significantly attenuated seizure severity. Finally, the brain PTZ levels in adult zebrafish immersed into the chemoconvulsant solution at 5 and 10 mM were comparable to those described for the rodent model, with a peak after a 20-min of exposure. The PTZ brain levels observed after 2.5-min PTZ exposure and after 60-min removal from exposure were similar. Altogether, our results showed a detailed temporal behavioral characterization of a PTZ epileptic seizure model in adult zebrafish. These behavioral analyses and the simple method for PTZ quantification could be considered as important tools for future investigations and translational research. PMID:23349914

  20. Wnt/β-catenin signaling promotes regeneration after adult zebrafish spinal cord injury.

    PubMed

    Strand, Nicholas S; Hoi, Kimberly K; Phan, Tien M T; Ray, Catherine A; Berndt, Jason D; Moon, Randall T

    2016-09-01

    Unlike mammals, zebrafish can regenerate their injured spinal cord and regain control of caudal tissues. It was recently shown that Wnt/β-catenin signaling is necessary for spinal cord regeneration in the larval zebrafish. However, the molecular mechanisms of regeneration may or may not be conserved between larval and adult zebrafish. To test this, we assessed the role of Wnt/β-catenin signaling after spinal cord injury in the adult zebrafish. We show that Wnt/β-catenin signaling is increased after spinal cord injury in the adult zebrafish. Moreover, overexpression of Dkk1b inhibited Wnt/β-catenin signaling in the regenerating spinal cord of adult zebrafish. Dkk1b overexpression also inhibited locomotor recovery, axon regeneration, and glial bridge formation in the injured spinal cord. Thus, our data illustrate a conserved role for Wnt/β-catenin signaling in adult and larval zebrafish spinal cord regeneration. PMID:27387232

  1. Sexual dimorphisms in swimming behavior, cerebral metabolic activity and adrenoceptors in adult zebrafish (Danio rerio).

    PubMed

    Ampatzis, Konstantinos; Dermon, Catherine R

    2016-10-01

    Sexually dimorphic behaviors and brain sex differences, not only restricted to reproduction, are considered to be evolutionary preserved. Specifically, anxiety related behavioral repertoire is suggested to exhibit sex-specific characteristics in rodents and primates. The present study investigated whether behavioral responses to novelty, have sex-specific characteristics in the neurogenetic model organism zebrafish (Danio rerio), lacking chromosomal sex determination. For this, aspects of anxiety-like behavior (including reduced exploration, increased freezing behavior and erratic movement) of male and female adult zebrafish were tested in a novel tank paradigm and after habituation. Male and female zebrafish showed significant differences in their swimming activity in response to novelty, with females showing less anxiety spending more time in the upper tank level. When fish have habituated, regional cerebral glucose uptake, an index of neuronal activity, and brain adrenoceptors' (ARs) expression (α2-ARs and β-ARs) were determined using in vivo 2-[(14)C]-deoxyglucose methodology and in vitro neurotransmitter receptors quantitative autoradiography, respectively. Intriguingly, females exhibited higher glucose utilization than males in hypothalamic brain areas. Adrenoceptor's expression pattern was dimorphic in zebrafish telencephalic, preoptic, hypothalamic nuclei, central gray, and cerebellum, similarly to birds and mammals. Specifically, the lateral zone of dorsal telencephalon (Dl), an area related to spatial cognition, homologous to the mammalian hippocampus, showed higher α2-AR densities in females. In contrast, male cerebellum included higher densities of β-ARs in comparison to female. Taken together, our data demonstrate a well-defined sex discriminant cerebral metabolic activity and ARs' pattern in zebrafish, possibly contributing to male-female differences in the swimming behavior. PMID:27363927

  2. A three-dimensional digital atlas of the zebrafish brain.

    PubMed

    Ullmann, Jeremy F P; Cowin, Gary; Kurniawan, Nyoman D; Collin, Shaun P

    2010-05-15

    In the past three decades, the zebrafish has become a vital animal model in a range of biological sciences. To augment current neurobiological research, we have developed the first three-dimensional digital atlas of the zebrafish brain from T2-weighted magnetic resonance histology (MRH) images acquired on a 16.4-T superconducting magnet. We achieved an isotropic resolution of 10 microm, which is the highest resolution achieved in a vertebrate brain and, for the first time, is comparable in slice thickness to conventional histology. By using manual segmentation, 53 anatomical structures, including fiber tracts as small as 40 microm, were delineated. Using Amira software, structures were also individually segmented and reconstructed to create three-dimensional animations. Additional quantitative information including, volume, surface areas, and mean gray scale intensities were also determined. Finally, we established a stereotaxic coordinate system as a framework in which maps created from other modalities can be incorporated into the atlas. PMID:20139016

  3. Embryonic oxidative stress results in reproductive impairment for adult zebrafish

    PubMed Central

    Newman, Trent A.C.; Carleton, Catherine R.; Leeke, Bryony; Hampton, Mark B.; Horsfield, Julia A.

    2015-01-01

    Exposure to environmental stressors during embryo development can have long-term effects on the adult organism. This study used the thioredoxin reductase inhibitor auranofin to investigate the consequences of oxidative stress during zebrafish development. Auranofin at low doses triggered upregulation of the antioxidant genes gstp1 and prdx1. As the dose was increased, acute developmental abnormalities, including cerebral hemorrhaging and jaw malformation, were observed. To determine whether transient disruption of redox homeostasis during development could have long-term consequences, zebrafish embryos were exposed to a low dose of auranofin from 6–24 hours post fertilization, and then raised to adulthood. The adult fish were outwardly normal in their appearance with no gross physical differences compared to the control group. However, these adult fish had reduced odds of breeding and a lower incidence of egg fertilization. This study shows that a suboptimal early life environment can reduce the chances of reproductive success in adulthood. PMID:26584358

  4. Localization of BDNF expression in the developing brain of zebrafish

    PubMed Central

    De Felice, E; Porreca, I; Alleva, E; De Girolamo, P; Ambrosino, C; Ciriaco, E; Germanà, A; Sordino, P

    2014-01-01

    The brain-derived neurotrophic factor (BDNF) gene is expressed in differentiating and post-mitotic neurons of the zebrafish embryo, where it has been implicated in Huntington's disease. Little is known, however, about the full complement of neuronal cell types that express BDNF in this important vertebrate model. Here, we further explored the transcriptional profiles during the first week of development using real-time quantitative polymerase chain reaction (RT-qPCR) and whole-mount in situ hybridization (WISH). RT-qPCR results revealed a high level of maternal contribution followed by a steady increase of zygotic transcription, consistent with the notion of a prominent role of BDNF in neuronal maturation and maintenance. Based on WISH, we demonstrate for the first time that BDNF expression in the developing brain of zebrafish is structure specific. Anatomical criteria and co-staining with genetic markers (shh, pax2a, emx1, krox20, lhx2b and lhx9) visualized major topological domains of BDNF-positive cells in the pallium, hypothalamus, posterior tuberculum and optic tectum. Moreover, the relative timing of BDNF transcription in the eye and tectum may illustrate a mechanism for coordinated development of the retinotectal system. Taken together, our results are compatible with a local delivery and early role of BDNF in the developing brain of zebrafish, adding basic knowledge to the study of neurotrophin functions in neural development and disease. PMID:24588510

  5. G-protein-coupled estrogen receptor 1 is involved in brain development during zebrafish (Danio rerio) embryogenesis

    SciTech Connect

    Shi, Yanan; Liu, Xiaochun; Zhu, Pei; Li, Jianzhen; Sham, Kathy W.Y.; Cheng, Shuk Han; Li, Shuisheng; Zhang, Yong; Cheng, Christopher H.K.; Lin, Haoran

    2013-05-24

    Highlights: •The Gper expression was detected in the developing brain of zebrafish. •Gper morpholino knockdown induced apoptosis of brain cells. •Gper morpholino knockdown reduced expression in neuron markers. •Zebrafish Gper may be involved in neuronal development. -- Abstract: G-protein-coupled estrogen receptor 1 (Gper, formerly known as GPR30) is found to be a trophic and protective factor in mediating action of estrogen in adult brain, while its role in developing brain remains to be elucidated. Here we present the expression pattern of Gper and its functions during embryogenesis in zebrafish. Both the mRNA and protein of Gper were detected throughout embryogenesis. Whole mount in situ hybridization (WISH) revealed a wide distribution of gper mRNAs in various regions of the developing brain. Gper knockdown by specific morpholinos resulted in growth retardation in embryos and morphological defects in the developing brain. In addition, induced apoptosis, decreased proliferation of the brain cells and maldevelopment of sensory and motor neurons were also found in the morphants. Our results provide novel insights into Gper functions in the developing brain, revealing that Gper can maintain the survival of the brain cells, and formation and/or differentiation of the sensory and motor neurons.

  6. Explant culture of adult zebrafish hearts for epicardial regeneration studies.

    PubMed

    Cao, Jingli; Poss, Kenneth D

    2016-05-01

    Here we describe how to culture adult zebrafish hearts as explants and study the regeneration of epicardial tissue ex vivo, as a means to identify therapeutic targets for heart disease. Uninjured or injured adult hearts are excised, washed and cultured in an incubator with gentle agitation. Heart explants can be prepared within 2 h, and they can be maintained in culture for 30 d or longer. If explants are prepared from appropriate transgenic lines, dynamic behaviors of epicardial cells can be monitored by live imaging using stereofluorescence microscopy. We also describe ex vivo procedures for genetic ablation of the epicardium, cell proliferation assays, tissue grafts and bead grafts. Basic cell culture and surgical skills are required to carry out this protocol. Unlike existing protocols for culturing isolated zebrafish epicardial cells on matrices, procedures described here maintain epicardial cells on an intact cardiac surface, thereby better supporting in vivo cell behaviors. Our protocols complement and extend in vivo studies of heart regeneration. PMID:27055096

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

    PubMed Central

    2013-01-01

    Background Teleost fish display widespread post-embryonic neurogenesis originating from many different proliferative niches that are distributed along the brain axis. During the development of the central nervous system (CNS) different cell types are produced in a strict temporal order from increasingly committed progenitors. However, it is not known whether diverse neural stem and progenitor cell types with restricted potential or stem cells with broad potential are maintained in the teleost fish brain. Results To study the diversity and output of neural stem and progenitor cell populations in the zebrafish brain the cerebellum was used as a model brain region, because of its well-known architecture and development. Transgenic zebrafish lines, in vivo imaging and molecular markers were used to follow and quantify how the proliferative activity and output of cerebellar progenitor populations progress. This analysis revealed that the proliferative activity and progenitor marker expression declines in juvenile zebrafish before they reach sexual maturity. Furthermore, this correlated with the diminished repertoire of cell types produced in the adult. The stem and progenitor cells derived from the upper rhombic lip were maintained into adulthood and they actively produced granule cells. Ventricular zone derived progenitor cells were largely quiescent in the adult cerebellum and produced a very limited number of glia and inhibitory inter-neurons. No Purkinje or Eurydendroid cells were produced in fish older than 3 months. This suggests that cerebellar cell types are produced in a strict temporal order from distinct pools of increasingly committed stem and progenitor cells. Conclusions Our results in the zebrafish cerebellum show that neural stem and progenitor cell types are specified and they produce distinct cell lineages and sub-types of brain cells. We propose that only specific subtypes of brain cells are continuously produced throughout life in the teleost fish

  8. Adult neural stem cell behavior underlying constitutive and restorative neurogenesis in zebrafish.

    PubMed

    Barbosa, Joana S; Ninkovic, Jovica

    2016-01-01

    Adult Neural Stem Cells (aNSCs) generate new neurons that integrate into the pre-existing networks in specific locations of the Vertebrate brain. Moreover, aNSCs contribute with new neurons to brain regeneration in some non-mammalian Vertebrates. The similarities and the differences in the cellular and molecular processes governing neurogenesis in the intact and regenerating brain are still to be assessed. Toward this end, we recently established a protocol for non-invasive imaging of aNSC behavior in their niche in vivo in the adult intact and regenerating zebrafish telencephalon. We observed different modes of aNSC division in the intact brain and a novel mode of neurogenesis by direct conversion, which contributes to stem cell depletion with age. After injury, the generation of neurons is increased both by the activation of additional aNSCs and a shift in the division mode of aNSCs, thereby contributing to the successful neuronal regeneration. The cellular behavior we observed opens new questions regarding long-term aNSC maintenance in homeostasis and in regeneration. In this commentary we discuss our data and new questions arising in the context of aNSC behavior, not only in zebrafish but also in other species, including mammals. PMID:27606336

  9. Integration of vascular systems between the brain and spinal cord in zebrafish.

    PubMed

    Kimura, Eiji; Isogai, Sumio; Hitomi, Jiro

    2015-10-01

    Cerebral and spinal vascular systems are organized individually, and they then conjugate at their border, through the integration of basilar artery and vertebral arteries. Zebrafish (Danio rerio) is an ideal organism for studying early vascular development, and the precise procedure of cranial and truncal vascular formation has been previously demonstrated using this model. However, the stepwise process of the integration between the brain and spinal cord has not been clearly elucidated. In this study, we describe the integration of the independent vascular systems for the brain and spinal cord, using transgenic zebrafish expressing enhanced green fluorescent protein in endothelial cells. Initially, basilar artery and primordial hindbrain channels, into which internal carotid arteries supplied blood, were connected with dorsal longitudinal anastomose vessels, via the first intersegmental artery. This initial connection was not influenced by flow dynamics, suggesting that vascular integration in this region is controlled by genetic cues. Vertebral arteries were formed individually as longitudinal vessels beneath the spinal cord, and became integrated with the basilar artery during subsequent remodeling. Furthermore, we confirmed the basal vasculature was well conserved in adult zebrafish. Observations of vascular integration presented herein will contribute to an understanding of regulatory mechanisms behind this process. PMID:26234750

  10. Differential expression of id genes and their potential regulator znf238 in zebrafish adult neural progenitor cells and neurons suggests distinct functions in adult neurogenesis.

    PubMed

    Diotel, Nicolas; Beil, Tanja; Strähle, Uwe; Rastegar, Sepand

    2015-01-01

    Teleost fish display a remarkable ability to generate new neurons and to repair brain lesions during adulthood. They are, therefore, a very popular model to investigate the molecular mechanisms of constitutive and induced neurogenesis in adult vertebrates. In this study, we investigated the expression patterns of inhibitor of DNA binding (id) genes and of their potential transcriptional repressor, znf238, in the whole brain of adult zebrafish. We show that while id1 is exclusively expressed in ventricular cells in the whole brain, id2a, id3 and id4 genes are expressed in broader areas. Interestingly, znf238 was also detected in these regions, its expression overlapping with id2a, id3 and id4 expression. Further detailed characterization of the id-expressing cells demonstrated that (a) id1 is expressed in type 1 and type 2 neural progenitors as previously published, (b) id2a in type 1, 2 and 3 neural progenitors, (c) id3 in type 3 neural progenitors and (d) id4 in postmitotic neurons. Our data provide a detailed map of id and znf238 expression in the brain of adult zebrafish, supplying a framework for studies of id genes function during adult neurogenesis and brain regeneration in the zebrafish. PMID:26107416

  11. In vivo cell tracking and quantification method in adult zebrafish

    NASA Astrophysics Data System (ADS)

    Zhang, Li; Alt, Clemens; Li, Pulin; White, Richard M.; Zon, Leonard I.; Wei, Xunbin; Lin, Charles P.

    2012-03-01

    Zebrafish have become a powerful vertebrate model organism for drug discovery, cancer and stem cell research. A recently developed transparent adult zebrafish using double pigmentation mutant, called casper, provide unparalleled imaging power in in vivo longitudinal analysis of biological processes at an anatomic resolution not readily achievable in murine or other systems. In this paper we introduce an optical method for simultaneous visualization and cell quantification, which combines the laser scanning confocal microscopy (LSCM) and the in vivo flow cytometry (IVFC). The system is designed specifically for non-invasive tracking of both stationary and circulating cells in adult zebrafish casper, under physiological conditions in the same fish over time. The confocal imaging part in this system serves the dual purposes of imaging fish tissue microstructure and a 3D navigation tool to locate a suitable vessel for circulating cell counting. The multi-color, multi-channel instrument allows the detection of multiple cell populations or different tissues or organs simultaneously. We demonstrate initial testing of this novel instrument by imaging vasculature and tracking circulating cells in CD41: GFP/Gata1: DsRed transgenic casper fish whose thrombocytes/erythrocytes express the green and red fluorescent proteins. Circulating fluorescent cell incidents were recorded and counted repeatedly over time and in different types of vessels. Great application opportunities in cancer and stem cell researches are discussed.

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

    PubMed

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

    2016-09-01

    Fibroblast growth factors (Fgfs) regulate critical biological processes such as embryonic development, tissue homeostasis, wound healing, and tissue regeneration. In zebrafish, Fgf signaling plays an important role in the regeneration of the spinal cord, liver, heart, fin, and photoreceptors, although its exact mechanism of action is not fully understood. Utilizing an adult zebrafish extraocular muscle (EOM) regeneration model, we demonstrate that blocking Fgf receptor function using either a chemical inhibitor (SU5402) or a dominant-negative transgenic construct (dnFGFR1a:EGFP) impairs muscle regeneration. Adult zebrafish EOMs regenerate through a myocyte dedifferentiation process, which involves a muscle-to-mesenchyme transition and cell cycle reentry by differentiated myocytes. Blocking Fgf signaling reduced cell proliferation and active caspase 3 levels in the regenerating muscle with no detectable levels of apoptosis, supporting the hypothesis that Fgf signaling is involved in the early steps of dedifferentiation. Fgf signaling in regenerating myocytes involves the MAPK/ERK pathway: inhibition of MEK activity with U0126 mimicked the phenotype of the Fgf receptor inhibition on both muscle regeneration and cell proliferation, and activated ERK (p-ERK) was detected in injured muscles by immunofluorescence and western blot. Interestingly, following injury, ERK2 expression is specifically induced and activated by phosphorylation, suggesting a key role in muscle regeneration. We conclude that the critical early steps of myocyte dedifferentiation in EOM regeneration are dependent on Fgf signaling. PMID:27267062

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

    PubMed Central

    Hui, Subhra Prakash; Nag, Tapas Chandra; Ghosh, Sukla

    2015-01-01

    Zebrafish can repair their injured brain and spinal cord after injury unlike adult mammalian central nervous system. Any injury to zebrafish spinal cord would lead to increased proliferation and neurogenesis. There are presences of proliferating progenitors from which both neuronal and glial loss can be reversed by appropriately generating new neurons and glia. We have demonstrated the presence of multiple progenitors, which are different types of proliferating populations like Sox2+ neural progenitor, A2B5+ astrocyte/ glial progenitor, NG2+ oligodendrocyte progenitor, radial glia and Schwann cell like progenitor. We analyzed the expression levels of two common markers of dedifferentiation like msx-b and vimentin during regeneration along with some of the pluripotency associated factors to explore the possible role of these two processes. Among the several key factors related to pluripotency, pou5f1 and sox2 are upregulated during regeneration and associated with activation of neural progenitor cells. Uncovering the molecular mechanism for endogenous regeneration of adult zebrafish spinal cord would give us more clues on important targets for future therapeutic approach in mammalian spinal cord repair and regeneration. PMID:26630262

  14. Gene expression changes in female zebrafish (Danio rerio) brain in response to acute exposure to methylmercury

    USGS Publications Warehouse

    Richter, Catherine A.; Garcia-Reyero, Natàlia; Martyniuk, Chris; Knoebl, Iris; Pope, Marie; Wright-Osment, Maureen K.; Denslow, Nancy D.; Tillitt, Donald E.

    2011-01-01

    Methylmercury (MeHg) is a potent neurotoxicant and endocrine disruptor that accumulates in aquatic systems. Previous studies have shown suppression of hormone levels in both male and female fish, suggesting effects on gonadotropin regulation in the brain. The gene expression profile in adult female zebrafish whole brain induced by acute (96 h) MeHg exposure was investigated. Fish were exposed by injection to 0 or 0.5(mu or u)g MeHg/g. Gene expression changes in the brain were examined using a 22,000-feature zebrafish microarray. At a significance level of pbrain. Future studies will compare the gene expression profile induced in response to MeHg with that induced by other toxicants and will investigate responsive genes as potential biomarkers of MeHg exposure.

  15. GENE EXPRESSION CHANGES IN FEMALE ZEBRAFISH (DANIO RERIO) BRAIN IN RESPONSE TO ACUTE EXPOSURE TO METHYLMERCURY

    PubMed Central

    Richter, Catherine A.; Garcia-Reyero, Natàlia; Martyniuk, Chris; Knoebl, Iris; Pope, Marie; Wright-Osment, Maureen K.; Denslow, Nancy D.; Tillitt, Donald E.

    2010-01-01

    Methylmercury (MeHg) is a potent neurotoxicant and endocrine disruptor that accumulates in aquatic systems. Previous studies have shown suppression of hormone levels in both male and female fish, suggesting effects on gonadotropin regulation in the brain. The gene expression profile in adult female zebrafish whole brain induced by acute (96 hr) MeHg exposure was investigated. Fish were exposed by injection to 0 or 0.5 μg MeHg/g. Gene expression changes in the brain were examined using a 22,000 feature zebrafish microarray. At a significance level of p<0.01, 79 genes were up-regulated and 76 genes were down-regulated in response to MeHg exposure. Individual genes exhibiting altered expression in response to MeHg exposure implicate effects on glutathione metabolism in the mechanism of MeHg neurotoxicity. Gene ontology (GO) terms significantly enriched among altered genes included protein folding, cell redox homeostasis, and steroid biosynthetic process. The most affected biological functions were related to the nervous system development and function, as well as lipid metabolism and molecular transport. These results support the involvement of oxidative stress and effects on protein structure in the mechanism of action of MeHg in the female brain. Future studies will compare the gene expression profile induced in response to MeHg with that induced by other toxicants and investigate responsive genes as potential biomarkers of MeHg exposure. PMID:21082716

  16. Targeted Electroporation in Embryonic, Larval, and Adult Zebrafish.

    PubMed

    Zou, Ming; Friedrich, Rainer W; Bianco, Isaac H

    2016-01-01

    This chapter describes three fast and straightforward methods to introduce nucleic acids, dyes, and other molecules into small numbers of cells of zebrafish embryos, larvae, and adults using electroporation. These reagents are delivered through a glass micropipette and electrical pulses are given through electrodes to permeabilize cell membranes and promote uptake of the reagent. This technique allows the experimenter to target cells of their choice at a particular time of development and at a particular location in the zebrafish with high precision and facilitates long-term noninvasive measurement of biological activities in vivo. Applications include cell fate mapping, neural circuit mapping, neuronal activity measurement, manipulation of activity, ectopic gene expression, and genetic knockdown experiments. PMID:27464813

  17. Preconditioning boosts regenerative programmes in the adult zebrafish heart

    PubMed Central

    de Preux Charles, Anne-Sophie; Bise, Thomas; Baier, Felix; Sallin, Pauline; Jaźwińska, Anna

    2016-01-01

    During preconditioning, exposure to a non-lethal harmful stimulus triggers a body-wide increase of survival and pro-regenerative programmes that enable the organism to better withstand the deleterious effects of subsequent injuries. This phenomenon has first been described in the mammalian heart, where it leads to a reduction of infarct size and limits the dysfunction of the injured organ. Despite its important clinical outcome, the actual mechanisms underlying preconditioning-induced cardioprotection remain unclear. Here, we describe two independent models of cardiac preconditioning in the adult zebrafish. As noxious stimuli, we used either a thoracotomy procedure or an induction of sterile inflammation by intraperitoneal injection of immunogenic particles. Similar to mammalian preconditioning, the zebrafish heart displayed increased expression of cardioprotective genes in response to these stimuli. As zebrafish cardiomyocytes have an endogenous proliferative capacity, preconditioning further elevated the re-entry into the cell cycle in the intact heart. This enhanced cycling activity led to a long-term modification of the myocardium architecture. Importantly, the protected phenotype brought beneficial effects for heart regeneration within one week after cryoinjury, such as a more effective cell-cycle reentry, enhanced reactivation of embryonic gene expression at the injury border, and improved cell survival shortly after injury. This study reveals that exposure to antecedent stimuli induces adaptive responses that render the fish more efficient in the activation of the regenerative programmes following heart damage. Our results open a new field of research by providing the adult zebrafish as a model system to study remote cardiac preconditioning. PMID:27440423

  18. Molecular psychiatry of zebrafish

    PubMed Central

    Stewart, Adam Michael; Ullmann, Jeremy F.P.; Norton, William H.J.; Brennan, Caroline H.; Parker, Matthew O.; Gerlai, Robert; Kalueff, Allan V.

    2014-01-01

    Due to their well-characterized neural development and high genetic homology to mammals, zebrafish (Danio rerio) have emerged as a powerful model organism in the field of biological psychiatry. Here, we discuss the molecular psychiatry of zebrafish, and its implications for translational neuroscience research and modeling CNS disorders. In particular, we outline recent genetic and technological developments allowing for in-vivo examinations, high-throughput screening and whole-brain analyses in larval and adult zebrafish. We also summarize the application of these molecular techniques to the understanding of neuropsychiatric disease, outlining the potential of zebrafish for modeling complex brain disorders, including attention-deficit/hyperactivity disorder (ADHD), aggression, post-traumatic stress and substance abuse. Critically evaluating the advantages and limitations of larval and adult fish tests, we suggest that zebrafish models become a rapidly emerging new field in modern biological psychiatry research. PMID:25349164

  19. Molecular psychiatry of zebrafish.

    PubMed

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

    2015-02-01

    Due to their well-characterized neural development and high genetic homology to mammals, zebrafish (Danio rerio) have emerged as a powerful model organism in the field of biological psychiatry. Here, we discuss the molecular psychiatry of zebrafish, and its implications for translational neuroscience research and modeling central nervous system (CNS) disorders. In particular, we outline recent genetic and technological developments allowing for in vivo examinations, high-throughput screening and whole-brain analyses in larval and adult zebrafish. We also summarize the application of these molecular techniques to the understanding of neuropsychiatric disease, outlining the potential of zebrafish for modeling complex brain disorders, including attention-deficit/hyperactivity disorder (ADHD), aggression, post-traumatic stress and substance abuse. Critically evaluating the advantages and limitations of larval and adult fish tests, we suggest that zebrafish models become a rapidly emerging new field in modern molecular psychiatry research. PMID:25349164

  20. Photopic and scotopic spatiotemporal tuning of adult zebrafish vision

    PubMed Central

    Hollbach, Nadine; Tappeiner, Christoph; Jazwinska, Anna; Enzmann, Volker; Tschopp, Markus

    2015-01-01

    Sensitivity to spatial and temporal patterns is a fundamental aspect of vision. Herein, we investigated this sensitivity in adult zebrafish for a wide range of spatial (0.014 to 0.511 cycles/degree [c/d]) and temporal frequencies (0.025 to 6 cycles/s) to better understand their visual system. Measurements were performed at photopic (1.8 log cd m−2) and scotopic (−4.5 log cd m−2) light levels to assess the optokinetic response (OKR). The resulting spatiotemporal contrast sensitivity (CS) functions revealed that the OKR of zebrafish is tuned to spatial frequency and speed but not to temporal frequencies. Thereby, optimal test parameters for CS measurements were identified. At photopic light levels, a spatial frequency of 0.116 ± 0.01 c/d (mean ± SD) and a grating speed of 8.42 ± 2.15 degrees/second (d/s) was ideal; at scotopic light levels, these values were 0.110 ± 0.02 c/d and 5.45 ± 1.31 d/s, respectively. This study allows to better characterize zebrafish mutants with altered vision and to distinguish between defects of rod and cone photoreceptors as measurements were performed under different light conditions. PMID:25788878

  1. In Vivo Whole-Cell Patch-Clamp Recording in the Zebrafish Brain.

    PubMed

    Zhang, Rong-Wei; Du, Jiu-Lin

    2016-01-01

    Zebrafish (Danio rerio) is a newly emerged vertebrate animal model with a conserved gross architecture of the brain and a rich repertoire of behaviors. Due to the optical transparency and structural simplicity of its brain, larval zebrafish has become an ideal in vivo model for dissecting neural mechanisms of brain functions at a whole-brain scale based on a strategy that spans scales from synapses, neurons, and circuits to behaviors. Whole-cell patch-clamp recording is an indispensable approach for studying synaptic and circuit mechanisms of brain functions. Due to the small size of neurons in the zebrafish brain, it is challenging to get whole-cell recordings from these cells. Here, we describe a protocol for obtaining in vivo whole-cell patch-clamp recordings from neurons in larval zebrafish. PMID:27464815

  2. Myocyte Dedifferentiation Drives Extraocular Muscle Regeneration in Adult Zebrafish

    PubMed Central

    Saera-Vila, Alfonso; Kasprick, Daniel S.; Junttila, Tyler L.; Grzegorski, Steven J.; Louie, Ke'ale W.; Chiari, Estelle F.; Kish, Phillip E.; Kahana, Alon

    2015-01-01

    Purpose The purpose of this study was to characterize the injury response of extraocular muscles (EOMs) in adult zebrafish. Methods Adult zebrafish underwent lateral rectus (LR) muscle myectomy surgery to remove 50% of the muscle, followed by molecular and cellular characterization of the tissue response to the injury. Results Following myectomy, the LR muscle regenerated an anatomically correct and functional muscle within 7 to 10 days post injury (DPI). Following injury, the residual muscle stump was replaced by a mesenchymal cell population that lost cell polarity and expressed mesenchymal markers. Next, a robust proliferative burst repopulated the area of the regenerating muscle. Regenerating cells expressed myod, identifying them as myoblasts. However, both immunofluorescence and electron microscopy failed to identify classic Pax7-positive satellite cells in control or injured EOMs. Instead, some proliferating nuclei were noted to express mef2c at the very earliest point in the proliferative burst, suggesting myonuclear reprogramming and dedifferentiation. Bromodeoxyuridine (BrdU) labeling of regenerating cells followed by a second myectomy without repeat labeling resulted in a twice-regenerated muscle broadly populated by BrdU-labeled nuclei with minimal apparent dilution of the BrdU signal. A double-pulse experiment using BrdU and 5-ethynyl-2′-deoxyuridine (EdU) identified double-labeled nuclei, confirming the shared progenitor lineage. Rapid regeneration occurred despite a cell cycle length of 19.1 hours, whereas 72% of the regenerating muscle nuclei entered the cell cycle by 48 hours post injury (HPI). Dextran lineage tracing revealed that residual myocytes were responsible for muscle regeneration. Conclusions EOM regeneration in adult zebrafish occurs by dedifferentiation of residual myocytes involving a muscle-to-mesenchyme transition. A mechanistic understanding of myocyte reprogramming may facilitate novel approaches to the development of molecular

  3. In vivo Electroporation of Morpholinos into the Adult Zebrafish Retina

    PubMed Central

    Thummel, Ryan; Bailey, Travis J.; Hyde, David R.

    2011-01-01

    Many devastating inherited eye diseases result in progressive and irreversible blindness because humans cannot regenerate dying or diseased retinal neurons. In contrast, the adult zebrafish retina possesses the robust ability to spontaneously regenerate any neuronal class that is lost in a variety of different retinal damage models, including retinal puncture, chemical ablation, concentrated high temperature, and intense light treatment 1-8. Our lab extensively characterized regeneration of photoreceptors following constant intense light treatment and inner retinal neurons after intravitreal ouabain injection 2, 5, 9. In all cases, resident Müller glia re-enter the cell cycle to produce neuronal progenitors, which continue to proliferate and migrate to the proper retinal layer, where they differentiate into the deficient neurons. We characterized five different stages during regeneration of the light-damaged retina that were highlighted by specific cellular responses. We identified several differentially expressed genes at each stage of retinal regeneration by mRNA microarray analysis 10. Many of these genes are also critical for ocular development. To test the role of each candidate gene/protein during retinal regeneration, we needed to develop a method to conditionally limit the expression of a candidate protein only at times during regeneration of the adult retina. Morpholino oligos are widely used to study loss of function of specific proteins during the development of zebrafish, Xenopus, chick, mouse, and tumors in human xenografts 11-14. These modified oligos basepair with complementary RNA sequence to either block the splicing or translation of the target RNA. Morpholinos are stable in the cell and can eliminate or "knockdown" protein expression for three to five days 12. Here, we describe a method to efficiently knockdown target protein expression in the adult zebrafish retina. This method employs lissamine-tagged antisense morpholinos that are injected

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

    PubMed Central

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

    2016-01-01

    Enantioselective enrichment of chiral PCB149 (2,2’,3,4’,5’,6-hexachlorobiphenyl) was analysed in adult zebrafish (Danio rerio) exposed to the racemate, (−)-PCB149, and (+)-PCB149. Greater enrichment of (−)-PCB149 compared to (+) PCB149 was observed following 0.5 ng/L exposure; however, as the exposure time and concentration increased, racemic enrichment was observed in adult fish exposed to the racemate. No biotransformation between the two isomers was observed in fish exposed to single enantiomers. When zebrafish were exposed to different forms of chiral PCB149, enantioselective expression of genes associated with polychlorinated biphenyls (PCBs) was observed in brain and liver tissues and enantioselective correlations between bioconcentration and target gene expression levels were observed in brain and liver tissues. The strong positive correlations between expression levels of target genes (alox5a and alox12) and PCB149 bioconcentration suggest that prolonged exposure to the racemate of chiral PCB149 may result in inflammation-associated diseases. Prolonged exposure to (−)-PCB149 may also affect metabolic pathways such as dehydrogenation and methylation in the brain tissues of adult zebrafish. Hepatic expression levels of genes related to the antioxidant system were significantly negatively correlated with bioconcentration following exposure to (+)-PCB149. PMID:26786282

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

    PubMed

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

    2016-01-01

    Enantioselective enrichment of chiral PCB149 (2,2',3,4',5',6-hexachlorobiphenyl) was analysed in adult zebrafish (Danio rerio) exposed to the racemate, (-)-PCB149, and (+)-PCB149. Greater enrichment of (-)-PCB149 compared to (+) PCB149 was observed following 0.5 ng/L exposure; however, as the exposure time and concentration increased, racemic enrichment was observed in adult fish exposed to the racemate. No biotransformation between the two isomers was observed in fish exposed to single enantiomers. When zebrafish were exposed to different forms of chiral PCB149, enantioselective expression of genes associated with polychlorinated biphenyls (PCBs) was observed in brain and liver tissues and enantioselective correlations between bioconcentration and target gene expression levels were observed in brain and liver tissues. The strong positive correlations between expression levels of target genes (alox5a and alox12) and PCB149 bioconcentration suggest that prolonged exposure to the racemate of chiral PCB149 may result in inflammation-associated diseases. Prolonged exposure to (-)-PCB149 may also affect metabolic pathways such as dehydrogenation and methylation in the brain tissues of adult zebrafish. Hepatic expression levels of genes related to the antioxidant system were significantly negatively correlated with bioconcentration following exposure to (+)-PCB149. PMID:26786282

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

    NASA Astrophysics Data System (ADS)

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

    2016-01-01

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

  7. Brain and Spinal Cord Tumors in Adults

    MedlinePlus

    ... saved articles window. My Saved Articles » My ACS » Brain and Spinal Cord Tumors in Adults Download Printable ... the topics below to get started. What Is Brain/CNS Tumors In Adults? What are adult brain ...

  8. Behavioral effects of bidirectional modulation of brain monoamines by reserpine and d-amphetamine in zebrafish

    PubMed Central

    Kyzar, Evan; Stewart, Adam Michael; Landsman, Samuel; Collins, Christopher; Gebhardt, Michael; Robinson, Kyle; Kalueff, Allan V.

    2013-01-01

    Brain monoamines play a key role in the regulation of behavior. Reserpine depletes monoamines, and causes depression and hypoactivity in humans and rodents. In contrast, d-amphetamine increases brain monoamines’ levels, and evokes hyperactivity and anxiety. However, the effects of these agents on behavior and in relation to monoamine levels remain poorly understood, necessitating further experimental studies to understand their psychotropic action. Zebrafish (Danio rerio) are rapidly emerging as a promising model organism for drug screening and translational neuroscience research. Here, we have examined the acute and long-term effects of reserpine and d-amphetamine on zebrafish behavior in the novel tank test. Overall, d-amphetamine (5 and 10 mg/L) evokes anxiogenic-like effects in zebrafish acutely, but not 7 days later. In contrast, reserpine (20 and 40 mg/L) did not evoke overt acute behavioral effects, but markedly reduced activity 7 days later, resembling motor retardation observed in depression and/or Parkinson’s disease. Three-dimensional ‘temporal’ (X, Y, Time) reconstructions of zebrafish locomotion further supports these findings, confirming the utility of 3D-based video-tracking analyses in zebrafish models of drug action. Our results show that zebrafish are highly sensitive to drugs bi-directionally modulating brain monoamines, generally paralleling rodent and clinical findings. Collectively, this emphasizes the potential of zebrafish tests to model complex brain disorders associated with monoamine dysregulation. PMID:23827499

  9. Brain Transcriptomic Response to Social Eavesdropping in Zebrafish (Danio rerio)

    PubMed Central

    Oliveira, Rui F.

    2015-01-01

    Public information is widely available at low cost to animals living in social groups. For instance, bystanders may eavesdrop on signaling interactions between conspecifics and use it to adapt their subsequent behavior towards the observed individuals. This social eavesdropping ability is expected to require specialized mechanisms such as social attention, which selects social information available for learning. To begin exploring the genetic basis of social eavesdropping, we used a previously established attention paradigm in the lab to study the brain gene expression profile of male zebrafish (Danio rerio) in relation to the attention they paid towards conspecifics involved or not involved in agonistic interactions. Microarray gene chips were used to characterize their brain transcriptomes based on differential expression of single genes and gene sets. These analyses were complemented by promoter region-based techniques. Using data from both approaches, we further drafted protein interaction networks. Our results suggest that attentiveness towards conspecifics, whether interacting or not, activates pathways linked to neuronal plasticity and memory formation. The network analyses suggested that fos and jun are key players on this response, and that npas4a, nr4a1 and egr4 may also play an important role. Furthermore, specifically observing fighting interactions further triggered pathways associated to a change in the alertness status (dnajb5) and to other genes related to memory formation (btg2, npas4b), which suggests that the acquisition of eavesdropped information about social relationships activates specific processes on top of those already activated just by observing conspecifics. PMID:26713440

  10. Brain Transcriptomic Response to Social Eavesdropping in Zebrafish (Danio rerio).

    PubMed

    Lopes, João Sollari; Abril-de-Abreu, Rodrigo; Oliveira, Rui F

    2015-01-01

    Public information is widely available at low cost to animals living in social groups. For instance, bystanders may eavesdrop on signaling interactions between conspecifics and use it to adapt their subsequent behavior towards the observed individuals. This social eavesdropping ability is expected to require specialized mechanisms such as social attention, which selects social information available for learning. To begin exploring the genetic basis of social eavesdropping, we used a previously established attention paradigm in the lab to study the brain gene expression profile of male zebrafish (Danio rerio) in relation to the attention they paid towards conspecifics involved or not involved in agonistic interactions. Microarray gene chips were used to characterize their brain transcriptomes based on differential expression of single genes and gene sets. These analyses were complemented by promoter region-based techniques. Using data from both approaches, we further drafted protein interaction networks. Our results suggest that attentiveness towards conspecifics, whether interacting or not, activates pathways linked to neuronal plasticity and memory formation. The network analyses suggested that fos and jun are key players on this response, and that npas4a, nr4a1 and egr4 may also play an important role. Furthermore, specifically observing fighting interactions further triggered pathways associated to a change in the alertness status (dnajb5) and to other genes related to memory formation (btg2, npas4b), which suggests that the acquisition of eavesdropped information about social relationships activates specific processes on top of those already activated just by observing conspecifics. PMID:26713440

  11. Zebrafish brain mapping--standardized spaces, length scales, and the power of N and n.

    PubMed

    Hunter, Paul R; Hendry, Aenea C; Lowe, Andrew S

    2015-06-01

    Mapping anatomical and functional parameters of the zebrafish brain is moving apace. Research communities undertaking such studies are becoming ever larger and more diverse. The unique features, tools, and technologies associated with zebrafish are propelling them as the 21st century model organism for brain mapping. Uniquely positioned as a vertebrate model system, the zebrafish enables imaging of anatomy and function at different length scales from intraneuronal compartments to sparsely distributed whole brain patterns. With a variety of diverse and established statistical modeling and analytic methods available from the wider brain mapping communities, the richness of zebrafish neuroimaging data is being realized. The statistical power of population observations (N) within and across many samples (n) projected onto a standardized space will provide vast databases for data-driven biological approaches. This article reviews key brain mapping initiatives at different levels of scale that highlight the potential of zebrafish brain mapping. By way of introduction to the next wave of brain mappers, an accessible introduction to the key concepts and caveats associated with neuroimaging are outlined and discussed. PMID:25418847

  12. Sex differences in DNA methylation and expression in zebrafish brain: a test of an extended 'male sex drive' hypothesis.

    PubMed

    Chatterjee, Aniruddha; Lagisz, Malgorzata; Rodger, Euan J; Zhen, Li; Stockwell, Peter A; Duncan, Elizabeth J; Horsfield, Julia A; Jeyakani, Justin; Mathavan, Sinnakaruppan; Ozaki, Yuichi; Nakagawa, Shinichi

    2016-09-30

    The sex drive hypothesis predicts that stronger selection on male traits has resulted in masculinization of the genome. Here we test whether such masculinizing effects can be detected at the level of the transcriptome and methylome in the adult zebrafish brain. Although methylation is globally similar, we identified 914 specific differentially methylated CpGs (DMCs) between males and females (435 were hypermethylated and 479 were hypomethylated in males compared to females). These DMCs were prevalent in gene body, intergenic regions and CpG island shores. We also discovered 15 distinct CpG clusters with striking sex-specific DNA methylation differences. In contrast, at transcriptome level, more female-biased genes than male-biased genes were expressed, giving little support for the male sex drive hypothesis. Our study provides genome-wide methylome and transcriptome assessment and sheds light on sex-specific epigenetic patterns and in zebrafish for the first time. PMID:27259666

  13. Advanced Echocardiography in Adult Zebrafish Reveals Delayed Recovery of Heart Function after Myocardial Cryoinjury

    PubMed Central

    Kossack, Mandy; Juergensen, Lonny; Fuchs, Dieter; Katus, Hugo A.; Hassel, David

    2015-01-01

    Translucent zebrafish larvae represent an established model to analyze genetics of cardiac development and human cardiac disease. More recently adult zebrafish are utilized to evaluate mechanisms of cardiac regeneration and by benefiting from recent genome editing technologies, including TALEN and CRISPR, adult zebrafish are emerging as a valuable in vivo model to evaluate novel disease genes and specifically validate disease causing mutations and their underlying pathomechanisms. However, methods to sensitively and non-invasively assess cardiac morphology and performance in adult zebrafish are still limited. We here present a standardized examination protocol to broadly assess cardiac performance in adult zebrafish by advancing conventional echocardiography with modern speckle-tracking analyses. This allows accurate detection of changes in cardiac performance and further enables highly sensitive assessment of regional myocardial motion and deformation in high spatio-temporal resolution. Combining conventional echocardiography measurements with radial and longitudinal velocity, displacement, strain, strain rate and myocardial wall delay rates after myocardial cryoinjury permitted to non-invasively determine injury dimensions and to longitudinally follow functional recovery during cardiac regeneration. We show that functional recovery of cryoinjured hearts occurs in three distinct phases. Importantly, the regeneration process after cryoinjury extends far beyond the proposed 45 days described for ventricular resection with reconstitution of myocardial performance up to 180 days post-injury (dpi). The imaging modalities evaluated here allow sensitive cardiac phenotyping and contribute to further establish adult zebrafish as valuable cardiac disease model beyond the larval developmental stage. PMID:25853735

  14. Probiotic modulation of the microbiota-gut-brain axis and behaviour in zebrafish

    PubMed Central

    Borrelli, Luca; Aceto, Serena; Agnisola, Claudio; De Paolo, Sofia; Dipineto, Ludovico; Stilling, Roman M.; Dinan, Timothy G.; Cryan, John F.; Menna, Lucia F.; Fioretti, Alessandro

    2016-01-01

    The gut microbiota plays a crucial role in the bi-directional gut–brain axis, a communication that integrates the gut and central nervous system (CNS) activities. Animal studies reveal that gut bacteria influence behaviour, Brain-Derived Neurotrophic Factor (BDNF) levels and serotonin metabolism. In the present study, we report for the first time an analysis of the microbiota–gut–brain axis in zebrafish (Danio rerio). After 28 days of dietary administration with the probiotic Lactobacillus rhamnosus IMC 501, we found differences in shoaling behaviour, brain expression levels of bdnf and of genes involved in serotonin signalling/metabolism between control and treated zebrafish group. In addition, in microbiota we found a significant increase of Firmicutes and a trending reduction of Proteobacteria. This study demonstrates that selected microbes can be used to modulate endogenous neuroactive molecules in zebrafish. PMID:27416816

  15. Probiotic modulation of the microbiota-gut-brain axis and behaviour in zebrafish.

    PubMed

    Borrelli, Luca; Aceto, Serena; Agnisola, Claudio; De Paolo, Sofia; Dipineto, Ludovico; Stilling, Roman M; Dinan, Timothy G; Cryan, John F; Menna, Lucia F; Fioretti, Alessandro

    2016-01-01

    The gut microbiota plays a crucial role in the bi-directional gut-brain axis, a communication that integrates the gut and central nervous system (CNS) activities. Animal studies reveal that gut bacteria influence behaviour, Brain-Derived Neurotrophic Factor (BDNF) levels and serotonin metabolism. In the present study, we report for the first time an analysis of the microbiota-gut-brain axis in zebrafish (Danio rerio). After 28 days of dietary administration with the probiotic Lactobacillus rhamnosus IMC 501, we found differences in shoaling behaviour, brain expression levels of bdnf and of genes involved in serotonin signalling/metabolism between control and treated zebrafish group. In addition, in microbiota we found a significant increase of Firmicutes and a trending reduction of Proteobacteria. This study demonstrates that selected microbes can be used to modulate endogenous neuroactive molecules in zebrafish. PMID:27416816

  16. Increased cell proliferation and neural activity by physostigmine in the telencephalon of adult zebrafish.

    PubMed

    Lee, Yunkyoung; Lee, Bongkyu; Jeong, Sumin; Park, Ji-Won; Han, Inn-Oc; Lee, Chang-Joong

    2016-08-26

    Physostigmine, an acetylcholinesterase inhibitor, is known to affect the brain function in various aspects. This study was conducted to test whether physostigmine affects cell proliferation in the telencephalon of zebrafish. BrdU-labeled cells was prominently observed in the ventral zone of the ventral telencephalon of zebrafish. The increased number of BrdU- and proliferating cell nuclear antigen-labeled cells were shown in zebrafish treated with 200μM physostigmine, which was inhibited by pretreatment with 200μM scopolamine. iNOS mRNA expression was increased in the brain of zebrafish treated with 200μM physostigmine. Consistently, aminoguanidine, an iNOS inhibitor, attenuated the increase in the number of BrdU-labeled cells by physostigmine treatment. Zebrafish also showed seizure-like locomotor activity characterized by a rapid and abrupt movement during a 30min treatment with 200μM physostigmine. Neural activity in response to an electrical stimulus was increased in the isolated telencephalon of zebrafish continuously perfused with 200μM physostigmine. None of the number of BrdU-labeled cells, neural activity, or locomotor activity was affected by treatment with 20μM physostigmine. These results suggest that 200μM physostigmine increased neural activity and induced cell proliferation via nitric oxide production in zebrafish. PMID:27378362

  17. Copper at low levels impairs memory of adult zebrafish (Danio rerio) and affects swimming performance of larvae.

    PubMed

    Acosta, Daiane da Silva; Danielle, Naissa Maria; Altenhofen, Stefani; Luzardo, Milene Dornelles; Costa, Patrícia Gomes; Bianchini, Adalto; Bonan, Carla Denise; da Silva, Rosane Souza; Dafre, Alcir Luiz

    2016-01-01

    Metal contamination at low levels is an important issue because it usually produces health and environmental effects, either positive or deleterious. Contamination of surface waters with copper (Cu) is a worldwide event, usually originated by mining, agricultural, industrial, commercial, and residential activities. Water quality criteria for Cu are variable among countries but allowed limits are generally in the μg/L range, which can disrupt several functions in the early life-stages of fish species. Behavioral and biochemical alterations after Cu exposure have also been described at concentrations close to the allowed limits. Aiming to search for the effects of Cu in the range of the allowed limits, larvae and adult zebrafish (Danio rerio) were exposed to different concentrations of dissolved Cu (nominally: 0, 5, 9, 20 and 60μg/L; measured: 0.4, 5.7, 7.2 16.6 and 42.3μg/L, respectively) for 96h. Larvae swimming and body length, and adult behavior and biochemical biomarkers (activity of glutathione-related enzymes in gills, muscle, and brain) were assessed after Cu exposure. Several effects were observed in fish exposed to 9μg/L nominal Cu, including increased larvae swimming distance and velocity, abolishment of adult inhibitory avoidance memory, and decreased glutathione S-transferase (GST) activity in gills of adult fish. At the highest Cu concentration tested (nominally: 60μg/L), body length of larvae, spatial memory of adults, and gill GST activity were decreased. Social behavior (aggressiveness and conspecific interaction), and glutathione reductase (GR) activity were not affected in adult zebrafish. Exposure to Cu, at concentrations close to the water quality criteria for this metal in fresh water, was able to alter larvae swimming performance and to induce detrimental effects on the behavior of adult zebrafish, thus indicating the need for further studies to reevaluate the currently allowed limits for Cu in fresh water. PMID:27012768

  18. Embryonic Atrazine Exposure Elicits Alterations in Genes Associated with Neuroendocrine Function in Adult Male Zebrafish.

    PubMed

    Wirbisky, Sara E; Sepúlveda, Maria S; Weber, Gregory J; Jannasch, Amber S; Horzmann, Katharine A; Freeman, Jennifer L

    2016-09-01

    The developmental origins of health and disease (DOHaD) hypothesis states that exposure to environmental stressors early in life can elicit genome and epigenome changes resulting in an increased susceptibility of a disease state during adulthood. Atrazine, a common agricultural herbicide used throughout the Midwestern United States, frequently contaminates potable water supplies and is a suspected endocrine disrupting chemical. In our previous studies, zebrafish was exposed to 0, 0.3, 3, or 30 parts per billion (μg/l) atrazine through embryogenesis, rinsed, and allowed to mature to adulthood. A decrease in spawning was observed with morphological alterations in offspring. In addition, adult females displayed an increase in ovarian progesterone and follicular atresia, alterations in levels of a serotonin metabolite and serotonin turnover in brain tissue, and transcriptome changes in brain and ovarian tissue supporting neuroendocrine alterations. As reproductive dysfunction is also influenced by males, this study assessed testes histology, hormone levels, and transcriptomic profiles of testes and brain tissue in the adult males. The embryonic atrazine exposure resulted in no alterations in body or testes weight, gonadosomatic index, testes histology, or levels of 11-ketotestosterone or testosterone. To further investigate potential alterations, transcriptomic profiles of adult male testes and brain tissue was completed. This analysis demonstrated alterations in genes associated with abnormal cell and neuronal growth and morphology; molecular transport, quantity, and production of steroid hormones; and neurotransmission with an emphasis on the hypothalamus-pituitary-adrenal and hypothalamus-pituitary-thyroid axes. Overall, this data indicate future studies should focus on additional neuroendocrine endpoints to determine potential functional impairments. PMID:27413107

  19. High Cholesterol Diet Induces IL-1β Expression in Adult but Not Larval Zebrafish

    PubMed Central

    Jang, Man-Young; Na, Yirang; Ko, Youngho; Choi, Jae-Hoon; Seok, Seung Hyeok

    2013-01-01

    Recently, it has been demonstrated that high cholesterol diet induced hypercholesterolemia and vascular lipid oxidation and accumulation in zebrafish larvae, suggesting that zebrafish is a new promising atherosclerosis model in addition to mouse models. However, up to date, there was no report regarding inflammatory cytokine expression during the lipid accumulation in zebrafish larva and adult fish. In this study, we first demonstrated the expression levels of IL-1β and TNF-α in high cholesterol diet (HCD)-fed zebrafish larvae, and found that although HCD induced vascular lipid accumulation, the cytokine expressions in the larvae were not changed by HCD. Furthermore, there was no significant difference in leukocyte accumulation in vessels between control and HCD fed group. But prolonged HCD induced IL-1β expression in spleen and liver compared to those of control zebrafish, and produced very early stage of fatty streak lesion in dorsal aorta of 19 week HCD-fed zebrafish. These results indicate that HCD induced hypercholesterolemia and atherosclerotic changes in zebrafish are very early stage, and suggest the necessity of the generation of mutant zebrafish having a disruption in a lipid metabolism-related gene leading to severe hypercholesterolemia and advanced atherosclerosis. PMID:23825600

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

    USGS Publications Warehouse

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

    2009-01-01

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

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

    PubMed Central

    Raymond, Pamela A; Barthel, Linda K; Bernardos, Rebecca L; Perkowski, John J

    2006-01-01

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

  2. Long-term drug administration in the adult zebrafish using oral gavage for cancer preclinical studies

    PubMed Central

    Dang, Michelle; Henderson, Rachel E.; Garraway, Levi A.

    2016-01-01

    ABSTRACT Zebrafish are a major model for chemical genetics, and most studies use embryos when investigating small molecules that cause interesting phenotypes or that can rescue disease models. Limited studies have dosed adults with small molecules by means of water-borne exposure or injection techniques. Challenges in the form of drug delivery-related trauma and anesthesia-related toxicity have excluded the adult zebrafish from long-term drug efficacy studies. Here, we introduce a novel anesthetic combination of MS-222 and isoflurane to an oral gavage technique for a non-toxic, non-invasive and long-term drug administration platform. As a proof of principle, we established drug efficacy of the FDA-approved BRAFV600E inhibitor, Vemurafenib, in adult zebrafish harboring BRAFV600E melanoma tumors. In the model, adult casper zebrafish intraperitoneally transplanted with a zebrafish melanoma cell line (ZMEL1) and exposed to daily sub-lethal dosing at 100 mg/kg of Vemurafenib for 2 weeks via oral gavage resulted in an average 65% decrease in tumor burden and a 15% mortality rate. In contrast, Vemurafenib-resistant ZMEL1 cell lines, generated in culture from low-dose drug exposure for 4 months, did not respond to the oral gavage treatment regimen. Similarly, this drug treatment regimen can be applied for treatment of primary melanoma tumors in the zebrafish. Taken together, we developed an effective long-term drug treatment system that will allow the adult zebrafish to be used to identify more effective anti-melanoma combination therapies and opens up possibilities for treating adult models of other diseases. PMID:27482819

  3. Morphologic and functional characterization of granulocytes and macrophages in embryonic and adult zebrafish.

    PubMed

    Lieschke, G J; Oates, A C; Crowhurst, M O; Ward, A C; Layton, J E

    2001-11-15

    The zebrafish is a useful model organism for developmental and genetic studies. The morphology and function of zebrafish myeloid cells were characterized. Adult zebrafish contain 2 distinct granulocytes, a heterophil and a rarer eosinophil, both of which circulate and are generated in the kidney, the adult hematopoietic organ. Heterophils show strong histochemical myeloperoxidasic activity, although weaker peroxidase activity was observed under some conditions in eosinophils and erythrocytes. Embryonic zebrafish have circulating immature heterophils by 48 hours after fertilization (hpf). A zebrafish myeloperoxidase homologue (myeloid-specific peroxidase; mpx) was isolated. Phylogenetic analysis suggested it represented a gene ancestral to the mammalian myeloperoxidase gene family. It was expressed in adult granulocytes and in embryos from 18 hpf, first diffusely in the axial intermediate cell mass and then discretely in a dispersed cell population. Comparison of hemoglobinized cell distribution, mpx gene expression, and myeloperoxidase histochemistry in wild-type and mutant embryos confirmed that the latter reliably identified a population of myeloid cells. Studies in embryos after tail transection demonstrated that mpx- and peroxidase-expressing cells were mobile and localized to a site of inflammation, indicating functional capability of these embryonic granulocytes. Embryonic macrophages removed carbon particles from the circulation by phagocytosis. Collectively, these observations have demonstrated the early onset of zebrafish granulopoiesis, have proved that granulocytes circulate by 48 hpf, and have demonstrated the functional activity of embryonic granulocytes and macrophages. These observations will facilitate the application of this genetically tractable organism to the study of myelopoiesis. PMID:11698295

  4. Proline-induced changes in acetylcholinesterase activity and gene expression in zebrafish brain: reversal by antipsychotic drugs.

    PubMed

    Savio, L E B; Vuaden, F C; Kist, L W; Pereira, T C; Rosemberg, D B; Bogo, M R; Bonan, C D; Wyse, A T S

    2013-10-10

    Hyperprolinemia is an inherited disorder of proline metabolism and hyperprolinemic patients can present neurological manifestations, such as seizures, cognitive dysfunctions, and schizoaffective disorders. However, the mechanisms related to these symptoms are still unclear. In the present study, we evaluated the in vivo and in vitro effects of proline on acetylcholinesterase (AChE) activity and gene expression in the zebrafish brain. For the in vivo studies, animals were exposed at two proline concentrations (1.5 and 3.0mM) during 1h or 7 days (short- or long-term treatments, respectively). For the in vitro assays, different proline concentrations (ranging from 3.0 to 1000 μM) were tested. Long-term proline exposures significantly increased AChE activity for both treated groups when compared to the control (34% and 39%). Moreover, the proline-induced increase on AChE activity was completely reverted by acute administration of antipsychotic drugs (haloperidol and sulpiride), as well as the changes induced in ache expression. When assessed in vitro, proline did not promote significant changes in AChE activity. Altogether, these data indicate that the enzyme responsible for the control of acetylcholine levels might be altered after proline exposure in the adult zebrafish. These findings contribute for better understanding of the pathophysiology of hyperprolinemia and might reinforce the use of the zebrafish as a complementary vertebrate model for studying inborn errors of amino acid metabolism. PMID:23867765

  5. Directional and color preference in adult zebrafish: Implications in behavioral and learning assays in neurotoxicology studies.

    PubMed

    Bault, Zachary A; Peterson, Samuel M; Freeman, Jennifer L

    2015-12-01

    The zebrafish (Danio rerio) is a useful vertebrate model organism for neurological studies. While a number of behavior and learning assays are recently reported in the literature for zebrafish, many of these assays are still being refined. The initial purpose of this study was to apply a published T-maze assay for adult zebrafish that measures how quickly an organism can discriminate between different color stimuli after receiving reinforcement to measure learning in a study investigating the later life impacts of developmental Pb exposure. The original results were inconclusive as the control group showed a directional and color preference. To assess directional preference further, a three-chambered testing apparatus was constructed and rotated in several directions. The directional preference observed in males was alleviated by rotating the arms pointing west and east. In addition, color preference was investigated using all combinations of five different colors (orange, yellow, green, blue and purple). With directional preference alleviated results showed that both male and female zebrafish preferred colors of shorter wavelengths. An additional experiment tested changes in color preference due to developmental exposure to Pb in adult male zebrafish. Results revealed that Pb-exposed males gained and lost certain color preferences compared to control males and the preference for short wavelengths was decreased. Overall, these results show that consideration and pretesting should be completed before applying behavioral and learning assays involving adult zebrafish to avoid innate preferences and confounding changes in neurotoxicology studies and that developmental Pb exposure alters color preferences in adult male zebrafish. PMID:25993913

  6. Estrogenic Effects of Several BPA Analogs in the Developing Zebrafish Brain.

    PubMed

    Cano-Nicolau, Joel; Vaillant, Colette; Pellegrini, Elisabeth; Charlier, Thierry D; Kah, Olivier; Coumailleau, Pascal

    2016-01-01

    Important set of studies have demonstrated the endocrine disrupting activity of Bisphenol A (BPA). The present work aimed at defining estrogenic-like activity of several BPA structural analogs, including BPS, BPF, BPAF, and BPAP, on 4- or 7-day post-fertilization (dpf) zebrafish larva as an in vivo model. We measured the induction level of the estrogen-sensitive marker cyp19a1b gene (Aromatase B), expressed in the brain, using three different in situ/in vivo strategies: (1) Quantification of cyp19a1b transcripts using RT-qPCR in wild type 7-dpf larva brains exposed to bisphenols; (2) Detection and distribution of cyp19a1b transcripts using in situ hybridization on 7-dpf brain sections (hypothalamus); and (3) Quantification of the cyp19a1b promoter activity in live cyp19a1b-GFP transgenic zebrafish (EASZY assay) at 4-dpf larval stage. These three different experimental approaches demonstrated that BPS, BPF, or BPAF exposure, similarly to BPA, significantly activates the expression of the estrogenic marker in the brain of developing zebrafish. In vitro experiments using both reporter gene assay in a glial cell context and competitive ligand binding assays strongly suggested that up-regulation of cyp19a1b is largely mediated by the zebrafish estrogen nuclear receptor alpha (zfERα). Importantly, and in contrast to other tested bisphenol A analogs, the bisphenol AP (BPAP) did not show estrogenic activity in our model. PMID:27047331

  7. Estrogenic Effects of Several BPA Analogs in the Developing Zebrafish Brain

    PubMed Central

    Cano-Nicolau, Joel; Vaillant, Colette; Pellegrini, Elisabeth; Charlier, Thierry D.; Kah, Olivier; Coumailleau, Pascal

    2016-01-01

    Important set of studies have demonstrated the endocrine disrupting activity of Bisphenol A (BPA). The present work aimed at defining estrogenic-like activity of several BPA structural analogs, including BPS, BPF, BPAF, and BPAP, on 4- or 7-day post-fertilization (dpf) zebrafish larva as an in vivo model. We measured the induction level of the estrogen-sensitive marker cyp19a1b gene (Aromatase B), expressed in the brain, using three different in situ/in vivo strategies: (1) Quantification of cyp19a1b transcripts using RT-qPCR in wild type 7-dpf larva brains exposed to bisphenols; (2) Detection and distribution of cyp19a1b transcripts using in situ hybridization on 7-dpf brain sections (hypothalamus); and (3) Quantification of the cyp19a1b promoter activity in live cyp19a1b-GFP transgenic zebrafish (EASZY assay) at 4-dpf larval stage. These three different experimental approaches demonstrated that BPS, BPF, or BPAF exposure, similarly to BPA, significantly activates the expression of the estrogenic marker in the brain of developing zebrafish. In vitro experiments using both reporter gene assay in a glial cell context and competitive ligand binding assays strongly suggested that up-regulation of cyp19a1b is largely mediated by the zebrafish estrogen nuclear receptor alpha (zfERα). Importantly, and in contrast to other tested bisphenol A analogs, the bisphenol AP (BPAP) did not show estrogenic activity in our model. PMID:27047331

  8. In vivo imaging of zebrafish from embryo to adult stage with optical projection tomography

    NASA Astrophysics Data System (ADS)

    Bassi, Andrea; Fieramonti, Luca; D'Andrea, Cosimo; Valentini, Gianluca; Cubeddu, Rinaldo; De Silvestri, Sandro; Cerullo, Giulio; Foglia, Efrem; Cotelli, Franco

    2013-02-01

    Optical Projection Tomography (OPT) is a three dimensional imaging technique that is particularly suitable for studying millimeter sized biological samples and organisms. Similarly to x-ray computed tomography, OPT is based on the acquisition of a sequence of images taken through the sample at many angles (projections). Assuming the linearity of the optical absorption process, the projections are combined to reconstruct the 3-D volume of the sample, typically using a filtered back-projection algorithm. OPT has been applied to in-vivo imaging of zebrafish (Danio rerio). The instrument and the protocol for in vivo imaging of zebrafish embryos and juvenile specimens are described. Light scattering remains a challenge for in vivo OPT, especially when samples at the upper size limit, like zebrafish at the adult stage, are under study. We describe Time-Gated Optical Projection Tomography (TGOPT), a technique able to reconstruct adult zebrafish internal structures by counteracting the scattering effects through a fast time-gate. The time gating mechanism is based on non-linear optical upconversion of an infrared ultrashort laser pulse and allows the detection of quasi-ballistic photons within a 100 fs temporal gate. This results in a strong improvement in contrast and resolution with respect to conventional OPT. Artifacts in the reconstructed images are reduced as well. We show that TGOPT is suited for imaging the skeletal system and nervous structures of adult zebrafish.

  9. Single-cell in vivo imaging of adult neural stem cells in the zebrafish telencephalon.

    PubMed

    Barbosa, Joana S; Di Giaimo, Rossella; Götz, Magdalena; Ninkovic, Jovica

    2016-08-01

    Adult neural stem cells (aNSCs) in zebrafish produce mature neurons throughout their entire life span in both the intact and regenerating brain. An understanding of the behavior of aNSCs in their intact niche and during regeneration in vivo should facilitate the identification of the molecular mechanisms controlling regeneration-specific cellular events. A greater understanding of the process in regeneration-competent species may enable regeneration to be achieved in regeneration-incompetent species, including humans. Here we describe a protocol for labeling and repetitive imaging of aNSCs in vivo. We label single aNSCs, allowing nonambiguous re-identification of single cells in repetitive imaging sessions using electroporation of a red-reporter plasmid in Tg(gfap:GFP)mi2001 transgenic fish expressing GFP in aNSCs. We image using two-photon microscopy through the thinned skull of anesthetized and immobilized fish. Our protocol allows imaging every 2 d for a period of up to 1 month. This methodology allowed the visualization of aNSC behavior in vivo in their natural niche, in contrast to previously available technologies, which rely on the imaging of either dissociated cells or tissue slices. We used this protocol to follow the mode of aNSC division, fate changes and cell death in both the intact and injured zebrafish telencephalon. This experimental setup can be widely used, with minimal prior experience, to assess key factors for processes that modulate aNSC behavior. A typical experiment with data analysis takes up to 1.5 months. PMID:27362338

  10. Zebrafish locomotor capacity and brain acetylcholinesterase activity is altered by Aphanizomenon flos-aquae DC-1 aphantoxins.

    PubMed

    Zhang, De Lu; Hu, Chun Xiang; Li, Dun Hai; Liu, Yong Ding

    2013-08-15

    Aphanizomenon flos-aquae (A. flos-aquae) is a source of neurotoxins known as aphantoxins or paralytic shellfish poisons (PSPs) that present a major threat to the environment and to human health. Generally, altered neurological function is reflected in behavior. Although the molecular mechanism of action of PSPs is well known, its neurobehavioral effects on adult zebrafish and its relationship with altered neurological functions are poorly understood. Aphantoxins purified from a natural isolate of A. flos-aquae DC-1 were analyzed by HPLC. The major analogs found in the toxins were the gonyautoxins 1 and 5 (GTX1 and GTX5; 34.04% and 21.28%, respectively) and the neosaxitoxin (neoSTX, 12.77%). Zebrafish (Danio rerio) were intraperitoneally injected with 5.3 and 7.61 μg STXeq/kg (low and high dose, respectively) of A. flos-aquae DC-1 aphantoxins. The swimming activity was investigated by observation combined with video at 6 timepoints from 1 to 24 h post-exposure. Both aphantoxin doses were associated with delayed touch responses, reduced head-tail locomotory abilities, inflexible turning of head, and a tailward-shifted center of gravity. The normal S-pattern (or undulating) locomotor trajectory was replaced by a mechanical motor pattern of swinging the head after wagging the tail. Finally, these fish principally distributed at the top and/or bottom water of the aquarium, and showed a clear polarized distribution pattern at 12 h post-exposure. Further analysis of neurological function demonstrated that both aphantoxin doses inhibited brain acetylcholinesterase activity. All these changes were dose- and time-dependent. These results demonstrate that aphantoxins can alter locomotor capacity, touch responses and distribution patterns by damaging the cholinergic system of zebrafish, and suggest that zebrafish locomotor behavior and acetylcholinesterase can be used as indicators for investigating aphantoxins and blooms in nature. PMID:23792258

  11. Analysis of axon tract formation in the zebrafish brain: the role of territories of gene expression and their boundaries.

    PubMed

    Wilson, S W; Brennan, C; Macdonald, R; Brand, M; Holder, N

    1997-11-01

    Mutant analysis in the zebrafish is revealing the genes that are expressed in the early neuroepithelium and that regulate factors responsible for the guidance of commissural axons. We review work on the developing zebrafish brain illustrating the way in which territories of regulatory gene expression influence the formation and positioning of axon pathways. PMID:9321679

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

    PubMed Central

    Kovrižnych, Jevgenij A.; Zeljenková, Dagmar; Rollerová, Eva; Szabová, Elena

    2014-01-01

    Nickel oxide in the form of nanoparticles (NiO NPs) is extensively used in different industrial branches. In a test on adult zebrafish, the acute toxicity of NiO NPs was shown to be low, however longlasting contact with this compound can lead to its accumulation in the tissues and to increased toxicity. In this work we determined the 30-day toxicity of NiO NPs using a static test for zebrafish Danio rerio. We found the 30-day LC50 value to be 45.0 mg/L, LC100 (minimum concentration causing 100% mortality) was 100.0 mg/L, and LC0 (maximum concentration causing no mortality) was 6.25 mg/L for adult individuals of zebrafish. Considering a broad use of Ni in the industry, NiO NPs chronic toxicity may have a negative impact on the population of aquatic organisms and on food web dynamics in aquatic systems. PMID:26038672

  13. Phylostratigraphic Profiles in Zebrafish Uncover Chordate Origins of the Vertebrate Brain

    PubMed Central

    Šestak, Martin Sebastijan; Domazet-Lošo, Tomislav

    2015-01-01

    An elaborated tripartite brain is considered one of the important innovations of vertebrates. Other extant chordate groups have a more basic brain organization. For instance, cephalochordates possess a relatively simple brain possibly homologous to the vertebrate forebrain and hindbrain, whereas tunicates display the tripartite organization, but without the specialized brain centers. The difference in anatomical complexity is even more pronounced if one compares chordates with other deuterostomes that have only a diffuse nerve net or alternatively a rather simple central nervous system. To gain a new perspective on the evolutionary roots of the complex vertebrate brain, we made here a phylostratigraphic analysis of gene expression patterns in the developing zebrafish (Danio rerio). The recovered adaptive landscape revealed three important periods in the evolutionary history of the zebrafish brain. The oldest period corresponds to preadaptive events in the first metazoans and the emergence of the nervous system at the metazoan–eumetazoan transition. The origin of chordates marks the next phase, where we found the overall strongest adaptive imprint in almost all analyzed brain regions. This finding supports the idea that the vertebrate brain evolved independently of the brains within the protostome lineage. Finally, at the origin of vertebrates we detected a pronounced signal coming from the dorsal telencephalon, in agreement with classical theories that consider this part of the cerebrum a genuine vertebrate innovation. Taken together, these results reveal a stepwise adaptive history of the vertebrate brain where most of its extant organization was already present in the chordate ancestor. PMID:25415965

  14. Phylostratigraphic profiles in zebrafish uncover chordate origins of the vertebrate brain.

    PubMed

    Šestak, Martin Sebastijan; Domazet-Lošo, Tomislav

    2015-02-01

    An elaborated tripartite brain is considered one of the important innovations of vertebrates. Other extant chordate groups have a more basic brain organization. For instance, cephalochordates possess a relatively simple brain possibly homologous to the vertebrate forebrain and hindbrain, whereas tunicates display the tripartite organization, but without the specialized brain centers. The difference in anatomical complexity is even more pronounced if one compares chordates with other deuterostomes that have only a diffuse nerve net or alternatively a rather simple central nervous system. To gain a new perspective on the evolutionary roots of the complex vertebrate brain, we made here a phylostratigraphic analysis of gene expression patterns in the developing zebrafish (Danio rerio). The recovered adaptive landscape revealed three important periods in the evolutionary history of the zebrafish brain. The oldest period corresponds to preadaptive events in the first metazoans and the emergence of the nervous system at the metazoan-eumetazoan transition. The origin of chordates marks the next phase, where we found the overall strongest adaptive imprint in almost all analyzed brain regions. This finding supports the idea that the vertebrate brain evolved independently of the brains within the protostome lineage. Finally, at the origin of vertebrates we detected a pronounced signal coming from the dorsal telencephalon, in agreement with classical theories that consider this part of the cerebrum a genuine vertebrate innovation. Taken together, these results reveal a stepwise adaptive history of the vertebrate brain where most of its extant organization was already present in the chordate ancestor. PMID:25415965

  15. A Statistically Enhanced Spectral Counting Approach to TCDD Cardiac Toxicity in the Adult Zebrafish Heart

    PubMed Central

    Zhang, Jiang; Lanham, Kevin A; Heideman, Warren; Peterson, Richard E.; Li, Lingjun

    2013-01-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a persistent environmental pollutant and teratogen that produces cardiac toxicity in the developing zebrafish. Here we adopted a label free quantitative proteomic approach based on normalized spectral abundance factor (NSAF) to investigate the disturbance of the cardiac proteome induced by TCDD in the adult zebrafish heart. The protein expression level changes between heart samples from TCDD treated and control zebrafish were systematically evaluated by a large scale MudPIT analysis which incorporated triplicate analyses for both control and TCDD exposed heart proteomic samples to overcome the data dependant variation in shotgun proteomic experiments and obtain a statistically significant protein dataset with improved quantification confidence. A total of 519 and 443 proteins were identified in hearts collected from control and TCDD treated zebrafish, respectively, among which 106 proteins showed statistically significant expression changes. After correcting for the experimental variation between replicate analyses by statistical evaluation, 55 proteins exhibited NSAF ratio above 2 and 43 proteins displayed NSAF ratio smaller than 0.5, with statistical significance by t-test (p < 0.05). The proteins identified as altered by TCDD encompass a wide range of biological functions including calcium handling, myocardium cell architecture, energy production and metabolism, mitochondrial homeostasis, and stress response. Collectively, our results indicate that TCDD exposure alters the adult zebrafish heart in a way that could result in cardiac hypertrophy and heart failure, and suggests a potential mechanism for the diastolic dysfunction observed in TCDD exposed embryos. PMID:23682714

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

    PubMed

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

    2015-12-01

    Feeding Artemia nauplii as the main nutrition source for zebrafish is a common practice for many research facilities. Culturing live feed can be time-consuming and requires additional equipment to be purchased, maintained, and cleaned. Nonhatching decapsulated Artemia cysts (decaps) are a commercially available product that can be fed directly to fish. Several other ornamental fish species have been successfully cultured using decaps. Replacing Artemia nauplii with decaps could reduce the overall time and costs associated with the operation of a zebrafish facility. The objective of this study was to determine if decaps could be a suitable replacement to Artemia nauplii in juvenile and adult zebrafish culture. Wild-type zebrafish were fed one of three dietary treatments: decaps only, nauplii only, or a standard consisting of nauplii plus a commercially prepared pellet food. Survival, growth (length and weight), and embryo production were analyzed between the treatments. Fish receiving the decap diet demonstrated a significantly higher growth and embryo production when compared to the fish receiving the nauplii-only diet. When comparing the decap fish to the standard fish, no significant difference was found in mean survival, mean weight at 90 days postfertilization, or mean embryo production. It was determined that nonhatching decapsulated Artemia cysts can be used as a suitable replacement to Artemia nauplii in juvenile and adult zebrafish culture. PMID:25495227

  17. Cardiac morphology and blood pressure in the adult zebrafish.

    PubMed

    Hu, N; Yost, H J; Clark, E B

    2001-09-01

    Zebrafish has become a popular model for the study of cardiovascular development. We performed morphologic analysis on 3 months postfertilization zebrafish hearts (n > or = 20) with scanning electron microscopy, hematoxylin and eosin staining and Masson's trichrome staining, and morphometric analysis on cell organelles with transmission electron photomicrographs. We measured atrial, ventricular, ventral, and dorsal aortic blood pressures (n > or = 5) with a servonull system. The atrioventricular orifice was positioned on the dorsomedial side of the anterior ventricle, surmounted by the single-chambered atrium. The atrioventricular valve was free of tension apparati but supported by papillary bands to prevent retrograde flow. The ventricle was spanned with fine trabeculae perpendicular to the compact layer and perforated with a subepicardial network of coronary arteries, which originated from the efferent branchial arteries by means of the main coronary vessel. Ventricular myocytes were larger than those in the atrium (P < 0.05) with abundant mitochondria close to the sarcolemmal. Sarcoplasmic reticulum was sparse in zebrafish ventricle. Bulbus arteriosus was located anterior to the ventricle, and functioned as an elastic reservoir to absorb the rapid rise of pressure during ventricular contraction. The dense matrix of collagen interspersed across the entire bulbus arteriosus exemplified the characteristics of vasculature smooth muscle. There were pressure gradients from atrium to ventricle, and from ventral to dorsal aorta, indicating that the valves and the branchial arteries, respectively, were points of resistance to blood flow. These data serve as a framework for structure-function investigations of the zebrafish cardiovascular system. PMID:11505366

  18. Developmental Apoptosis Mediates Entry and Positioning of Microglia in the Zebrafish Brain.

    PubMed

    Casano, Alessandra Maria; Albert, Marvin; Peri, Francesca

    2016-07-26

    In the brain, neurons that fail to assemble into functional circuits are eliminated. Their clearance depends on microglia, immune cells that colonize the CNS during embryogenesis. Despite the importance of these cells in development and disease, the mechanisms that target and position microglia within the brain are unclear. Here we show that, in zebrafish, attraction of microglia into the brain exploits differences in developmental neuronal apoptosis and that these provide a mechanism for microglial distribution. Reducing neuronal cell death results in fewer microglia, whereas increased apoptosis enhances brain colonization, resulting in more microglia at later stages. Interestingly, attraction into the brain depends on nucleotide signaling, the same signaling system used to guide microglia toward brain injuries. Finally, this work uncovers a cell-non-autonomous role for developmental apoptosis. Classically considered a wasteful process, programmed cell death is exploited here to configure the immune-neuronal interface of the brain. PMID:27425604

  19. New Tools for the Identification of Developmentally Regulated Enhancer Regions in Embryonic and Adult Zebrafish

    PubMed Central

    Krauss, Jana; Koehler, Carla; Boden, Cindy; Harris, Matthew P.

    2013-01-01

    Abstract We have conducted a screen to identify developmentally regulated enhancers that drive tissue-specific Gal4 expression in zebrafish. We obtained 63 stable transgenic lines with expression patterns in embryonic or adult zebrafish. The use of a newly identified minimal promoter from the medaka edar locus resulted in a relatively unbiased set of expression patterns representing many tissue types derived from all germ layers. Subsequent detailed characterization of selected lines showed strong and reproducible Gal4-driven GFP expression in diverse tissues, including neurons from the central and peripheral nervous systems, pigment cells, erythrocytes, and peridermal cells. By screening adults for GFP expression, we also isolated lines expressed in tissues of the adult zebrafish, including scales, fin rays, and joints. The new and efficient minimal promoter and large number of transactivating driver-lines we identified will provide the zebrafish community with a useful resource for further enhancer trap screening, as well as precise investigation of tissue-specific processes in vivo. PMID:23461416

  20. Distinct effects of inflammation on preconditioning and regeneration of the adult zebrafish heart

    PubMed Central

    de Preux Charles, Anne-Sophie; Bise, Thomas; Baier, Felix; Marro, Jan; Jaźwińska, Anna

    2016-01-01

    The adult heart is able to activate cardioprotective programmes and modifies its architecture in response to physiological or pathological changes. While mammalian cardiac remodelling often involves hypertrophic expansion, the adult zebrafish heart exploits hyperplastic growth. This capacity depends on the responsiveness of zebrafish cardiomyocytes to mitogenic signals throughout their entire life. Here, we have examined the role of inflammation on the stimulation of cell cycle activity in the context of heart preconditioning and regeneration. We used thoracotomy as a cardiac preconditioning model and cryoinjury as a model of cardiac infarction in the adult zebrafish. First, we performed a spatio-temporal characterization of leucocytes and cycling cardiac cells after thoracotomy. This analysis revealed a concomitance between the infiltration of inflammatory cells and the stimulation of the mitotic activity. However, decreasing the immune response using clodronate liposome injection, PLX3397 treatment or anti-inflammatory drugs surprisingly had no effect on the re-entry of cardiac cells into the cell cycle. In contrast, reducing inflammation using the same strategies after cryoinjury strongly impaired cardiac cell mitotic activity and the regenerative process. Taken together, our results show that, while the immune response is not necessary to induce cell-cycle activity in intact preconditioned hearts, inflammation is required for the regeneration of injured hearts in zebrafish. PMID:27440424

  1. Identification of Wnt Genes Expressed in Neural Progenitor Zones during Zebrafish Brain Development.

    PubMed

    Duncan, Robert N; Panahi, Samin; Piotrowski, Tatjana; Dorsky, Richard I

    2015-01-01

    Wnt signaling regulates multiple aspects of vertebrate central nervous system (CNS) development, including neurogenesis. However, vertebrate genomes can contain up to 25 Wnt genes, the functions of which are poorly characterized partly due to redundancy in their expression. To identify candidate Wnt genes as candidate mediators of pathway activity in specific brain progenitor zones, we have performed a comprehensive expression analysis at three different stages during zebrafish development. Antisense RNA probes for 21 Wnt genes were generated from existing and newly synthesized cDNA clones and used for in situ hybridization on whole embryos and dissected brains. As in other species, we found that Wnt expression patterns in the embryonic zebrafish CNS are complex and often redundant. We observed that progenitor zones in the telencephalon, dorsal diencephalon, hypothalamus, midbrain, midbrain-hindbrain boundary, cerebellum and retina all express multiple Wnt genes. Our data identify 12 specific ligands that can now be tested using loss-of-function approaches. PMID:26713625

  2. Identification of Wnt Genes Expressed in Neural Progenitor Zones during Zebrafish Brain Development

    PubMed Central

    Piotrowski, Tatjana; Dorsky, Richard I.

    2015-01-01

    Wnt signaling regulates multiple aspects of vertebrate central nervous system (CNS) development, including neurogenesis. However, vertebrate genomes can contain up to 25 Wnt genes, the functions of which are poorly characterized partly due to redundancy in their expression. To identify candidate Wnt genes as candidate mediators of pathway activity in specific brain progenitor zones, we have performed a comprehensive expression analysis at three different stages during zebrafish development. Antisense RNA probes for 21 Wnt genes were generated from existing and newly synthesized cDNA clones and used for in situ hybridization on whole embryos and dissected brains. As in other species, we found that Wnt expression patterns in the embryonic zebrafish CNS are complex and often redundant. We observed that progenitor zones in the telencephalon, dorsal diencephalon, hypothalamus, midbrain, midbrain-hindbrain boundary, cerebellum and retina all express multiple Wnt genes. Our data identify 12 specific ligands that can now be tested using loss-of-function approaches. PMID:26713625

  3. Patterns of olfactory bulb neurogenesis in the adult zebrafish are altered following reversible deafferentation.

    PubMed

    Trimpe, Darcy M; Byrd-Jacobs, Christine A

    2016-09-01

    Adult brain plasticity can be investigated using reversible methods that remove afferent innervation but allow return of sensory input. Repeated intranasal irrigation with Triton X-100 in adult zebrafish diminishes innervation to the olfactory bulb, resulting in a number of alterations in bulb structure and function, and cessation of the treatment allows for reinnervation and recovery. Using bromodeoxyuridine, Hu, and caspase-3 immunoreactivity we examined cell proliferation, differentiation, migration, and survival under conditions of acute and chronic deafferentation and reafferentation. Cell proliferation within the olfactory bulb was not influenced by acute or chronic deafferentation or reafferentation, but cell fate (including differentiation, migration, and/or survival of newly formed cells) was affected. We found that chronic deafferentation caused a bilateral increase in the number of newly formed cells that migrated into the bulb, although the amount of cell death of these new cells was significantly increased compared to untreated fish. Reafferentation also increased the number of newly formed cells migrating into both bulbs, suggesting that the deafferentation effect on cell fate was maintained. Reafferentation resulted in a decrease in newly formed cells that became neurons and, although death of newly formed cells was not altered from control levels, survival was reduced in relation to that seen in chronically deafferented fish. The potential effect of age on cell genesis was also examined. While the amount of cell migration into the olfactory bulbs was not affected by fish age, more of the newly formed cells became neurons in older fish. Younger fish displayed more cell death under conditions of chronic deafferentation. In sum, our results show that reversible deafferentation affects several aspects of cell fate, including cell differentiation, migration, and survival, and age of the fish influences the response to deafferentation. PMID:27343831

  4. Developmental lead acetate exposure induces embryonic toxicity and memory deficit in adult zebrafish.

    PubMed

    Chen, Jiangfei; Chen, Yuanhong; Liu, Wei; Bai, Chenglian; Liu, Xuexia; Liu, Kai; Li, Rong; Zhu, Jian-Hong; Huang, Changjiang

    2012-01-01

    Lead is a persistent metal and commonly present in our living environment. The present study was aimed to investigate lead-induced embryonic toxicity, behavioral responses, and adult learning/memory deficit in zebrafish. Lead acetate (PbAc) induced malformations such as uninflated swim bladder, bent spine and yolk-sac edema with an EC₅₀ of 0.29 mg/L at 120 h post fertilization (hpf). Spontaneous movement as characterized by tail bend frequency was significantly altered in zebrafish embryos following exposure to PbAc. Behavior assessment demonstrated that lead exposure changed behavioral responses in zebrafish larvae, as hyperactivity was detected within the first minute of light-to-dark transition in the fish exposed to PbAc from 6 to 96 hpf, and a different dose-dependent change was found in swimming speeds in the dark and in the light at 120 hpf following lead exposure. Learning/memory task assay showed that embryos exposed to PbAc from 6 to 120 hpf developed learning/memory deficit at adulthood as exhibited by a significant decrease in accuracy rate to find the food and a significant increase in finding time. Overall, our results suggested that low dose of developmental lead exposure resulted in embryonic toxicity, behavioral alteration, and adult learning/memory deficit in zebrafish. PMID:22975620

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

    PubMed Central

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

    2014-01-01

    Exposure to moderate levels of ethanol during brain development has a number of effects on social behavior but the molecular mechanisms that mediate this are not well understood. Gaining a better understanding of these factors may help to develop therapeutic interventions in the future. Zebrafish offer a potentially useful model in this regard. Here, we introduce a zebrafish model of moderate prenatal ethanol exposure. Embryos were exposed to 20 mM ethanol for seven days (48hpfs–9dpf) and tested as adults for individual social behavior and shoaling. We also tested their basal anxiety with the novel tank diving test. We found that the ethanol-exposed fish displayed reductions in social approach and shoaling, and an increase in anxiety in the novel tank test. These behavioral differences corresponded to differences in hrt1aa, slc6a4 and oxtr expression. Namely, acute ethanol caused a spike in oxtr and ht1aa mRNA expression, which was followed by down-regulation at 7dpf, and an up-regulation in slc6a4 at 72hpf. This study confirms the utility of zebrafish as a model system for studying the molecular basis of developmental ethanol exposure. Furthermore, it proposes a putative developmental mechanism characterized by ethanol-induced OT inhibition leading to suppression of 5-HT and up-regulation of 5-HT1A, which leads, in turn, to possible homeostatic up-regulation of 5-HTT at 72hpf and subsequent imbalance of the 5-HT system. PMID:24690524

  6. Improvement of surface ECG recording in adult zebrafish reveals that the value of this model exceeds our expectation.

    PubMed

    Liu, Chi Chi; Li, Li; Lam, Yun Wah; Siu, Chung Wah; Cheng, Shuk Han

    2016-01-01

    The adult zebrafish has been used to model the electrocardiogram (ECG) for human cardiovascular studies. Nonetheless huge variations are observed among studies probably because of the lack of a reliable and reproducible recording method. In our study, an adult zebrafish surface ECG recording technique was improved using a multi-electrode method and by pre-opening the pericardial sac. A convenient ECG data analysis method without wavelet transform was also established. Intraperitoneal injection of KCl in zebrafish induced an arrhythmia similar to that of humans, and the arrhythmia was partially rescued by calcium gluconate. Amputation and cryoinjury of the zebrafish heart induced ST segment depression and affected QRS duration after injury. Only cryoinjury decelerated the heart rate. Different changes were also observed in the QT interval during heart regeneration in these two injury models. We also characterized the electrocardiophysiology of breakdance zebrafish mutant with a prolonged QT interval, that has not been well described in previous studies. Our study provided a reliable and reproducible means to record zebrafish ECG and analyse data. The detailed characterization of the cardiac electrophysiology of zebrafish and its mutant revealed that the potential of the zebrafish in modeling the human cardiovascular system exceeds expectations. PMID:27125643

  7. Improvement of surface ECG recording in adult zebrafish reveals that the value of this model exceeds our expectation

    PubMed Central

    Liu, Chi Chi; Li, Li; Lam, Yun Wah; Siu, Chung Wah; Cheng, Shuk Han

    2016-01-01

    The adult zebrafish has been used to model the electrocardiogram (ECG) for human cardiovascular studies. Nonetheless huge variations are observed among studies probably because of the lack of a reliable and reproducible recording method. In our study, an adult zebrafish surface ECG recording technique was improved using a multi-electrode method and by pre-opening the pericardial sac. A convenient ECG data analysis method without wavelet transform was also established. Intraperitoneal injection of KCl in zebrafish induced an arrhythmia similar to that of humans, and the arrhythmia was partially rescued by calcium gluconate. Amputation and cryoinjury of the zebrafish heart induced ST segment depression and affected QRS duration after injury. Only cryoinjury decelerated the heart rate. Different changes were also observed in the QT interval during heart regeneration in these two injury models. We also characterized the electrocardiophysiology of breakdance zebrafish mutant with a prolonged QT interval, that has not been well described in previous studies. Our study provided a reliable and reproducible means to record zebrafish ECG and analyse data. The detailed characterization of the cardiac electrophysiology of zebrafish and its mutant revealed that the potential of the zebrafish in modeling the human cardiovascular system exceeds expectations. PMID:27125643

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

    PubMed Central

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

    2013-01-01

    The zebrafish has become a standard model system for stem cell and tissue regeneration research, based on powerful genetics, high tissue regenerative capacity and low maintenance costs. Yet, these studies can be challenged by current limitations of tissue visualization techniques in adult animals. Here we describe new imaging methodology and present several ubiquitous and tissue-specific luciferase-based transgenic lines, which we have termed zebraflash, that facilitate the assessment of regeneration and engraftment in freely moving adult zebrafish. We show that luciferase-based live imaging reliably estimates muscle quantity in an internal organ, the heart, and can longitudinally follow cardiac regeneration in individual animals after major injury. Furthermore, luciferase-based detection enables visualization and quantification of engraftment in live recipients of transplanted hematopoietic stem cell progeny, with advantages in sensitivity and gross spatial resolution over fluorescence detection. Our findings present a versatile resource for monitoring and dissecting vertebrate stem cell and regeneration biology. PMID:24198277

  9. Fast functional imaging of multiple brain regions in intact zebrafish larvae using selective plane illumination microscopy.

    PubMed

    Panier, Thomas; Romano, Sebastián A; Olive, Raphaël; Pietri, Thomas; Sumbre, Germán; Candelier, Raphaël; Debrégeas, Georges

    2013-01-01

    The optical transparency and the small dimensions of zebrafish at the larval stage make it a vertebrate model of choice for brain-wide in-vivo functional imaging. However, current point-scanning imaging techniques, such as two-photon or confocal microscopy, impose a strong limit on acquisition speed which in turn sets the number of neurons that can be simultaneously recorded. At 5 Hz, this number is of the order of one thousand, i.e., approximately 1-2% of the brain. Here we demonstrate that this limitation can be greatly overcome by using Selective-plane Illumination Microscopy (SPIM). Zebrafish larvae expressing the genetically encoded calcium indicator GCaMP3 were illuminated with a scanned laser sheet and imaged with a camera whose optical axis was oriented orthogonally to the illumination plane. This optical sectioning approach was shown to permit functional imaging of a very large fraction of the brain volume of 5-9-day-old larvae with single- or near single-cell resolution. The spontaneous activity of up to 5,000 neurons was recorded at 20 Hz for 20-60 min. By rapidly scanning the specimen in the axial direction, the activity of 25,000 individual neurons from 5 different z-planes (approximately 30% of the entire brain) could be simultaneously monitored at 4 Hz. Compared to point-scanning techniques, this imaging strategy thus yields a ≃20-fold increase in data throughput (number of recorded neurons times acquisition rate) without compromising the signal-to-noise ratio (SNR). The extended field of view offered by the SPIM method allowed us to directly identify large scale ensembles of neurons, spanning several brain regions, that displayed correlated activity and were thus likely to participate in common neural processes. The benefits and limitations of SPIM for functional imaging in zebrafish as well as future developments are briefly discussed. PMID:23576959

  10. Amigo Adhesion Protein Regulates Development of Neural Circuits in Zebrafish Brain*

    PubMed Central

    Zhao, Xiang; Kuja-Panula, Juha; Sundvik, Maria; Chen, Yu-Chia; Aho, Vilma; Peltola, Marjaana A.; Porkka-Heiskanen, Tarja; Panula, Pertti; Rauvala, Heikki

    2014-01-01

    The Amigo protein family consists of three transmembrane proteins characterized by six leucine-rich repeat domains and one immunoglobulin-like domain in their extracellular moieties. Previous in vitro studies have suggested a role as homophilic adhesion molecules in brain neurons, but the in vivo functions remain unknown. Here we have cloned all three zebrafish amigos and show that amigo1 is the predominant family member expressed during nervous system development in zebrafish. Knockdown of amigo1 expression using morpholino oligonucleotides impairs the formation of fasciculated tracts in early fiber scaffolds of brain. A similar defect in fiber tract development is caused by mRNA-mediated expression of the Amigo1 ectodomain that inhibits adhesion mediated by the full-length protein. Analysis of differentiated neural circuits reveals defects in the catecholaminergic system. At the behavioral level, the disturbed formation of neural circuitry is reflected in enhanced locomotor activity and in the inability of the larvae to perform normal escape responses. We suggest that Amigo1 is essential for the development of neural circuits of zebrafish, where its mechanism involves homophilic interactions within the developing fiber tracts and regulation of the Kv2.1 potassium channel to form functional neural circuitry that controls locomotion. PMID:24904058

  11. Delayed effects of developmental exposure to low levels of the aryl hydrocarbon receptor agonist 3,3',4,4',5-pentachlorobiphenyl (PCB126) on adult zebrafish behavior.

    PubMed

    Glazer, Lilah; Hahn, Mark E; Aluru, Neelakanteswar

    2016-01-01

    Polychlorinated biphenyls (PCBs) are ubiquitous environmental contaminants. The most toxic PCBs are the non-ortho-substituted ("dioxin-like") congeners that act through the aryl hydrocarbon receptor (AHR) pathway. In humans, perinatal exposure to dioxin-like PCBs is associated with neurodevelopmental toxicity in children. Yet, the full potential for later-life neurobehavioral effects that result from early-life low level exposure to dioxin-like PCBs is not well understood. The objective of this study was to determine the effects of developmental exposure to low levels of dioxin-like PCBs on early- and later-life behavioral phenotypes using zebrafish as a model system. We exposed zebrafish embryos to either vehicle (DMSO) or low concentrations of PCB126 (0.3, 0.6, 1.2nM) for 20h (4-24h post fertilization), and then reared them to adulthood in clean water. Locomotor activity was tested at two larval stages (7 and 14 days post fertilization). Adult fish were tested for anxiety-related behavior using the novel tank and shoaling assays. Adult behavioral assays were repeated several times on the same group of fish and effects on intra- and inter-trial habituation were determined. While there was no effect of PCB126 on larval locomotor activity in response to changes in light conditions, developmental exposure to PCB126 resulted in impaired short- and long-term habituation to a novel environment in adult zebrafish. Cyp1a induction was measured as an indicator for AHR activation. Despite high induction at early stages, cyp1a expression was not induced in the brains of developmentally exposed adult fish that showed altered behavior, suggesting that AHR was not activated at this stage. Our results demonstrate the effectiveness of the zebrafish model in detecting subtle and delayed behavioral effects resulting from developmental exposure to an environmental contaminant. PMID:26616910

  12. Differential requirement for irf8 in formation of embryonic and adult macrophages in zebrafish

    DOE PAGESBeta

    Shiau, Celia E.; Kaufman, Zoe; Meireles, Ana M.; Talbot, William S.

    2015-01-23

    Interferon regulatory factor 8 (Irf8) is critical for mammalian macrophage development and innate immunity, but its role in teleost myelopoiesis remains incompletely understood. Specifically, genetic tools to analyze the role of irf8 in zebrafish macrophage development at larval and adult stages are lacking. In this study, we generated irf8 null mutants in zebrafish using TALEN-mediated targeting. Our analysis defines different requirements for irf8 at different stages. irf8 is required for formation of all macrophages during primitive and transient definitive hematopoiesis, but not during adult-phase definitive hematopoiesis starting at 5-6 days postfertilization. At early stages, irf8 mutants have excess neutrophils andmore » excess cell death in pu.1-expressing myeloid cells. Macrophage fates were recovered in irf8 mutants after wildtype irf8 expression in neutrophil and macrophage lineages, suggesting that irf8 regulates macrophage specification and survival. In juvenile irf8 mutant fish, mature macrophages are present, but at numbers significantly reduced compared to wildtype, indicating an ongoing requirement for irf8 after embryogenesis. As development progresses, tissue macrophages become apparent in zebrafish irf8 mutants, with the possible exception of microglia. Our study defines distinct requirement for irf8 in myelopoiesis before and after transition to the adult hematopoietic system.« less

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

    PubMed Central

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

    2015-01-01

    Environmental enrichment provides laboratory-housed species the opportunity to express natural behavior and exert control over their home environment, thereby minimizing stress. We sought to determine whether providing an artificial plant in the holding tank as enrichment influenced anxiety-like behaviors and place-preference choice in adult zebrafish. Fish were housed singly or in social groups of 5 for 3 wk in 1 of 4 experimental housing environments: single-housed enriched (n = 30), single-housed barren (n = 30), group-housed enriched (n = 30), and group-housed barren (n = 30). On week 4, individual fish were selected randomly from each of the experimental housing environments and tested by using novel-tank, light–dark, and place-preference tests. Housing fish singly in a barren environment increased anxiety-like behaviors in the novel-tank and light–dark behavioral tests. Single-housed zebrafish in barren tanks as well as zebrafish group-housed with conspecifics, both with and without plant enrichment, spent more time associating with conspecifics than with the artificial plant enrichment device during the place-preference test. Single-housed fish maintained in enriched tanks displayed no preference between a compartment with conspecifics or an artificial plant. Our results suggest the addition of an artificial plant as enrichment may benefit single-housed zebrafish when social housing is not possible. PMID:26045453

  14. Development of the histaminergic neurons and expression of histidine decarboxylase mRNA in the zebrafish brain in the absence of all peripheral histaminergic systems.

    PubMed

    Eriksson, K S; Peitsaro, N; Karlstedt, K; Kaslin, J; Panula, P

    1998-12-01

    The histamine-storing neural system in adult and developing zebrafish (Danio rerio) was studied with immunocytochemical and chromatographical methods. Furthermore, the gene for histidine decarboxylase was partially cloned and its expression mapped with in situ hybridization. The histamine-storing neurons were only seen in the caudal hypothalamus, around the posterior recess of the diencephalic ventricle. Almost all parts of the brain, except the cerebellum, contained at least some histamine-immunoreactive fibres. The ascending projections had the rostral part of the dorsal telencephalon as a major target. Descending projections terminated in the torus semicircularis, central grey and inferior olive. A prominent innervation of the optic tectum, which has not been reported in other fish, was seen. The in situ hybridization gave a strong signal in cells with the same anatomical position as the histamine-immunoreactive neurons. The first histamine-immunoreactive neurons appeared in the ventral hypothalamus at about 85 h post-fertilization, and at 90 h, immunoreactive fibres terminated in the dorsal telencephalon. The embryonic histamine production described in mammals was lacking in this species. Both immunocytochemical and chromatographical studies indicated that histamine is absent in all other parts of the zebrafish body, and no specific hybridization was seen in any other part of the fish than the hypothalamus. The zebrafish could therefore be a very useful model for pharmacological in vivo studies of the histaminergic system of the brain, since the powerful peripheral actions of histamine should be lacking in this species. PMID:9875358

  15. Novel function of vitamin E in regulation of zebrafish (Danio rerio) brain lysophospholipids discovered using lipidomics

    PubMed Central

    Choi, Jaewoo; Leonard, Scott W.; Kasper, Katherine; McDougall, Melissa; Stevens, Jan F.; Tanguay, Robert L.; Traber, Maret G.

    2015-01-01

    We hypothesized that brains from vitamin E-deficient (E−) zebrafish (Danio rerio) would undergo increased lipid peroxidation because they contain highly polyunsaturated fatty acids, thus susceptible lipids could be identified. Brains from zebrafish fed for 9 months defined diets without (E−) or with (E+) added vitamin E (500 mg RRR-α-tocopheryl acetate per kilogram diet) were studied. Using an untargeted approach, 1-hexadecanoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine [DHA-PC 38:6, PC 16:0/22:6]was the lipid that showed the most significant and greatest fold-differences between groups. DHA-PC concentrations were approximately 1/3 lower in E− (4.3 ± 0.6 mg/g) compared with E+ brains (6.5 ± 0.9 mg/g, mean ± SEM, n = 10 per group, P = 0.04). Using lipidomics, 155 lipids in brain extracts were identified. Only four phospholipids (PLs) were different (P < 0.05) between groups; they were lower in E− brains and contained DHA with DHA-PC 38:6 at the highest abundances. Moreover, hydroxy-DHA-PC 38:6 was increased in E− brains (P = 0.0341) supporting the hypothesis of DHA peroxidation. More striking was the depletion in E− brains of nearly 60% of 19 different lysophospholipids (lysoPLs) (combined P = 0.0003), which are critical for membrane PL remodeling. Thus, E− brains contained fewer DHA-PLs, more hydroxy-DHA-PCs, and fewer lysoPLs, suggesting that lipid peroxidation depletes membrane DHA-PC and homeostatic mechanisms to repair the damage resulting in lysoPL depletion. PMID:25855633

  16. Novel function of vitamin E in regulation of zebrafish (Danio rerio) brain lysophospholipids discovered using lipidomics.

    PubMed

    Choi, Jaewoo; Leonard, Scott W; Kasper, Katherine; McDougall, Melissa; Stevens, Jan F; Tanguay, Robert L; Traber, Maret G

    2015-06-01

    We hypothesized that brains from vitamin E-deficient (E-) zebrafish (Danio rerio) would undergo increased lipid peroxidation because they contain highly polyunsaturated fatty acids, thus susceptible lipids could be identified. Brains from zebrafish fed for 9 months defined diets without (E-) or with (E+) added vitamin E (500 mg RRR-α-tocopheryl acetate per kilogram diet) were studied. Using an untargeted approach, 1-hexadecanoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine [DHA-PC 38:6, PC 16:0/22:6]was the lipid that showed the most significant and greatest fold-differences between groups. DHA-PC concentrations were approximately 1/3 lower in E- (4.3 ± 0.6 mg/g) compared with E+ brains (6.5 ± 0.9 mg/g, mean ± SEM, n = 10 per group, P = 0.04). Using lipidomics, 155 lipids in brain extracts were identified. Only four phospholipids (PLs) were different (P < 0.05) between groups; they were lower in E- brains and contained DHA with DHA-PC 38:6 at the highest abundances. Moreover, hydroxy-DHA-PC 38:6 was increased in E- brains (P = 0.0341) supporting the hypothesis of DHA peroxidation. More striking was the depletion in E- brains of nearly 60% of 19 different lysophospholipids (lysoPLs) (combined P = 0.0003), which are critical for membrane PL remodeling. Thus, E- brains contained fewer DHA-PLs, more hydroxy-DHA-PCs, and fewer lysoPLs, suggesting that lipid peroxidation depletes membrane DHA-PC and homeostatic mechanisms to repair the damage resulting in lysoPL depletion. PMID:25855633

  17. Brain zinc chelation by diethyldithiocarbamate increased the behavioral and mitochondrial damages in zebrafish subjected to hypoxia

    PubMed Central

    Braga, Marcos M.; Silva, Emerson S.; Moraes, Tarsila B.; Schirmbeck, Gabriel Henrique; Rico, Eduardo P.; Pinto, Charles B.; Rosemberg, Denis B.; Dutra-Filho, Carlos S.; Dias, Renato D.; Oliveira, Diogo L.; T. Rocha, João Batista; Souza, Diogo O.

    2016-01-01

    The increase in brain levels of chelatable zinc (Zn) in dysfunctions involving oxygen deprivation has stimulated the treatment with Zn chelators, such as diethyldithiocarbamate (DEDTC). However, DEDTC is a redox-active compound and it should be better evaluated during hypoxia. We use the hypoxia model in zebrafish to evaluate DEDTC effects. The exploratory behavior, chelatable Zn content, activities of mitochondrial dehydrogenases, reactive species levels (nitric oxide, superoxide anion, hydroxyl radical scavenger capacity) and cellular antioxidants (sulfhydryl, superoxide dismutase) of zebrafish brain were assessed after recovery, with or without 0.2 mM DEDTC. The increased brain levels of chelatable Zn induced by hypoxia were mitigated by DEDTC. However, the novel tank task indicated that DEDTC did further enhance the exploratory deficit caused by hypoxia. Furthermore, these behavioral impairments caused by DEDTC were more associated with a negative action on mitochondrial activity and brain oxidative balance. Thus, due to apparent pro-oxidant action of DEDTC, our data do not support its use for neuroprotection in neuropathologies involving oxygen deprivation. PMID:26854133

  18. Optical mapping of the electrical activity of isolated adult zebrafish hearts: acute effects of temperature

    PubMed Central

    Lin, Eric; Ribeiro, Amanda; Ding, Weiguang; Hove-Madsen, Leif; Sarunic, Marinko V.; Beg, Mirza Faisal

    2014-01-01

    The zebrafish (Danio rerio) has emerged as an important model for developmental cardiovascular (CV) biology; however, little is known about the cardiac function of the adult zebrafish enabling it to be used as a model of teleost CV biology. Here, we describe electrophysiological parameters, such as heart rate (HR), action potential duration (APD), and atrioventricular (AV) delay, in the zebrafish heart over a range of physiological temperatures (18–28°C). Hearts were isolated and incubated in a potentiometric dye, RH-237, enabling electrical activity assessment in several distinct regions of the heart simultaneously. Integration of a rapid thermoelectric cooling system facilitated the investigation of acute changes in temperature on critical electrophysiological parameters in the zebrafish heart. While intrinsic HR varied considerably between fish, the ex vivo preparation exhibited impressively stable HRs and sinus rhythm for more than 5 h, with a mean HR of 158 ± 9 bpm (means ± SE; n = 20) at 28°C. Atrial and ventricular APDs at 50% repolarization (APD50) were 33 ± 1 ms and 98 ± 2 ms, respectively. Excitation originated in the atrium, and there was an AV delay of 61 ± 3 ms prior to activation of the ventricle at 28°C. APD and AV delay varied between hearts beating at unique HRs; however, APD and AV delay did not appear to be statistically dependent on intrinsic basal HR, likely due to the innate beat-to-beat variability within each heart. As hearts were cooled to 18°C (by 1°C increments), HR decreased by ∼40%, and atrial and ventricular APD50 increased by a factor of ∼3 and 2, respectively. The increase in APD with cooling was disproportionate at different levels of repolarization, indicating unique temperature sensitivities for ion currents at different phases of the action potential. The effect of temperature was more apparent at lower levels of repolarization and, as a whole, the atrial APD was the cardiac parameter most affected by acute

  19. Hypoxia/Reoxygenation Cardiac Injury and Regeneration in Zebrafish Adult Heart

    PubMed Central

    Pompilio, Giulio; Verduci, Lorena; Colombo, Gualtiero I.; Milano, Giuseppina; Guerrini, Uliano; Squadroni, Lidia; Cotelli, Franco; Pozzoli, Ombretta; Capogrossi, Maurizio C.

    2013-01-01

    Aims the adult zebrafish heart regenerates spontaneously after injury and has been used to study the mechanisms of cardiac repair. However, no zebrafish model is available that mimics ischemic injury in mammalian heart. We developed and characterized zebrafish cardiac injury induced by hypoxia/reoxygenation (H/R) and the regeneration that followed it. Methods and Results adult zebrafish were kept either in hypoxic (H) or normoxic control (C) water for 15 min; thereafter fishes were returned to C water. Within 2–6 hours (h) after reoxygenation there was evidence of cardiac oxidative stress by dihydroethidium fluorescence and protein nitrosylation, as well as of inflammation. We used Tg(cmlc2:nucDsRed) transgenic zebrafish to identify myocardial cell nuclei. Cardiomyocyte apoptosis and necrosis were evidenced by TUNEL and Acridine Orange (AO) staining, respectively; 18 h after H/R, 9.9±2.6% of myocardial cell nuclei were TUNEL+ and 15.0±2.5% were AO+. At the 30-day (d) time point myocardial cell death was back to baseline (n = 3 at each time point). We evaluated cardiomyocyte proliferation by Phospho Histone H3 (pHH3) or Proliferating Cell Nuclear Antigen (PCNA) expression. Cardiomyocyte proliferation was apparent 18–24 h after H/R, it achieved its peak 3–7d later, and was back to baseline at 30d. 7d after H/R 17.4±2.3% of all cardiomyocytes were pHH3+ and 7.4±0.6% were PCNA+ (n = 3 at each time point). Cardiac function was assessed by 2D-echocardiography and Ventricular Diastolic and Systolic Areas were used to compute Fractional Area Change (FAC). FAC decreased from 29.3±2.0% in normoxia to 16.4±1.8% at 18 h after H/R; one month later ventricular function was back to baseline (n = 12 at each time point). Conclusions zebrafish exposed to H/R exhibit evidence of cardiac oxidative stress and inflammation, myocardial cell death and proliferation. The initial decrease in ventricular function is followed by full recovery. This model more closely

  20. The microcephaly gene aspm is involved in brain development in zebrafish

    SciTech Connect

    Kim, Hyun-Taek; Lee, Mi-Sun; Choi, Jung-Hwa; Jung, Ju-Yeon; Ahn, Dae-Gwon; Yeo, Sang-Yeob; Choi, Dong-Kug; Kim, Cheol-Hee

    2011-06-17

    Highlights: {yields} We identified a zebrafish aspm/mcph5 gene that is expressed in proliferating cells in the CNS during early development. {yields} Embryos injected with the aspm MO consistently showed a reduced head and eye size but were otherwise grossly normal, closely mimicking the known phenotypes of human microcephaly patients. {yields} Knock-down of aspm causes cell cycle arrest and apoptotic cell death during early development. -- Abstract: MCPH is a neurodevelopmental disorder characterized by a global reduction in cerebral cortical volume. Homozygous mutation of the MCPH5 gene, also known as ASPM, is the most common cause of the MCPH phenotype. To elucidate the roles of ASPM during embryonic development, the zebrafish aspm was identified, which is specifically expressed in proliferating cells in the CNS. Morpholino-mediated knock-down of aspm resulted in a significant reduction in head size. Furthermore, aspm-deficient embryos exhibited a mitotic arrest during early development. These findings suggest that the reduction in brain size in MCPH might be caused by lack of aspm function in the mitotic cell cycle and demonstrate that the zebrafish can provide a model system for congenital diseases of the human nervous system.

  1. Large-scale Scanning Transmission Electron Microscopy (Nanotomy) of Healthy and Injured Zebrafish Brain.

    PubMed

    Kuipers, Jeroen; Kalicharan, Ruby D; Wolters, Anouk H G; van Ham, Tjakko J; Giepmans, Ben N G

    2016-01-01

    Large-scale 2D electron microscopy (EM), or nanotomy, is the tissue-wide application of nanoscale resolution electron microscopy. Others and we previously applied large scale EM to human skin pancreatic islets, tissue culture and whole zebrafish larvae(1-7). Here we describe a universally applicable method for tissue-scale scanning EM for unbiased detection of sub-cellular and molecular features. Nanotomy was applied to investigate the healthy and a neurodegenerative zebrafish brain. Our method is based on standardized EM sample preparation protocols: Fixation with glutaraldehyde and osmium, followed by epoxy-resin embedding, ultrathin sectioning and mounting of ultrathin-sections on one-hole grids, followed by post staining with uranyl and lead. Large-scale 2D EM mosaic images are acquired using a scanning EM connected to an external large area scan generator using scanning transmission EM (STEM). Large scale EM images are typically ~ 5 - 50 G pixels in size, and best viewed using zoomable HTML files, which can be opened in any web browser, similar to online geographical HTML maps. This method can be applied to (human) tissue, cross sections of whole animals as well as tissue culture(1-5). Here, zebrafish brains were analyzed in a non-invasive neuronal ablation model. We visualize within a single dataset tissue, cellular and subcellular changes which can be quantified in various cell types including neurons and microglia, the brain's macrophages. In addition, nanotomy facilitates the correlation of EM with light microscopy (CLEM)(8) on the same tissue, as large surface areas previously imaged using fluorescent microscopy, can subsequently be subjected to large area EM, resulting in the nano-anatomy (nanotomy) of tissues. In all, nanotomy allows unbiased detection of features at EM level in a tissue-wide quantifiable manner. PMID:27285162

  2. Large-scale Scanning Transmission Electron Microscopy (Nanotomy) of Healthy and Injured Zebrafish Brain

    PubMed Central

    Kuipers, Jeroen; Kalicharan, Ruby D.; Wolters, Anouk H. G.

    2016-01-01

    Large-scale 2D electron microscopy (EM), or nanotomy, is the tissue-wide application of nanoscale resolution electron microscopy. Others and we previously applied large scale EM to human skin pancreatic islets, tissue culture and whole zebrafish larvae1-7. Here we describe a universally applicable method for tissue-scale scanning EM for unbiased detection of sub-cellular and molecular features. Nanotomy was applied to investigate the healthy and a neurodegenerative zebrafish brain. Our method is based on standardized EM sample preparation protocols: Fixation with glutaraldehyde and osmium, followed by epoxy-resin embedding, ultrathin sectioning and mounting of ultrathin-sections on one-hole grids, followed by post staining with uranyl and lead. Large-scale 2D EM mosaic images are acquired using a scanning EM connected to an external large area scan generator using scanning transmission EM (STEM). Large scale EM images are typically ~ 5 - 50 G pixels in size, and best viewed using zoomable HTML files, which can be opened in any web browser, similar to online geographical HTML maps. This method can be applied to (human) tissue, cross sections of whole animals as well as tissue culture1-5. Here, zebrafish brains were analyzed in a non-invasive neuronal ablation model. We visualize within a single dataset tissue, cellular and subcellular changes which can be quantified in various cell types including neurons and microglia, the brain's macrophages. In addition, nanotomy facilitates the correlation of EM with light microscopy (CLEM)8 on the same tissue, as large surface areas previously imaged using fluorescent microscopy, can subsequently be subjected to large area EM, resulting in the nano-anatomy (nanotomy) of tissues. In all, nanotomy allows unbiased detection of features at EM level in a tissue-wide quantifiable manner. PMID:27285162

  3. Exercise quantity-dependent muscle hypertrophy in adult zebrafish (Danio rerio).

    PubMed

    Hasumura, Takahiro; Meguro, Shinichi

    2016-07-01

    Exercise is very important for maintaining and increasing skeletal muscle mass, and is particularly important to prevent and care for sarcopenia and muscle disuse atrophy. However, the dose-response relationship between exercise quantity, duration/day, and overall duration and muscle mass is poorly understood. Therefore, we investigated the effect of exercise duration on skeletal muscle to reveal the relationship between exercise quantity and muscle hypertrophy in zebrafish forced to exercise. Adult male zebrafish were exercised 6 h/day for 4 weeks, 6 h/day for 2 weeks, or 3 h/day for 2 weeks. Flow velocity was adjusted to maximum velocity during continual swimming (initial 43 cm/s). High-speed consecutive photographs revealed that zebrafish mainly drove the caudal part. Additionally, X-ray micro computed tomography measurements indicated muscle hypertrophy of the mid-caudal half compared with the mid-cranial half part. The cross-sectional analysis of the mid-caudal half muscle revealed that skeletal muscle (red, white, or total) mass increased with increasing exercise quantity, whereas that of white muscle and total muscle increased only under the maximum exercise load condition of 6 h/day for 4 weeks. Additionally, the muscle fiver size distributions of exercised fish were larger than those from non-exercised fish. We revealed that exercise quantity, duration/day, and overall duration were correlated with skeletal muscle hypertrophy. The forced exercise model enabled us to investigate the relationship between exercise quantity and skeletal muscle mass. These results open up the possibility for further investigations on the effects of exercise on skeletal muscle in adult zebrafish. PMID:26951149

  4. Lipid peroxidation and antioxidant responses in zebrafish brain induced by Aphanizomenon flos-aquae DC-1 aphantoxins.

    PubMed

    Zhang, De Lu; Hu, Chun Xiang; Li, Dun Hai; Liu, Yong Ding

    2013-11-15

    Aphanizomenon flos-aquae is a cyanobacterium that is frequently encountered in eutrophic waters worldwide. It is source of neurotoxins known as aphantoxins or paralytic shellfish poisons (PSPs), which present a major threat to the environment and human health. The molecular mechanism of PSP action is known, however the in vivo effects of this neurotoxin on oxidative stress, lipid peroxidation and the antioxidant defense responses in zebrafish brain remain to be understood. Aphantoxins purified from a natural isolate of A. flos-aquae DC-1 were analyzed using high performance liquid chromatography. The major components of the toxins were gonyautoxins 1 and 5 (GTX1 and GTX5, 34.04% and 21.28%, respectively) and neosaxitoxin (neoSTX, 12.77%). Zebrafish (Danio rerio) were injected intraperitoneally with 7.73 μg/kg (low dose) and 11.13 μg/kg (high dose) of A. flos-aquae DC-1 aphantoxins. Oxidative stress, lipid peroxidation and antioxidant defense responses in the zebrafish brain were investigated at various timepoints at 1-24h post-exposure. Aphantoxin exposure was associated with significantly increased (>1-2 times) reactive oxygen species (ROS) and malondialdehyde (MDA) in zebrafish brain compared with the controls at 1-12h postexposure, suggestive of oxidative stress and lipid peroxidation. In contrast, reduced glutathione (GSH) levels in the zebrafish brain exposed to high or low doses of aphantoxins decreased by 44.88% and 41.33%, respectively, after 1-12h compared with the controls, suggesting that GSH participated in detoxification to ROS and MDA. Further analysis showed a significant increase in the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) compared with the controls, suggesting elimination of oxidative stress by the antioxidant response in zebrafish brain. All these changes were dose and time dependent. These results suggested that aphantoxins or PSPs increased ROS and MDA and decreased GSH in zebrafish brain

  5. Circadian rhythms in the pineal organ persist in zebrafish larvae that lack ventral brain

    PubMed Central

    2011-01-01

    Background The mammalian suprachiasmatic nucleus (SCN), located in the ventral hypothalamus, is a major regulator of circadian rhythms in mammals and birds. However, the role of the SCN in lower vertebrates remains poorly understood. Zebrafish cyclops (cyc) mutants lack ventral brain, including the region that gives rise to the SCN. We have used cyc embryos to define the function of the zebrafish SCN in regulating circadian rhythms in the developing pineal organ. The pineal organ is the major source of the circadian hormone melatonin, which regulates rhythms such as daily rest/activity cycles. Mammalian pineal rhythms are controlled almost exclusively by the SCN. In zebrafish and many other lower vertebrates, the pineal has an endogenous clock that is responsible in part for cyclic melatonin biosynthesis and gene expression. Results We find that pineal rhythms are present in cyc mutants despite the absence of an SCN. The arginine vasopressin-like protein (Avpl, formerly called Vasotocin) is a peptide hormone expressed in and around the SCN. We find avpl mRNA is absent in cyc mutants, supporting previous work suggesting the SCN is missing. In contrast, expression of the putative circadian clock genes, cryptochrome 1b (cry1b) and cryptochrome 3 (cry3), in the brain of the developing fish is unaltered. Expression of two pineal rhythmic genes, exo-rhodopsin (exorh) and serotonin-N-acetyltransferase (aanat2), involved in photoreception and melatonin synthesis, respectively, is also similar between cyc embryos and their wildtype (WT) siblings. The timing of the peaks and troughs of expression are the same, although the amplitude of expression is slightly decreased in the mutants. Cyclic gene expression persists for two days in cyc embryos transferred to constant light or constant dark, suggesting a circadian clock is driving the rhythms. However, the amplitude of rhythms in cyc mutants kept in constant conditions decreased more quickly than in their WT siblings

  6. Retinal Vasculature of Adult Zebrafish: In Vivo Imaging Using Confocal Scanning Laser Ophthalmoscopy

    PubMed Central

    Bell, Brent A.; Xie, Jing; Yuan, Alex; Kaul, Charles; Hollyfield, Joe G.; Anand-Apte, Bela

    2014-01-01

    Over the past 3 decades the zebrafish (Danio rerio) has become an important biomedical research species. As their use continues to grow additional techniques and tools will be required to keep pace with ongoing research using this species. In this paper we describe a novel method for in vivo imaging of the retinal vasculature in adult animals using a commercially available confocal scanning laser ophthalmoscope (SLO). With this instrumentation, we demonstrate the ability to distinguish diverse vascular phenotypes in different transgenic GFP lines. In addition this technology allows repeated visualization of the vasculature in individual zebrafish over time to document vascular leakage progression and recovery induced by intraocular delivery of proteins that induce vascular permeability. SLO of the retinal vasculature was found to be highly informative, providing images of high contrast and resolution that were capable of resolving individual vascular endothelial cells. Finally, the procedures required to acquire SLO images from zebrafish are non-invasive, simple to perform and can be achieved with low animal mortality, allowing repeated imaging of individual fish. PMID:25447564

  7. Using an Automated 3D-tracking System to Record Individual and Shoals of Adult Zebrafish

    PubMed Central

    Maaswinkel, Hans; Zhu, Liqun; Weng, Wei

    2013-01-01

    Like many aquatic animals, zebrafish (Danio rerio) moves in a 3D space. It is thus preferable to use a 3D recording system to study its behavior. The presented automatic video tracking system accomplishes this by using a mirror system and a calibration procedure that corrects for the considerable error introduced by the transition of light from water to air. With this system it is possible to record both single and groups of adult zebrafish. Before use, the system has to be calibrated. The system consists of three modules: Recording, Path Reconstruction, and Data Processing. The step-by-step protocols for calibration and using the three modules are presented. Depending on the experimental setup, the system can be used for testing neophobia, white aversion, social cohesion, motor impairments, novel object exploration etc. It is especially promising as a first-step tool to study the effects of drugs or mutations on basic behavioral patterns. The system provides information about vertical and horizontal distribution of the zebrafish, about the xyz-components of kinematic parameters (such as locomotion, velocity, acceleration, and turning angle) and it provides the data necessary to calculate parameters for social cohesions when testing shoals. PMID:24336189

  8. A novel protocol for the oral administration of test chemicals to adult zebrafish.

    PubMed

    Zang, Liqing; Morikane, Daizo; Shimada, Yasuhito; Tanaka, Toshio; Nishimura, Norihiro

    2011-12-01

    A novel protocol using gluten as a carrier material was developed to administer chemicals to adult zebrafish, per os (p.o.). To evaluate the capacity of gluten to retain chemicals, we prepared gluten granules containing eight types of chemicals with different Log P(ow) values and immersed them in water. Less than 5% of chemicals were eluted from gluten granules within 5 min, a standard feeding time for zebrafish. Although retention capability was dependent on the hydrophilicity and hydrophobicity of the chemicals, the gluten granules retained 62%-99% of the total amount of chemical, even after immersion in water for 60 min. Vital staining dyes, such as 4-Di-2-Asp and Nile red, administered p.o., were delivered into the gastrointestinal tract where they were digested and secreted. Subsequently, we conducted a pharmacokinetic study of oral administration of felbinac and confirmed that it was successfully delivered into the blood of zebrafish. This indicates that chemicals administered using gluten granules are satisfactorily absorbed from the digestive tract and delivered into the metabolic system. The absorption, distribution, and pharmacokinetics of chemicals given by oral administration were also compared with those of chemicals given by alternative administration routes such as intraperitoneal injection and exposure to chemical solution. PMID:22181663

  9. Assessment of fight outcome is needed to activate socially driven transcriptional changes in the zebrafish brain

    PubMed Central

    Oliveira, Rui F.; Simões, José M.; Teles, Magda C.; Oliveira, Catarina R.; Lopes, João S.

    2016-01-01

    Group living animals must be able to express different behavior profiles depending on their social status. Therefore, the same genotype may translate into different behavioral phenotypes through socially driven differential gene expression. However, how social information is translated into a neurogenomic response and what are the specific cues in a social interaction that signal a change in social status are questions that have remained unanswered. Here, we show for the first time, to our knowledge, that the switch between status-specific neurogenomic states relies on the assessment of fight outcome rather than just on self- or opponent-only assessment of fighting ability. For this purpose, we manipulated the perception of fight outcome in male zebrafish and measured its impact on the brain transcriptome using a zebrafish whole genome gene chip. Males fought either a real opponent, and a winner and a loser were identified, or their own image on a mirror, in which case, despite expressing aggressive behavior, males did not experience either a victory or a defeat. Massive changes in the brain transcriptome were observed in real opponent fighters, with losers displaying both a higher number of differentially expressed genes and of coexpressed gene modules than winners. In contrast, mirror fighters expressed a neurogenomic state similar to that of noninteracting fish. The genes that responded to fight outcome included immediate early genes and genes involved in neuroplasticity and epigenetic modifications. These results indicate that, even in cognitively simple organisms such as zebrafish, neurogenomic responses underlying changes in social status rely on mutual assessment of fighting ability. PMID:26787876

  10. Assessment of fight outcome is needed to activate socially driven transcriptional changes in the zebrafish brain.

    PubMed

    Oliveira, Rui F; Simões, José M; Teles, Magda C; Oliveira, Catarina R; Becker, Jorg D; Lopes, João S

    2016-02-01

    Group living animals must be able to express different behavior profiles depending on their social status. Therefore, the same genotype may translate into different behavioral phenotypes through socially driven differential gene expression. However, how social information is translated into a neurogenomic response and what are the specific cues in a social interaction that signal a change in social status are questions that have remained unanswered. Here, we show for the first time, to our knowledge, that the switch between status-specific neurogenomic states relies on the assessment of fight outcome rather than just on self- or opponent-only assessment of fighting ability. For this purpose, we manipulated the perception of fight outcome in male zebrafish and measured its impact on the brain transcriptome using a zebrafish whole genome gene chip. Males fought either a real opponent, and a winner and a loser were identified, or their own image on a mirror, in which case, despite expressing aggressive behavior, males did not experience either a victory or a defeat. Massive changes in the brain transcriptome were observed in real opponent fighters, with losers displaying both a higher number of differentially expressed genes and of coexpressed gene modules than winners. In contrast, mirror fighters expressed a neurogenomic state similar to that of noninteracting fish. The genes that responded to fight outcome included immediate early genes and genes involved in neuroplasticity and epigenetic modifications. These results indicate that, even in cognitively simple organisms such as zebrafish, neurogenomic responses underlying changes in social status rely on mutual assessment of fighting ability. PMID:26787876

  11. Aminoglycoside-Induced Hair Cell Death of Inner Ear Organs Causes Functional Deficits in Adult Zebrafish (Danio rerio)

    PubMed Central

    Uribe, Phillip M.; Sun, Huifang; Wang, Kevin; Asuncion, James D.; Wang, Qi; Chen, Chien-Wei; Steyger, Peter S.; Smith, Michael E.; Matsui, Jonathan I.

    2013-01-01

    Aminoglycoside antibiotics, like gentamicin, kill inner ear sensory hair cells in a variety of species including chickens, mice, and humans. The zebrafish (Danio rerio) has been used to study hair cell cytotoxicity in the lateral line organs of larval and adult animals. Little is known about whether aminoglycosides kill the hair cells within the inner ear of adult zebrafish. We report here the ototoxic effects of gentamicin on hair cells in the saccule, the putative hearing organ, and utricle of zebrafish. First, adult zebrafish received a single 30 mg/kg intraperitoneal injection of fluorescently-tagged gentamicin (GTTR) to determine the distribution of gentamicin within inner ear sensory epithelia. After 4 hours, GTTR was observed in hair cells throughout the saccular and utriclar sensory epithelia. To assess the ototoxic effects of gentamicin, adult zebrafish received a single 250 mg/kg intraperitoneal injection of gentamicin and, 24 hours later, auditory evoked potential recordings (AEPs) revealed significant shifts in auditory thresholds compared to untreated controls. Zebrafish were then euthanized, the inner ear fixed, and labeled for apoptotic cells (TUNEL reaction), and the stereociliary bundles of hair cells labeled with fluorescently-tagged phalloidin. Whole mounts of the saccule and utricle were imaged and cells counted. There were significantly more TUNEL-labeled cells found in both organs 4 hours after gentamicin injection compared to vehicle-injected controls. As expected, significantly fewer hair cell bundles were found along the rostral-caudal axis of the saccule and in the extrastriolar and striolar regions of the utricle in gentamicin-treated animals compared to untreated controls. Therefore, as in other species, gentamicin causes significant inner ear sensory hair cell death and auditory dysfunction in zebrafish. PMID:23533589

  12. Strong Static Magnetic Fields Elicit Swimming Behaviors Consistent with Direct Vestibular Stimulation in Adult Zebrafish

    PubMed Central

    Ward, Bryan K.; Tan, Grace X-J; Roberts, Dale C.; Della Santina, Charles C.; Zee, David S.; Carey, John P.

    2014-01-01

    Zebrafish (Danio rerio) offer advantages as model animals for studies of inner ear development, genetics and ototoxicity. However, traditional assessment of vestibular function in this species using the vestibulo-ocular reflex requires agar-immobilization of individual fish and specialized video, which are difficult and labor-intensive. We report that using a static magnetic field to directly stimulate the zebrafish labyrinth results in an efficient, quantitative behavioral assay in free-swimming fish. We recently observed that humans have sustained nystagmus in high strength magnetic fields, and we attributed this observation to magnetohydrodynamic forces acting on the labyrinths. Here, fish were individually introduced into the center of a vertical 11.7T magnetic field bore for 2-minute intervals, and their movements were tracked. To assess for heading preference relative to a magnetic field, fish were also placed in a horizontally oriented 4.7T magnet in infrared (IR) light. A sub-population was tested again in the magnet after gentamicin bath to ablate lateral line hair cell function. Free-swimming adult zebrafish exhibited markedly altered swimming behavior while in strong static magnetic fields, independent of vision or lateral line function. Two-thirds of fish showed increased swimming velocity or consistent looping/rolling behavior throughout exposure to a strong, vertically oriented magnetic field. Fish also demonstrated altered swimming behavior in a strong horizontally oriented field, demonstrating in most cases preferred swimming direction with respect to the field. These findings could be adapted for ‘high-throughput’ investigations of the effects of environmental manipulations as well as for changes that occur during development on vestibular function in zebrafish. PMID:24647586

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

    PubMed

    Ward, Bryan K; Tan, Grace X-J; Roberts, Dale C; Della Santina, Charles C; Zee, David S; Carey, John P

    2014-01-01

    Zebrafish (Danio rerio) offer advantages as model animals for studies of inner ear development, genetics and ototoxicity. However, traditional assessment of vestibular function in this species using the vestibulo-ocular reflex requires agar-immobilization of individual fish and specialized video, which are difficult and labor-intensive. We report that using a static magnetic field to directly stimulate the zebrafish labyrinth results in an efficient, quantitative behavioral assay in free-swimming fish. We recently observed that humans have sustained nystagmus in high strength magnetic fields, and we attributed this observation to magnetohydrodynamic forces acting on the labyrinths. Here, fish were individually introduced into the center of a vertical 11.7T magnetic field bore for 2-minute intervals, and their movements were tracked. To assess for heading preference relative to a magnetic field, fish were also placed in a horizontally oriented 4.7T magnet in infrared (IR) light. A sub-population was tested again in the magnet after gentamicin bath to ablate lateral line hair cell function. Free-swimming adult zebrafish exhibited markedly altered swimming behavior while in strong static magnetic fields, independent of vision or lateral line function. Two-thirds of fish showed increased swimming velocity or consistent looping/rolling behavior throughout exposure to a strong, vertically oriented magnetic field. Fish also demonstrated altered swimming behavior in a strong horizontally oriented field, demonstrating in most cases preferred swimming direction with respect to the field. These findings could be adapted for 'high-throughput' investigations of the effects of environmental manipulations as well as for changes that occur during development on vestibular function in zebrafish. PMID:24647586

  14. Inhibitory effect of cadmium on estrogen signaling in zebrafish brain and protection by zinc.

    PubMed

    Chouchene, Lina; Pellegrini, Elisabeth; Gueguen, Marie-Madeleine; Hinfray, Nathalie; Brion, François; Piccini, Benjamin; Kah, Olivier; Saïd, Khaled; Messaoudi, Imed; Pakdel, Farzad

    2016-06-01

    The present study was conducted to assess the effects of Cd exposure on estrogen signaling in the zebrafish brain, as well as the potential protective role of Zn against Cd-induced toxicity. For this purpose, the effects on transcriptional activation of the estrogen receptors (ERs), aromatase B (Aro-B) protein expression and molecular expression of related genes were examined in vivo using wild-type and transgenic zebrafish embryos. For in vitro studies, an ER-negative glial cell line (U251MG) transfected with different zebrafish ER subtypes (ERα, ERβ1 and ERβ2) was also used. Embryos were exposed either to estradiol (E2 ), Cd, E2 +Cd or E2 +Cd+Zn for 72 h and cells were exposed to the same treatments for 30 h. Our results show that E2 treatment promoted the transcriptional activation of ERs and increased Aro-B expression, at both the protein and mRNA levels. Although exposure to Cd, does not affect the studied parameters when administered alone, it significantly abolished the E2 -stimulated transcriptional response of the reporter gene for the three ER subtypes in U251-MG cells, and clearly inhibited the E2 induction of Aro-B in radial glial cells of zebrafish embryos. These inhibitory effects were accompanied by a significant downregulation of the expression of esr1, esr2a, esr2b and cyp19a1b genes compared to the E2 -treated group used as a positive control. Zn administration during simultaneous exposure to E2 and Cd strongly stimulated zebrafish ERs transactivation and increased Aro-B protein expression, whereas mRNA levels of the three ERs as well as the cyp19a1b remained unchanged in comparison with Cd-treated embryos. In conclusion, our results clearly demonstrate that Cd acts as a potent anti-estrogen in vivo and in vitro, and that Cd-induced E2 antagonism can be reversed, at the protein level, by Zn supplement. Copyright © 2016 John Wiley & Sons, Ltd. PMID:26857037

  15. Re-epithelialization of cutaneous wounds in adult zebrafish combines mechanisms of wound closure in embryonic and adult mammals.

    PubMed

    Richardson, Rebecca; Metzger, Manuel; Knyphausen, Philipp; Ramezani, Thomas; Slanchev, Krasimir; Kraus, Christopher; Schmelzer, Elmon; Hammerschmidt, Matthias

    2016-06-15

    Re-epithelialization of cutaneous wounds in adult mammals takes days to complete and relies on numerous signalling cues and multiple overlapping cellular processes that take place both within the epidermis and in other participating tissues. Re-epithelialization of partial- or full-thickness skin wounds of adult zebrafish, however, is extremely rapid and largely independent of the other processes of wound healing. Live imaging after treatment with transgene-encoded or chemical inhibitors reveals that re-epithelializing keratinocytes repopulate wounds by TGF-β- and integrin-dependent lamellipodial crawling at the leading edges of the epidermal tongue. In addition, re-epithelialization requires long-range epithelial rearrangements, involving radial intercalations, flattening and directed elongation of cells - processes that are dependent on Rho kinase, JNK and, to some extent, planar cell polarity within the epidermis. These rearrangements lead to a massive recruitment of keratinocytes from the adjacent epidermis and make re-epithelialization independent of keratinocyte proliferation and the mitogenic effect of FGF signalling, which are only required after wound closure, allowing the epidermis outside the wound to re-establish its normal thickness. Together, these results demonstrate that the adult zebrafish is a valuable in vivo model for studying and visualizing the processes involved in cutaneous wound closure, facilitating the dissection of direct from indirect and motogenic from mitogenic effects of genes and molecules affecting wound re-epithelialization. PMID:27122176

  16. Investigation into effects of antipsychotics on ectonucleotidase and adenosine deaminase in zebrafish brain.

    PubMed

    Seibt, Kelly Juliana; Oliveira, Renata da Luz; Bogo, Mauricio Reis; Senger, Mario Roberto; Bonan, Carla Denise

    2015-12-01

    Antipsychotic agents are used for the treatment of psychotic symptoms in patients with several brain disorders, such as schizophrenia. Atypical and typical antipsychotics differ regarding their clinical and side-effects profile. Haloperidol is a representative typical antipsychotic drug and has potent dopamine receptor antagonistic functions; however, atypical antipsychotics have been developed and characterized an important advance in the treatment of schizophrenia and other psychotic disorders. Purine nucleotides and nucleosides, such as ATP and adenosine, constitute a ubiquitous class of extracellular signaling molecules crucial for normal functioning of the nervous system. Indirect findings suggest that changes in the purinergic system, more specifically in adenosinergic activity, could be involved in the pathophysiology of schizophrenia. We investigated the effects of typical and atypical antipsychotics on ectonucleotidase and adenosine deaminase (ADA) activities, followed by an analysis of gene expression patterns in zebrafish brain. Haloperidol treatment (9 µM) was able to decrease ATP hydrolysis (35%), whereas there were no changes in hydrolysis of ADP and AMP in brain membranes after antipsychotic exposure. Adenosine deamination in membrane fractions was inhibited (38%) after haloperidol treatment when compared to the control; however, no changes were observed in ADA soluble fractions after haloperidol exposure. Sulpiride (250 µM) and olanzapine (100 µM) did not alter ectonucleotidase and ADA activities. Haloperidol also led to a decrease in entpd2_mq, entpd3 and adal mRNA transcripts. These findings demonstrate that haloperidol is an inhibitor of NTPDase and ADA activities in zebrafish brain, suggesting that purinergic signaling may also be a target of pharmacological effects promoted by this drug. PMID:26156500

  17. Contrast-Enhanced X-Ray Micro-Computed Tomography as a Versatile Method for Anatomical Studies of Adult Zebrafish.

    PubMed

    Babaei, Fatemeh; Hong, Tony Liu Chi; Yeung, Kelvin; Cheng, Shuk Han; Lam, Yun Wah

    2016-08-01

    One attractive quality of zebrafish as a model organism for biological research is that transparency at early developmental stages allows the optical imaging of cellular and molecular events. However, this advantage cannot be applied to adult zebrafish. In this study, we explored the use of contrast-enhanced X-ray micro-computed tomography (microCT) on adult zebrafish in which the organism was stained with iodine, a simple and economical contrasting agent, after fixation. Tomographic reconstruction of the microCT data allowed the three-dimensional (3D) volumetric analyses of individual organs in adult zebrafish. Adipose tissues showed a higher affinity to iodine and were more strongly contrasted in microCT. As traditional histological techniques often involve dehydration steps that remove tissue lipids, iodine-contrasted microCT offers a convenient method for visualizing fat deposition in fish. Utilizing this advantage, we discovered a transient accumulation of lipids around the heart after ventricular amputation, suggesting a correlation between lipid distribution and heart regeneration. Taken together, microCT is a versatile technique that enables the 3D visualization of zebrafish organs, as well as other fish models, in their anatomical context. This simple method is a valuable new addition to the arsenal of techniques available to this model organism. PMID:27058023

  18. The role of acid-sensing ion channels in epithelial Na+ uptake in adult zebrafish (Danio rerio).

    PubMed

    Dymowska, Agnieszka K; Boyle, David; Schultz, Aaron G; Goss, Greg G

    2015-04-15

    Acid-sensing ion channels (ASICs) are epithelial Na(+) channels gated by external H(+). Recently, it has been demonstrated that ASICs play a role in Na(+) uptake in freshwater rainbow trout. Here, we investigate the potential involvement of ASICs in Na(+) transport in another freshwater fish species, the zebrafish (Danio rerio). Using molecular and histological techniques we found that asic genes and the ASIC4.2 protein are expressed in the gill of adult zebrafish. Immunohistochemistry revealed that mitochondrion-rich cells positive for ASIC4.2 do not co-localize with Na(+)/K(+)-ATPase-rich cells, but co-localize with cells expressing vacuolar-type H(+)-ATPase. Furthermore, pharmacological inhibitors of ASIC and Na(+)/H(+)-exchanger significantly reduced uptake of Na(+) in adult zebrafish exposed to low-Na(+) media, but did not cause the same response in individuals exposed to ultra-low-Na(+) water. Our results suggest that in adult zebrafish ASICs play a role in branchial Na(+) uptake in media with low Na(+) concentrations and that mechanisms used for Na(+) uptake by zebrafish may depend on the Na(+) concentration in the acclimation medium. PMID:25722005

  19. The common neural parasite Pseudoloma neurophilia is associated with altered startle response habituation in adult zebrafish (Danio rerio): Implications for the zebrafish as a model organism.

    PubMed

    Spagnoli, Sean; Xue, Lan; Kent, Michael L

    2015-09-15

    The zebrafish's potential as a model for human neurobehavioral research appears nearly limitless despite its relatively recent emergence as an experimental organism. Since the zebrafish has only been part of the research community for a handful of decades, pathogens from its commercial origins continue to plague laboratory stocks. One such pathogen is Pseudoloma neurophilia, a common microparasite in zebrafish laboratories world-wide that generally produces subclinical infections. Given its high prevalence, its predilection for the host's brain and spinal cord, and the delicate nature of neurobehavioral research, the behavioral consequences of subclinical P. neurophilia infection must be explored. Fish infected via cohabitation were tested for startle response habituation in parallel with controls in a device that administered ten taps over 10 min along with taps at 18 and 60 min to evaluate habituation extinction. After testing, fish were euthanized and evaluated for infection via histopathology. Infected fish had a significantly smaller reduction in startle velocity during habituation compared to uninfected tankmates and controls. Habituation was eliminated in infected and control fish at 18 min, whereas exposed negative fish retained partial habituation at 18 min. Infection was also associated with enhanced capture evasion: Despite the absence of external symptoms, infected fish tended to be caught later than uninfected fish netted from the same tank. The combination of decreased overall habituation, early extinction of habituation compared to uninfected cohorts, and enhanced netting evasion indicates that P. neurophilia infection is associated with a behavioral phenotype distinct from that of controls and uninfected cohorts. Because of its prevalence in zebrafish facilities, P. neurophilia has the potential to insidiously influence a wide range of neurobehavioral studies if these associations are causative. Rigorous health screening is therefore vital to the

  20. TBBPA chronic exposure produces sex-specific neurobehavioral and social interaction changes in adult zebrafish.

    PubMed

    Chen, Jiangfei; Tanguay, Robert L; Simonich, Michael; Nie, Shangfei; Zhao, Yuxin; Li, Lelin; Bai, Chenglian; Dong, Qiaoxiang; Huang, Changjiang; Lin, Kuangfei

    2016-01-01

    The toxicity of tetrabromobisphenol A (TBBPA) has been extensively studied because of its high production volume. TBBPA is toxic to aquatic fish based on acute high concentration exposure tests, and few studies have assessed the behavioral effects of low concentration chronic TBBPA exposures in aquatic organisms. The present study defined the developmental and neurobehavioral effects associated with exposure of zebrafish to 0, 5 and 50nM TBBPA during 1-120days post-fertilization (dpf) following by detoxification for four months before the behaviors assessment. These low concentration TBBPA exposures were not associated with malformations and did not alter sex ratio, but resulted in reduced zebrafish body weight and length. Adult behavioral assays indicated that TBBPA exposed males had significantly higher average swim speeds and spent significantly more time in high speed darting mode and less time in medium cruising mode compared to control males. In an adult photomotor response assay, TBBPA exposure was associated with hyperactivity in male fish. Female zebrafish responses in these assays followed a similar trend, but the magnitude of TBBPA effects was generally smaller than in males. Social interaction evaluated using a mirror attack test showed that 50nM TBBPA exposed males had heightened aggression. Females exposed to 50nM TBBPA spent more time in the vicinity of the mirror, but did not show increased aggression toward the mirror compared to unexposed control fish. Overall, the hyperactivity and social behavior deficits ascribed here to chronic TBBPA exposure was most profound in males. Our findings indicate that TBBPA can cause developmental and neurobehavioral deficits, and may pose significant health risk to humans. PMID:27221227

  1. Neuronal labeling patterns in the spinal cord of adult transgenic Zebrafish.

    PubMed

    Stil, Aurélie; Drapeau, Pierre

    2016-06-01

    We describe neuronal patterns in the spinal cord of adult zebrafish. We studied the distribution of cells and processes in the three spinal regions reported in the literature: the 8th vertebra used as a transection injury site, the 15th vertebra mainly used for motor cell recordings and also for crush injury, and the 24th vertebra used to record motor nerve activity. We used well-known transgenic lines in which expression of green fluorescent protein (GFP) is driven by promoters to hb9 and isl1 in motoneurons, alx/chx10 and evx1 interneurons, ngn1 in sensory neurons and olig2 in oligodendrocytes, as well as antibodies for neurons (HuC/D, NF and SV2) and glia (GFAP). In isl1:GFP fish, GFP-positive processes are retained in the upper part of ventral horns and two subsets of cell bodies are observed. The pattern of the transgene in hb9:GFP adults is more diffuse and fibers are present broadly through the adult spinal cord. In alx/chx10 and evx1 lines we respectively observed two and three different GFP-positive populations. Finally, the ngn1:GFP transgene identifies dorsal root ganglion and some cells in dorsal horns. Interestingly some GFP positive fibers in ngn1:GFP fish are located around Mauthner axons and their density seems to be related to a rostrocaudal gradient. Many other cell types have been described in embryos and need to be studied in adults. Our findings provide a reference for further studies on spinal cytoarchitecture. Combined with physiological, histological and pathological/traumatic approaches, these studies will help clarify the operation of spinal locomotor circuits of adult zebrafish. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 642-660, 2016. PMID:26408263

  2. Neural repair in the adult brain

    PubMed Central

    Jessberger, Sebastian

    2016-01-01

    Acute or chronic injury to the adult brain often results in substantial loss of neural tissue and subsequent permanent functional impairment. Over the last two decades, a number of approaches have been developed to harness the regenerative potential of neural stem cells and the existing fate plasticity of neural cells in the nervous system to prevent tissue loss or to enhance structural and functional regeneration upon injury. Here, we review recent advances of stem cell-associated neural repair in the adult brain, discuss current challenges and limitations, and suggest potential directions to foster the translation of experimental stem cell therapies into the clinic. PMID:26918167

  3. Subchronic atrazine exposure changes defensive behaviour profile and disrupts brain acetylcholinesterase activity of zebrafish.

    PubMed

    Schmidel, Ademir J; Assmann, Karla L; Werlang, Chariane C; Bertoncello, Kanandra T; Francescon, Francini; Rambo, Cassiano L; Beltrame, Gabriela M; Calegari, Daiane; Batista, Cibele B; Blaser, Rachel E; Roman Júnior, Walter A; Conterato, Greicy M M; Piato, Angelo L; Zanatta, Leila; Magro, Jacir Dal; Rosemberg, Denis B

    2014-01-01

    Animal behaviour is the interaction between environment and an individual organism, which also can be influenced by its neighbours. Variations in environmental conditions, as those caused by contaminants, may lead to neurochemical impairments altering the pattern of the behavioural repertoire of the species. Atrazine (ATZ) is an herbicide widely used in agriculture that is frequently detected in surface water, affecting non-target species. The zebrafish is a valuable model organism to assess behavioural and neurochemical effects of different contaminants since it presents a robust behavioural repertoire and also all major neurotransmitter systems described for mammalian species. The goal of this study was to evaluate the effects of subchronic ATZ exposure in defensive behaviours of zebrafish (shoaling, thigmotaxis, and depth preference) using the split depth tank. Furthermore, to investigate a putative role of cholinergic signalling on ATZ-mediated effects, we tested whether this herbicide alters acetylcholinesterase (AChE) activity in brain and muscle preparations. Fish were exposed to ATZ for 14days and the following groups were tested: control (0.2% acetone) and ATZ (10 and 1000μg/L). The behaviour of four animals in the same tank was recorded for 6min and biological samples were prepared. Our results showed that 1000μg/L ATZ significantly increased the inter-fish distance, as well as the nearest and farthest neighbour distances. This group also presented an increase in the shoal area with decreased social interaction. No significant differences were detected for the number of animals in the shallow area, latency to enter the shallow and time spent in shallow and deep areas of the apparatus, but the ATZ 1000 group spent significantly more time near the walls. Although ATZ did not affect muscular AChE, it significantly reduced AChE activity in brain. Exposure to 10μg/L ATZ did not affect behaviour or AChE activity. These data suggest that ATZ impairs defensive

  4. Crossing the blood-brain-barrier with transferrin conjugated carbon dots: A zebrafish model study.

    PubMed

    Li, Shanghao; Peng, Zhili; Dallman, Julia; Baker, James; Othman, Abdelhameed M; Blackwelder, Patrica L; Leblanc, Roger M

    2016-09-01

    Drug delivery to the central nervous system (CNS) in biological systems remains a major medical challenge due to the tight junctions between endothelial cells known as the blood-brain-barrier (BBB). Here we use a zebrafish model to explore the possibility of using transferrin-conjugated carbon dots (C-Dots) to ferry compounds across the BBB. C-Dots have previously been reported to inhibit protein fibrillation, and they are also used to deliver drugs for disease treatment. In terms of the potential medical application of C-Dots for the treatment of CNS diseases, one of the most formidable challenges is how to deliver them inside the CNS. To achieve this in this study, human transferrin was covalently conjugated to C-Dots. The conjugates were then injected into the vasculature of zebrafish to examine the possibility of crossing the BBB in vivo via transferrin receptor-mediated endocytosis. The experimental observations suggest that the transferrin-C-Dots can enter the CNS while C-Dots alone cannot. PMID:27187189

  5. Stereoselective induction by 2,2',3,4',6-pentachlorobiphenyl in adult zebrafish (Danio rerio): Implication of chirality in oxidative stress and bioaccumulation.

    PubMed

    Chai, Tingting; Cui, Feng; Mu, Xiyan; Yang, Yang; Qi, Suzhen; Zhu, Lizhen; Wang, Chengju; Qiu, Jing

    2016-08-01

    This study aimed to investigate the oxidative stress process and bioaccumulation the racemic/(-)-/(+)- 2,2',3,4',6-pentachlorobiphenyl were administered to adult zebrafish (Danio rerio) after prolonged exposure of 56-days uptake and 49-days depuration experiments. Stereoselective accumulation was observed in adult samples after racemic exposure as revealed by decreased enantiomer fractions. The two enantiomers of PCB91 accumulated at different rates with logBCFk values close to 3.7, suggesting that they were highly hazardous and persistent pollutants. Exposure to racemic/(-)-/(+)- PCB91 stereoselectively induced oxidative stress owing to changes in reactive oxygen species, malondialdehyde contents, antioxidant enzyme activities and gene expressions in brain and liver tissues. In addition, the stereoselective relationship between bioconcentration and oxidative stress were also presented in this study. Our findings might be helpful for elucidating the environmental risk of the two enantiomers of PCB91 that induce toxicity in aquatic organisms. PMID:27179325

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

    PubMed

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

    2016-04-01

    Recently, zebrafish have been used to examine hyperglycemia-induced complications (retinopathy and neuropathy), as would occur in individuals with diabetes. Current models to induce hyperglycemia in zebrafish include glucose immersion and streptozotocin injections. Both are effective, although neither is reported to elevate blood sugar values for more than 1 month. In this article, we report differences in hyperglycemia induction and maintenance in young (4-11 months) versus old (1-3 years) zebrafish adults. In particular, older fish immersed in an alternating constant external glucose solution (2%) for 2 months displayed elevated blood glucose levels for the entire experimental duration. In contrast, younger adults displayed only transient hyperglycemia, suggesting the fish were acclimating to the glucose exposure protocol. However, modifying the immersion protocol to include a stepwise increasing glucose concentration (from 1% → 2%→3%) resulted in maintained hyperglycemia in younger zebrafish adults for up to 2 months. Glucose-exposed younger fish collected after 8 weeks of exposure also displayed a significant decrease in wet weight. Taken together, these data suggest different susceptibilities to hyperglycemia in older and younger fish and that stepwise increasing glucose concentrations of 1% are required for maintenance of hyperglycemia in younger adults, with higher concentrations of glucose resulting in greater increases in blood sugar values. PMID:26771444

  7. Two knockdown models of the autism genes SYNGAP1 and SHANK3 in zebrafish produce similar behavioral phenotypes associated with embryonic disruptions of brain morphogenesis.

    PubMed

    Kozol, Robert A; Cukier, Holly N; Zou, Bing; Mayo, Vera; De Rubeis, Silvia; Cai, Guiqing; Griswold, Anthony J; Whitehead, Patrice L; Haines, Jonathan L; Gilbert, John R; Cuccaro, Michael L; Martin, Eden R; Baker, James D; Buxbaum, Joseph D; Pericak-Vance, Margaret A; Dallman, Julia E

    2015-07-15

    Despite significant progress in the genetics of autism spectrum disorder (ASD), how genetic mutations translate to the behavioral changes characteristic of ASD remains largely unknown. ASD affects 1-2% of children and adults, and is characterized by deficits in verbal and non-verbal communication, and social interactions, as well as the presence of repetitive behaviors and/or stereotyped interests. ASD is clinically and etiologically heterogeneous, with a strong genetic component. Here, we present functional data from syngap1 and shank3 zebrafish loss-of-function models of ASD. SYNGAP1, a synaptic Ras GTPase activating protein, and SHANK3, a synaptic scaffolding protein, were chosen because of mounting evidence that haploinsufficiency in these genes is highly penetrant for ASD and intellectual disability (ID). Orthologs of both SYNGAP1 and SHANK3 are duplicated in the zebrafish genome and we find that all four transcripts (syngap1a, syngap1b, shank3a and shank3b) are expressed at the earliest stages of nervous system development with pronounced expression in the larval brain. Consistent with early expression of these genes, knockdown of syngap1b or shank3a cause common embryonic phenotypes including delayed mid- and hindbrain development, disruptions in motor behaviors that manifest as unproductive swim attempts, and spontaneous, seizure-like behaviors. Our findings indicate that both syngap1b and shank3a play novel roles in morphogenesis resulting in common brain and behavioral phenotypes. PMID:25882707

  8. Two knockdown models of the autism genes SYNGAP1 and SHANK3 in zebrafish produce similar behavioral phenotypes associated with embryonic disruptions of brain morphogenesis

    PubMed Central

    Kozol, Robert A.; Cukier, Holly N.; Zou, Bing; Mayo, Vera; De Rubeis, Silvia; Cai, Guiqing; Griswold, Anthony J.; Whitehead, Patrice L.; Haines, Jonathan L.; Gilbert, John R.; Cuccaro, Michael L.; Martin, Eden R.; Baker, James D.; Buxbaum, Joseph D.; Pericak-Vance, Margaret A.; Dallman, Julia E.

    2015-01-01

    Despite significant progress in the genetics of autism spectrum disorder (ASD), how genetic mutations translate to the behavioral changes characteristic of ASD remains largely unknown. ASD affects 1–2% of children and adults, and is characterized by deficits in verbal and non-verbal communication, and social interactions, as well as the presence of repetitive behaviors and/or stereotyped interests. ASD is clinically and etiologically heterogeneous, with a strong genetic component. Here, we present functional data from syngap1 and shank3 zebrafish loss-of-function models of ASD. SYNGAP1, a synaptic Ras GTPase activating protein, and SHANK3, a synaptic scaffolding protein, were chosen because of mounting evidence that haploinsufficiency in these genes is highly penetrant for ASD and intellectual disability (ID). Orthologs of both SYNGAP1 and SHANK3 are duplicated in the zebrafish genome and we find that all four transcripts (syngap1a, syngap1b, shank3a and shank3b) are expressed at the earliest stages of nervous system development with pronounced expression in the larval brain. Consistent with early expression of these genes, knockdown of syngap1b or shank3a cause common embryonic phenotypes including delayed mid- and hindbrain development, disruptions in motor behaviors that manifest as unproductive swim attempts, and spontaneous, seizure-like behaviors. Our findings indicate that both syngap1b and shank3a play novel roles in morphogenesis resulting in common brain and behavioral phenotypes. PMID:25882707

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

    PubMed

    Gaulke, Christopher A; Barton, Carrie L; Proffitt, Sarah; Tanguay, Robert L; Sharpton, Thomas J

    2016-01-01

    Growing evidence indicates that disrupting the microbial community that comprises the intestinal tract, known as the gut microbiome, can contribute to the development or severity of disease. As a result, it is important to discern the agents responsible for microbiome disruption. While animals are frequently exposed to a diverse array of environmental chemicals, little is known about their effects on gut microbiome stability and structure. Here, we demonstrate how zebrafish can be used to glean insight into the effects of environmental chemical exposure on the structure and ecological dynamics of the gut microbiome. Specifically, we exposed forty-five adult zebrafish to triclosan-laden food for four or seven days or a control diet, and analyzed their microbial communities using 16S rRNA amplicon sequencing. Triclosan exposure was associated with rapid shifts in microbiome structure and diversity. We find evidence that several operational taxonomic units (OTUs) associated with the family Enterobacteriaceae appear to be susceptible to triclosan exposure, while OTUs associated with the genus Pseudomonas appeared to be more resilient and resistant to exposure. We also found that triclosan exposure is associated with topological alterations to microbial interaction networks and results in an overall increase in the number of negative interactions per microbe in these networks. Together these data indicate that triclosan exposure results in altered composition and ecological dynamics of microbial communities in the gut. Our work demonstrates that because zebrafish afford rapid and inexpensive interrogation of a large number of individuals, it is a useful experimental system for the discovery of the gut microbiome's interaction with environmental chemicals. PMID:27191725

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

    PubMed Central

    Barton, Carrie L.; Proffitt, Sarah; Tanguay, Robert L.; Sharpton, Thomas J.

    2016-01-01

    Growing evidence indicates that disrupting the microbial community that comprises the intestinal tract, known as the gut microbiome, can contribute to the development or severity of disease. As a result, it is important to discern the agents responsible for microbiome disruption. While animals are frequently exposed to a diverse array of environmental chemicals, little is known about their effects on gut microbiome stability and structure. Here, we demonstrate how zebrafish can be used to glean insight into the effects of environmental chemical exposure on the structure and ecological dynamics of the gut microbiome. Specifically, we exposed forty-five adult zebrafish to triclosan-laden food for four or seven days or a control diet, and analyzed their microbial communities using 16S rRNA amplicon sequencing. Triclosan exposure was associated with rapid shifts in microbiome structure and diversity. We find evidence that several operational taxonomic units (OTUs) associated with the family Enterobacteriaceae appear to be susceptible to triclosan exposure, while OTUs associated with the genus Pseudomonas appeared to be more resilient and resistant to exposure. We also found that triclosan exposure is associated with topological alterations to microbial interaction networks and results in an overall increase in the number of negative interactions per microbe in these networks. Together these data indicate that triclosan exposure results in altered composition and ecological dynamics of microbial communities in the gut. Our work demonstrates that because zebrafish afford rapid and inexpensive interrogation of a large number of individuals, it is a useful experimental system for the discovery of the gut microbiome’s interaction with environmental chemicals. PMID:27191725

  11. Mitral cells in the olfactory bulb of adult zebrafish (Danio rerio): morphology and distribution.

    PubMed

    Fuller, Cynthia L; Yettaw, Holly K; Byrd, Christine A

    2006-11-10

    The mitral cell is the primary output neuron and central relay in the olfactory bulb of vertebrates. The morphology of these cells has been studied extensively in mammalian systems and to a lesser degree in teleosts. This study uses retrograde tract tracing and other techniques to characterize the morphology and distribution of mitral cells in the olfactory bulb of adult zebrafish, Danio rerio. These output neurons, located primarily in the glomerular layer and superficial internal cell layer, had variable-shaped somata that ranged in size from 4-18 microm in diameter and 31-96 microm2 in cross-sectional area. The mitral cells exhibited two main types of morphologies with regard to their dendrites: the unidendritic morphology was a single primary dendrite with one or more tufts, but multidendritic cells with several dendritic projections also were seen. The axons of these cells projected to either the medial or the lateral olfactory tract and, in general, the location of the cell on the medial or lateral side of the bulb was indicative of the tract to which it would project. Further, this study shows that the majority of zebrafish mitral cells likely innervate a single glomerulus rather than multiple glomeruli. This information is contrary to the multiple innervation pattern suggested for all teleost mitral cells. Our findings suggest that mitral cells in zebrafish may be more similar to mammalian mitral cells than previously believed, despite variation in size and structure. This information provides a revised anatomical framework for olfactory processing studies in this key model system. PMID:16977629

  12. Brain size and limits to adult neurogenesis.

    PubMed

    Paredes, Mercedes F; Sorrells, Shawn F; Garcia-Verdugo, Jose M; Alvarez-Buylla, Arturo

    2016-02-15

    The walls of the cerebral ventricles in the developing embryo harbor the primary neural stem cells from which most neurons and glia derive. In many vertebrates, neurogenesis continues postnatally and into adulthood in this region. Adult neurogenesis at the ventricle has been most extensively studied in organisms with small brains, such as reptiles, birds, and rodents. In reptiles and birds, these progenitor cells give rise to young neurons that migrate into many regions of the forebrain. Neurogenesis in adult rodents is also relatively widespread along the lateral ventricles, but migration is largely restricted to the rostral migratory stream into the olfactory bulb. Recent work indicates that the wall of the lateral ventricle is highly regionalized, with progenitor cells giving rise to different types of neurons depending on their location. In species with larger brains, young neurons born in these spatially specified domains become dramatically separated from potential final destinations. Here we hypothesize that the increase in size and topographical complexity (e.g., intervening white matter tracts) in larger brains may severely limit the long-term contribution of new neurons born close to, or in, the ventricular wall. We compare the process of adult neuronal birth, migration, and integration across species with different brain sizes, and discuss how early regional specification of progenitor cells may interact with brain size and affect where and when new neurons are added. PMID:26417888

  13. The Asparaginyl Endopeptidase Legumain Is Essential for Functional Recovery after Spinal Cord Injury in Adult Zebrafish

    PubMed Central

    Ma, Liping; Shen, Yan-Qin; Khatri, Harsh P.; Schachner, Melitta

    2014-01-01

    Unlike mammals, adult zebrafish are capable of regenerating severed axons and regaining locomotor function after spinal cord injury. A key factor for this regenerative capacity is the innate ability of neurons to re-express growth-associated genes and regrow their axons after injury in a permissive environment. By microarray analysis, we have previously shown that the expression of legumain (also known as asparaginyl endopeptidase) is upregulated after complete transection of the spinal cord. In situ hybridization showed upregulation of legumain expression in neurons of regenerative nuclei during the phase of axon regrowth/sprouting after spinal cord injury. Upregulation of Legumain protein expression was confirmed by immunohistochemistry. Interestingly, upregulation of legumain expression was also observed in macrophages/microglia and neurons in the spinal cord caudal to the lesion site after injury. The role of legumain in locomotor function after spinal cord injury was tested by reducing Legumain expression by application of anti-sense morpholino oligonucleotides. Using two independent anti-sense morpholinos, locomotor recovery and axonal regrowth were impaired when compared with a standard control morpholino. We conclude that upregulation of legumain expression after spinal cord injury in the adult zebrafish is an essential component of the capacity of injured neurons to regrow their axons. Another feature contributing to functional recovery implicates upregulation of legumain expression in the spinal cord caudal to the injury site. In conclusion, we established for the first time a function for an unusual protease, the asparaginyl endopeptidase, in the nervous system. This study is also the first to demonstrate the importance of legumain for repair of an injured adult central nervous system of a spontaneously regenerating vertebrate and is expected to yield insights into its potential in nervous system regeneration in mammals. PMID:24747977

  14. What is the Thalamus in Zebrafish?

    PubMed Central

    Mueller, Thomas

    2012-01-01

    Current research on the thalamus and related structures in the zebrafish diencephalon identifies an increasing number of both neurological structures and ontogenetic processes as evolutionary conserved between teleosts and mammals. The patterning processes, for example, which during the embryonic development of zebrafish form the thalamus proper appear largely conserved. Yet also striking differences between zebrafish and other vertebrates have been observed, particularly when we look at mature and histologically differentiated brains. A case in point is the migrated preglomerular complex of zebrafish which evolved only within the lineage of ray-finned fish and has no counterpart in mammals or tetrapod vertebrates. Based on its function as a sensory relay station with projections to pallial zones, the preglomerular complex has been compared to specific thalamic nuclei in mammals. However, no thalamic projections to the zebrafish dorsal pallium, which corresponds topologically to the mammalian isocortex, have been identified. Merely one teleostean thalamic nucleus proper, the auditory nucleus, projects to a part of the dorsal telencephalon, the pallial amygdala. Studies on patterning mechanisms identify a rostral and caudal domain in the embryonic thalamus proper. In both, teleosts and mammals, the rostral domain gives rise to GABAergic neurons, whereas glutamatergic neurons originate in the caudal domain of the zebrafish thalamus. The distribution of GABAergic derivatives in the adult zebrafish brain, furthermore, revealed previously overlooked thalamic nuclei and redefined already established ones. These findings require some reconsideration regarding the topological origin of these adult structures. In what follows, I discuss how evolutionary conserved and newly acquired features of the developing and adult zebrafish thalamus can be compared to the mammalian situation. PMID:22586363

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

    PubMed

    Hentig, James T; Byrd-Jacobs, Christine A

    2016-01-01

    Zinc sulfate is a known olfactory toxicant, although its specific effects on the olfactory epithelium of zebrafish are unknown. Olfactory organs of adult zebrafish were exposed to zinc sulfate and, after 2, 3, 5, 7, 10 or 14 days, fish were processed for histological, immunohistochemical, ultrastructural, and behavioral analyses. Severe morphological disruption of the olfactory organ was observed two days following zinc sulfate exposure, including fusion of lamellae, epithelial inflammation, and significant loss of anti-calretinin labeling. Scanning electron microscopy revealed the apical surface of the sensory region was absent of ciliated structures, but microvilli were still present. Behavioral analysis showed significant loss of the ability to perceive bile salts and some fish also had no response to amino acids. Over the next several days, olfactory organ morphology, epithelial structure, and anti-calretinin labeling returned to control-like conditions, although the ability to perceive bile salts remained lost until day 14. Thus, exposure to zinc sulfate results in rapid degeneration of the olfactory organ, followed by restoration of morphology and function within two weeks. Zinc sulfate appears to have a greater effect on ciliated olfactory sensory neurons than on microvillous olfactory sensory neurons, suggesting differential effects on sensory neuron subtypes. PMID:27589738

  16. Agonistic interactions elicit rapid changes in brain nonapeptide levels in zebrafish.

    PubMed

    Teles, Magda C; Gozdowska, Magdalena; Kalamarz-Kubiak, Hanna; Kulczykowska, Ewa; Oliveira, Rui F

    2016-08-01

    The teleost fish nonapeptides, arginine vasotocin (AVT) and isotocin (IT), have been implicated in the regulation of social behavior. These peptides are expected to be involved in acute and transient changes in social context, in order to be efficient in modulating the expression of social behavior according to changes in the social environment. Here we tested the hypothesis that short-term social interactions are related to changes in the level of both nonapeptides across different brain regions. For this purpose we exposed male zebrafish to two types of social interactions: (1) real opponent interactions, from which a Winner and a Loser emerged; and (2) mirror-elicited interactions, that produced individuals that did not experience a change in social status despite expressing similar levels of aggressive behavior to those of participants in real-opponent fights. Non-interacting individuals were used as a reference group. Each social phenotype (i.e. Winners, Losers, Mirror-fighters) presented a specific brain profile of nonapeptides when compared to the reference group. Moreover, the comparison between the different social phenotypes allowed to address the specific aspects of the interaction (e.g. assessment of opponent aggressive behavior vs. self-assessment of expressed aggressive behavior) that are linked with neuropeptide responses. Overall, agonistic interactions seem to be more associated with the changes in brain AVT than IT, which highlights the preferential role of AVT in the regulation of aggressive behavior already described for other species. PMID:27235811

  17. Acid-sensing ion channels (ASICs) 2 and 4.2 are expressed in the retina of the adult zebrafish.

    PubMed

    Viña, E; Parisi, V; Sánchez-Ramos, C; Cabo, R; Guerrera, M C; Quirós, L M; Germanà, A; Vega, J A; García-Suárez, O

    2015-05-01

    Acid-sensing ion channels (ASICs) are H(+)-gated, voltage-insensitive cation channels involved in synaptic transmission, mechanosensation and nociception. Different ASICs have been detected in the retina of mammals but it is not known whether they are expressed in adult zebrafish, a commonly used animal model to study the retina in both normal and pathological conditions. We study the expression and distribution of ASIC2 and ASIC4 in the retina of adult zebrafish and its regulation by light using PCR, in situ hybridization, western blot and immunohistochemistry. We detected mRNA encoding zASIC2 and zASIC4.2 but not zASIC4.1. ASIC2, at the mRNA or protein level, was detected in the outer nuclear layer, the outer plexiform layer, the inner plexiform layer, the retinal ganglion cell layer and the optic nerve. ASIC4 was expressed in the photoreceptors layer and to a lesser extent in the retinal ganglion cell layer. Furthermore, the expression of both ASIC2 and ASIC4.2 was down-regulated by light and darkness. These results are the first demonstration that ASIC2 and ASIC4 are expressed in the adult zebrafish retina and suggest that zebrafish could be used as a model organism for studying retinal pathologies involving ASICs. PMID:25585988

  18. Effects of Chronic Dietary Selenomethionine Exposure on the Visual System of Adult and F1 Generation Zebrafish (Danio rerio).

    PubMed

    Raine, Jason C; Lallemand, Lise; Pettem, Connor M; Janz, David M

    2016-09-01

    The effects of chronic dietary selenomethionine (SeMet) exposure on the visual system of adult zebrafish and their progeny were investigated. Adult zebrafish were exposed to measured concentrations of 1.1 (control) and 10.3 µg Se/g dry mass as SeMet for 57 days, then encouraged to breed. Progeny were reared to swim-up and differences in mortality, eye size and visual behaviour were determined. Adult vision was also investigated using behavioural assays. Adults fed the SeMet-spiked diet exhibited significantly fewer positive reactions in the escape response assay when compared to controls. Larvae from adults fed elevated SeMet had smaller eyes and a lower proportion of positive responses in phototaxis, oculomotor and optokinetic response assays compared to controls. These results demonstrate that environmentally relevant elevated dietary SeMet exposure can affect the visual system of both exposed adult zebrafish and their progeny, which could affect fitness and survivability. PMID:27312825

  19. High-frequency dual mode pulsed wave Doppler imaging for monitoring the functional regeneration of adult zebrafish hearts

    PubMed Central

    Kang, Bong Jin; Park, Jinhyoung; Kim, Jieun; Kim, Hyung Ham; Lee, Changyang; Hwang, Jae Youn; Lien, Ching-Ling; Shung, K. Kirk

    2015-01-01

    Adult zebrafish is a well-known small animal model for studying heart regeneration. Although the regeneration of scars made by resecting the ventricular apex has been visualized with histological methods, there is no adequate imaging tool for tracking the functional recovery of the damaged heart. For this reason, high-frequency Doppler echocardiography using dual mode pulsed wave Doppler, which provides both tissue Doppler (TD) and Doppler flow in a same cardiac cycle, is developed with a 30 MHz high-frequency array ultrasound imaging system. Phantom studies show that the Doppler flow mode of the dual mode is capable of measuring the flow velocity from 0.1 to 15 cm s−1 with high accuracy (p-value = 0.974 > 0.05). In the in vivo study of zebrafish, both TD and Doppler flow signals were simultaneously obtained from the zebrafish heart for the first time, and the synchronized valve motions with the blood flow signals were identified. In the longitudinal study on the zebrafish heart regeneration, the parameters for diagnosing the diastolic dysfunction, for example, E/Em < 10, E/A < 0.14 for wild-type zebrafish, were measured, and the type of diastolic dysfunction caused by the amputation was found to be similar to the restrictive filling. The diastolic function was fully recovered within four weeks post-amputation. PMID:25505135

  20. Startle response memory and hippocampal changes in adult zebrafish pharmacologically-induced to exhibit anxiety/depression-like behaviors.

    PubMed

    Pittman, Julian T; Lott, Chad S

    2014-01-17

    Zebrafish (Danio rerio) are rapidly becoming a popular animal model for neurobehavioral and psychopharmacological research. While startle testing is a well-established assay to investigate anxiety-like behaviors in different species, screening of the startle response and its habituation in zebrafish is a new direction of translational biomedical research. This study focuses on a novel behavioral protocol to assess a tapping-induced startle response and its habituation in adult zebrafish that have been pharmacologically-induced to exhibit anxiety/depression-like behaviors. We demonstrated that zebrafish exhibit robust learning performance in a task adapted from the mammalian literature, a modified plus maze, and showed that ethanol and fluoxetine impair memory performance in this maze when administered after training at a dose that does not impair motor function, however, leads to significant upregulation of hippocampal serotoninergic neurons. These results suggest that the maze associative learning paradigm has face and construct validity and that zebrafish may become a translationally relevant study species for the analysis of the mechanisms of learning and memory changes associated with psychopharmacological treatment of anxiety/depression. PMID:24184510

  1. Cortisol-treated zebrafish embryos develop into pro-inflammatory adults with aberrant immune gene regulation.

    PubMed

    Hartig, Ellen I; Zhu, Shusen; King, Benjamin L; Coffman, James A

    2016-01-01

    Chronic early-life stress increases adult susceptibility to numerous health problems linked to chronic inflammation. One way that this may occur is via glucocorticoid-induced developmental programming. To gain insight into such programming we treated zebrafish embryos with cortisol and examined the effects on both larvae and adults. Treated larvae had elevated whole-body cortisol and glucocorticoid signaling, and upregulated genes associated with defense response and immune system processes. In adulthood the treated fish maintained elevated basal cortisol levels in the absence of exogenous cortisol, and constitutively mis-expressed genes involved in defense response and its regulation. Adults derived from cortisol-treated embryos displayed defective tailfin regeneration, heightened basal expression of pro-inflammatory genes, and failure to appropriately regulate those genes following injury or immunological challenge. These results support the hypothesis that chronically elevated glucocorticoid signaling early in life directs development of a pro-inflammatory adult phenotype, at the expense of immunoregulation and somatic regenerative capacity. PMID:27444789

  2. V-ATPase Proton Pumping Activity Is Required for Adult Zebrafish Appendage Regeneration

    PubMed Central

    Monteiro, Joana; Aires, Rita; Becker, Jörg D.; Jacinto, António; Certal, Ana C.; Rodríguez-León, Joaquín

    2014-01-01

    The activity of ion channels and transporters generates ion-specific fluxes that encode electrical and/or chemical signals with biological significance. Even though it is long known that some of those signals are crucial for regeneration, only in recent years the corresponding molecular sources started to be identified using mainly invertebrate or larval vertebrate models. We used adult zebrafish caudal fin as a model to investigate which and how ion transporters affect regeneration in an adult vertebrate model. Through the combined use of biophysical and molecular approaches, we show that V-ATPase activity contributes to a regeneration-specific H+ ef`flux. The onset and intensity of both V-ATPase expression and H+ efflux correlate with the different regeneration rate along the proximal-distal axis. Moreover, we show that V-ATPase inhibition impairs regeneration in adult vertebrate. Notably, the activity of this H+ pump is necessary for aldh1a2 and mkp3 expression, blastema cell proliferation and fin innervation. To the best of our knowledge, this is the first report on the role of V-ATPase during adult vertebrate regeneration. PMID:24671205

  3. Cortisol-treated zebrafish embryos develop into pro-inflammatory adults with aberrant immune gene regulation

    PubMed Central

    Hartig, Ellen I.; Zhu, Shusen; King, Benjamin L.

    2016-01-01

    ABSTRACT Chronic early-life stress increases adult susceptibility to numerous health problems linked to chronic inflammation. One way that this may occur is via glucocorticoid-induced developmental programming. To gain insight into such programming we treated zebrafish embryos with cortisol and examined the effects on both larvae and adults. Treated larvae had elevated whole-body cortisol and glucocorticoid signaling, and upregulated genes associated with defense response and immune system processes. In adulthood the treated fish maintained elevated basal cortisol levels in the absence of exogenous cortisol, and constitutively mis-expressed genes involved in defense response and its regulation. Adults derived from cortisol-treated embryos displayed defective tailfin regeneration, heightened basal expression of pro-inflammatory genes, and failure to appropriately regulate those genes following injury or immunological challenge. These results support the hypothesis that chronically elevated glucocorticoid signaling early in life directs development of a pro-inflammatory adult phenotype, at the expense of immunoregulation and somatic regenerative capacity. PMID:27444789

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

    PubMed Central

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

    2014-01-01

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

  5. Efficacy and Safety of 5 Anesthetics in Adult Zebrafish (Danio rerio)

    PubMed Central

    Collymore, Chereen; Tolwani, Angela; Lieggi, Christine; Rasmussen, Skye

    2014-01-01

    Although the safety and efficacy of tricaine methanesulfonate (MS222) for anesthesia of fish are well established, other anesthetics used less commonly in fish have been less extensively evaluated. Therefore, we compared gradual cooling, lidocaine hydrochloride (300, 325, and 350 mg/L), metomidate hydrochloride (2, 4, 6, 8, and 10 mg/L), and isoflurane (0.5 mL/L) with MS222 (150 mg/L) for anesthesia of adult zebrafish. The efficacy and safety of each agent was evaluated by observing loss of equilibrium, slowing of opercular movement, response to tail-fin pinch, recovery time, and anesthesia-associated mortality rates. At 15 min after anesthetic recovery, we used a novel-tank test to evaluate whether anesthetic exposure influenced short-term anxiety-like behavior. Behavioral parameters measured included latency to enter and number of transitions to the upper half of the tank, number of erratic movements, and number of freezing bouts. Behavior after anesthesia was unaltered regardless of the anesthetic used. Efficacy and safety differed among the anesthetics evaluated. Gradual cooling was useful for short procedures requiring immobilization only, but all instrumentation and surfaces that come in contact with fish must be maintained at approximately 10 °C. MS222 and lidocaine hydrochloride at 325 mg/L were effective as anesthetic agents for surgical procedures in adult zebrafish, but isoflurane and high-dose lidocaine hydrochloride were unsuitable as sole anesthetic agents due to high (30%) mortality rates. Although MS222 remains the best choice for generating a surgical plane of anesthesia, metomidate hydrochloride and gradual cooling were useful for sedation and immobilization for nonpainful procedures. PMID:24602548

  6. Peripheral Axons of the Adult Zebrafish Maxillary Barbel Extensively Remyelinate During Sensory Appendage Regeneration

    PubMed Central

    Moore, Alex C.; Mark, Tiffany E.; Hogan, Ann K.; Topczewski, Jacek; LeClair, Elizabeth E.

    2013-01-01

    Myelination is a cellular adaptation allowing rapid conduction along axons. We have investigated peripheral axons of the zebrafish maxillary barbel (ZMB), an optically clear sensory appendage. Each barbel carries taste buds, solitary chemosensory cells, and epithelial nerve endings, all of which regenerate after amputation (LeClair and Topczewski [2010] PLoS One 5:e8737). The ZMB contains axons from the facial nerve; however, myelination within the barbel itself has not been established. Transcripts of myelin basic protein (mbp) are expressed in normal and regenerating adult barbels, indicating activity in both maintenance and repair. Myelin was confirmed in situ by using toluidine blue, an anti-MBP antibody, and transmission electron microscopy (TEM). The adult ZMB contains ~180 small-diameter axons (<2 μm), approximately 60% of which are myelinated. Developmental myelination was observed via whole-mount immunohistochemistry 4-6 weeks postfertilization, showing myelin sheaths lagging behind growing axons. Early-regenerating axons (10 days postsurgery), having no or few myelin layers, were disorganized within a fibroblast-rich collagenous scar. Twenty-eight days postsurgery, barbel axons had grown out several millimeters and were organized with compact myelin sheaths. Fiber types and axon areas were similar between normal and regenerated tissue; within 4 weeks, regenerating axons restored ~85% of normal myelin thickness. Regenerating barbels express multiple promyelinating transcription factors (sox10, oct6 = pou3f1; krox20a/b = egr2a/b) typical of Schwann cells. These observations extend our understanding of the zebrafish peripheral nervous system within a little-studied sensory appendage. The accessible ZMB provides a novel context for studying axon regeneration, Schwann cell migration, and remyelination in a model vertebrate. PMID:22592645

  7. Activity-dependent expression of neuronal PAS domain-containing protein 4 (npas4a) in the developing zebrafish brain

    PubMed Central

    Klarić, Thomas; Lardelli, Michael; Key, Brian; Koblar, Simon; Lewis, Martin

    2014-01-01

    In rodents, the Npas4 gene has recently been identified as being an important regulator of synaptic plasticity and memory. Homologs of Npas4 have been found in invertebrate species though their functions appear to be too divergent for them to be studied as a proxy for the mammalian proteins. The aim of this study, therefore, was to ascertain the suitability of the zebrafish as a model organism for investigating the function of Npas4 genes. We show here that the expression and regulation of the zebrafish Npas4 homolog, npas4a, is remarkably similar to that of the rodent Npas4 genes. As in mammals, expression of the zebrafish npas4a gene is restricted to the brain where it is up-regulated in response to neuronal activity. Furthermore, we also show that knockdown of npas4a during embryonic development results in a number of forebrain-specific defects including increased apoptosis and misexpression of the forebrain marker genes dlx1a and shha. Our work demonstrates that the zebrafish is a suitable model organism for investigating the role of the npas4a gene and one that is likely to provide valuable insights into the function of the mammalian homologs. Furthermore, our findings highlight a potential role for npas4a in forebrain development. PMID:25538572

  8. Depleted uranium induces sex- and tissue-specific methylation patterns in adult zebrafish.

    PubMed

    Gombeau, Kewin; Pereira, Sandrine; Ravanat, Jean-Luc; Camilleri, Virginie; Cavalie, Isabelle; Bourdineaud, Jean-Paul; Adam-Guillermin, Christelle

    2016-04-01

    We examined the effects of chronic exposure to different concentrations (2 and 20 μg L(-)(1)) of environmentally relevant waterborne depleted uranium (DU) on the DNA methylation patterns both at HpaII restriction sites (5'-CCGG-3') and across the whole genome in the zebrafish brain, gonads, and eyes. We first identified sex-dependent differences in the methylation level of HpaII sites after exposure. In males, these effects were present as early as 7 days after exposure to 20 μg L(-)(1) DU, and were even more pronounced in the brain, gonads, and eyes after 24 days. However, in females, hypomethylation was only observed in the gonads after exposure to 20 μg L(-)(1) DU for 24 days. Sex-specific effects of DU were also apparent at the whole-genome level, because in males, exposure to 20 μg L(-)(1) DU for 24 days resulted in cytosine hypermethylation in the brain and eyes and hypomethylation in the gonads. In contrast, in females, hypermethylation was observed in the brain after exposure to both concentrations of DU for 7 days. Based on our current knowledge of uranium toxicity, several hypotheses are proposed to explain these findings, including the involvement of oxidative stress, alteration of demethylation enzymes and the calcium signaling pathway. This study reports, for the first time, the sex- and tissue-specific epigenetic changes that occur in a nonhuman organism after exposure to environmentally relevant concentrations of uranium, which could induce transgenerational epigenetic effects. PMID:26829549

  9. Neurobehavioural Changes and Brain Oxidative Stress Induced by Acute Exposure to GSM900 Mobile Phone Radiations in Zebrafish (Danio rerio)

    PubMed Central

    Nirwane, Abhijit; Sridhar, Vinay; Majumdar, Anuradha

    2016-01-01

    The impact of mobile phone (MP) radiation on the brain is of specific interest to the scientific community and warrants investigations, as MP is held close to the head. Studies on humans and rodents revealed hazards MP radiation associated such as brain tumors, impairment in cognition, hearing etc. Melatonin (MT) is an important modulator of CNS functioning and is a neural antioxidant hormone. Zebrafish has emerged as a popular model organism for CNS studies. Herein, we evaluated the impact of GSM900MP (GSM900MP) radiation exposure daily for 1 hr for 14 days with the SAR of 1.34W/Kg on neurobehavioral and oxidative stress parameters in zebrafish. Our study revealed that, GSM900MP radiation exposure, significantly decreased time spent near social stimulus zone and increased total distance travelled, in social interaction test. In the novel tank dive test, the GSM900MP radiation exposure elicited anxiety as revealed by significantly increased time spent in bottom half; freezing bouts and duration and decreased distance travelled, average velocity, and number of entries to upper half of the tank. Exposed zebrafish spent less time in the novel arm of the Y-Maze, corroborating significant impairment in learning as compared to the control group. Exposure decreased superoxide dismutase (SOD), catalase (CAT) activities whereas, increased levels of reduced glutathione (GSH) and lipid peroxidation (LPO) was encountered showing compromised antioxidant defense. Treatment with MT significantly reversed the above neurobehavioral and oxidative derangements induced by GSM900MP radiation exposure. This study traced GSM900MP radiation exposure induced neurobehavioral aberrations and alterations in brain oxidative status. Furthermore, MT proved to be a promising therapeutic candidate in ameliorating such outcomes in zebrafish. PMID:27123163

  10. Neurobehavioural Changes and Brain Oxidative Stress Induced by Acute Exposure to GSM900 Mobile Phone Radiations in Zebrafish (Danio rerio).

    PubMed

    Nirwane, Abhijit; Sridhar, Vinay; Majumdar, Anuradha

    2016-04-01

    The impact of mobile phone (MP) radiation on the brain is of specific interest to the scientific community and warrants investigations, as MP is held close to the head. Studies on humans and rodents revealed hazards MP radiation associated such as brain tumors, impairment in cognition, hearing etc. Melatonin (MT) is an important modulator of CNS functioning and is a neural antioxidant hormone. Zebrafish has emerged as a popular model organism for CNS studies. Herein, we evaluated the impact of GSM900MP (GSM900MP) radiation exposure daily for 1 hr for 14 days with the SAR of 1.34W/Kg on neurobehavioral and oxidative stress parameters in zebrafish. Our study revealed that, GSM900MP radiation exposure, significantly decreased time spent near social stimulus zone and increased total distance travelled, in social interaction test. In the novel tank dive test, the GSM900MP radiation exposure elicited anxiety as revealed by significantly increased time spent in bottom half; freezing bouts and duration and decreased distance travelled, average velocity, and number of entries to upper half of the tank. Exposed zebrafish spent less time in the novel arm of the Y-Maze, corroborating significant impairment in learning as compared to the control group. Exposure decreased superoxide dismutase (SOD), catalase (CAT) activities whereas, increased levels of reduced glutathione (GSH) and lipid peroxidation (LPO) was encountered showing compromised antioxidant defense. Treatment with MT significantly reversed the above neurobehavioral and oxidative derangements induced by GSM900MP radiation exposure. This study traced GSM900MP radiation exposure induced neurobehavioral aberrations and alterations in brain oxidative status. Furthermore, MT proved to be a promising therapeutic candidate in ameliorating such outcomes in zebrafish. PMID:27123163

  11. Reversible Deafferentation of the Adult Zebrafish Olfactory Bulb Affects Glomerular Distribution and Olfactory-Mediated Behavior

    PubMed Central

    Paskin, Taylor R.; Byrd-Jacobs, Christine A.

    2012-01-01

    The olfactory system is a useful model for studying central nervous system recovery from damage due to its neuroplasticity. We recently developed a novel method of deafferentation by repeated exposure of Triton X-100 to the olfactory organ of adult zebrafish. This long-term, reversible method of deafferentation allows both degeneration and regeneration to be observed in the olfactory bulb. The aim of the present study is to examine olfactory bulb innervation, glomerular patterns, and olfactory-mediated behavior with repeated Triton X-100 treatment and the potential for recovery following cessation of treatment. Olfactory bulbs of control, chronic-treated, and recovery animals were examined for the presence or absence of glomeruli that have been identified in the zebrafish glomerular map. Following chronic treatment, the number of glomeruli was dramatically reduced; however, partial innervation remained in the lateral region of the bulb. When animals were given time to recover, complete glomerular distribution returned. A behavioral assay was developed to determine if innervation remaining correlated with behavior of the fish. Chronic-treated fish did not respond to odorants involved with social behavior but continued to react to odorants that mediate feeding behavior. Following recovery, responses to odorants involved with social behavior returned. The morphological and behavioral effects of chronic Triton X-100 treatment in the olfactory system suggest there may be differential susceptibility or resistance to external damage in a subset of sensory neurons. The results of this study demonstrate the remarkable regenerative ability of the olfactory system following extensive and long-term injury. PMID:22963994

  12. Reversible deafferentation of the adult zebrafish olfactory bulb affects glomerular distribution and olfactory-mediated behavior.

    PubMed

    Paskin, Taylor R; Byrd-Jacobs, Christine A

    2012-12-01

    The olfactory system is a useful model for studying central nervous system recovery from damage due to its neuroplasticity. We recently developed a novel method of deafferentation by repeated exposure of Triton X-100 to the olfactory organ of adult zebrafish. This long-term, reversible method of deafferentation allows both degeneration and regeneration to be observed in the olfactory bulb. The aim of the present study is to examine olfactory bulb innervation, glomerular patterns, and olfactory-mediated behavior with repeated Triton X-100 treatment and the potential for recovery following cessation of treatment. Olfactory bulbs of control, chronic-treated, and recovery animals were examined for the presence or absence of glomeruli that have been identified in the zebrafish glomerular map. Following chronic treatment, the number of glomeruli was dramatically reduced; however, partial innervation remained in the lateral region of the bulb. When animals were given time to recover, complete glomerular distribution returned. A behavioral assay was developed to determine if innervation remaining correlated with behavior of the fish. Chronic-treated fish did not respond to odorants involved with social behavior but continued to react to odorants that mediate feeding behavior. Following recovery, responses to odorants involved with social behavior returned. The morphological and behavioral effects of chronic Triton X-100 treatment in the olfactory system suggest there may be differential susceptibility or resistance to external damage in a subset of sensory neurons. The results of this study demonstrate the remarkable regenerative ability of the olfactory system following extensive and long-term injury. PMID:22963994

  13. A Model of Excitotoxic Brain Injury in Larval Zebrafish: Potential Application for High-Throughput Drug Evaluation to Treat Traumatic Brain Injury.

    PubMed

    McCutcheon, Victoria; Park, Eugene; Liu, Elaine; Wang, Youdong; Wen, Xiao-Yan; Baker, Andrew J

    2016-06-01

    Traumatic brain injury (TBI) is a leading cause of death and morbidity with no effective therapeutic treatments for secondary injury. Preclinical drug evaluation in rodent models of TBI is a lengthy process. In this regard, the zebrafish has numerous advantages to address the technical and time-dependent obstacles associated with drug evaluation. We developed a reproducible brain injury using glutamate excitoxicity in zebrafish larvae, a known initiator of delayed cell death in TBI. Glutamate challenge resulted in dose-dependent lethality over an 84-h observation period. We report significant decrease in locomotion (p < 0.0001) and mean velocity (p < 0.001) with 10 μM glutamate application as measured through automated 96-well plate behavioral analysis. Application of the NMDA receptor antagonist MK-801 (400 nM) or the calpain inhibitor, MDL-28170 (20 μM), resulted in significant recovery of locomotor function. A secA5-YFP transgenic line was used to visualize the localization of cell death due to glutamate exposure in vivo using confocal fluorescence microscopy. Our results indicate that zebrafish larvae exhibit responses to excitotoxic injury and pharmacotherapeutic intervention with pathophysiological relevance to mammalian excitotoxic brain injury. This system has potential to be applied as a high-throughput drug screening model to quickly identify candidate lead compounds for further evaluation. PMID:27028704

  14. Malformation of certain brain blood vessels caused by TCDD activation of Ahr2/Arnt1 signaling in developing zebrafish

    PubMed Central

    Teraoka, Hiroki; Ogawa, Akira; Kubota, Akira; Stegeman, John J.; Peterson, Richard E.; Hiraga, Takeo

    2011-01-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) causes various signs of toxicity in early life stages of vertebrates through activation of the aryl hydrocarbon receptor (AHR). The AHR also plays important roles in normal development in mice, and AHR-/- mice show abnormal development of vascular structures in various blood vessels. Our previous studies revealed that Ahr type 2 (Ahr2) activation by TCDD and β-naphthoflavone (BNF) caused a significant decrease in blood flow in the dorsal midbrain of zebrafish embryos. Here we report effects of TCDD exposure on the morphology of some blood vessels in the head of developing zebrafish. TCDD caused concentration-dependent anatomical rearrangements in the shape of the prosencephalic artery in zebrafish larvae. In contrast, no major vascular defects were recognized in the trunk and tail regions following exposure to TCDD at least at the concentrations used. Essentially, the same observations were also confirmed in BNF-exposed larvae. Knock-down of either Ahr2 or Ahr nuclear translocator type 1 (Arnt1) by morpholino oligonucleotides (MOs) protected larvae against abnormal shape of the prosencephalic artery caused by TCDD and BNF. On the other hand, knock-down of Ahr2 or Arnt1 in vehicle-exposed zebrafish larvae had no clear effect on morphology of the prosencephalic artery or trunk vessels. Ascorbic acid, an antioxidant, protected against the TCDD-induced decrease in blood flow through the prosencephalic artery, but not the abnormal morphological changes in the shape of this artery. These results indicate that activation of Ahr2/Arnt1 pathway by TCDD and BNF affects the shape of certain blood vessels in the brain of developing zebrafish. PMID:20554057

  15. Differential requirement for irf8 in formation of embryonic and adult macrophages in zebrafish

    SciTech Connect

    Shiau, Celia E.; Kaufman, Zoe; Meireles, Ana M.; Talbot, William S.

    2015-01-23

    Interferon regulatory factor 8 (Irf8) is critical for mammalian macrophage development and innate immunity, but its role in teleost myelopoiesis remains incompletely understood. Specifically, genetic tools to analyze the role of irf8 in zebrafish macrophage development at larval and adult stages are lacking. In this study, we generated irf8 null mutants in zebrafish using TALEN-mediated targeting. Our analysis defines different requirements for irf8 at different stages. irf8 is required for formation of all macrophages during primitive and transient definitive hematopoiesis, but not during adult-phase definitive hematopoiesis starting at 5-6 days postfertilization. At early stages, irf8 mutants have excess neutrophils and excess cell death in pu.1-expressing myeloid cells. Macrophage fates were recovered in irf8 mutants after wildtype irf8 expression in neutrophil and macrophage lineages, suggesting that irf8 regulates macrophage specification and survival. In juvenile irf8 mutant fish, mature macrophages are present, but at numbers significantly reduced compared to wildtype, indicating an ongoing requirement for irf8 after embryogenesis. As development progresses, tissue macrophages become apparent in zebrafish irf8 mutants, with the possible exception of microglia. Our study defines distinct requirement for irf8 in myelopoiesis before and after transition to the adult hematopoietic system.

  16. Feed and Feeding Regime Affect Growth Rate and Gonadosomatic Index of Adult Zebrafish (Danio Rerio)

    PubMed Central

    Law, Sheran Hiu Wan

    2013-01-01

    Abstract A 5-week study was conducted to evaluate commercially available Artemia, Ziegler zebrafish diet, and Calamac diet fed in five different feeding regimes on the growth and reproductive development of 7-month-old zebrafish. Zebrafish were fed to satiation three times daily during the normal work week and twice daily during the weekend and holidays. Zebrafish in dietary groups CCC (Calamac three times daily) and CCA (Calamac twice daily, Artemia once daily) had a significantly (p<0.05) greater weight gain and specific growth rate as compared to all other dietary groups. Male zebrafish in dietary group 5 had significantly larger gonadosomatic index (GSI) values than all other groups, while female zebrafish in dietary group CCC had significantly larger GSI values than all other groups. No differences in the fatty acid content of female gonads were detected. Zebrafish fed solely Artemia had the greatest weight loss and lowest GSI values. Preliminary evidence of protein sparing in zebrafish is reported. Collectively, this study sheds more light into the effects of the use of commercially available feeds and feeding regime on the rearing of zebrafish. PMID:23902461

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

    PubMed

    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. PMID:23590400

  18. Towards a Comprehensive Catalog of Zebrafish Behavior 1.0 and Beyond

    PubMed Central

    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

    2013-01-01

    Abstract 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. PMID:23590400

  19. Behind melanocortin antagonist overexpression in the zebrafish brain: A behavioral and transcriptomic approach.

    PubMed

    Guillot, Raúl; Cortés, Raúl; Navarro, Sandra; Mischitelli, Morena; García-Herranz, Víctor; Sánchez, Elisa; Cal, Laura; Navarro, Juan Carlos; Míguez, Jesús M; Afanasyev, Sergey; Krasnov, Aleksei; Cone, Roger D; Rotllant, Josep; Cerdá-Reverter, Jose Miguel

    2016-06-01

    Melanocortin signaling is regulated by the binding of naturally occurring antagonists, agouti-signaling protein (ASIP) and agouti-related protein (AGRP) that compete with melanocortin peptides by binding to melanocortin receptors to regulate energy balance and growth. Using a transgenic model overexpressing ASIP, we studied the involvement of melanocortin system in the feeding behaviour, growth and stress response of zebrafish. Our data demonstrate that ASIP overexpression results in enhanced growth but not obesity. The differential growth is explained by increased food intake and feeding efficiency mediated by a differential sensitivity of the satiety system that seems to involve the cocaine- and amphetamine- related transcript (CART). Stress response was similar in both genotypes. Brain transcriptome of transgenic (ASIP) vs wild type (WT) fish was compared using microarrays. WT females and males exhibited 255 genes differentially expressed (DEG) but this difference was reduced to 31 after ASIP overexpression. Statistical analysis revealed 1122 DEG when considering only fish genotype but 1066 and 981 DEG when comparing ASIP males or females with their WT counterparts, respectively. Interaction between genotype and sex significantly affected the expression of 97 genes. Several neuronal systems involved in the control of food intake were identified which displayed a differential expression according to the genotype of the fish that unravelling the flow of melanocortinergic information through the central pathways that controls the energy balance. The information provided herein will help to elucidate new central systems involved in control of obesity and should be of invaluable use for sustaining fish production systems. PMID:27156808

  20. Brain-wide mapping of neural activity controlling zebrafish exploratory locomotion

    PubMed Central

    Dunn, Timothy W; Mu, Yu; Narayan, Sujatha; Randlett, Owen; Naumann, Eva A; Yang, Chao-Tsung; Schier, Alexander F

    2016-01-01

    In the absence of salient sensory cues to guide behavior, animals must still execute sequences of motor actions in order to forage and explore. How such successive motor actions are coordinated to form global locomotion trajectories is unknown. We mapped the structure of larval zebrafish swim trajectories in homogeneous environments and found that trajectories were characterized by alternating sequences of repeated turns to the left and to the right. Using whole-brain light-sheet imaging, we identified activity relating to the behavior in specific neural populations that we termed the anterior rhombencephalic turning region (ARTR). ARTR perturbations biased swim direction and reduced the dependence of turn direction on turn history, indicating that the ARTR is part of a network generating the temporal correlations in turn direction. We also find suggestive evidence for ARTR mutual inhibition and ARTR projections to premotor neurons. Finally, simulations suggest the observed turn sequences may underlie efficient exploration of local environments. DOI: http://dx.doi.org/10.7554/eLife.12741.001 PMID:27003593

  1. The UV-absorber benzophenone-4 alters transcripts of genes involved in hormonal pathways in zebrafish (Danio rerio) eleuthero-embryos and adult males

    SciTech Connect

    Zucchi, Sara; Bluethgen, Nancy; Ieronimo, Andrea; Fent, Karl

    2011-01-15

    Benzophenone-4 (BP-4) is frequently used as UV-absorber in cosmetics and materials protection. Despite its frequent detection in the aquatic environment potential effects on aquatic life are unknown. In this study, we evaluate the effects of BP-4 in eleuthero-embryos and in the liver, testis and brain of adult male fish on the transcriptional level by focusing on target genes involved in hormonal pathways to provide a more complete toxicological profile of this important UV-absorber. Eleuthero-embryos and males of zebrafish were exposed up to 3 days after hatching and for 14 days, respectively, to BP-4 concentrations between 30 and 3000 {mu}g/L. In eleuthero-embryos transcripts of vtg1, vtg3, esr1, esr2b, hsd17ss3, cyp19b cyp19a, hhex and pax8 were induced at 3000 {mu}g/L BP-4, which points to a low estrogenic activity and interference with early thyroid development, respectively. In adult males BP-4 displayed multiple effects on gene expression in different tissues. In the liver vtg1, vtg3, esr1 and esr2b were down-regulated, while in the brain, vtg1, vtg3 and cyp19b transcripts were up-regulated. In conclusion, the transcription profile revealed that BP-4 interferes with the expression of genes involved in hormonal pathways and steroidogenesis. The effects of BP-4 differ in life stages and adult tissues and point to an estrogenic activity in eleuthero-embryos and adult brain, and an antiestrogenic activity in the liver. The results indicate that BP-4 interferes with the sex hormone system of fish, which is important for the risk assessment of this UV-absorber.

  2. Spectral-Domain Optical Coherence Tomography as a Noninvasive Method to Assess Damaged and Regenerating Adult Zebrafish Retinas

    PubMed Central

    Bailey, Travis J.; Davis, Darin H.; Vance, Joseph E.; Hyde, David R.

    2012-01-01

    Purpose. These experiments assessed the ability of spectral-domain optical coherence tomography (SD-OCT) to accurately represent the structural organization of the adult zebrafish retina and reveal the dynamic morphologic changes during either light-induced damage and regeneration of photoreceptors or ouabain-induced inner retinal damage. Methods. Retinas of control dark-adapted adult albino zebrafish were compared with retinas subjected to 24 hours of constant intense light and recovered for up to 8 weeks or ouabain-damaged retinas that recovered for up to 3 weeks. Images were captured and the measurements of retinal morphology were made by SD-OCT, and then compared with those obtained by histology of the same eyes. Results. Measurements between SD-OCT and histology were very similar for the undamaged, damaged, and regenerating retinas. Axial measurements of SD-OCT also revealed vitreal morphology that was not readily visualized by histology. Conclusions. SD-OCT accurately represented retinal lamination and photoreceptor loss and recovery during light-induced damage and subsequent regeneration. SD-OCT was less accurate at detecting the inner nuclear layer in ouabain-damaged retinas, but accurately detected the undamaged outer nuclear layer. Thus, SD-OCT provides a noninvasive and quantitative method to assess the morphology and the extent of damage and repair in the zebrafish retina. PMID:22499984

  3. Acid-sensing ion channel immunoreactivities in the cephalic neuromasts of adult zebrafish.

    PubMed

    Abbate, F; Madrigrano, M; Scopitteri, T; Levanti, M; Cobo, J L; Germanà, A; Vega, J A; Laurà, R

    2016-09-01

    The neuromasts are the morphofunctional unit of the lateral line system serving as mechanosensors for water flow and movement. The mechanisms underlying the detection of the mechanical stimuli in the vertebrate mechanosensory cells remain poorly understood at the molecular level, and no information is available on neuromasts. Mechanotransduction is the conversion of a mechanical stimulus into an electrical signal via activation of ion channels. The acid-sensing ion channels (ASICs) are presumably involved in mechanosensation, and therefore are expected to be expressed in the mechanoreceptors. Here we used immunohistochemistry to investigate the occurrence and distribution of ASICs in the cephalic neuromasts of the adult zebrafish. Specific immunoreactivity for ASIC1 and ASIC4 was detected in the hair cells while ASIC2 was restricted to the nerves supplying neuromasts. Moreover, supporting and mantle cells; i.e., the non-sensory cells of the neuromasts, also displayed ASIC4. For the first time, these results demonstrate the presence of the putative mechanoproteins ASIC1, ASIC2 and ASIC4 in neuromasts, suggesting a role for these ion channels in mechanosensation. PMID:27443821

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

    PubMed

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

    2013-11-01

    The neurotoxic effects of a commercial formulation of Azadirachta indica A. Juss, also called neem or nim, in adult zebrafish were determined using behavioral models. General activity, anxiety-like effects, and learning and memory in a passive avoidance task were assessed after exposure to 20 or 40 μl/L neem. The results showed that 20 μl/L neem reduced the number of runs. Both neem concentrations increased the number of climbs to the water surface, and 40 μl/L increased the number of tremors. In the anxiety test, the 20 μl/L dose increased the number of entries in the light side compared with controls, but the latency to enter the dark side and the freezing behavior in this side did not changed. In relation to controls, the 40 μl/L neem reduced the latency to enter in the light side, did not change the number of entries in this side and increased freezing behavior in the light side. In the passive avoidance test, pre-training and pre-test neem exposure to 40 μl/L decreased the response to the learning task. Thus, no impairment was observed in this behavioral test. We conclude that neem reduced general activity and increased anxiety-like behavior but did not affect learning and memory. PMID:24211596

  5. Acid-sensing ion channels (ASICs) in the taste buds of adult zebrafish.

    PubMed

    Viña, E; Parisi, V; Cabo, R; Laurà, R; López-Velasco, S; López-Muñiz, A; García-Suárez, O; Germanà, A; Vega, J A

    2013-03-01

    In detecting chemical properties of food, different molecules and ion channels are involved including members of the acid-sensing ion channels (ASICs) family. Consistently ASICs are present in sensory cells of taste buds of mammals. In the present study the presence of ASICs (ASIC1, ASIC2, ASIC3 and ASIC4) was investigated in the taste buds of adult zebrafish (zASICs) using Western blot and immunohistochemistry. zASIC1 and zASIC3 were regularly absent from taste buds, whereas faint zASIC2 and robust zASIC4 immunoreactivities were detected in sensory cells. Moreover, zASIC2 also immunolabelled nerves supplying taste buds. The present results demonstrate for the first time the presence of zASICs in taste buds of teleosts, with different patterns to that occurring in mammals, probably due to the function of taste buds in aquatic environment and feeding. Nevertheless, the role of zASICs in taste remains to be demonstrated. PMID:23328442

  6. Persistent Adult Zebrafish Behavioral Deficits Results from Acute Embryonic Exposure to Gold Nanoparticles

    PubMed Central

    Truong, Lisa; Saili, Katerine S.; Miller, John M.; Hutchison, James E.; Tanguay, Robert L.

    2011-01-01

    As the number of products containing nanomaterials increase, human exposure to nanoparticles (NPs) is unavoidable. Presently, few studies focus on the potential long-term consequences of developmental NP exposure. In this study, zebrafish embryos were acutely exposed to three gold NPs that possess functional groups with differing surface charge. Embryos were exposed to 50 μg/mL of 1.5 nm gold nanoparticles (AuNPs) possessing negatively charged 2-mercaptoethanesulfonic acid (MES) or neutral 2-(2-(2-mercaptoethoxy)ethoxy)ethanol (MEEE) ligands or 10 μg/mL of the AuNPs possessing positively charged trimethylammoniumethanethiol (TMAT). Both MES- and TMAT-AuNP exposed embryos exhibited hypo-locomotor activity, while those exposed to MEEE-AuNPs did not. A subset of embryos that were exposed to 1.5 nm MES- and TMAT-AuNPs during development from 6–120 hours post fertilization were raised to adulthood. Behavioral abnormalities and the number of survivors into adulthood were evaluated at 122 days post fertilization. We found that both treatments induced abnormal startle behavior following a tap stimulus. However, the MES-AuNPs exposed group also exhibited abnormal adult behavior in the light and had a lower survivorship into adulthood. This study demonstrates that acute, developmental exposure to 1.5 nm MES- and TMAT- AuNPs, two NPs differing only in the functional group, affects larval behavior, with behavioral effects persisting into adulthood. PMID:21946249

  7. A rapid throughput approach identifies cognitive deficits in adult zebrafish from developmental exposure to polybrominated flame retardants

    PubMed Central

    Truong, Lisa; Mandrell, David; Mandrell, Rick; Simonich, Michael; Tanguay, Robert L.

    2014-01-01

    A substantial body of evidence has correlated the human body burdens of some polybrominated diphenyl ether (PBDE) flame retardants with cognitive and other behavioral deficits. Adult zebrafish exhibit testable learning and memory, making them an increasingly attractive model for neurotoxicology. Our goal was to develop a rapid throughput means of identifying the cognitive impact of developmental exposure to flame retardants in the zebrafish model. We exposed embryos from 6 hours post fertilization to 5 days post fertilization to either PBDE 47 (0.1 uM), PBDE 99 (0.1 uM) or PBDE 153 (0.1 uM), vehicle (0.1% DMSO), or embryo medium (EM). The larvae were grown to adulthood and evaluated for the rate at which they learned an active-avoidance response in an automated shuttle box array. Zebrafish developmentally exposed to PBDE 47 learned the active avoidance paradigm significantly faster than the 0.1% DMSO control fish (P < 0.0001), but exhibited significantly poorer performance when retested suggestive of impaired memory retention or altered neuromotor activity. Learning in the PBDE 153 group was not significantly different from the DMSO group. Developmental exposure to 0.1% DMSO impaired adult active avoidance learning relative to the sham group (n = 39; P < 0.0001). PBDE 99 prevented the DMSO effect, yielding a learning rate not significantly different from the sham group (n = 36; P > 0.9). Our results underscore the importance of vehicle choice in accurately assessing chemical effects on behavior. Active avoidance response in zebrafish is an effective model of learning that, combined with automated shuttle box testing, will provide a highly efficient platform for evaluating persistent neurotoxic hazard from many chemicals. PMID:24674958

  8. Molecular analysis, developmental function and heavy metal-induced expression of ABCC5 in zebrafish.

    PubMed

    Long, Yong; Li, Qing; Li, Jie; Cui, Zongbin

    2011-01-01

    ABCC5/MRP5 is an organic anion transporter that participates in tissue defense and cellular signal transduction through efflux of anticancer drugs, toxicants and a second messenger cGMP, but its physiological functions in zebrafish remain to be defined. Herein, we report the characterization, spatiotemporal expression and developmental function of zebrafish ABCC5 and its transcriptional responses to heavy metals. Zebrafish abcc5 gene is located on chromosome 18 and comprised of 28 exons. The deduced polypeptide of zebrafish ABCC5 consists of 1426 amino acids, which shares high sequence identity with those from other species. Zebrafish abcc5 is maternally expressed and its transcripts are mainly distributed in brain, lens, liver and intestine of developing embryos. In adults, zebrafish abcc5 is extensively expressed, at higher levels in testis, brain, eye, ovary, intestine and kidney, but at relatively lower levels in gill, liver, heart and muscle. Blockage of endogenous ABCC5 activity by its dominant-negative led to the developmental retardation of zebrafish embryos in which activity of p21 signaling was markedly stimulated and cellular cGMP content was significantly increased. In addition, expression of abcc5 in ZF4 cells and zebrafish embryos was significantly induced by cadmium (Cd), lead (Pb), mercury (Hg) or arsenic (As). The induced expression of ABCC5 by heavy metals was mainly detected in the liver of embryos at 96-h post-fertilization (hpf). In adult zebrafish, expression of abcc5 in brain, intestine, liver, kidney and ovary was significantly induced by one or more of these heavy metals. Furthermore, overexpression of ABCC5 attenuated the toxicity of Cd to zebrafish embryos, but did not affect the toxicity of Hg or As. Thus, ABCC5 is likely to play an active role in embryonic development and heavy metal detoxification through the export of second messenger molecules and toxicants out of cells in zebrafish. PMID:20869459

  9. Midkine-a Protein Localization in the Developing and Adult Retina of the Zebrafish and Its Function During Photoreceptor Regeneration

    PubMed Central

    Taylor, Scott; Thummel, Ryan; Hitchcock, Peter F.

    2015-01-01

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

  10. Molecular characterization and functions of zebrafish ABCC2 in cellular efflux of heavy metals.

    PubMed

    Long, Yong; Li, Qing; Zhong, Shan; Wang, Youhui; Cui, Zongbin

    2011-05-01

    Multidrug-resistance associated protein 2 (MRP2/ABCC2) plays crucial roles in bile formation and detoxification by transporting a wide variety of endogenous compounds and xenobiotics, but its functions in zebrafish (Danio rerio) remain to be characterized. In this study, we obtained the full-length cDNA of zebrafish abcc2, analyzed its expression in developing embryos and adult tissues, investigated its transcriptional response to heavy metals, and evaluated its roles in efflux of heavy metals including cadmium, mercury and lead. Zebrafish abcc2 gene is located on chromosome 13 and composed of 32 exons. The deduced polypeptide of zebrafish ABCC2 consists of 1567 amino acids and possesses most of functional domains and critical residues defined in human ABCC2. Zebrafish abcc2 gene is not maternally expressed and its earliest expression was detected in embryos at 72hpf. In larval zebrafish, abcc2 gene was found to be exclusively expressed in liver, intestine and pronephric tubules. In adult zebrafish, the highest expression of abcc2 gene was found in intestine followed by those in liver and kidney, while relative low expression was detected in brain and muscle. Expression of abcc2 in excretory organs including kidney, liver and intestine of zebrafish larvae was induced by exposure to 0.5μM mercury or 5μM lead. Moreover, exposure to 0.125-1μM of mercury or lead also significantly induced abcc2 expression in these excretory organs of adult zebrafish. Furthermore, overexpression of zebrafish ABCC2 in ZF4 cells and zebrafish embryos decreased the cellular accumulation of heavy metals including cadmium, mercury and lead as determined by MRE (metal responsive element)- or EPRE (electrophile response element)-driven luciferase reporters and atomic absorption spectrometry. These results suggest that zebrafish ABCC2/MRP2 is capable of effluxing heavy metals from cells and may play important roles in the detoxification of toxic metals. PMID:21266201

  11. Experience-Dependent Neural Plasticity in the Adult Damaged Brain

    ERIC Educational Resources Information Center

    Kerr, Abigail L.; Cheng, Shao-Ying; Jones, Theresa A.

    2011-01-01

    Behavioral experience is at work modifying the structure and function of the brain throughout the lifespan, but it has a particularly dramatic influence after brain injury. This review summarizes recent findings on the role of experience in reorganizing the adult damaged brain, with a focus on findings from rodent stroke models of chronic upper…

  12. Decreased thyroid hormone signaling accelerates the reinnervation of the optic tectum following optic nerve crush in adult zebrafish.

    PubMed

    Bhumika, Stitipragyan; Lemmens, Kim; Vancamp, Pieter; Moons, Lieve; Darras, Veerle M

    2015-09-01

    The regenerative capacity of the adult mammalian central nervous system (CNS) is poor and finding ways to stimulate long distance axonal regeneration in humans remains a challenge for neuroscientists. Thyroid hormones, well known for their key function in CNS development and maturation, more recently also emerged as molecules influencing regeneration. While several studies investigated their influence on peripheral nerve regeneration, in vivo studies on their role in adult CNS regeneration remain scarce. We therefore investigated the effect of lowering T3 signaling on the regeneration of the optic nerve (ON) following crush in zebrafish, a species where full recovery occurs spontaneously. Adult zebrafish were exposed to iopanoic acid (IOP), which lowered intracellular 3,5,3'-triiodothyronine (T3) availability, or to the thyroid hormone receptor β antagonist methylsulfonylnitrobenzoate (C1). Both treatments accelerated optic tectum (OT) reinnervation. At 7days post injury (7dpi) there was a clear increase in the biocytin labeled area in the OT following anterograde tracing as well as an increased immunostaining of Gap43, a protein expressed in outgrowing axons. This effect was attenuated by T3 supplementation to IOP-treated fish. ON crush induced very limited cell death and proliferation at the level of the retina in control, IOP- and C1-treated fish. The treatments also had no effect on the mRNA upregulation of the regeneration markers gap43, tub1a, and socs3b at the level of the retina at 4 and 7dpi. We did, however, find a correlation between the accelerated OT reinnervation and a more rapid resolution of microglia/macrophages in the ON and the OT of IOP-treated fish. Taken together these data indicate that lowering T3 signaling accelerates OT reinnervation following ON crush in zebrafish and that this is accompanied by a more rapid resolution of the inflammatory response. PMID:25913150

  13. Chronic PFOS exposures induce life stage-specific behavioral deficits in adult zebrafish and produce malformations in F1 offspring

    PubMed Central

    Chen, Jiangfei; Huang, Changjiang; Das, Siba R.; La Du, Jane; Corvi, Margaret M.; Bai, Chenglian; Chen, Yuanhong; Tanguay, Robert L.; Dong, Qiaoxiang

    2014-01-01

    Perfluorooctanesulphonicacid (PFOS) is an organic contaminant that is ubiquitous in the environment, wildlife, and humans. Few studies have assessed the effects of chronic PFOS exposure on central nervous system function in aquatic organisms. The present study defined the behavioral effects of varying life span chronic exposures to low dose PFOS in zebrafish. The zebrafish were treated with vehicle control or 0.5μM PFOS during 1–21, 21–120, or 1–120 day post fertilization (dpf). Chronic PFOS exposure impaired the adult zebrafish behavior mode under the tapping stimulus. The movement speed of 1–120 dpf exposed fish was significantly increased compared with control, while 1–21 and 21–120 dpf exposed groups were not severely affected. PFOS residues in F1 embryos derived from parental exposure during both the 1–120 and 21–120 dpf groups was significantly higher than control, and F1 embryos in these two groups showed obvious malformations, such as uninflated swim bladder (USB) and bent spine (BS). Larvae of the parental exposed to PFOS from 1–21 or 21–120 dpf elicited a higher swim rate than control in both the light and dark periods. Embryos derived from the 1–120 dpf group showed a statistically lower speed in the light period and a higher speed in the dark period as compared with control. Though there is little PFOS residue in 1–21 dpf group, the adverse behavioral effects on both adult and F1 larvae indicate that exposure during the first 21 dpf induce long-term neurobehavior toxicity. Our findings demonstrate that chronic exposure to low dose PFOS in different life stage adversely impacts adult behavior, subsequent offspring malformation, and larval behavior. PMID:23059794

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

    PubMed Central

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

    2015-01-01

    Loss of significant intestinal length from congenital anomaly or disease may lead to short bowel syndrome (SBS); intestinal failure may be partially offset by a gain in epithelial surface area, termed adaptation. Current in vivo models of SBS are costly and technically challenging. Operative times and survival rates have slowed extension to transgenic models. We created a new reproducible in vivo model of SBS in zebrafish, a tractable vertebrate model, to facilitate investigation of the mechanisms of intestinal adaptation. Proximal intestinal diversion at segment 1 (S1, equivalent to jejunum) was performed in adult male zebrafish. SBS fish emptied distal intestinal contents via stoma as in the human disease. After 2 wk, S1 was dilated compared with controls and villus ridges had increased complexity, contributing to greater villus epithelial perimeter. The number of intervillus pockets, the intestinal stem cell zone of the zebrafish increased and contained a higher number of bromodeoxyuridine (BrdU)-labeled cells after 2 wk of SBS. Egf receptor and a subset of its ligands, also drivers of adaptation, were upregulated in SBS fish. Igf has been reported as a driver of intestinal adaptation in other animal models, and SBS fish exposed to a pharmacological inhibitor of the Igf receptor failed to demonstrate signs of intestinal adaptation, such as increased inner epithelial perimeter and BrdU incorporation. We describe a technically feasible model of human SBS in the zebrafish, a faster and less expensive tool to investigate intestinal stem cell plasticity as well as the mechanisms that drive intestinal adaptation. PMID:26089336

  15. Monitoring of Single-Cell Responses in the Optic Tectum of Adult Zebrafish with Dextran-Coupled Calcium Dyes Delivered via Local Electroporation

    PubMed Central

    Kassing, Vanessa

    2013-01-01

    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. PMID:23667529

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

    PubMed

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

    2016-01-01

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

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

    PubMed Central

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

    2016-01-01

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

  18. Matrix metalloproteinases as promising regulators of axonal regrowth in the injured adult zebrafish retinotectal system.

    PubMed

    Lemmens, Kim; Bollaerts, Ilse; Bhumika, Stitipragyan; de Groef, Lies; Van Houcke, Jessie; Darras, Veerle M; Van Hove, Inge; Moons, Lieve

    2016-05-01

    Overcoming the failure of axon regeneration in the mammalian central nervous system (CNS) after injury remains a major challenge, which makes the search for proregenerative molecules essential. Matrix metalloproteinases (MMPs) have been implicated in axonal outgrowth during CNS development and show increased expression levels during vertebrate CNS repair. In mammals, MMPs are believed to alter the suppressive extracellular matrix to become more permissive for axon regrowth. We investigated the role of MMPs in axonal regeneration following optic nerve crush (ONC) in adult zebrafish, which fully recover from such injuries due to a high intrinsic axon growth capacity and a less inhibitory environment. Lowering general retinal MMP activity through intravitreal injections of GM6001 after ONC strongly reduced retinal ganglion cell (RGC) axonal regrowth, without influencing RGC survival. Based on a recently performed transcriptome profiling study, the expression pattern of four MMPs after ONC was determined via combined use of western blotting and immunostainings. Mmp-2 and -13a were increasingly present in RGC somata during axonal regrowth. Moreover, Mmp-2 and -9 became upregulated in regrowing RGC axons and inner plexiform layer (IPL) synapses, respectively. In contrast, after an initial rise in IPL neurites and RGC axons during the injury response, Mmp-14 expression decreased during regeneration. Altogether, a phase-dependent expression pattern for each specific MMP was observed, implicating them in axonal regrowth and inner retina remodeling after injury. In conclusion, these data suggest a novel, neuron-intrinsic function for multiple MMPs in axon regrowth that is distinct from breaking down environmental barriers. J. Comp. Neurol. 524:1472-1493, 2016. © 2015 Wiley Periodicals, Inc. PMID:26509469

  19. Recovery of pigmentation following selective photothermolysis in adult zebrafish skin: clinical implications for laser toning treatment of melasma.

    PubMed

    Kim, Jae Hwan; Kim, Do Hyun; Kim, Ji Hae; Lee, Sang Geun; Kim, Hyeon Soo; Park, Hae Chul; Kim, Il-Hwan

    2012-12-01

    In recent years, laser toning has gained popularity for the treatment of melasma, and tyrosinase inhibitors are conventionally used to prevent its recurrence after this treatment. The effectiveness of this treatment modality, however, is still questionable, and additional in vivo studies are needed to validate the method. In this study, we used adult zebrafish skin as an adult melanocyte regenerative system and examined the simulated human skin response to laser toning. Melanosomes regenerated after selective photothermolysis, and these organelles showed a bi-directional translocation pattern in accordance with the changes of intact melanosome patterns. Furthermore, a tyrosinase inhibitor, 1-phenyl-2-thiourea (PTU), completely blocked melanosome regeneration after laser irradiation, but this inhibitor failed to prevent melanosome regeneration after the medication was discontinued. Finally, arbutin and kojic acid, the commercially available tyrosinase inhibitors, slowed down but did not completely block melanosome regeneration. Based on these findings, we describe the limitations of laser toning treatment of melasma and the combined use of tyrosinase inhibitors. We suggest that melanosomes in adult zebrafish skin can be utilized for studying melanosome regeneration response to laser irradiation and for developing a system to assess the comparative efficacy of melanogenic regulatory compounds. PMID:23057411

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

    PubMed

    Cronan, Mark R; Rosenberg, Allison F; Oehlers, Stefan H; Saelens, Joseph W; Sisk, Dana M; Jurcic Smith, Kristen L; Lee, Sunhee; Tobin, David M

    2015-12-01

    Visualization of infection and the associated host response has been challenging in adult vertebrates. Owing to their transparency, zebrafish larvae have been used to directly observe infection in vivo; however, such larvae have not yet developed a functional adaptive immune system. Cells involved in adaptive immunity mature later and have therefore been difficult to access optically in intact animals. Thus, the study of many aspects of vertebrate infection requires dissection of adult organs or ex vivo isolation of immune cells. Recently, CLARITY and PACT (passive clarity technique) methodologies have enabled clearing and direct visualization of dissected organs. Here, we show that these techniques can be applied to image host-pathogen interactions directly in whole animals. CLARITY and PACT-based clearing of whole adult zebrafish and Mycobacterium tuberculosis-infected mouse lungs enables imaging of mycobacterial granulomas deep within tissue to a depth of more than 1 mm. Using established transgenic lines, we were able to image normal and pathogenic structures and their surrounding host context at high resolution. We identified the three-dimensional organization of granuloma-associated angiogenesis, an important feature of mycobacterial infection, and characterized the induction of the cytokine tumor necrosis factor (TNF) within the granuloma using an established fluorescent reporter line. We observed heterogeneity in TNF induction within granuloma macrophages, consistent with an evolving view of the tuberculous granuloma as a non-uniform, heterogeneous structure. Broad application of this technique will enable new understanding of host-pathogen interactions in situ. PMID:26449262

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

    PubMed Central

    Cronan, Mark R.; Rosenberg, Allison F.; Oehlers, Stefan H.; Saelens, Joseph W.; Sisk, Dana M.; Jurcic Smith, Kristen L.; Lee, Sunhee; Tobin, David M.

    2015-01-01

    ABSTRACT Visualization of infection and the associated host response has been challenging in adult vertebrates. Owing to their transparency, zebrafish larvae have been used to directly observe infection in vivo; however, such larvae have not yet developed a functional adaptive immune system. Cells involved in adaptive immunity mature later and have therefore been difficult to access optically in intact animals. Thus, the study of many aspects of vertebrate infection requires dissection of adult organs or ex vivo isolation of immune cells. Recently, CLARITY and PACT (passive clarity technique) methodologies have enabled clearing and direct visualization of dissected organs. Here, we show that these techniques can be applied to image host-pathogen interactions directly in whole animals. CLARITY and PACT-based clearing of whole adult zebrafish and Mycobacterium tuberculosis-infected mouse lungs enables imaging of mycobacterial granulomas deep within tissue to a depth of more than 1 mm. Using established transgenic lines, we were able to image normal and pathogenic structures and their surrounding host context at high resolution. We identified the three-dimensional organization of granuloma-associated angiogenesis, an important feature of mycobacterial infection, and characterized the induction of the cytokine tumor necrosis factor (TNF) within the granuloma using an established fluorescent reporter line. We observed heterogeneity in TNF induction within granuloma macrophages, consistent with an evolving view of the tuberculous granuloma as a non-uniform, heterogeneous structure. Broad application of this technique will enable new understanding of host-pathogen interactions in situ. PMID:26449262

  2. Efficient Cargo Delivery into Adult Brain Tissue Using Short Cell-Penetrating Peptides

    PubMed Central

    Thomas, Alvin Kuriakose; Bhattarai, Prabesh; Zhang, Yixin; Brand, Michael

    2015-01-01

    Zebrafish brains can regenerate lost neurons upon neurogenic activity of the radial glial progenitor cells (RGCs) that reside at the ventricular region. Understanding the molecular events underlying this ability is of great interest for translational studies of regenerative medicine. Therefore, functional analyses of gene function in RGCs and neurons are essential. Using cerebroventricular microinjection (CVMI), RGCs can be targeted efficiently but the penetration capacity of the injected molecules reduces dramatically in deeper parts of the brain tissue, such as the parenchymal regions that contain the neurons. In this report, we tested the penetration efficiency of five known cell-penetrating peptides (CPPs) and identified two– polyR and Trans – that efficiently penetrate the brain tissue without overt toxicity in a dose-dependent manner as determined by TUNEL staining and L-Plastin immunohistochemistry. We also found that polyR peptide can help carry plasmid DNA several cell diameters into the brain tissue after a series of coupling reactions using DBCO-PEG4-maleimide-based Michael’s addition and azide-mediated copper-free click reaction. Combined with the advantages of CVMI, such as rapidness, reproducibility, and ability to be used in adult animals, CPPs improve the applicability of the CVMI technique to deeper parts of the central nervous system tissues. PMID:25894337

  3. A novel cyclic nucleotide-gated ion channel enriched in synaptic terminals of isotocin neurons in zebrafish brain and pituitary

    PubMed Central

    Khan, Sakina; Perry, Christine; Tetreault, Michelle L.; Henry, Diane; Trimmer, James S.; Zimmerman, Anita L.; Matthews, Gary

    2009-01-01

    Cyclic nucleotide-gated (CNG) channels are nonselective cation channels opened by binding of intracellular cyclic GMP or cyclic AMP. CNG channels mediate sensory transduction in the rods and cones of the retina and in olfactory sensory neurons, but in addition, CNG channels are also expressed elsewhere in the central nervous system, where their physiological roles have not yet been well defined. Besides the CNG channel subtypes that mediate vision and olfaction, zebrafish has an additional subtype, CNGA5, which is expressed almost exclusively in the brain. We have generated CNGA5-specific monoclonal antibodies, which we use here to show that immunoreactivity for CNGA5 channels is highly enriched in synaptic terminals of a discrete set of neurons that project to a subregion of the pituitary, as well as diffusely in the brain and spinal cord. Double labeling with a variety of antibodies against pituitary hormones revealed that CNGA5 is located in the terminals of neuroendocrine cells that secrete the nonapeptide hormone/transmitter isotocin in the neurohypophysis, brain, and spinal cord. Furthermore, we show that CNGA5 channels expressed in Xenopus oocytes are highly permeable to Ca2+, which suggests that the channels are capable of modulating isotocin release in the zebrafish brain and pituitary. Isotocin is the teleost homolog of the mammalian hormone oxytocin, and like oxytocin, it regulates reproductive and social behavior. Therefore, the high calcium permeability of CNGA5 channels and their strategic location in isotocin-secreting synaptic terminals suggest that activation of CNGA5 channels in response to cyclic nucleotide signaling may have wide-ranging neuroendocrine and behavioral effects. PMID:19778592

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

    PubMed Central

    Lin, Li-Jen; Chiang, Chung-Jen; Chao, Yun-Peng; Wang, Shulhn-Der; Chiou, Yu-Ting; Wang, Han-Yu; Kao, Shung-Te

    2016-01-01

    Oral administration of Traditional Chinese Medicine (TCM) by patients is the common way to treat health problems. Zebrafish emerges as an excellent animal model for the pharmacology investigation. However, the oral delivery system of TCM in zebrafish has not been established so far. This issue was addressed by development of alginate microparticles for oral delivery of chuanxiong, a TCM that displays antifibrotic and antiproliferative effects on hepatocytes. The delivery microparticles were prepared from gelification of alginate containing various levels of chuanxiong. The chuanxiong-encapsulated alginate microparticles were characterized for their solubility, structure, encapsulation efficiency, the cargo release profile, and digestion in gastrointestinal tract of zebrafish. Encapsulation of chuanxiong resulted in more compact structure and the smaller size of microparticles. The release rate of chuanxiong increased for alginate microparticles carrying more chuanxiong in simulated intestinal fluid. This remarkable feature ensures the controlled release of encapsulated cargos in the gastrointestinal tract of zebrafish. Moreover, chuanxiong-loaded alginate microparticles were moved to the end of gastrointestinal tract after oral administration for 6 hr and excreted from the body after 16 hr. Therefore, our developed method for oral administration of TCM in zebrafish is useful for easy and rapid evaluation of the drug effect on disease. PMID:27403425

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

    PubMed

    Lin, Li-Jen; Chiang, Chung-Jen; Chao, Yun-Peng; Wang, Shulhn-Der; Chiou, Yu-Ting; Wang, Han-Yu; Kao, Shung-Te

    2016-01-01

    Oral administration of Traditional Chinese Medicine (TCM) by patients is the common way to treat health problems. Zebrafish emerges as an excellent animal model for the pharmacology investigation. However, the oral delivery system of TCM in zebrafish has not been established so far. This issue was addressed by development of alginate microparticles for oral delivery of chuanxiong, a TCM that displays antifibrotic and antiproliferative effects on hepatocytes. The delivery microparticles were prepared from gelification of alginate containing various levels of chuanxiong. The chuanxiong-encapsulated alginate microparticles were characterized for their solubility, structure, encapsulation efficiency, the cargo release profile, and digestion in gastrointestinal tract of zebrafish. Encapsulation of chuanxiong resulted in more compact structure and the smaller size of microparticles. The release rate of chuanxiong increased for alginate microparticles carrying more chuanxiong in simulated intestinal fluid. This remarkable feature ensures the controlled release of encapsulated cargos in the gastrointestinal tract of zebrafish. Moreover, chuanxiong-loaded alginate microparticles were moved to the end of gastrointestinal tract after oral administration for 6 hr and excreted from the body after 16 hr. Therefore, our developed method for oral administration of TCM in zebrafish is useful for easy and rapid evaluation of the drug effect on disease. PMID:27403425

  6. Establishment of a neuroepithelial barrier by Claudin5a is essential for zebrafish brain ventricular lumen expansion

    PubMed Central

    Zhang, Jingjing; Piontek, Jörg; Wolburg, Hartwig; Piehl, Christian; Liss, Martin; Otten, Cécile; Christ, Annabel; Willnow, Thomas E.; Blasig, Ingolf E.; Abdelilah-Seyfried, Salim

    2010-01-01

    Lumen expansion driven by hydrostatic pressure occurs during many morphogenetic processes. Although it is well established that members of the Claudin family of transmembrane tight junction proteins determine paracellular tightness within epithelial/endothelial barrier systems, functional evidence for their role in the morphogenesis of lumenized organs has been scarce. Here, we identify Claudin5a as a core component of an early cerebral-ventricular barrier system that is required for ventricular lumen expansion in the zebrafish embryonic brain before the establishment of the embryonic blood–brain barrier. Loss of Claudin5a or expression of a tight junction-opening Claudin5a mutant reduces brain ventricular volume expansion without disrupting the polarized organization of the neuroepithelium. Perfusion experiments with the electron-dense small molecule lanthanum nitrate reveal that paracellular tightness of the cerebral-ventricular barrier decreases upon loss of Claudin5a. Genetic analyses show that the apical neuroepithelial localization of Claudin5a depends on epithelial cell polarity and provide evidence for concerted activities between Claudin5a and Na+,K+-ATPase during luminal expansion of brain ventricles. These data establish an essential role of a barrier-forming Claudin in ventricular lumen expansion, thereby contributing to brain morphogenesis. PMID:20080584

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

    PubMed

    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. PMID:27464817

  8. Reproductive toxicity of inorganic mercury exposure in adult zebrafish: Histological damage, oxidative stress, and alterations of sex hormone and gene expression in the hypothalamic-pituitary-gonadal axis.

    PubMed

    Zhang, Qun-Fang; Li, Ying-Wen; Liu, Zhi-Hao; Chen, Qi-Liang

    2016-08-01

    Mercury (Hg) is a prominent environmental contaminant that causes a variety of adverse effects on aquatic organisms. However, the mechanisms underlying inorganic Hg-induced reproductive impairment in fish remains largely unknown. In this study, adult zebrafish were exposed to 0 (control), 15 and 30μg Hg/l (added as mercuric chloride, HgCl2) for 30days, and the effects on histological structure, antioxidant status and sex hormone levels in the ovary and testis, as well as the mRNA expression of genes involved in the hypothalamic-pituitary-gonadal (HPG) axis were analyzed. Exposure to Hg caused pathological lesions in zebrafish gonads, and changed the activities and mRNA levels of antioxidant enzymes (catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPx)) as well as the content of glutathione (GSH) and malondialdehyde (MDA). In females, although ovarian 17β-estradiol (E2) content remained relatively stable, significant down-regulation of lhβ, gnrh2, gnrh3, lhr and erα were observed. In males, testosterone (T) levels in the testis significantly decreased after Hg exposure, accompanied by down-regulated expression of gnrh2, gnrh3, fshβ and lhβ in the brain as well as fshr, lhr, ar, cyp17 and cyp11b in the testis. Thus, our study indicated that waterborne inorganic Hg exposure caused histological damage and oxidative stress in the gonads of zebrafish, and altered sex hormone levels by disrupting the transcription of related HPG-axis genes, which could subsequently impair the reproduction of fish. Different response of the antioxidant defense system, sex hormone and HPG-axis genes between females and males exposed to inorganic Hg indicated the gender-specific regulatory effect by Hg. To our knowledge, this is the first time to explore the effects and mechanisms of inorganic Hg exposure on reproduction at the histological, enzymatic and molecular levels, which will greatly extend our understanding on the mechanisms underlying of reproductive

  9. Dynamics of axonal regeneration in adult and aging zebrafish reveal the promoting effect of a first lesion

    PubMed Central

    Graciarena, Mariana; Dambly-Chaudière, Christine; Ghysen, Alain

    2014-01-01

    Axonal regeneration is a major issue in the maintenance of adult nervous systems, both after nerve injuries and in neurodegenerative diseases. However, studying this process in vivo is difficult or even impossible in most vertebrates. Here we show that the posterior lateral line (PLL) of zebrafish is a suitable system to study axonal regeneration in vivo because of both the superficial location and reproducible spatial arrangement of neurons and targets, and the possibility of following reinnervation in live fish on a daily basis. Axonal regeneration after nerve cut has been demonstrated in this system during the first few days of life, leading to complete regeneration within 24 h. However, the potential for PLL nerve regeneration has not been tested yet beyond the early larval stage. We explore the regeneration potential and dynamics of the PLL nerve in adult zebrafish and report that regeneration occurs throughout adulthood. We observed that irregularities in the original branching pattern are faithfully reproduced after regeneration, suggesting that regenerating axons follow the path laid down by the original nerve branches. We quantified the extent of target reinnervation after a nerve cut and found that the latency before the nerve regenerates increases with age. This latency is reduced after a second nerve cut at all ages, suggesting that a regeneration-promoting factor induced by the first cut facilitates regeneration on a second cut. We provide evidence that this factor remains present at the site of the first lesion for several days and is intrinsic to the neurons. PMID:24474787

  10. Actin-Cytoskeleton- and Rock-Mediated INM Are Required for Photoreceptor Regeneration in the Adult Zebrafish Retina

    PubMed Central

    Lahne, Manuela; Li, Jingling; Marton, Rebecca M.

    2015-01-01

    Loss of retinal neurons in adult zebrafish (Danio rerio) induces a robust regenerative response mediated by the reentry of the resident Müller glia into the cell cycle. Upon initiating Müller glia proliferation, their nuclei migrate along the apicobasal axis of the retina in phase with the cell cycle in a process termed interkinetic nuclear migration (INM). We examined the mechanisms governing this cellular process and explored its function in regenerating the adult zebrafish retina. Live-cell imaging revealed that the majority of Müller glia nuclei migrated to the outer nuclear layer (ONL) to divide. These Müller glia formed prominent actin filaments at the rear of nuclei that had migrated to the ONL. Inhibiting actin filament formation or Rho-associated coiled-coil kinase (Rock) activity, which is necessary for phosphorylation of myosin light chain and actin myosin-mediated contraction, disrupted INM with increased numbers of mitotic nuclei remaining in the basal inner nuclear layer, the region where Müller glia typically reside. Double knockdown of Rho-associated coiled-coil kinase 2a (Rock2a) and Rho-associated coiled-coil kinase 2b (Rock2b) similarly disrupted INM and reduced Müller glial cell cycle reentry. In contrast, Rock inhibition immediately before the onset of INM did not affect Müller glia proliferation, but subsequently reduced neuronal progenitor cell proliferation due to early cell cycle exit. Long-term, Rock inhibition increased the generation of mislocalized ganglion/amacrine cells at the expense of rod and cone photoreceptors. In summary, INM is driven by an actin-myosin-mediated process controlled by Rock2a and Rock2b activity, which is required for sufficient proliferation and regeneration of photoreceptors after light damage. SIGNIFICANCE STATEMENT The human retina does not replace lost or damaged neurons, ultimately causing vision impairment. In contrast, zebrafish are capable of regenerating lost neurons. Understanding the mechanisms

  11. Guidelines for Better Communication with Brain Impaired Adults

    MedlinePlus

    ... A You are here Home Guidelines for Better Communication with Brain Impaired Adults Printer-friendly version Communicating ... easy solutions, following some basic guidelines should ease communication, and lower levels of stress both for you ...

  12. Additive effects of levonorgestrel and ethinylestradiol on brain aromatase (cyp19a1b) in zebrafish specific in vitro and in vivo bioassays.

    PubMed

    Hinfray, N; Tebby, C; Garoche, C; Piccini, B; Bourgine, G; Aït-Aïssa, S; Kah, O; Pakdel, F; Brion, F

    2016-09-15

    Estrogens and progestins are widely used in combination in human medicine and both are present in aquatic environment. Despite the joint exposure of aquatic wildlife to estrogens and progestins, very little information is available on their combined effects. In the present study we investigated the effect of ethinylestradiol (EE2) and Levonorgestrel (LNG), alone and in mixtures, on the expression of the brain specific ER-regulated cyp19a1b gene. For that purpose, recently established zebrafish-derived tools were used: (i) an in vitro transient reporter gene assay in a human glial cell line (U251-MG) co-transfected with zebrafish estrogen receptors (zfERs) and the luciferase gene under the control of the zebrafish cyp19a1b gene promoter and (ii) an in vivo bioassay using a transgenic zebrafish expressing GFP under the control of the zebrafish cyp19a1b gene promoter (cyp19a1b-GFP). Concentration-response relationships for single chemicals were modeled and used to design the mixture experiments following a ray design. The results from mixture experiments were analyzed to predict joint effects according to concentration addition and statistical approaches were used to characterize the potential interactions between the components of the mixtures (synergism/antagonism). We confirmed that some progestins could elicit estrogenic effects in fish brain. In mixtures, EE2 and LNG exerted additive estrogenic effects both in vitro and in vivo, suggesting that some environmental progestin could exert effects that will add to those of environmental (xeno-)estrogens. Moreover, our zebrafish specific assays are valuable tools that could be used in risk assessment for both single chemicals and their mixtures. PMID:27491593

  13. Nutrient status modulates the expression of nesfatin-1 encoding nucleobindin 2A and 2B mRNAs in zebrafish gut, liver and brain.

    PubMed

    Hatef, Azadeh; Shajan, Sooraj; Unniappan, Suraj

    2015-05-01

    Nesfatin-1 is a naturally occurring, 82-amino acid peptide processed from the precursor nucleobindin 2 (NUCB2), a highly conserved protein among vertebrates. In fish, two isoforms of NUCB2 (NUCB2A and NUCB2B) exist, and nesfatin-1 has been identified in goldfish and Ya fish. We recently reported the presence and appetite suppressing effects of nesfatin-1 in goldfish. The main objectives of this study were to characterize NUCB2 in zebrafish, and determine whether NUCB2 mRNAs are affected by food availability. Tissue distribution of NUCB2A and NUCB2B mRNAs, and NUCB2/nesfatin-1-like immunoreactivity (ir) in the gut of zebrafish were also investigated. In zebrafish, nesfatin-1 region (1-82 amino acids) in NUCB2A is 78% identical to NUCB2B. Both NUCB2A and NUCB2 mRNAs were most abundant in the liver, while less expression was found in other tissues including the brain and gut. NUCB2/nesfatin-1-like immunoreactivity was detected in the mucosal layer cells of zebrafish anterior gastrointestinal tract. NUCB2A and NUCB2B mRNA expression were decreased in the brain of zebrafish 3h after feeding, and after a 7-day food deprivation. Both NUCB2A and NUCB2B mRNAs in the gut were also decreased following 7 days of food deprivation, while NUCB2B mRNA was increased in the liver. Our results provide molecular and functional evidences to support potential anorectic and metabolic roles for endogenous nesfatin-1 in zebrafish. To our knowledge, this is the first report on NUCB2B characterization in vertebrates. PMID:25260251

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

    NASA Astrophysics Data System (ADS)

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

    2014-03-01

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

  15. Memory and Brain Volume in Adults Prenatally Exposed to Alcohol

    ERIC Educational Resources Information Center

    Coles, Claire D.; Goldstein, Felicia C.; Lynch, Mary Ellen; Chen, Xiangchuan; Kable, Julie A.; Johnson, Katrina C.; Hu, Xiaoping

    2011-01-01

    The impact of prenatal alcohol exposure on memory and brain development was investigated in 92 African-American, young adults who were first identified in the prenatal period. Three groups (Control, n = 26; Alcohol-related Neurodevelopmental Disorder, n = 36; and Dysmorphic, n = 30) were imaged using structural MRI with brain volume calculated for…

  16. Childhood Onset Schizophrenia: Cortical Brain Abnormalities as Young Adults

    ERIC Educational Resources Information Center

    Greenstein, Deanna; Lerch, Jason; Shaw, Philip; Clasen, Liv; Giedd, Jay; Gochman, Peter; Rapoport, Judith; Gogtay, Nitin

    2006-01-01

    Background: Childhood onset schizophrenia (COS) is a rare but severe form of the adult onset disorder. While structural brain imaging studies show robust, widespread, and progressive gray matter loss in COS during adolescence, there have been no longitudinal studies of sufficient duration to examine comparability with the more common adult onset…

  17. A New Anaesthetic Protocol for Adult Zebrafish (Danio rerio): Propofol Combined with Lidocaine

    PubMed Central

    Valentim, Ana M.; Félix, Luís M.; Carvalho, Leonor; Diniz, Enoque; Antunes, Luís M.

    2016-01-01

    Background The increasing use of zebrafish model has not been accompanied by the evolution of proper anaesthesia for this species in research. The most used anaesthetic in fishes, MS222, may induce aversion, reduction of heart rate, and consequently high mortality, especially during long exposures. Therefore, we aim to explore new anaesthetic protocols to be used in zebrafish by studying the quality of anaesthesia and recovery induced by different concentrations of propofol alone and in combination with different concentrations of lidocaine. Material and Methods In experiment A, eighty-three AB zebrafish were randomly assigned to 7 different groups: control, 2.5 (2.5P), 5 (5P) or 7.5 μg/ml (7.5P) of propofol; and 2.5 μg/ml of propofol combined with 50, (P/50L), 100 (P/100L) or 150 μg/ml (P/150L) of lidocaine. Zebrafish were placed in an anaesthetic water bath and time to lose the equilibrium, reflex to touch, reflex to a tail pinch, and respiratory rate were measured. Time to gain equilibrium was also assessed in a clean tank. Five and 24 hours after anaesthesia recovery, zebrafish were evaluated concerning activity and reactivity. Afterwards, in a second phase of experiments (experiment B), the best protocol of the experiment A was compared with a new group of 8 fishes treated with 100 mg/L of MS222 (100M). Results In experiment A, only different concentrations of propofol/lidocaine combination induced full anaesthesia in all animals. Thus only these groups were compared with a standard dose of MS222 in experiment B. Propofol/lidocaine induced a quicker loss of equilibrium, and loss of response to light and painful stimuli compared with MS222. However zebrafish treated with MS222 recovered quickly than the ones treated with propofol/lidocaine. Conclusion In conclusion, propofol/lidocaine combination and MS222 have advantages in different situations. MS222 is ideal for minor procedures when a quick recovery is important, while propofol/lidocaine is best to

  18. Temperature- and exercise-induced gene expression and metabolic enzyme changes in skeletal muscle of adult zebrafish (Danio rerio)

    PubMed Central

    McClelland, Grant B; Craig, Paul M; Dhekney, Kalindi; Dipardo, Shawn

    2006-01-01

    Both exercise training and cold acclimatization induce muscle remodelling in vertebrates, producing a more aerobic phenotype. In ectothermic species exercise training and cold-acclimatization represent distinct stimuli. It is currently unclear if these stimuli act through a common mechanism or if different mechanisms lead to a common phenotype. The goal of this study was to survey responses that represent potential mechanisms responsible for contraction- and temperature-induced muscle remodelling, using an ectothermic vertebrate. Separate groups of adult zebrafish (Danio rerio) were either swim trained or cold acclimatized for 4 weeks. We found that the mitochondrial marker enzyme citrate synthase (CS) was increased by 1.5× in cold and by 1.3× with exercise (P < 0.05). Cytochrome c oxidase (COx) was increased by 1.2× following exercise training (P < 0.05) and 1.2× (P = 0.07) with cold acclimatization. However, only cold acclimatization increased β-hydroxyacyl-CoA dehydrogenase (HOAD) compared to exercise-trained (by 1.3×) and pyruvate kinase (PK) relative to control zebrafish. We assessed the whole-animal performance outcomes of these treatments. Maximum absolute sustained swimming speed (Ucrit) was increased in the exercise trained group but not in the cold acclimatized group. Real-time PCR analysis indicated that increases in CS are primarily transcriptionally regulated with exercise but not with cold treatments. Both treatments showed increases in nuclear respiratory factor (NRF)-1 mRNA which was increased by 2.3× in cold-acclimatized and 4× in exercise-trained zebrafish above controls. In contrast, peroxisome proliferator-activated receptor (PPAR)-α mRNA levels were decreased in both experimental groups while PPAR-β1 declined in exercise training only. Moreover, PPAR-γ coactivator (PGC)-1α mRNA was not changed by either treatment. In zebrafish, both temperature and exercise produce a more aerobic phenotype, but there are stimulus-dependent responses

  19. Brain stem auditory evoked responses in human infants and adults

    NASA Technical Reports Server (NTRS)

    Hecox, K.; Galambos, R.

    1974-01-01

    Brain stem evoked potentials were recorded by conventional scalp electrodes in infants (3 weeks to 3 years of age) and adults. The latency of one of the major response components (wave V) is shown to be a function both of click intensity and the age of the subject; this latency at a given signal strength shortens postnatally to reach the adult value (about 6 msec) by 12 to 18 months of age. The demonstrated reliability and limited variability of these brain stem electrophysiological responses provide the basis for an optimistic estimate of their usefulness as an objective method for assessing hearing in infants and adults.

  20. Tetraspanin 3c requirement for pigment cell interactions and boundary formation in zebrafish adult pigment stripes

    PubMed Central

    Inoue, Shinya; Kondo, Shigeru; Parichy, David M.; Watanabe, Masakatsu

    2014-01-01

    Summary Skin pigment pattern formation in zebrafish requires pigment-cell autonomous interactions between melanophores and xanthophores, yet the molecular bases for these interactions remain largely unknown. Here, we examined the dali mutant that exhibits stripes in which melanophores are intermingled abnormally with xanthophores. By in vitro cell culture, we found that melanophores of dali mutants have a defect in motility and that interactions between melanophores and xanthophores are defective as well. Positional cloning and rescue identified dali as tetraspanin 3c (tspan3c), encoding a transmembrane scaffolding protein expressed by melanophores and xanthophores. We further showed that dali mutant Tspan3c expressed in HeLa cell exhibits a defect in N-glycosylation and is retained inappropriately in the endoplasmic reticulum. Our results are the first to identify roles for a tetraspanin superfamily protein in skin pigment pattern formation and suggest new mechanisms for the establishment and maintenance of zebrafish stripe boundaries. PMID:24734316

  1. Role of eosinophils and apoptosis in PDIMs/PGLs deficient mycobacterium elimination in adult zebrafish.

    PubMed

    Huang, Xinhua; Wang, Hui; Meng, Lu; Wang, Qinglan; Yu, Jia; Gao, Qian; Wang, Decheng

    2016-06-01

    The cell wall lipids phthiocerol dimycocerosates (PDIMs) and its structurally-related compound, phenolic glycolipids (PGLs) are major virulence factors of mycobacterium, as shown by the reduced growth of PDIMs/PGLs deficient mutants in various animal models. PDIMs/PGLs play active roles in modulating host immune responses. However, the cellular and molecular mechanisms of how PDIMs/PGLs deficient mutant was eliminated in vivo are still elusive. Our aim was to investigate what host immune responses have effect on mycobacterium elimination in vivo. Using microarray, we find PDIMs/PGLs modulate divergent host responses, including chemotaxis and focal adhesion's downstream pathway and apoptosis. We examine these two host responses by Diff-Quik stain, coupled with transmission electron microscopy and TUNEL stain respectively. The ultrastructure observation showed that eosinophils appeared in WT-infected zebrafish at day 1, however eosinophils arrived was delayed to day 7 in PDIMs/PGLs-deficient mutant-infected animals. More intriguingly, apoptosis was markedly increased in PDIMs/PGLs-mutant infected zebrafish at day 1 after infection, compared to WT-infected fishes at this time. However, apoptosis trend was fully reversed by day 7, with increased apoptosis were detected in WT-infected zebrafish compared with the PDIMs/PGLs-deficient mutant, especially more apoptosis within the granuloma. This study shows that the anti-apoptotic effects of PDIMs/PGLs and the recruitment of eosinophils in tissue during the early infection in zebrafish might promote bacterium growth in vivo. PMID:26855012

  2. New Nerve Cells for the Adult Brain.

    ERIC Educational Resources Information Center

    Kempermann, Gerd; Gage, Fred H.

    1999-01-01

    Contrary to dogma, the human brain does produce new nerve cells in adulthood. The mature human brain spawns neurons routinely in the hippocampus, an area important to memory and learning. This research can make it possible to ease any number of disorders involving neurological damage and death. (CCM)

  3. Atomoxetine reduces anticipatory responding in a 5-choice serial reaction time task for adult zebrafish

    PubMed Central

    Parker, Matthew O.; Brock, Alistair J.; Sudwarts, Ari; Brennan, Caroline H.

    2014-01-01

    Deficits in impulse control are related to a number of psychiatric diagnoses, including attention deficit hyperactivity disorder, addiction, and pathological gambling. Despite increases in our knowledge about the underlying neurochemical and neuroanatomical correlates, understanding of the molecular and cellular mechanisms is less well established. Understanding these mechanisms is essential in order to move towards individualized treatment programs and increase efficacy of interventions. Zebrafish are a very useful vertebrate model for exploring molecular processes underlying disease owing to their small size and genetic tractability. Their utility in terms of behavioral neuroscience, however, hinges on the validation and publication of reliable assays with adequate translational relevance. Here, we report an initial pharmacological validation of a fully automated zebrafish version of the commonly used five-choice serial reaction time task using a variable interval pre-stimulus interval. We found that atomoxetine reduced anticipatory responses (0.6 mg/kg), whereas a high-dose (4 mg/kg) methylphenidate increased anticipatory responses and the number of trials completed in a session. On the basis of these results, we argue that similar neurochemical processes in fish as in mammals may control impulsivity, as operationally defined by anticipatory responses on a continuous performance task such as this, making zebrafish potentially a good model for exploring the molecular basis of impulse control disorders and for first-round drug screening. PMID:24481568

  4. Substance use and brain reward mechanisms in older adults.

    PubMed

    Snyder, Marsha; Platt, Lois

    2013-07-01

    Substance use among older adults is on the rise, with statistics indicating this to be a growing health problem. Brain changes in the reward center of the brain that naturally occur with aging are offered as one source of these statistics. Aging is generally associated with increased prevalence of chronic disease, disability, and death, and therefore a public health goal for older adults is to maintain health, independence, and function. Psychiatric-mental health nurses are uniquely positioned to assist older adults in achievement of these goals through health assessment and promotion. The use of client-centered counseling approaches that recognize the older adult's developmental need for autonomy and choice in decision making have been shown to be effective in increasing motivation in this adult population. PMID:23758223

  5. Associative learning in the multichamber tank: A new learning paradigm for zebrafish.

    PubMed

    Fernandes, Yohaan M; Rampersad, Mindy; Luchiari, Ana C; Gerlai, Robert

    2016-10-01

    The zebrafish has been gaining prominence in the field of behavioural brain research as this species offers a good balance between system complexity and practical simplicity. While the number of studies examining the behaviour of zebrafish has exponentially increased over the past decade, the need is still substantial for paradigms capable of assessing cognitive and mnemonic characteristics of this species. Here we describe and utilize a novel visual discrimination task with which we evaluated acquisition of CS (colour)-US (sight of conspecifics) association in adult zebrafish. We report significant acquisition of CS-US association indicated by the increased time the fish spent in and the increased frequency of visits of the target chamber during a probe trial in the absence of reward. Given the simplicity of the apparatus and procedure, we conclude that the new task may be employed to assay learning and memory in adult zebrafish in an efficient manner. PMID:27345425

  6. Toxicity to embryo and adult zebrafish of copper complexes with two malonic acids as models for dissolved organic matter

    SciTech Connect

    Palmer, F.B.; Evans, C.W.; Butler, C.A.; Timperley, M.H.

    1998-08-01

    The toxicity to embryo and adult zebrafish (Danio rerio) of Cu complexes with two substituted malonic acids, benzyl- and n-hexadecyl-, chosen as models for low-molecular-weight natural dissolved organic matter, were investigated. Toxicity test solutions at pH 6.5 {+-} 0.1 with the required Cu ion-specific electrode. In the absence of malonic acids, concentrations of Cu{sup 2+} up to 1.13 {mu}mol/L increased the embryo hatching time from approx. 2 d in control solutions (no Cu or malonic acid) and solutions containing malonic acids without Cu to approx. 8 d. The Cu-benzylmalonic acid complex in the presence of inorganic Cu species did not delay hatching beyond that attributable to Cu{sup 2+}. In contrast, 0.60 {mu}mol/L Cu-n-hexadecylmalonic complexes delayed hatching by 5.5 d in excess of that attributable to 1.13 {mu}mol/L Cu{sup 2+}, assuming that the hatching delays caused by the different Cu species were additive, possibly because of Cu entry into the embryo as the lipophilic Cu-n-hexadecylmalonic complex. None of the Cu-malonic acid complexes was acutely toxic to adult zebrafish at concentrations up to 1.4 {mu}mol/L, possibly because Cu was removed from the Cu-malonic acid complexes by stronger chelating groups at the gill surface. Substituted malonic acids with similar proton and Cu association constants can be readily prepared with a variety of simple substituents, radiolabeled if required. Their results show that these acids could be useful ligands for investigating intracellular transport and metabolism of metal-organic complexes.

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

    PubMed

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

    2015-10-27

    We studied adult zebrafish to determine whether the size of 20 and 110 nm citrate-coated silver nanoparticles (AgC NPs) differentially impact the gills and intestines, known target organs for Ag toxicity in fish. Following exposure for 4 h, 4 days, or 4 days plus a 7 day depuration period, we obtained different toxicokinetic profiles for different particle sizes, as determined by Ag content of the tissues. Ionic AgNO3 served as a positive control. The gills showed a significantly higher Ag content for the 20 nm particles at 4 h and 4 days than the 110 nm particles, while the values were more similar in the intestines. Both particle types were retained in the intestines even after depuration. These toxicokinetics were accompanied by striking size-dependent differences in the ultrastructural features and histopathology in the target organs in response to the particulates. Ag staining of the gills and intestines confirmed prominent Ag deposition in the basolateral membranes for the 20 nm but not for the 110 nm particles. Furthermore, it was possible to link the site of tissue deposition to disruption of the Na(+)/K(+) ion channel, which is also localized to the basolateral membrane. This was confirmed by a reduction in ATPase activity and immunohistochemical detection of the α subunit of this channel in both target organs, with the 20 nm particles causing significantly higher inhibition and disruption than the larger size particles or AgNO3. These results demonstrate the importance of particle size in determining the hazardous impact of AgNPs in the gills and intestines of adult zebrafish. PMID:26327297

  8. Histomorphological Phenotyping of the Adult Mouse Brain.

    PubMed

    Mikhaleva, Anna; Kannan, Meghna; Wagner, Christel; Yalcin, Binnaz

    2016-01-01

    This article describes a series of standard operating procedures for morphological phenotyping of the mouse brain using basic histology. Many histological studies of the mouse brain use qualitative approaches based on what the human eye can detect. Consequently, some phenotypic information may be missed. Here we describe a quantitative approach for the assessment of brain morphology that is simple and robust. A total of 78 measurements are made throughout the brain at specific and well-defined regions, including the cortex, the hippocampus, and the cerebellum. Experimental design and timeline considerations, including strain background effects, the importance of sectioning quality, measurement variability, and efforts to correct human errors are discussed. © 2016 by John Wiley & Sons, Inc. PMID:27584555

  9. Adult mouse brain gene expression patterns bear an embryologic imprint

    PubMed Central

    Zapala, Matthew A.; Hovatta, Iiris; Ellison, Julie A.; Wodicka, Lisa; Del Rio, Jo A.; Tennant, Richard; Tynan, Wendy; Broide, Ron S.; Helton, Rob; Stoveken, Barbara S.; Winrow, Christopher; Lockhart, Daniel J.; Reilly, John F.; Young, Warren G.; Bloom, Floyd E.; Lockhart, David J.; Barlow, Carrolee

    2005-01-01

    The current model to explain the organization of the mammalian nervous system is based on studies of anatomy, embryology, and evolution. To further investigate the molecular organization of the adult mammalian brain, we have built a gene expression-based brain map. We measured gene expression patterns for 24 neural tissues covering the mouse central nervous system and found, surprisingly, that the adult brain bears a transcriptional “imprint” consistent with both embryological origins and classic evolutionary relationships. Embryonic cellular position along the anterior–posterior axis of the neural tube was shown to be closely associated with, and possibly a determinant of, the gene expression patterns in adult structures. We also observed a significant number of embryonic patterning and homeobox genes with region-specific expression in the adult nervous system. The relationships between global expression patterns for different anatomical regions and the nature of the observed region-specific genes suggest that the adult brain retains a degree of overall gene expression established during embryogenesis that is important for regional specificity and the functional relationships between regions in the adult. The complete collection of extensively annotated gene expression data along with data mining and visualization tools have been made available on a publicly accessible web site (www.barlow-lockhart-brainmapnimhgrant.org). PMID:16002470

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

    PubMed Central

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

    2008-01-01

    Comparison between related species is a successful approach to uncover conserved and divergent principles of development. Here, we studied the pattern of epithalamic asymmetry in zebrafish (Danio rerio) and medaka (Oryzias latipes), two related teleost species with 115–200 Myr of independent evolution. We found that these species share a strikingly conserved overall pattern of asymmetry in the parapineal–habenular–interpeduncular system. Nodal signalling exhibits comparable spatial and temporal asymmetric expressions in the presumptive epithalamus preceding the development of morphological asymmetries. Neuroanatomical asymmetries consist of left-sided asymmetric positioning and connectivity of the parapineal organ, enlargement of neuropil in the left habenula compared with the right habenula and segregation of left–right habenular efferents along the dorsoventral axis of the interpeduncular nucleus. Despite the overall conservation of asymmetry, we observed heterotopic changes in the topology of parapineal efferent connectivity, heterochronic shifts in the timing of developmental events underlying the establishment of asymmetry and divergent degrees of canalization of embryo laterality. We offer new tools for developmental time comparison among species and propose, for each of these transformations, novel hypotheses of ontogenic mechanisms that explain interspecies variations that can be tested experimentally. Together, these findings highlight the usefulness of zebrafish and medaka as comparative models to study the developmental mechanisms of epithalamic asymmetry in vertebrates. PMID:19064351

  11. In vitro CYP1A activity in the zebrafish: temporal but low metabolite levels during organogenesis and lack of gender differences in the adult stage.

    PubMed

    Saad, Moayad; Verbueken, Evy; Pype, Casper; Casteleyn, Christophe; Van Ginneken, Chris; Maes, Louis; Cos, Paul; Van Cruchten, Steven

    2016-09-01

    The zebrafish (Danio rerio) is increasingly used as a screening model for acute, chronic and developmental toxicity. More specifically, the embryo is currently investigated as a replacement of in vivo developmental toxicity studies, although its biotransformation capacity remains a point of debate. As the cytochrome P450 1 (CYP1) family plays an important role in the biotransformation of several pollutants and drugs, a quantitative in vitro protocol was refined to assess gender- and age-related CYP1A activity in the zebrafish using the ethoxyresorufin-o-deethylase (EROD) assay. Microsomal protein fractions were prepared from livers of adult males and females, ovaries and whole embryo homogenates of different developmental stages. A large biological variation but no gender-related difference in CYP1A activity was observed in adult zebrafish. Embryos showed distinct temporal but low CYP1A activity during organogenesis. These in vitro data raise questions on the bioactivation capacity of zebrafish embryos in developmental toxicity studies. PMID:27046732

  12. The effects of vitamin D on brain development and adult brain function.

    PubMed

    Kesby, James P; Eyles, Darryl W; Burne, Thomas H J; McGrath, John J

    2011-12-01

    A role for vitamin D in brain development and function has been gaining support over the last decade. Multiple lines of evidence suggest that this vitamin is actually a neuroactive steroid that acts on brain development, leading to alterations in brain neurochemistry and adult brain function. Early deficiencies have been linked with neuropsychiatric disorders, such as schizophrenia, and adult deficiencies have been associated with a host of adverse brain outcomes, including Parkinson's disease, Alzheimer's disease, depression and cognitive decline. This review summarises the current state of research on the actions of vitamin D in the brain and the consequences of deficiencies in this vitamin. Furthermore, we discuss specific implications of vitamin D status on the neurotransmitter, dopamine. PMID:21664231

  13. Effects of bisphenol A and triclocarban on brain-specific expression of aromatase in early zebrafish embryos

    PubMed Central

    Chung, Eunah; Genco, Maria C.; Megrelis, Laura; Ruderman, Joan V.

    2011-01-01

    Estrogen regulates numerous developmental and physiological processes. Most effects are mediated by estrogen receptors (ERs), which function as ligand-regulated transcription factors. Estrogen also regulates the activity of GPR30, a membrane-associated G protein-coupled receptor. Many different types of environmental contaminants can activate ERs; some can bind GPR30 as well. There is growing concern that exposure to some of these compounds, termed xenoestrogens, is interfering with the behavior and reproductive potential of numerous wildlife species, as well as affecting human health. Here, we investigated how two common, environmentally pervasive chemicals affect the in vivo expression of a known estrogen target gene in the brain of developing zebrafish embryos, aromatase AroB, which converts androgens to estrogens. We confirm that, like estrogen, the well-studied xenoestrogen bisphenol A (BPA, a plastics monomer), induces strong brain-specific overexpression of aromatase. Experiments using ER- and GPR30-selective modulators argue that this induction is largely through nuclear ERs. BPA induces dramatic overexpression of AroB RNA in the same subregions of the developing brain as estrogen. The antibacterial triclocarban (TCC) by itself stimulates AroB expression only slightly, but TCC strongly enhances the overexpression of AroB that is induced by exogenous estrogen. Thus, both BPA and TCC have the potential to elevate levels of aromatase and, thereby, levels of endogenous estrogens in the developing brain. In contrast to estrogen, BPA-induced AroB overexpression was suppressed by TCC. These results indicate that exposures to combinations of certain hormonally active pollutants can have outcomes that are not easily predicted from their individual effects. PMID:22006313

  14. Progesterone alters global transcription profiles at environmental concentrations in brain and ovary of female zebrafish (Danio rerio).

    PubMed

    Zucchi, Sara; Castiglioni, Sara; Fent, Karl

    2013-01-01

    Progesterone (P4) is a natural steroid hormone excreted by humans and animals. Noncomplete degradation in treatment plants result in levels in the ng/L range in surface waters. Very little is known of the effects on fish at such concentrations. Here we determine the global expression profile in the brain and ovary of female zebrafish exposed for 14 days to 3.5, 33 and 306 ng/L P4 to elucidate molecular effects. For validation selected transcripts were determined by RT-qPCR. In the brain, 54 and 255 transcripts were altered at 3.5 and 306 ng/L, respectively. Genes related to circadian rhythm (nr1d2b, per1b), cell cycle and reproduction (cdc20, ccnb1) were down-regulated. In the ovary, transcriptional changes occurred in 200, 84 and 196 genes at 3.5, 33 and 306 ng/L, respectively. The genes belong to different pathways including cardiac hypertrophy, cell cycle and its regulation. P4 slightly influenced oocyte maturation as revealed by histology of the ovaries. In the liver, vtg1 was down-regulated at all concentrations and VTG protein at 306 ng/L in the blood. The data show molecular effects and the modes of action of P4 at environmental concentrations. Ultimately they may translate to adverse effects on reproduction. PMID:24083816

  15. In vivo assessment of the permeability of the blood-brain barrier and blood-retinal barrier to fluorescent indoline derivatives in zebrafish

    PubMed Central

    2012-01-01

    Background Successful delivery of compounds to the brain and retina is a challenge in the development of therapeutic drugs and imaging agents. This challenge arises because internalization of compounds into the brain and retina is restricted by the blood–brain barrier (BBB) and blood-retinal barrier (BRB), respectively. Simple and reliable in vivo assays are necessary to identify compounds that can easily cross the BBB and BRB. Methods We developed six fluorescent indoline derivatives (IDs) and examined their ability to cross the BBB and BRB in zebrafish by in vivo fluorescence imaging. These fluorescent IDs were administered to live zebrafish by immersing the zebrafish larvae at 7-8 days post fertilization in medium containing the ID, or by intracardiac injection. We also examined the effect of multidrug resistance proteins (MRPs) on the permeability of the BBB and BRB to the ID using MK571, a selective inhibitor of MRPs. Results The permeability of these barriers to fluorescent IDs administered by simple immersion was comparable to when administered by intracardiac injection. Thus, this finding supports the validity of drug administration by simple immersion for the assessment of BBB and BRB permeability to fluorescent IDs. Using this zebrafish model, we demonstrated that the length of the methylene chain in these fluorescent IDs significantly affected their ability to cross the BBB and BRB via MRPs. Conclusions We demonstrated that in vivo assessment of the permeability of the BBB and BRB to fluorescent IDs could be simply and reliably performed using zebrafish. The structure of fluorescent IDs can be flexibly modified and, thus, the permeability of the BBB and BRB to a large number of IDs can be assessed using this zebrafish-based assay. The large amount of data acquired might be useful for in silico analysis to elucidate the precise mechanisms underlying the interactions between chemical structure and the efflux transporters at the BBB and BRB. In turn

  16. A brain sexual dimorphism controlled by adult circulating androgens.

    PubMed

    Cooke, B M; Tabibnia, G; Breedlove, S M

    1999-06-22

    Reports of structural differences between the brains of men and women, heterosexual and homosexual men, and male-to-female transsexuals and other men have been offered as evidence that the behavioral differences between these groups are likely caused by differences in the early development of the brain. However, a possible confounding variable is the concentration of circulating hormones seen in these groups in adulthood. Evaluation of this possibility hinges on the extent to which circulating hormones can alter the size of mammalian brain regions as revealed by Nissl stains. We now report a sexual dimorphism in the volume of a brain nucleus in rats that can be completely accounted for by adult sex differences in circulating androgen. The posterodorsal nucleus of the medial amygdala (MePD) has a greater volume in male rats than in females, but adult castration of males causes the volume to shrink to female values within four weeks, whereas androgen treatment of adult females for that period enlarges the MePD to levels equivalent to normal males. This report demonstrates that adult hormone manipulations can completely reverse a sexual dimorphism in brain regional volume in a mammalian species. The sex difference and androgen responsiveness of MePD volume is reflected in the soma size of neurons there. PMID:10377450

  17. Expansion of Multipotent Stem Cells from the Adult Human Brain

    PubMed Central

    Murrell, Wayne; Palmero, Emily; Bianco, John; Stangeland, Biljana; Joel, Mrinal; Paulson, Linda; Thiede, Bernd; Grieg, Zanina; Ramsnes, Ingunn; Skjellegrind, Håvard K.; Nygård, Ståle; Brandal, Petter; Sandberg, Cecilie; Vik-Mo, Einar; Palmero, Sheryl; Langmoen, Iver A.

    2013-01-01

    The discovery of stem cells in the adult human brain has revealed new possible scenarios for treatment of the sick or injured brain. Both clinical use of and preclinical research on human adult neural stem cells have, however, been seriously hampered by the fact that it has been impossible to passage these cells more than a very few times and with little expansion of cell numbers. Having explored a number of alternative culturing conditions we here present an efficient method for the establishment and propagation of human brain stem cells from whatever brain tissue samples we have tried. We describe virtually unlimited expansion of an authentic stem cell phenotype. Pluripotency proteins Sox2 and Oct4 are expressed without artificial induction. For the first time multipotency of adult human brain-derived stem cells is demonstrated beyond tissue boundaries. We characterize these cells in detail in vitro including microarray and proteomic approaches. Whilst clarification of these cells’ behavior is ongoing, results so far portend well for the future repair of tissues by transplantation of an adult patient’s own-derived stem cells. PMID:23967194

  18. Biochemical response of diverse organs in adult Danio rerio (zebrafish) exposed to sub-lethal concentrations of microcystin-LR and microcystin-RR: a balneation study.

    PubMed

    Pavagadhi, Shruti; Gong, Zhiyuan; Hande, M Prakash; Dionysiou, Dionysios D; de la Cruz, Armah A; Balasubramanian, Rajasekhar

    2012-03-01

    The present study was carried out to examine the dose-response of microcystin-LR (MC-LR) and microcystin-RR (MC-RR) toxicity in adult Danio rerio (zebrafish) under balneation conditions at various time points. The differential responses of superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione-S-transferase (GST) as biomarkers were assessed for oxygen mediated toxicity in liver, gills, intestine and brain tissues of zebrafish exposed to dissolved MC-LR and MC-RR (0.1-10.0 μgl(-1)). To investigate the time related response of biomarkers, fish were sampled after 4, 7 and 15 days of exposure. Responses varied (i) between MC-LR and MC-RR (for certain groups), (ii) for different enzymes at all time points, and (iii) for different tissues. In general, most of the enzymes followed a bell shaped curve, with an abrupt increase in activity at a particular concentration. It was observed that upon exposure to MC-LR and MC-RR, some enzymes showed an adaptive response after the first time point wherein the enzyme activity increased in some tissues. The increase in enzyme activity is suggestive of their cellular and metabolic adaptations to the continued stress and toxin exposure. Enzyme activities in general increased at lower concentrations (≤ 5.0 μgl(-1)) and decreased at higher concentrations (≥ 5.0 μgl(-1)). An abrupt change in enzyme activities was observed at a particular concentration in all the tissue enzymes. For GPx and GR, there was a differential response in the case of fish exposed to MC-LR and MC-RR, which could be due to the difference in toxicity potentials of these cyanotoxins. In general, initial stress conditions were observed in most of the tissue enzymes following the exposure to microcystins (MCs). This observation suggests that MCs found in trace levels are likely to have deleterious effects on aquatic organisms and can trigger a variety of biochemical responses depending on their specific toxicity

  19. Global DNA methylation in gonads of adult zebrafish Danio rerio under bisphenol A exposure.

    PubMed

    Liu, Yan; Zhang, Yingying; Tao, Shiyu; Guan, Yongjing; Zhang, Ting; Wang, Zaizhao

    2016-08-01

    Altered DNA methylation is pervasively associated with changes in gene expression and signal transduction after exposure to a wide range of endocrine disrupting chemicals. As a weak estrogenic chemical, bisphenol A (BPA) has been extensively studied for reproductive toxicity. In order to explore the effects of BPA on epigenetic modification in gonads of zebrafish Danio rerio, we measured the global DNA methylation together with the gene expression of DNA methyltransferase (dnmts), glycine N-methyltransferase (gnmt), and ten-eleven translocation (tets) in gonads of D. rerio under BPA exposure by ELISA and quantitative real-time PCR method, respectively. The global level of DNA methylation was significantly decreased in ovaries after exposed to BPA for 7 days, and testes following 35-day exposure. Moreover, the global level of DNA methylation was also significantly reduced in testes after exposed to 15μg/L BPA for 7 days. Besides the alteration of the global level of DNA methylation, varying degrees of transcriptional changes of dnmts, gnmt and tets were detected in gonads of D. rerio under BPA exposure. The present study suggested that BPA might cause the global DNA demethylation in gonads of zebrafish by regulating the transcriptional changes of the DNA methylation/demethylation-associated genes (dnmts, gnmt, and tets). PMID:27101439

  20. Significance of metabolite extraction method for evaluating sulfamethazine toxicity in adult zebrafish using metabolomics.

    PubMed

    De Sotto, Ryan; Medriano, Carl; Cho, Yunchul; Seok, Kwang-Seol; Park, Youngja; Kim, Sungpyo

    2016-05-01

    Recently, environmental metabolomics has been introduced as a next generation environmental toxicity method which helps in evaluating toxicity of bioactive compounds to non-target organisms. In general, efficient metabolite extraction from target cells is one of the keys to success to better understand the effects of toxic substances to organisms. In this regard, the aim of this study is (1) to compare two sample extraction methods in terms of abundance and quality of metabolites and (2) investigate how this could lead to difference in data interpretation using pathway analysis. For this purpose, the antibiotic sulfamethazine and zebrafish (Danio rerio) were selected as model toxic substance and target organism, respectively. The zebrafish was exposed to four different sulfamethazine concentrations (0, 10, 30, and 50mg/L) for 72h. Metabolites were extracted using two different methods (Bligh and Dyer and solid-phase extraction). A total of 13,538 and 12,469 features were detected using quadrupole time-of-flight liquid chromatography mass spectrometry (QTOF LC-MS). Of these metabolites, 4278 (Bligh and Dyer) and 332 (solid phase extraction) were found to be significant after false discovery rate adjustment at a significance threshold of 0.01. Metlin and KEGG pathway analysis showed comprehensive information from fish samples extracted using Bligh and Dyer compared to solid phase extraction. This study shows that proper selection of sample extraction method is critically important for interpreting and analyzing the toxicity data of organisms when metabolomics is applied. PMID:26827276

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

    PubMed

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

    2015-08-01

    Cannabidiol (CBD) has been investigated in a wide spectrum of clinical approaches due to its psychopharmacological properties. CBD has low affinity for cannabinoid neuroreceptors and agonistic properties to 5-HT receptors. An interaction between cannabinoid and purinergic receptor systems has been proposed. The purpose of this study is to evaluate CBD properties on memory behavioral and locomotor parameters and the effects of pre-treatment of adenosine receptor blockers on CBD impacts on memory using adult zebrafish. CBD (0.1, 0.5, 5, and 10mg/kg) was tested in the avoidance inhibitory paradigm and anxiety task. We analyzed the effect of a long-term caffeine pre-treatment (~20mg/L - four months). Also, acute block of adenosine receptors was performed in co-administration with CBD exposure in the memory assessment. CBD promoted an inverted U-shaped dose-response curve in the anxiety task; in the memory assessment, CBD in the dose of 5mg/Kg promoted the strongest effects without interfering with social and aggressive behavior. Caffeine treatment was able to prevent CBD (5mg/kg) effects on memory when CBD was given after the training session. CBD effects on memory were partially prevented by co-treatment with a specific A2A adenosine receptor antagonist when given prior to or after the training session, while CBD effects after the training session were fully prevented by adenosine A1 receptor antagonist. These results indicated that zebrafish have responses to CBD anxiolytic properties that are comparable to other animal models, and high doses changed memory retention in a way dependent on adenosine. PMID:26099242

  2. Sustained Action of Developmental Ethanol Exposure on the Cortisol Response to Stress in Zebrafish Larvae and Adults

    PubMed Central

    Baiamonte, Matteo; Brennan, Caroline H.; Vinson, Gavin P.

    2015-01-01

    Background Ethanol exposure during pregnancy is one of the leading causes of preventable birth defects, leading to a range of symptoms collectively known as fetal alcohol spectrum disorder. More moderate levels of prenatal ethanol exposure lead to a range of behavioural deficits including aggression, poor social interaction, poor cognitive performance and increased likelihood of addiction in later life. Current theories suggest that adaptation in the hypothalamo-pituitary-adrenal (HPA) axis and neuroendocrine systems contributes to mood alterations underlying behavioural deficits and vulnerability to addiction. In using zebrafish (Danio rerio), the aim is to determine whether developmental ethanol exposure provokes changes in the hypothalamo-pituitary-interrenal (HPI) axis (the teleost equivalent of the HPA), as it does in mammalian models, therefore opening the possibilities of using zebrafish to elucidate the mechanisms involved, and to test novel therapeutics to alleviate deleterious symptoms. Results and Conclusions The results showed that developmental exposure to ambient ethanol, 20mM-50mM 1-9 days post fertilisation, had immediate effects on the HPI, markedly reducing the cortisol response to air exposure stress, as measured by whole body cortisol content. This effect was sustained in adults 6 months later. Morphology, growth and locomotor activity of the animals were unaffected, suggesting a specific action of ethanol on the HPI. In this respect the data are consistent with mammalian results, although they contrast with the higher corticosteroid stress response reported in rats after developmental ethanol exposure. The mechanisms that underlie the specific sensitivity of the HPI to ethanol require elucidation. PMID:25875496

  3. Inflammation is detrimental for neurogenesis in adult brain

    NASA Astrophysics Data System (ADS)

    Ekdahl, Christine T.; Claasen, Jan-Hendrik; Bonde, Sara; Kokaia, Zaal; Lindvall, Olle

    2003-11-01

    New hippocampal neurons are continuously generated in the adult brain. Here, we demonstrate that lipopolysaccharide-induced inflammation, which gives rise to microglia activation in the area where the new neurons are born, strongly impairs basal hippocampal neurogenesis in rats. The increased neurogenesis triggered by a brain insult is also attenuated if it is associated with microglia activation caused by tissue damage or lipopolysaccharide infusion. The impaired neurogenesis in inflammation is restored by systemic administration of minocycline, which inhibits microglia activation. Our data raise the possibility that suppression of hippocampal neurogenesis by activated microglia contributes to cognitive dysfunction in aging, dementia, epilepsy, and other conditions leading to brain inflammation.

  4. Zebrafish models of Tauopathy

    PubMed Central

    Bai, Qing; Burton, Edward A.

    2016-01-01

    Tauopathies are a group of incurable neurodegenerative diseases, in which loss of neurons is accompanied by intracellular deposition of fibrillar material composed of hyper phosphorylated forms of the microtubule associated protein Tau. A zebrafish model of Tauopathy could complement existing murine models by providing a platform for genetic and chemical screens, in order to identify novel therapeutic targets and compounds with disease-modifying potential. In addition, Tauopathy zebrafish would be useful for hypothesis-driven experiments, especially those exploiting the potential to deploy in vivo imaging modalities. Several considerations, including conservation of specialized neuronal and other cellular populations, and biochemical pathways implicated in disease pathogenesis, suggest that the zebrafish brain is an appropriate setting in which to model these complex disorders. Novel transgenic zebrafish lines expressing wild-type and mutant forms of human Tau inCNS neurons have recently been reported. These studies show evidence that human Tau undergoes disease-relevant changes in zebrafish neurons, including somato-dendritic relocalization, hyper phosphorylation and aggregation. In addition, preliminary evidence suggests that Tau transgene expression can precipitate neuronal dysfunction and death. These initial studies are encouraging that the zebrafish holds considerable promise as a model in which to study Tauopathies. Further studies are necessary to clarify the phenotypes of transgenic lines and to develop assays and models suitable for unbiased high-throughput screening approaches. This article is part of a Special Issue entitled Zebrafish Models of Neurological Diseases. PMID:20849952

  5. Effects of embryonic ethanol exposure at low doses on neuronal development, voluntary ethanol consumption and related behaviors in larval and adult zebrafish: Role of hypothalamic orexigenic peptides.

    PubMed

    Sterling, M E; Chang, G-Q; Karatayev, O; Chang, S Y; Leibowitz, S F

    2016-05-01

    Embryonic exposure to ethanol is known to affect neurochemical systems in rodents and increase alcohol drinking and related behaviors in humans and rodents. With zebrafish emerging as a powerful tool for uncovering neural mechanisms of numerous diseases and exhibiting similarities to rodents, the present report building on our rat studies examined in zebrafish the effects of embryonic ethanol exposure on hypothalamic neurogenesis, expression of orexigenic neuropeptides, and voluntary ethanol consumption and locomotor behaviors in larval and adult zebrafish, and also effects of central neuropeptide injections on these behaviors affected by ethanol. At 24h post-fertilization, zebrafish embryos were exposed for 2h to ethanol, at low concentrations of 0.25% and 0.5%, in the tank water. Embryonic ethanol compared to control dose-dependently increased hypothalamic neurogenesis and the proliferation and expression of the orexigenic peptides, galanin (GAL) and orexin (OX), in the anterior hypothalamus. These changes in hypothalamic peptide neurons were accompanied by an increase in voluntary consumption of 10% ethanol-gelatin and in novelty-induced locomotor and exploratory behavior in adult zebrafish and locomotor activity in larvae. After intracerebroventricular injection, these peptides compared to vehicle had specific effects on these behaviors altered by ethanol, with GAL stimulating consumption of 10% ethanol-gelatin more than plain gelatin food and OX stimulating novelty-induced locomotor behavior while increasing intake of food and ethanol equally. These results, similar to those obtained in rats, suggest that the ethanol-induced increase in genesis and expression of these hypothalamic peptide neurons contribute to the behavioral changes induced by embryonic exposure to ethanol. PMID:26778786

  6. [Chemotherapy for brain tumors in adult patients].

    PubMed

    Weller, M

    2008-02-01

    Chemotherapy has become a third major treatment option for patients with brain tumors, in addition to surgery and radiotherapy. The role of chemotherapy in the treatment of gliomas is no longer limited to recurrent disease. Temozolomide has become the standard of care in newly diagnosed glioblastoma. Several ongoing trials seek to define the role of chemotherapy in the primary care of other gliomas. Some of these studies are no longer only based on histological diagnoses, but take into consideration molecular markers such as MGMT promoter methylation and loss of genetic material on chromosomal arms 1p and 19q. Outside such clinical trials chemotherapy is used in addition to radiotherapy, e.g., in anaplastic astrocytoma, medulloblastoma or germ cell tumors, or as an alternative to radiotherapy, e.g., in anaplastic oligodendroglial tumors or low-grade gliomas. In contrast, there is no established role for chemotherapy in other tumors such as ependymomas, meningiomas or neurinomas. Primary cerebral lymphomas are probably the only brain tumors which can be cured by chemotherapy alone and only by chemotherapy. The chemotherapy of brain metastases follows the recommendations for the respective primary tumors. Further, strategies of combined radiochemotherapy using mainly temozolomide or topotecan are currently explored. Leptomeningeal metastases are treated by radiotherapy or systemic or intrathecal chemotherapy depending on their pattern of growth. PMID:18253773

  7. Gelsolin is a dorsalizing factor in zebrafish

    PubMed Central

    Kanungo, Jyotshnabala; Kozmik, Zbynek; Swamynathan, Shivalingappa K.; Piatigorsky, Joram

    2003-01-01

    The gene for gelsolin (an actin-binding, cytoskeletal regulatory protein) was shown earlier to be specialized for high corneal expression in adult zebrafish. We show here that zebrafish gelsolin is required for proper dorsalization during embryogenesis. Inhibition of gelsolin expression by injecting fertilized eggs with a specific morpholino oligonucleotide resulted in a range of concentration-dependent ventralized phenotypes, including those lacking a brain and eyes. These were rescued by coinjection of zebrafish gelsolin or chordin (a known dorsalizing agent) mRNAs, or human gelsolin protein. Moreover, injection of gelsolin mRNA or human gelsolin protein by itself dorsalized the developing embryos, often resulting in axis duplication. Injection of the gelsolin-specific morpholino oligonucleotide enhanced the expression of Vent mRNA, a ventral marker downstream of bone morphogenetic proteins, whereas injection of gelsolin mRNA enhanced the expression of chordin and goosecoid mRNAs, both dorsal markers. Our results indicate that gelsolin also modulates embryonic dorsal/ventral pattern formation in zebrafish. PMID:12629212

  8. Bilateral Brain Regions Associated with Naming in Older Adults

    ERIC Educational Resources Information Center

    Obler, Loraine K.; Rykhlevskaia, Elena; Schnyer, David; Clark-Cotton, Manuella R.; Spiro, Avron, III; Hyun, JungMoon; Kim, Dae-Shik; Goral, Mira; Albert, Martin L.

    2010-01-01

    To determine structural brain correlates of naming abilities in older adults, we tested 24 individuals aged 56-79 on two confrontation-naming tests (the Boston Naming Test (BNT) and the Action Naming Test (ANT)), then collected from these individuals structural Magnetic-Resonance Imaging (MRI) and Diffusion Tensor Imaging (DTI) data. Overall,…

  9. Induction of Female-to-Male Sex Change in Adult Zebrafish by Aromatase Inhibitor Treatment

    NASA Astrophysics Data System (ADS)

    Takatsu, Kanae; Miyaoku, Kaori; Roy, Shimi Rani; Murono, Yuki; Sago, Tomohiro; Itagaki, Hideyuki; Nakamura, Masaru; Tokumoto, Toshinobu

    2013-12-01

    This study investigated whether undifferentiated germ and/or somatic stem cells remain in the differentiated ovary of a species that does not undergo sex changes under natural conditions and retain their sexual plasticity. The effect of aromatase inhibitor (AI)-treatment on sexually mature female zebrafish was examined. A 5-month AI treatment caused retraction of the ovaries after which testes-like organs appeared, and cyst structures filled with spermatozoa-like cells were observed in sections of these tissues. Electron microscopic observations revealed that these cells appeared as large sperm heads without tails. Sperm formation was re-examined after changing the diet to an AI-free food. A large number of normal sperm were obtained after eight weeks, and no formation of ovarian tissue was observed. Artificial fertilization using sperm from the sex-changed females was successful. These results demonstrated that sex plasticity remains in the mature ovaries of this species.

  10. Pedophilic brain potential responses to adult erotic stimuli.

    PubMed

    Knott, Verner; Impey, Danielle; Fisher, Derek; Delpero, Emily; Fedoroff, Paul

    2016-02-01

    Cognitive mechanisms associated with the relative lack of sexual interest in adults by pedophiles are poorly understood and may benefit from investigations examining how the brain processes adult erotic stimuli. The current study used event-related brain potentials (ERP) to investigate the time course of the explicit processing of erotic, emotional, and neutral pictures in 22 pedophilic patients and 22 healthy controls. Consistent with previous studies, early latency anterior ERP components were highly selective for erotic pictures. Although the ERPs elicited by emotional stimuli were similar in patients and controls, an early frontal positive (P2) component starting as early as 185 ms was significantly attenuated and slow to onset in pedophilia, and correlated with a clinical measure of cognitive distortions. Failure of rapid attentional capture by erotic stimuli suggests a relative reduction in early processing in pedophilic patients which may be associated with relatively diminished sexual interest in adults. PMID:26683083