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1
Goldsmith, P. "Zebrafish models of retinal degeneration." Thesis, University of Cambridge, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.599478.
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This project has explored whether zebrafish can model human neurological disease, using retinal degeneration as the prototype. The retinal degenerations are a major cause of morbidity, being the commonest cause of blindness in the Western world. Age-related macular degeneration, the commonest retinal degeneration, affects 1 in 20 of the population, and is not treatable. The project began with an F2 mutagenesis screen for candidate blind fish, using an optokinetic assay. Blind fish were characterised using a variety of histological techniques and transplantation studies. Two strains of fish were then selected for more detailed genetic analysis, as they appeared to have a photoreceptor degeneration resembling human retinal degenerations. This involved further mapping of the genes responsible for the phenotype in these two mutants and candidate gene selection.
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Smith, Laura Lindsay. "Zebrafish Models of Congenital Myopathy." Thesis, Harvard University, 2015. http://nrs.harvard.edu/urn-3:HUL.InstRepos:23845448.
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The congenital myopathies are a diverse group of inherited neuromuscular disorders that manifest as skeletal muscle weakness at birth or in infancy, and are classically defined by the predominant morphological features observed on muscle biopsy. The goals of this dissertation were to better understand the pathophysiology behind these devastating diseases and to identify new therapeutic approaches through the use of faithful vertebrate models.
Due to their proliferative capacity, transparency, and well-characterized genome, zebrafish represent a robust vertebrate model system to study muscle development. In the first part of this work, we created and characterized a novel zebrafish model of centronuclear myopathy using antisense morpholinos targeting the bridging integrator 1 (bin1) gene. Bin1 morphant skeletal muscles revealed structural defects reported in human biopsies, and live calcium imaging offered new mechanistic insights linking abnormal triads to impairments in intracellular signaling.
Later studies focused on two forms of core myopathy, and utilized stable zebrafish models to guide development of targeted and effective therapies. We began by using TALE nucleases to generate germ line mutations in the zebrafish selenoprotein N (sepn1) gene, and in doing so created the first vertebrate to accurately model human SEPN1-related myopathy (SEPN1-RM). Sepn1 zebrafish mutants exhibited morphological abnormalities, reduced contractile strength, and skeletal muscle “cores” under electron microscopy. We then showed that the sepn1 phenotype could be ameliorated by pharmacological inhibition of a thiol oxidase localized at the sarcoplasmic reticulum. These data served as the first in vivo evidence to indicate that reactive oxygen species significantly contribute to SEPN1-RM, and may do so by impairing calcium re-uptake following muscle contraction. Finally, we performed a medium-throughput chemical screen on the closely related relatively relaxed (ryr1b) zebrafish, and identified JAK-STAT cytokine signaling as a druggable molecular pathway relevant to these pathologies.
In summary, these studies increase our knowledge of the affected systems in both centronuclear and core myopathies, and provide strong in vivo support that these conditions arise from defects in skeletal muscle excitation-contraction coupling. This work also further establishes zebrafish-based small molecule screens as a powerful tool for lead compound identification and drug development in human genetic disease.Medical Sciences
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Pressley, Meagan E. "Infectious Disease Models for the Zebrafish." Fogler Library, University of Maine, 2004. http://www.library.umaine.edu/theses/pdf/PressleyME2004.pdf.
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Simms, Roslyn Jane. "Zebrafish models of cystic kidney disease related ciliopathies." Thesis, University of Newcastle upon Tyne, 2013. http://hdl.handle.net/10443/2473.
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Cystic kidney diseases are a fascinating cluster of discrete conditions and an important, common cause of established renal failure. Both isolated and syndromic inherited cystic kidney diseases are known to be linked by their pathogenesis involving ciliary dysfunction. Interestingly to date, all mutated genes which have been related to cystic kidney disease, encode proteins which are located on cilia, the basal body or centrosomes and are required for ciliary function. To date, over 50 causal genes have been identified and are capable of causing additional disease phenotypes, such as neurological disorders and blindness, often of variable severity. Understanding this clinical heterogeneity may considerably guide appropriate genetic counselling and screening of patients for relevant complications. Zebrafish are a well-recognised animal model, their advantages of: transparency; conserved genome; representative kidney and rapid external development; make them useful for studying organogenesis in the context of disease. Furthermore the ability to perform combined gene knockdown in zebrafish, to study the effect of oliogenicity, which was proposed to influence clinical phenotypes in cystic kidney disease related ciliopathies, was of interest. Using zebrafish models, this work studied the impact of four key genes, independently and in combination: ahi1, cc2d2a, nphp6 and mks3 on the development of cystic kidney disease and ciliopathy phenotypes, to resemble the human diseases nephronophthisis (NPHP), Joubert syndrome (JBTS) and Meckel Gruber syndrome, (MKS). A frequent finding in zebrafish morphants was a reduction in the number of cilia, which was usually associated with abnormal development of left-right body patterning and cystic kidney disease. Additionally, combined gene knockdown of: nphp6 and cc2d2a; ahi1 and cc2d2a; ahi1 and nphp6 was associated with a synergistic increase in disease phenotypes, suggesting an interaction between these genes. In conclusion, zebrafish are a powerful developmental model to study and ideally improve understanding of cystic kidney disease related ciliopathies.
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Novorol, Claire. "Microcephaly models in the developing zebrafish retinal neuroepithelium." Thesis, University of Cambridge, 2014. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.648860.
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Daggett, Jenny. "Evaluation and characterisation of two zebrafish models of schizophrenia." Thesis, University of St Andrews, 2016. http://hdl.handle.net/10023/9603.
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Cognitive deficits are the single strongest predictor of the functional outcome in patients with schizophrenia. Current treatments are largely ineffective in improving cognitive impairments and promising pre-clinical research has mostly failed to translate clinically. Despite the advances provided by rodent models, the neurobiological basis of cognitive deficits in schizophrenia is poorly understood. Therefore, this thesis proposes a zebrafish model for studying cognitive impairments of schizophrenia. Although more evolutionarily distant to humans compared to the rat, the zebrafish has emerged as a popular vertebrate model of human disorders due to its genetic tractability, complex nervous system and elaborate behavioural repertoire. We investigated the effects of genetic alterations and neurodevelopmental disruption on behaviour and learning in zebrafish. Using both disc1 mutant lines and sub-chronic phencyclidine (PCP) on larvae from 6-10 dpf, we were able to assess behavioural changes as a function of developmental age. In particular, this thesis aimed to develop appropriate behavioural assays to assess zebrafish learning and executive function relevant to disorders seen in human patients with schizophrenia. It was possible to demonstrate robust learning across several domains, namely, reversal, classical avoidance and non-associative learning, alongside locomotor and anxiety-related behaviours. There were varied deficits associated with each of the two – genetic (disc1 gene mutation) and environmental (sub-chronic PCP) – manipulations, consistent with observations in rat research. Together, the research in this thesis demonstrates that a zebrafish model exhibits behaviour resembling that of mammalian models of schizophrenia and provides a foundation for the utility of zebrafish in examining cognitive impairments associated with schizophrenia.
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Ganis, Jared Jason. "Regulators of hemoglobin switching in zebrafish and human models." Thesis, Harvard University, 2014. http://dissertations.umi.com/gsas.harvard:11391.
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Hemoglobin switching is a developmental process involving the dynamic transcriptional regulation of multiple globin genes. This molecular process involves multiple layer of complexity, and elucidating new mechanisms in this process will result in a more complete understanding of general gene regulation and will likely have direct clinical implications for hemoglobinopathies, such as sickle cell anemia. In this dissertation, I develop and characterize a new model for hemoglobin switching, the zebrafish. I defined and fully annotated the two zebrafish globin loci, termed major and minor loci. Both loci contain α– and β–genes oriented in a head–to–head fashion. Characterization of the globin expression pattern precisely defined the timing of normal switching and demonstrated that zebrafish, like humans, have two globin switches. The locus control region for the major locus was identified and in conjunction with a proximal promoter was able to generate robust, erythroid–specific expression in a transgenic line.
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Albacker, Colleen Elizabeth. "Chromatin-Modifying Factors in Zebrafish Models of Rhabdomyosarcoma and Hematopoiesis." Thesis, Harvard University, 2012. http://dissertations.umi.com/gsas.harvard:10098.
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Epigenetics, or the reversible and heritable marks of gene regulation not including DNA sequence, encompasses modifications on both the DNA and histones and is as important as the DNA sequence itself. Gene transcription, DNA repair, DNA replication, and the cell cycle are each impacted by the chromatin structure. A variety of enzymes modulate these modifications, and a suite of factors interacts with them to aid in promoting or inhibiting cellular functions. Many of these chromatin-modifying factors are deregulated in cancer, making them novel therapeutic targets. This dissertation describes the identification of an H3K9 histone methyltransferase, SUV39H1, as a suppressor of rhabdomyosarcoma formation in zebrafish. This suppressor is dependent on the methyltransferase domain of the enzyme, ruling out any scaffold effects since this enzyme is a part of a multiprotein complex. SUV39H1-overexpressing and control tumors share many of the same characteristics, including proliferation rate, muscle differentiation state, and tumor growth rate. The tumor suppressive phenotype cannot be rescued by alterations in the downstream muscle program alone. However, SUV39H1-overexpressing fish initiate fewer tumors, which results in the observed suppressive phenotype. This initiation defect occurs between 5 and 7 days of life in the zebrafish, likely by impacting cyclin B1 expression. This dissertation also describes the development of a novel F1 transgenic screening strategy in the zebrafish. This approach was utilized to screen a variety of chromatin-modifying factors for their effects on hematopoietic development. The developed strategy will have future applications as a zebrafish screening tool. Our data suggest that chromatin-modifying factors play an important role in rhabdomyosarcoma and illustrate the use of the zebrafish in discovering genes involved in tumorigenesis and hematopoiesis.
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Dong, Bo. "High-throughput image analysis of zebrafish models of Parkinson's disease." Thesis, University of Sheffield, 2017. http://etheses.whiterose.ac.uk/18090/.
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Light microscopy can be used to advance our understanding of the molecular and cellular biology related to human health and diseases. As a powerful new vertebrate model, zebrafish have been used in various research areas, particularly in cancer and Parkinson's disease research. Large-scale data extraction from microscopy is highly attractive because it enables unbiased multivariate analysis that could lead to systems medicine approaches. To obtain useful information from large-scale data, high-throughput image analysis methods and applications are desperately required. In this thesis, we have explored methods and developed applications for highthroughput light microscopy zebrafish image analysis, including addressing the key problems related to three-dimensional (3D) deconvolution/deblurring, robust feature detection and description, and object counting. In biological image analysis, dealing with out-of-focus light noise, low image quality, large-scale dataset, illumination, overlapping, occlusion and insufficient prior knowledge remains challenging. Methods to address the following problems have been presented in this thesis. The low image quality of fluorescence microscopy images is addressed in Chapter 3. Owing to the limitations of light microscopes, the whole imaging process can be considered as a convolution between the object and the point spread function. Out-of-focus light and noise cause loss of detail in captured images. Deconvolution is an ill-posed inverse problem; thus, regularization methods are required to obtain better results. A maximum a posterior approach with a novel regularisation strategy to remove out-of-focus blur in 3D fluorescence microscopy images is introduced in Chapter 3. The second problem is dyed dopaminergic neurone detection in zebrafish RGB optical sections (Chapter 4). Owing to the large-scale of the image, low image quality, irregular appearance of neurones and touching situations, manually counting individual neurones via the microscope can be labour-intensive, time-consuming, subjective, and error-prone. To solve this problem, this thesis explores different methods to detect individual neurones in 3D zebrafish RGB images, including using detectors with many different handcrafted features and features learned automatically from deep learning architectures. An additional class-imbalanced problem is discovered during the experiments involving the training of patch-based deep learning techniques using a large-scale dataset that contains a limited number of positive samples. To solve this problem, a dynamic cascade framework with deep learning architectures is designed. The last problem is cell counting in fluorescent microscopy images (Chapter 5). Detecting individual cells in two-dimensional (2D) fluorescence microscopy images is difficult owing to overlap. Rather than counting-by-detection methods, a countingby- regression method with an interactive interface for cell counting in a fluorescent image is proposed. Sparse Bayesian Poisson regression based on a Relevance Vector Machine framework is also proposed. The proposed framework enables accurate counting of a discrete number of cells and leads to much sparser models, which results in faster performance and maintains a comparable generalisation error.
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Metzner, Aylin. "A study of ADPKD pathogenesis and treatment in zebrafish models." Thesis, University of Sheffield, 2016. http://etheses.whiterose.ac.uk/16020/.
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Precazzini, Francesca. "Zebrafish models of uveal and cutaneous melanoma for preclinical studies." Doctoral thesis, Università degli studi di Trento, 2019. http://hdl.handle.net/11572/245749.
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Uveal melanoma (UM) is the most common primary cancer of the eye and its prognosis is strongly influenced by the occurrence of metastases, which are both rapidly developing and mostly fatal. The most frequent driver mutations occur in a small number of genes including GNAQ, GNA11, BAP1, CYSLTR2 and SF3B1. Due to a lack of suitable animal models, the mechanism through which mutations in these genes cause or cooperate in UM initiation and progression is still largely unknown. We aimed at generating transgenic strains expressing the human mutant proteins in zebrafish uveal melanocytes, using the kita promoter. We used the binary Gal4/UAS system to express the mutant genes mentioned above. Moreover, we performed xenotransplantation experiments with uveal melanoma human and zebrafish cell lines in optically-clear, immunocompromised, zebrafish larvae. Transplanted fish developed melanoma near the site of transplantation in two weeks and showed metastatic growth within one month of age. This approach could be used for short-term assays in larvae, and be further developed for long-term uveal melanoma studies. In parallel, we performed a chemical screen using a transgenic model previously generated in our laboratory, where oncogenic RAS is expressed under the kita promoter. As adults transgenic kita:RAS develop cutaneous melanoma with high frequency and uveal melanoma with a much lower percentage. Larvae showed an increased number of melanocytes already at 3 days post fertilization (dpf) as the earliest evidence of abnormal melanocyte growth. Using this model we performed a chemical screen based on automated detection of a reduction of melanocytes number caused by any of the 1280 FDA or EMA approved drugs of the Prestwick library. The analysis showed that 55 molecules were able to reduce by 60% or more the number of melanocytes per embryo. We identified clotrimazole, as the best candidate. The molecule is an azole derivative acting on the energetic metabolism of melanoma cells. We further tested two compounds for each of the 5 pharmacological classes, and a farnesyltransferase inhibitor (lonafarnib), that inhibits an essential post-translational modification of HRAS and suppresses the hyperpigmentation phenotype. Combinations of clotrimazole and lonafarnib showed the most promising results in zebrafish embryos, allowing a dose reduction of both drugs. We performed validation of these observations in the metastatic human melanoma cell line A375M, and in normal human epithelial melanocytes (NHEM) as control cells, in order to investigate the mechanism of action of clotrimazole in blocking the proliferation of transformed melanocytes. Viability assay and analysis of energy metabolism in clotrimazole treated cells show that this drug specifically affects melanoma cells in vitro and transformed melanocytes in vivo, having no effects on NHEM or wild type larvae. Similar effects were observed with another hit of the antifungal class, miconazole. Furthermore, we show that the effects of clotrimazole are mediated by the inhibition of hexokinase activity and suggest further testing of clotrimazole in combinatorial treatments. In conclusion, this thesis investigated different possibilities of modeling the rare cancer uveal melanoma in zebrafish, using both transgenic and transplantation approaches, and developed a pipeline for a high-throughput, semi-automated chemical screen in a zebrafish melanoma that identified clotrimazole and miconazole as targeting a metabolic vulnerability in melanoma cells.
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Lyon, Alison Nicole. "Generation and Analysis of Motor Neuron Disease Models in Zebrafish." The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1337276861.
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Toms, Maria. "Investigating the pathophysiology of KCNJ13 and USH2A retinopathies using zebrafish models." Thesis, University College London (University of London), 2018. http://discovery.ucl.ac.uk/10053968/.
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Inherited retinal diseases (IRDs) encompass a large group of clinically and genetically heterogeneous diseases that affect approximately 1 in 3000 people, representing an important cause of severe visual loss in the human population. In the past two decades, significant contributions towards our understanding of these disorders has included the identification of over 250 causative genes. In this thesis, the pathophysiology of IRDs associated with KCNJ13 and USH2A was investigated using relevant zebrafish models. Mutations in KCNJ13 are associated with Leber congenital amaurosis, the most severe IRD subtype that causes blindness in childhood. KCNJ13 encodes the Kir7.1 channel, a potassium channel expressed on the apical retinal pigment epithelium (RPE). Longitudinal assessment of the kcnj13 mutant zebrafish (obelixtd15) revealed a late onset retinal degeneration at 12 months with retinovascular abnormalities, and a corresponding decline of visual function. Ultrastructural examination of the obelixtd15 RPE uncovered changes in phagosome clearance and mitochondrial growth prior to notable degeneration which indicate that the disease may be a primary phagosome failure with a secondary failure in mitochondrial physiology. In addition, gene and protein expression changes consistent with altered mitochondrial activity and retinal stress were observed. The alterations in the RPE are reminiscent of those seen in age-related macular degeneration and highlight potential therapeutic targets for KCNJ13 retinopathy. USH2A mutations are the most common cause of Usher Syndrome, characterised by combined retinitis pigmentosa and sensorineural deafness. USH2A encodes the large transmembrane protein, usherin, expressed in the photoreceptors and cochlear hair cells. CRISPR/Cas9 gene targeting was used to create an ush2a mutant zebrafish line, which showed a slowly progressive photoreceptor degeneration from 6 months with mislocalisation of rhodopsin. Developmental abnormalities in the neuromast and inner ear hair cells were also identified in ush2a larvae. The findings suggest that ush2a is involved in rhodopsin trafficking and rod photoreceptor maintenance in the zebrafish retina, as well as playing a role in mechanosensory system development. Overall, the ush2a zebrafish phenotype showed consistency with the clinical phenotype in USH2A patients, indicating the value of this model for gaining further insight into the disease pathophysiology.
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Reimers, Mark J. "Ethanol-dependent developmental toxicity in zebrafish /." Connect to full text via ProQuest. IP filtered, 2005.
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Zaksauskaite, Ringaile. "Analysis of chromosomal protein-linked break repair in zebrafish and mammalian models." Thesis, University of Sheffield, 2018. http://etheses.whiterose.ac.uk/20765/.
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Nicolas, Hannah Almira. "Understanding Molecular Mechanisms of Striated Muscle Laminopathies Using Cellular and Zebrafish Models." Thesis, Université d'Ottawa / University of Ottawa, 2020. http://hdl.handle.net/10393/41008.
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Toulis, Vasileios. "Zebrafish and mouse models for studying deubiquitinating enzyme genes as candidates for retinal dystrophies." Doctoral thesis, Universitat de Barcelona, 2020. http://hdl.handle.net/10803/674031.
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The retina consists of several structured layers of highly specialized neurons that capture and process light stimuli enabling vision. Such a fine architecture turns retinal differentiation into an extremely complex event that must be accurately regulated. The ubiquitin-proteasome system (UPS) is considered one of the most dynamic and versatile mechanisms of protein regulation in eukaryotic cells. As ubiquitination is reversible, deubiquitinating enzymes (DUBs) play a major regulatory role in the UPS. Despite the importance of proteostasis and the UPS in health and disease, a more comprehensive in-depth analysis of DUB expression and function on particular tissues or organs, such as the retina, is still missing.
Combining expression quantification, mRNA localization assays and functional analyses in animal and cellular models, we analyzed the function of several DUB genes in the retina to identify DUBs that regulate important retinal cell mechanisms, explore their relevance in retinal function in health and disease, and finally, posit them as new potential candidate genes for retinal dystrophies.
Taking into consideration our results in the expression levels and pattern of DUBs in the retina, we first selected USP45 to perform functional assays in animal models in order to define its role and function in the retina. By morpholino-knockdown of usp45 in zebrafish embryos, our results showed moderate to severe eye morphological defects, eye size reduction, small body size with small tail or without tail, and disruption in notochord formation. There is also defective lamination and formation of the retinal
structures, with no distinguishable layers and smaller retinas.
Overall, our results supported the relevance of USP45 in the normal development and formation of the vertebrate retina, and we proposed this gene as a good candidate for causing hereditary retinal dystrophies, as later confirmed by other authors in several families.
We also selected ATXN3, a DUB gene that causes the dominant polyQ disease Spinocerebellar ataxia type 3 (SCA3), and we aimed to analyze its function in the retina. We showed that depletion of Atxn3 in zebrafish and mice caused retinal morphological and functional alterations with photoreceptor outer segment elongation, cone opsin mislocalization, and cone hyperexcitation upon light stimuli. A pool of ATXN3 resides at the basal body and axoneme of the photoreceptor cilium, where it controls the levels and recruitment of the regulatory proteins KEAP1 and HDAC6. Abrogation of Atxn3 expression causes delayed phagosome maturation in the retinal pigment epithelium. We propose that ATXN3 regulates two relevant biological processes in the retina, ciliogenesis and phagocytosis, by modulating microtubule polymerization and microtubule-dependent retrograde transport, and propose ATXN3 as a causative or modifier gene in retinal/macular dystrophies.
We further aimed to explore whether the SCA3 humanized mouse model showed specific retinal phenotype traits. We showed that polyQ-expanded ATXN3 protein formed a high number of progressive pathogenic aggregates in the retinal layers of transgenic Atxn3Q84 mice, and caused a decrease in the number of cone photoreceptors. Optical coherence tomography revealed a general decrease in the thickness of the retinal layers whereas retinal electrophysiological analyses showed a strong decrease in photoreceptor response to light, thus supporting severe retinal dysfunction in Atxn3Q84 mice. Similar analyses in human patients detected a correlation of retinal alterations with the number of CAG repeats and the age of onset of SCA3 symptoms. We propose that retinal alterations detected by non-invasive eye examination and electroretinography tests in SCA3 patients could serve as a valuable early-onset symptom and a biological marker of disease progression.
As a conclusion, our work posits several DUB genes as candidates for inherited retinal dystrophies, but further investigation is needed to dissect the function of DUBs in retinal cell differentiation, photoreceptor function, and retinal homeostasis.
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Hulbig, Veronica A. "Developing a Model for Bacterial Kidney Disease in the Zebrafish, Danio rerio." Fogler Library, University of Maine, 2010. http://www.library.umaine.edu/theses/pdf/HulbigVA2010.pdf.
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Tian, Li, and 田理. "Toxicological effects and mechanisms of selected foodborne toxins in medaka and zebrafish models." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2013. http://hdl.handle.net/10722/197111.
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Foodborne toxins include a variety of biologically produced toxins and process-induced toxicants. Among them, marine algal toxins in polluted seafood can induce serious harmful effects on human, while heat-induced toxicants remain in a wide range of food and chronically affect health. In this present study, several representative toxins from these two categories were selected and studied, they are, brevetoxins (PbTxs), saxitoxin (STX) and acrolein (ACR). During past decades, the molecular actions of these toxins have been well studied, however, their effects and mechanisms corresponding to their sublethal toxicity in vivo still need more investigation. Therefore, in our current study, we adopted medaka fish (Oryzias melastigma) and zebrafish (Danio rerio) as models to study the toxicological responses and pathways of these selected toxins at the molecular and cellular level.
In the first part of study, the targeted organs of medaka fish, gills and brains, were studied in medaka fish after exposure to the sublethal level of PbTx-1, the most potent neurotic shellfish poisoning (NSP) toxin. Facilitated by the two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ ionization tandem time-of-flight mass spectrometry (MALDI TOF/TOF MS), proteins affected by PbTx-1 in these organs were identified, including myosin like proteins, aldose reductase, gelsolin and keratin. Collectively, the proteins altered after exposure suggested the altered calcium ion binding process, and dysfunction in cytoskeleton assembly and metabolism.
After successfully applying the fish model and proteomic approaches in the NSP toxin study, we put emphasis on the developmental toxicity of foodborne toxins, as children are more sensitive and vulnerable to foodborne toxins. Among the marine algal toxins, paralytic shellfish poisoning (PSP) toxins are the commonest and most lethal ones with STX as the most potent PSP toxin. Therefore, we examined the developmental effects of STX using medaka model. By exposing medaka embryos from the early blastula stage onwards, which covered the main developmental stage of the central nervous system and somites, we found newly hatched medaka fish exhibit abnormal growth with longer body length and relatively smaller yolk sac size. High cell proliferation, neuron development, and metabolism were confirmed by whole-mount immunostaining and 2-DE. In summary, STX disturbs the normal growth of medaka embryos probably by affecting the metabolic rate in the exposed medaka embryos.
Opposite to STX, after exposure to ACR, a pollutant that ubiquitously exists in food and environment, the zebrafish exhibited delayed development. ACR is a known glutathione (GSH) depleting factor and oxidative stress inducer. Apart from developmental retardance, increased reactive oxygen species (ROS) and inflammation were found in newly hatched zebrafish, suggesting the developmental delay may be partly related to the ACR-induced oxidative stress.
In summary, findings from the present study highlighted the molecular responses and possible pathways of some selected foodborne toxins. Developmental effects and toxicity were also found after exposing to both the biologically produced toxin and the heat-produced toxicant. Our current study makes contributions to the knowledge on the hazardous effects of foodborne toxins in vivo, and provides useful information for the further study on the human health.published_or_final_version
Biological Sciences
Doctoral
Doctor of Philosophy
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Chhabria, Karishma. "Development of zebrafish and computational models of neurovascular coupling in health and disease." Thesis, University of Sheffield, 2018. http://etheses.whiterose.ac.uk/22833/.
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In this thesis, I have developed a novel zebrafish model of neurovascular coupling. Combining lightsheet imaging, compound transgenic zebrafish models and custom MATLAB based analysis pipelines, I characterised the neurovascular responses (neuronal calcium increases and change in red blood cell speed) in the optic tectum in response to visual stimulation. I determined the development stage at which neurovascular coupling in zebrafish larvae develops, followed by testing the requirement for nitric oxide or astrocyte cyclo-oxygenase in my model. I then used this model to investigate factors influencing neurovascular function. I first characterized the effect of glucose exposure and the role of nitric oxide in modulating neurovascular coupling. I then examined the effect of genetic mutation of Guanosine Triphosphate cyclohydrolase (an enzyme involved in nitric oxide and dopamine production in the brain) on neurovascular coupling. Finally, I have developed a minimal mathematical model of the neurovascular unit. To demonstrate the potential of this model I have simulated the effect of high blood glucose and low nitric oxide on neurovascular coupling and show this conforms with experimental data obtained in zebrafish.
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Rounding, Natalie. "Zebrafish C9orf72 loss-of-function models of Amyotrophic Lateral Sclerosis and Frontotemporal Dementia." Thesis, University of Sheffield, 2018. http://etheses.whiterose.ac.uk/19832/.
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Balow, Stephanie Ann. "Investigation of CHD7 Function in Developmental Models of CHARGE Syndrome." Case Western Reserve University School of Graduate Studies / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=case1396625345.
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Tekeli, Işil. "Bioengineering approach to study the role of cell migration during zebrafish heart regneration." Doctoral thesis, Universitat de Barcelona, 2016. http://hdl.handle.net/10803/396098.
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Zebrafish heart regeneration remains one of the most interesting phenomena of the 21st century. Considering the extremely high rate of deaths due to cardiovascular diseases in the developed countries, 1 out of every 3 people, understanding natural cardiac regeneration would address a worldwide challenge. Even though many aspects of zebrafish heart regeneration have been elucidated, there are still many open questions to be answered. Among these, the work presented here focuses on understanding cell migration mechanisms of cardiomyocytes and epicardial cells during heart regeneration.
The first approach involves the development of a cardiomyocyte-specific, photoinducible Cre/lox genetic labeling system and its use in lineage tracing of embryonic cardiomyocytes during heart development and regeneration. By using this method we showed that cardiomyocytes labeled in embryonic hearts survive and contribute to all myocardial layers of the adult zebrafish heart. Moreover, lineage tracing during regeneration showed that only cardiomyocytes immediately adjacent to the injury site contribute to the regeneration, and cardiomyocyte fate is extensively predetermined, with cells from each myocardial layer giving rise to cells that retain their layer identity in the regenerated myocardium.
Then, we showed that by coupling this labeling system to three-photon microscopy activation, we can perform prospective labeling, and increase the spatial resolution of our labeling system. Three-photon illumination has been used for in vivo imaging of deep structures, but whether it can be used for photo-activation had never been tested. Here we showed, theoretically and experimentally for the first time, that three-photon illumination is suitable for activating molecules in deep tissues and improving our system in terms of spatial resolution and prospective labeling.
The final approach consisted on developing an ex vivo experimental set up in order to investigate physical characteristics of epicardial cell migration during zebrafish heart regeneration. This method allowed us to measure physical features that are essential for cell migration such as migration velocity and traction forces of the epicardial explants obtained from zebrafish hearts.
All the approaches developed in this thesis offer new bioengineering tools to study zebrafish heart regeneration, and reveal new insights on this process. Moreover, these techniques present wide applicability to perform lineage tracing of other cell types during zebrafish heart regeneration or in other biological processes.Los humanos tienen una capacidad muy limitada para regenerar el corazón y, como consecuencia de ello, una de cada tres personas fallece debido a enfermedades cardíacas. En cambio, el pez cebra tiene una enorme capacidad intrínseca para restaurar grandes porciones de su corazón tras un daño, por lo que se ha convertido en uno de los modelos experimentales más usados para investigar la regeneración del corazón. Entender los mecanismos que controlan la regeneración cardíaca del pez cebra permitirá desarrollar estrategias para regenerar el corazón humano y buscar soluciones a estas enfermedades. Basándonos en este gran objetivo, el trabajo que forma el estudio de esta tesis doctoral detalla algunas aproximaciones de bioingeniería dirigidas a estudiar el papel de la migración celular de los cardiomiocitos y de las células del epicardio durante la regeneración del corazón del pez cebra. La primera aproximación es un sistema de seguimiento del linaje de los cardiomiocitos del pez cebra para investigar su destino durante la regeneración cardiaca. Para esto, se desarrolló un sistema de marcado genético de linaje con triple especificidad, de tipo celular, temporal y espacial, que permitió marcar los cardiomiocitos en diferentes partes del corazón. Utilizando esta técnica se demostró que durante la regeneración sólo los cardiomiocitos inmediatamente adyacentes a la lesión contribuyen al miocardio regenerado. A continuación, se utilizó la iluminación multifotón de tres fotones para fotoactivar cardiomiocitos in vivo con el objetivo de aumentar la resolución espacial del marcado genético. En este trabajo se demostró teóricamente y experimentalmente que la iluminación de tres-fotones supera problemas de dispersión y es capaz de realizar la fotoactivación, convirtiéndose en el primer ejemplo donde se consigue utilizar este tipo de iluminación para fotoactivar células in vivo. El uso de la iluminación de tres fotones en combinación con el sistema de marcado genético fotoinducible permitió marcar los cardiomiocitos de embriones de pez cebra de forma prospectiva. Por último, se desarrolló un sistema ex vivo para caracterizar el comportamiento migratorio de las células del epicardio del corazón del pez cebra con el fin de investigar las características físicas de la migración celular durante la regeneración. Este método permitió medir las características físicas esenciales para la migración celular, tales como la velocidad migratoria y las fuerzas de tracción en las células del epicardio.
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Pimentel, Angela Bairos. "Algorithm for the Parkinson’s disease behavioural models characterization using a biosensor." Master's thesis, Faculdade de Ciências e Tecnologia, 2012. http://hdl.handle.net/10362/8443.
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Dissertação para obtenção do Grau de Mestre em
Engenharia BiomédicaThe neurodegenerative disease, Parkinson’s Disease (PD) constitutes a major health problem in the modern world, and its impact on public health and society is expected to increase with the ongoing ageing of the human population. This disease is characterized by motor and non-motor manifestations that are progressive and ultimately refractory to therapeutic interventions. The degeneration of dopaminergic neurons emanating from the substantia nigra is largely responsible for the motor manifestations. Thus, understanding the behaviour related to this disease is an added value for the diagnosis and treatment of PD. Also, in vivo models are essential tools for deciphering the molecular mechanisms underpinning the neurodegenerative process. Zebrafish has several features that make this species a good candidate to study PD. In particular, the occurrence of behavioural phenotypes of treated animals with neurotoxin drugs that mimic the disease has been investigated. And, an electric biosensor, Marine On-line Biomonitor System (MOBS) is being used for the real-time quantification of such behaviour. This equipment allows quantifying the fish movements through signal processing algorithms. Specifically, the algorithm is used for the evaluation of fish locomotion detected by a series of bursts in the domain of MOBS that correspond to the zebrafish tail-flip activity. In this thesis we proceeded to the development of an algorithm affording a electrical signal discrimination between "healthy" and "ill" zebrafish and consequently improving the detection of parkinsonism-like phenotypes in zebrafish. The first approach was the improvement of the existent algorithm. However, the first analysis failed to distinguish between different behavioural phenotypes when fish were treated with the neurotoxin 6-hydroxydopamine (6-OHDA). Consequently, we generated a new algorithm based on Machine Learning techniques. As a result, the novel algorithm provided a classification over the health condition of the fish, if the same is "healthy" or "ill" with its respective probability and the level of activity of the fish in number of tail-flips per minute. The method Support Vector Machine (SVM)was useful for the classification of the fish events. The zero crossing rate parameter was used for the characterization of the swimming activities. The algorithm was also integrated in the platform Open Signals, and for a faster evaluation of the signals, the algorithm implementation included parallel programming methods. This algorithm is a useful tool to study behaviour in zebrafish. Not only it will allow a more realistic study over the PD research area but also test and assess new drugs that use zebrafish as animal model.
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Green, Jon Marc. "Development of novel transgenic zebrafish models and their application to studies on environmental oestrogens." Thesis, University of Exeter, 2016. http://hdl.handle.net/10871/27141.
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Oestrogenic chemicals have become increasingly associated with health effects in wildlife populations and humans. Transgenic animal models have been developed to understand the mechanisms by which these oestrogenic chemicals alter hormonal signalling pathways and how these alterations can lead to chronic health effects. The use of highly informative transgenic animal models will also result in better use and potential reduction of intact animals used in animal testing in line with the principles of the 3Rs. In this thesis work, two novel oestrogen responsive transgenic zebrafish models have been generated to investigate the effects of oestrogenic chemicals, identify their tissue targets and better understand the temporal dynamics of these responses. Both models express the pigment-free ‘Casper’ (a mutant line lacking skin pigment) phenotype, which facilitate identification of responding target tissues in the whole fish in all fish life stages (embryos to adults). The oestrogen response element green fluorescent (ERE-GFP)-Casper model was generated by crossing an established ERE-GFP line with the skin pigment free Casper line. The model generated is highly sensitive to oestrogenic chemicals, detecting responses to environmentally relevant concentrations of EE2, bisphenol A (BPA), genistein and nonylphenol. Use of the ERE-GFP- Casper model shows chemical type and concentration dependence for green fluorescent protein (GFP) induction and both spatial and temporal responses for different environmental oestrogens tested. A semi-automated (ArrayScan) imaging and image analysis system was also developed to quantify whole body fluorescence responses for a range of different oestrogenic chemicals in the new transgenic zebrafish model. The zebrafish model developed provides a sensitive and highly integrative system for identifying oestrogenic chemicals, their target tissues and effect concentrations for exposures in real time and across different life stages. It thus has application for chemical screening to better direct health effects analysis of environmental oestrogens and for investigating the functional roles of oestrogens in vertebrates. The second model generated was an ERE-Kaede-Casper line developed via crossing of the ERE-GFP-Casper line and a UAS-Kaede line and screening subsequent generations for a desired genotype and homozygous expression of the transgenes. Kaede is a photoconvertible fluorescent protein that initially fluoresces green in colour and can be permanently converted to red fluorescence upon short exposure to UV light. The model has a silenced skin pigmentation and high sensitivity to oestrogenic chemicals comparable with the previously developed ERE-GFP-Casper model. Use of this model has identified windows of tissue-specific sensitivity to ethinyloestradiol (EE2) for exposure during early-life (0-48hpf) and illustrated that exposure to oestrogen (EE2) during early life (0-48hpf) can enhance responsiveness (sensitivity) to different environmental oestrogens (EE2, genistein and bisphenol A) for subsequent exposures during development. These findings illustrate the importance of oestrogen exposure history in effects assessments and they have wider implications for the possible adverse effects associated with oestrogen exposure.
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Örn, Stefan. "The zebrafish as a model organism for evaluation of endocrine disrupters /." Uppsala : Department of Biomedical Sciences and Veterinary Public Health, Swedish University of Agricultural Sciences, 2006. http://epsilon.slu.se/200628.pdf.
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Wojciechowska, Sonia. "The conditional control of MITF reveals cellular subpopulations essential for melanoma survival and recurrence in new zebrafish models." Thesis, University of Edinburgh, 2018. http://hdl.handle.net/1842/29645.
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Melanoma is the most lethal type of skin cancer with over 132,000 cases occurring globally each year and continually rising incidence. BRAFV600E inhibitors have led to clinically significant improvements in outcomes for melanoma patients, yet many patients with metastatic melanoma rapidly succumb to the disease due to eventual chemoresistance or insensitivity to the drug. Thus, it is critical to identify new therapies that can act alone, or be combined with available treatments for enhanced efficacy and/or to overcome drug resistance. Evidence from human melanoma indicates that the melanocyte lineage is critical for melanoma survival and contributes to therapeutic resistance. MITF is a highly conserved “master melanocyte transcription factor” with a complex role in melanoma. Our lab has previously developed a temperature sensitive BRAFV600E mitfavc7 zebrafish melanoma model carrying a human oncogene and mitfavc7 splice site mutation that enables the conditional control of its endogenous activity by changes to water temperature. As part of my PhD project, I characterized and compared two new models developed since then: a very aggressive BRAFV600E mitfavc7p53M214K melanoma model with three driving mutations and a slower developing BRAF-independent mitfavc7p53M214K. I showed that the MITF activity is crucial for melanocyte survival in both models and that both mutated BRAF and p53 deficiency are oncogenic with low levels of MITF, and result in fish nevi and melanoma resembling the pathology of human disease. Both models are also relevant to a low-MITF subclass of human melanomas that emerged from a recent classification by The Cancer Genome Atlas Network. In addition, I established that, similarly to the BRAFV600Emitfavc7, complete inhibition of MITF activity leads to rapid tumour regression, but once its activity is restored the melanomas recur at the same site as the original tumour. I used histopathology studies and melanocyte lineage transgenes to identify and visualize subpopulations of cells remaining at the site of regression in these new zebrafish melanoma models. I hypothesised that these are the cells of origin for tumour recurrence (melanoma stem or progenitor cells), showed that some of them express a cancer stem cell marker aldehyde dehydrogenase, and attempted to target these subpopulations using 5-nitrofurans (a prodrug NFN1, shown previously by our lab to target ALDHhigh subpopulations in context of melanoma) in fish after melanoma regression. Finally, I also developed and described a new primary zebrafish melanoma cell line that I derived from one of these zebrafish tumours. This study is still in progress, but the cell line will be a useful tool for further investigation of these proposed melanoma progenitor cells in vitro, with potential applications for lineage tracing and transplantations.
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Ivarsson, Adam. "Expediting Gathering and Labeling of Data from Zebrafish Models of Tumor Progression and Metastasis Using Bespoke Software." Thesis, Linköpings universitet, Institutionen för datavetenskap, 2018. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-148691.
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In this paper I describe a set of algorithms used to partly automate the labeling and preparation of images of zebrafish embryos used as models of tumor progression and metastasis. These algorithms show promise for saving time for researchers using zebrafish in this way.
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Vishnolia, Krishan Kumar. "Development and charaterisation of 3 dimensional culture models for zebrafish (Danio rerio) skeletal muscle cells." Thesis, University of Bedfordshire, 2013. http://hdl.handle.net/10547/556396.
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Zebrafish (Danio rerio) have been extensively used over the past two decades to study muscle development, human myopathies and dystrophies, due to its higher degree of homology with human disease causing genes and genome. Despite its unique qualities, zebrafish have only been used as an in-vivo model for muscle development research, due to the limitations surrounding lack of a consistent isolation and culture protocol for zebrafish muscle progenitor cells in-vitro. Using different mammalian myoblast isolation protocols, a novel and robust protocol has been developed to successfully isolate and culture zebrafish skeletal muscle cells repeatedly and obtain differentiated long multi nucleated zebrafish myotubes. Commitment to myogenic lineage was confirmed by immuno-staining against muscle specific protein desmin, and expression pattern of different genetic markers regulating myogenesis. In order to recapitulate the in-vivo bio-physiological environment for zebrafish skeletal muscle cells in-vitro, these cells were successfully cultured in tissue engineered three dimensional (3D) constructs based on fibrin and collagen models. Maturation of tissue engineered collagen and fibrin based constructs was confirmed using the basic parameters described in the literature i.e. collagen three times greater contraction from the original width (Mudera, Smith et al. 2010) and fibrin constructs tightly coiled up to 4mm of diameter (Khodabukus, Paxton et al. 2007). In-vitro characterisation of zebrafish skeletal muscle cells showed hypertrophic growth of muscle mass compared to hyperplasic growth in-vivo as suggested for fish species in literature (Johnston 2006), which is different from human and other mammals. Comparative analysis of zebrafish muscle cells cultured in monolayer against cultured in 3D tissue engineered constructs showed significant increase in fusion index, nuclei per myotube (two-fold) and myotubes per microscopic frame (two-fold). Cells cultured in tissue engineered construct closely resembled in-vivo muscle in terms of their unidirectional orientation of myotubes. These tissue engineered 3D zebrafish skeletal muscle models could be used for various purposes such as drug screening, effect of different temperature extremes, studying underlined pathways involved in human diseases; and with further refinements it would potentially replace the need for studies on live fish in these areas.
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Yu, Xinge. "Stroke Study: Novel Animal Models and Innovative Treatment Strategy." Ohio University / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1450735023.
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Martin, Brit Leigh Martin. "Toward the use of whole, live developing zebrafish as models for skeletal and cardiac muscle contraction." The Ohio State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=osu1500611447591388.
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Avdesh, Avdesh. "Pharmacological modulation of learning and memory in mice and zebrafish: Implications for Alzheimer’s disease animal models." Thesis, Edith Cowan University, Research Online, Perth, Western Australia, 2013. https://ro.ecu.edu.au/theses/570.
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Learning and memory impairments are the clinical manifestations of several neurodegenerative disorders including Alzheimer’s disease (AD), the most common form of dementia affecting millions of people worldwide. Current AD treatments target cognitive decline or failure and provide only minor benefits across the array of clinical symptoms. The development of numerous treatment strategies have been trailed in pre-clinical, or human clinical trials. One of the potential candidates to be used for cognitive enhancement is CB1 cannabinoid receptor antagonist. CB1 cannabinoid receptor antagonists have been shown to improve learning and memory in rodents, whereas CB1 cannabinoid receptor agonists have been shown to impair memory. The project has overall aims of (1) investigating the role of CB1 receptors in learning and memory, and assessing the potential of a CB1 receptor antagonist (SR141716A) as a novel cognitive enhancer in mice models; (2) Assessing the potential of zebrafish as an alternative in vivo model for the high-through put assessment of learning and memory. The involvement of cannabinoid CB1 (CB1) receptors in associative learning in two strains of mice (i.e., C57BL/6J and C3H/HeJ) were first investigated. Here, the effects of the cannabinoid CB1 receptor agonist (i.e., CP55940) and antagonist (i.e., SR141716A) on associative learning were assessed in mice using eight-arm radial maze. The results showed that the CB1 receptor agonist impaired reference memory in C57BL/6J but not in C3H/HeJ strains, and the former effect was probably mediated by CB1 receptors since the selective antagonist, SR141716A, reversed these effects. The CP55940 agonist also impaired working memory selectively in the C57BL/6J mice, which was not affected by SR141716A. This study provided evidence for a strain- specific effect of CP55940 on working memory that is likely to be independent of the CB1 receptors. Cannabinoid receptor effects were also evaluated on non-associative learning in mice by measuring habituation of acoustic startle reflex (ASR). CB1 receptor agonist and antagonist's effects were assessed in two different strains of mice (i.e., C57BL/6J and C3H/HeJ). The findings further supported the strain-specific effects of CP55940. This agonist decreased habituation of the ASR in C57BL/6J, but increased habituation in C3H/HeJ. Both of these measures were reversed by SR141716A. This indicates that the cannabinoid agonist exerts its effects on non-associative learning and memory in both strains of mice via CB1 receptors. The advantages of zebrafish in comparison to other common vertebrate models including high fecundity, low maintenance cost, transparent embryos, and rapid development are some reasons for its popularity in genetics, pharmacological and behavioural research. As vertebrates, zebrafish share considerable sequence similarity with humans and are being used as an animal model for various human disease conditions. Here a video and written protocol is presented for the regular care and maintenance of a zebrafish laboratory and the optimisation of breeding conditions to enable efficient embryo production. In order to establish learning and memory procedures for a novel animal model, it is also important to assess potential confounds that may affect the results of the experiments. One such confound is the natural preference of zebrafish towards a particular colour if zebrafish are to be trained for a colour discrimination learning task. Therefore, the natural colour preference of zebrafish towards specific colours was evaluated using a place preference procedure and a T-maze with coloured sleeves. The results from this study showed a strong aversion towards blue colour compared to red, green, and yellow, with yellow being less preferred relative to red and green. Hence, the findings from this study may underpin any further designing of colour-based associative learning and memory paradigms in the zebrafish. The potential of zebrafish to be used for non-associative learning in a high-throughput manner was also evaluated in a 96-well plate by measuring habituation of the acoustic startle reflex in zebrafish larvae. It was found that larvae can be used for assessing non-associative learning and memory in a 96-well plate because zebrafish larvae startled significantly after the presentation of acoustic stimuli and this response decreased when the stimuli were presented continuously over a period of time. The reduction in response over time was consistent with the phenomenon of habituation, a form of non-associative learning. The non-associative learning and memory in zebrafish larvae was further validated by modulating with several pharmacological agents that are known to alter learning and memory in rodents and in humans such as donepezil and scopolamine. Donepezil was found to reduce the ASR and habituation of ASR in zebrafish larvae, consistent with some reports in rodents. Scopolamine treatment of zebrafish larvae produced a biphasic effect on non-cognitive measures such as ASR and locomotion such that lower doses decreased locomotion and ASR whereas the highest dose showed no effects. Similarly, scopolamine impaired non-associative learning with a lower dose but not with the higher dose. Therefore, the use of this drug for cognitive deficits model cannot be supported in this animal model before the reasons for the biphasic dose effect of scopolamine are elucidated. One hypothesis is suggested, that the differences in the effects of scopolamine are due to different affinities of scopolamine for different muscarinic acetylcholine receptor subtypes. The biphasic dose effect of the antagonist may be related to differential activation of these receptors by the endogenous agonist, rather than pharmacological properties of scopolamine itself. Finally, the effect of CP55940 was evaluated on non-associative learning and non-cognitive measures such as locomotor activity in zebrafish larvae. The solubility of CP55940 is a pragmatic issue when used for zebrafish research, as frequently used solvents are toxic to zebrafish. Therefore, CP55940 was dissolved in several different solvents and studied for its toxicity, locomotion effects, ASR and finally habituation of ASR. It was found that CP55940 was successfully dissolved in two solvents (i.e., ethanol + PBS and captisol) that were not toxic to zebrafish larvae, so CP55940 dissolved in these two solvents was tested for its effects on ASR and habituation of ASR. The results of these experiments showed that CP55940 dissolved in these two solvents produced differing effects on ASR and habituation. For instance, CP dissolved in ethanol+PBS had no effects on ASR but impaired habituation; however, CP dissolved in captisol reduced ASR but had no effects on habituation. CP dissolved in both these solvents decreased locomotor activity in zebrafish larvae, a well known effect of CP in rodents. In conclusion, the cannabinoid CB1 agonist may be used as a pharmacological model for inducing memory impairments with a caveat that effects can be strongly influenced by mouse strain. This project also validated the use of zebrafish for high-throughput screening of the effects of pharmacological agents on non-associative learning and showed potential confounds that may impact on zebrafish behaviour. Furthermore, similar to mice, cannabinoid CB1 agonist can induce habituation deficits in zebrafish. Overall, this project has evaluated novel CB1 receptor agonists and antagonists in modulating learning and memory in mice, and has established and validated non-associative memory tasks in zebrafish that can now be utilised in high-throughput screening for modulators of memory.
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Tang, Fan. "Protective effects of a novel derivative from danshensu tetramethylpyrazine on doxorubicin-induced cardiotoxicity in H9c2 and zebrafish models." Thesis, University of Macau, 2018. http://umaclib3.umac.mo/record=b3952166.
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Fenero, Camila Ideli Morales. "Estudo do modelo de inflamação intestinal induzida por TNBS em larvas de Zebrafish (Danio rerio)." Universidade de São Paulo, 2015. http://www.teses.usp.br/teses/disponiveis/42/42133/tde-09122015-123622/.
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As doenças inflamatórias intestinais são caracterizadas por uma desregulação na resposta imune contra a microbiota. O zebrafish, tem emergido como um novo modelo para o estudo de doenças inflamatórias. Os ácidos graxos de cadeia curta (AGCCs) são produtos da microbiota intestinal, que possuem papéis antiinflamatórios e aparecem como uma das possíveis terapias contra doenças inflamatórias. A implementação do modelo de inflamação intestinal induzido pelo ácido trinitrobenzenesulfônico (TNBS) em larvas de zebrafish aumentou a mortalidade de forma dose-dependente. Se observou dilatação do lúmen com alisamento das vilosidades intestinais. Não se detectaram mudanças na produção de muco nem no número de células caliciformes mas teve um leve aumento da apoptóse e diminuição da proliferação, nos expostos ao TNBS. Teve aumento de células mielóides infiltrantes e de citocinas inflamatórias, assim como disbiose da microbiota. O tratamento com AGCCs gero alta mortalidade a concentrações acima de 10 mM, mas existe ação anti-inflamatória a esta mesma concentração.Inflammatory bowel diseases are characterized by a dysregulation in immune response against microbiota. The zebrafish has emerged as a new model for the study of inflammatory diseases. Short chain fatty acids (SCFAs) are products of the intestinal microflora, which have anti-inflammatory roles and appear as a possible therapy against inflammatory diseases. The implementation of intestinal inflammation model induced by trinitrobencenesulfonic acid (TNBS) in zebrafish larvae, increased mortality in a dose-dependent manner. Was observe dilatation lumen and straightening of the intestinal villi. No changes were detected in the production of mucus or in the number of goblet cells but had a slight increase in apoptosis and decreased proliferation in exposed to TNBS. Also has, increased cytokines and infiltrating myeloid cells, and dysbiosis of the microbiota. Treatment with SCFAs generate high mortality above 10 mM concentrations, but there anti-inflammatory action to this same concentration.
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Akande, Motunrayo Ganiyat. "The toxicological evaluation of sewage effluents and pharmaceuticals with the use of zebrafish as a model organism /." Uppsala : Department of Biomedical Sciences and Veterinary Public Health, Swedish University of Agricultural Sciences, 2008. http://epsilon.slu.se/10691471.pdf.
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Knapp, Silvana Maria [Verfasser], and F. le [Akademischer Betreuer] Noble. "Esm1 modulates spinal cord vascularization in Vegfaa gain-of-function zebrafish models / Silvana Maria Knapp ; Betreuer: F. le Noble." Karlsruhe : KIT-Bibliothek, 2020. http://d-nb.info/1206646616/34.
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Hewamadduma, Channa. "Investigation of the role of TDP-43 in amyotrophic lateral sclerosis (ALS) using patient derived fibroblasts and zebrafish as models." Thesis, University of Sheffield, 2014. http://etheses.whiterose.ac.uk/7134/.
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Solchenberger, Barbara [Verfasser], and Christian [Akademischer Betreuer] Haass. "Generation of granulin knock out zebrafish models for frontotemporal lobar degeneration and related disorders by genome editing / Barbara Solchenberger. Betreuer: Christian Haass." München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2014. http://d-nb.info/1080663258/34.
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Kneipp, Moritz [Verfasser], Daniel [Akademischer Betreuer] Razansky, Gil [Gutachter] Westmeyer, Vasilis [Gutachter] Ntziachristos, and Arthur [Gutachter] Konnerth. "Rapid Functional Optoacoustic Imaging of Brain Activity in Murine and Zebrafish Models / Moritz Kneipp ; Gutachter: Gil Westmeyer, Vasilis Ntziachristos, Arthur Konnerth ; Betreuer: Daniel Razansky." München : Universitätsbibliothek der TU München, 2015. http://d-nb.info/1140835076/34.
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Bourefis, Annis-Rayan. "Novel FUS and CHCHD10 models to investigate pathogenic mechanisms in Amyotrophic Lateral Sclerosis." Thesis, Sorbonne université, 2019. http://www.theses.fr/2019SORUS177.
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La sclérose latérale amyotrophique (SLA) est une maladie neurodégénérative dévastatrice causée par la dégénérescence progressive des motoneurones (MNs) supérieurs et inférieurs menant à une faiblesse et une atrophie musculaire qui progresse jusqu’à la paralysie. Deux gènes majeurs identifiés chez les patients SLA sont le gène FUS (FUSed in sarcoma), impliqué dans le métabolisme de l’ARN, et CHCHD10, qui joue un rôle dans la stabilité des mitochondries. Ces deux gènes ont été étudiés à travers différents modèles, de petits modèles invertébrés aux biopsies de patients. Cependant, les différents traits phénotypiques observés sont complexes et parfois controversés. L’objectif de cette thèse est de fournir de nouvelles informations sur l’implication de ces deux gènes dans la SLA à travers l’utilisation de nouveaux modèles. Pour étudier les mécanismes pathologiques induits par FUS et CHCHD10, nous avons généré et caractérisé deux nouveaux modèles de poisson-zèbres présentant une mutation non-sens des orthologues de ces gènes, et nous avons mis en évidence différents traits phénotypiques propres à la SLA. Nous avons montré, pour FUS, une espérance de vie réduite, une locomotion altérée, des axones moteurs aberrants, une jonction neuromusculaire (JNM) désorganisée, une altération des muscles et mitochondries, ainsi que des changements moléculaires. Ces résultats montrent que la perte de fonction de fus est responsable de l’apparition de signes pathologiques distaux au niveau de la JNM, indiquant une neuronopathie en « dying-back », dans laquelle les traits pathologiques de la SLA commencent au niveau de la JNM et progressent vers les corps cellulaires des MNsAmyotrophic Lateral Sclerosis (ALS) is a devastating neurodegenerative disorder caused by progressive degeneration of upper and lower motor neurons (MNs), with a very rapid clinical course. It leads to muscle weakness and atrophy progressing to paralysis, with respiratory failure being the major cause of death within years following clinical diagnosis. Two major genes mutated in ALS patients are the RNA-binding protein FUS (FUSed in sarcoma), implicated in RNA metabolism, and coiled-coil-helix-coiled-coil-helix domain 10 (CHCHD10), which plays a role in mitochondria stability. Both these genes have been investigated through different model systems, from small invertebrate models to patient biopsies. However, the major phenotypic features obtained in these models are complex and often controversial. The objective of this work is to provide new insights on the implication of these genes in ALS through the use of new models.To investigate the pathogenic mechanisms induced by FUS and CHCHD10, we generated and characterized two novel stable non-sense mutant zebrafish models for the orthologues of these genes and highlighted several ALS phenotypic features. We demonstrated, for the FUS model but not for CHCHD10, reduced lifespan, locomotor disabilities, aberrant motor axons, disorganized neuromuscular junction (NMJ), muscle and mitochondrial alteration, as well as molecular changes. These findings indicate that loss of fus expression is responsible for the occurrence of distal pathological signs at the NMJ, thus supporting a “dying-back” neuronopathy, in which early disease hallmarks start at the level of the NMJ and progress towards MN cell bodies
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Söllner, Rosemarie Waltraud [Verfasser], Wolfgang [Akademischer Betreuer] Wurst, and Thomas [Akademischer Betreuer] Misgeld. "Establishment and cell biological analysis of neurodegeneration models in zebrafish based on Parkinson’s disease-linked genes LRRK2 and PINK1 / Rosemarie Waltraud Söllner. Gutachter: Wolfgang Wurst ; Thomas Misgeld. Betreuer: Wolfgang Wurst." München : Universitätsbibliothek der TU München, 2015. http://d-nb.info/1072758024/34.
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Hejmanowski, Ashley Q. "Allelic and genetic heterogeneity of two common genetic diseases." The Ohio State University, 2004. http://rave.ohiolink.edu/etdc/view?acc_num=osu1095309751.
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Leventea, Eleni. "Zebrafish as model for human ciliopathies." Thesis, University of Sheffield, 2016. http://etheses.whiterose.ac.uk/15413/.
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Karttunen, Marja Johanna. "Zebrafish model of demyelination and remyelination." Thesis, University of Edinburgh, 2017. http://hdl.handle.net/1842/25702.
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Myelin is a protective layer wrapped around axons which helps them conduct electrical signals rapidly, and provides them with metabolic support. In the central nervous system (CNS), myelin is produced by specialised glial cells called oligodendrocytes. Loss of myelin (demyelination) is associated with degeneration of axons and many neurodegenerative disorders, including multiple sclerosis (MS). The restoration of myelin sheaths by remyelination may protect axons and help functional recovery of patients, but achieving this requires better understanding of how the process unfolds at the cellular level. To investigate the processes of de- and remyelination in vivo, I have characterised a transgenic zebrafish line in which expression of the bacterial enzyme nitroreductase (NTR) is driven under the myelin basic protein promoter, thus in myelinating glia. I treat larvae with the NTR substrate metronidazole (Mtz). The reaction between NTR and Mtz results in a toxic metabolite which selectively kills NTR-expressing cells. The treatment with Mtz consistently ablates two-thirds of oligodendrocytes while not harming the animals otherwise. Myelin sheaths continue to deteriorate after the end of the treatment, such that seven days later, extensive demyelination is observed by electron microscopy. By 16 days after Mtz-treatment, robust recovery has occurred, with no discernible axon loss and myelin thickness restored to control levels. At this time point, oligodendrocyte numbers have also returned to control levels. During the demyelinated phase, I observe a striking increase in microglia and macrophages in the spinal cord. In order to study the role of the innate immune system in recovery, I used a mutant line, irf8-/- which lacks a transcription factor essential for development of microglia and macrophages. I am in the process of determining the ability of these mutants to regenerate their oligodendrocytes and myelin; preliminary results suggest that they are able to restore their myelin sheaths fully, but seem to have a delay in regenerating their oligodendrocytes compared to wild-types. The model I have established can be used in the future to better understand the consequences of demyelination to axon health, as well as chemical screening to identify compounds that could accelerate the remyelination process or enhance the thickness of myelin generated during remyelination. Insights arising from such studies will be useful in designing strategies to reduce axon loss and improve myelin regeneration in demyelinating diseases.
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Rose, Kristin. "A Zebrafish Model of Uveal Melanoma." Thesis, Harvard University, 2015. http://nrs.harvard.edu/urn-3:HUL.InstRepos:17467345.
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Uveal melanoma is the most common primary intraocular tumor in adults, and is often characterized by poor prognosis and few effective therapeutic options. The typical site of metastasis for uveal melanoma is the liver, and over 80% of patients with metastatic disease will die within one year of metastasis diagnosis. The vast majority of human uveal melanomas contain activating somatic mutations in the GPCR alpha subunits GNAQ or GNA11. To directly observe the in vivo effects of GNA11 Q209L (constitutively active) overexpression, I used a zebrafish cancer model in which plasmids can be injected into transgenic fish and melanoma formation can be assessed. Surprisingly, zebrafish injected with a construct overexpressing mitfa:GNA11 Q209L developed a significant incidence of uveal melanomas. A mini-screen of HOX genes using this system revealed a novel role for HOXB7, which also functioned as an inducer of uveal melanomas in our zebrafish model. Other plasmids containing oncogenes do not lead to uveal tumors, suggesting a specificity for GNA11 and HOXB7. Cell lines were derived from GNA11 Q209L-overexpressing zebrafish tumors and uveal melanoma cells were sensitive to PKC inhibition, which has been observed in human uveal melanoma cells as well. Additionally, RNAseq analysis of zebrafish uveal melanoma cell lines revealed high expression of genes such as cyr61 and ctgf, YAP pathway genes that are known to be induced by GNAQ in human uveal melanomas. I found that overexpression of catalytically inactive BAP1, a chromatin factor whose function is frequently lost in metastatic uveal melanomas, led to a significant acceleration of overall melanoma onset but did not induce a uveal melanoma phenotype in our model. Morpholino experiments showed that knockdown of either zebrafish gna11 or bap1 affected the spatial expression of hoxb7a, with a combined knockdown of gna11 and bap1 suggesting that bap1 is epistatic to gna11. My studies demonstrate the first known instance of spontaneous uveal melanoma formation in response to overexpression of a human uveal melanoma oncogene, and that the resulting uveal melanomas share genetic features and drug response tendencies with human uveal melanomas. This model is a useful new tool for the study of uveal melanoma pathogenesis, genetics, and therapeutics.Medical Sciences
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Costa, Marc Michael John Da. "A zebrafish model of motor neurone disease." Thesis, University of Sheffield, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.548470.
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Buckley, C. E. "Zebrafish : a transparent screening model of myelination." Thesis, University of Cambridge, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.597064.
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This thesis provides a description of the temporal dynamics of myelination in zebrafish larvae, demonstrating that oligodendrocytes are present in zebrafish larvae from ld.p.f. and myelination commences by 3d.p.f. The regulation of myelination is dependent on the growth of axons and not on their absolute size. Zebrafish larvae proved an accessible myelination model. A medium throughput in vivo reprofiling screen was consequently set up for pro-myelination compounds. Changes in oligodendrocyte lineage cell number in response to different compound treatments were assessed using the olig2:EGFP transgenic line of zebrafish larvae. As well as patentable compounds such as isoxicam, unknown for its effect on myelination, several compounds were identified that were known to affect oligodendrocyte differentiation and/or myelination, such as the sex steroid precursor, pregnenolone and the PPAR agonist, bezafibrate. The identification of these compounds therefore validated the screening techniques used. Analysis of mbp mRNA transcript levels by qPCR identified 3 compounds that decreased transcript levels, one of which was PP2. The effect of PP2 was specific to src kinase inhibition, confirming the importance of this known pathway in regulating myelination and again validating the screen. However, it was not possible to increase mbp levels with any of the compounds screened and there was no apparent correlation between the number of olig2+ cells and mbp transcript levels. Preliminary further analysis of some of the compounds identified in the olig2+ cell counting screen, such as isoxicam, also illustrated that the effects on olig2+ cell number were not always specific to oligodendrocytes. Compounds with potential pro-myelination effects should now be analysed further for their specificity of effect, both in zebrafish models of myelination and in remyelination models.
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Kishida, Marcia Gruppi. "Investigating non-canonical vertebral development in the zebrafish model system." Thesis, University of Cambridge, 2018. https://www.repository.cam.ac.uk/handle/1810/276830.
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A segmented vertebral column is one of the major innovations vertebrates. In mice and chicks – amniotes – a subpopulation of the somites, the sclerotome, is sole source of vertebral tissue. It is unclear, however, how applicable this amniote-based ‘canonical’ mechanism is across the vertebrates. In fact, the vast majority and diversity of vertebrates are not amniotes, but are members of ‘fish’ groups where there has been relatively little investigation into vertebral development. Indeed, there is great diversity in vertebra form throughout ‘fish’ groups and fossil evidence suggests that the components of the vertebra, the neural arches and the vertebral bodies, arose separately and that vertebrates have evolved multiple ways of building vertebral bodies. In teleosts fish, the vertebral bodies initially form as mineralised rings within the notochord sheath (chordacentra) and then secondarily, bone is deposited around this (perichordal centra and arches). Notochord cells (chordoblasts) have been implicated in chordacentrum mineralisation and patterning in zebrafish and Atlantic salmon, though the question of how the overtly unsegmented notochord could direct segmental mineralisation still remains. My project first aims to address this dual mechanism in the zebrafish model, by testing whether the chordoblasts can mineralise and pattern the chordacentra. The second aim is to elucidate the role of the sclerotome in teleost vertebral development. To do this, I explored CRISPR knock-in tools to label the sclerotome and used a Gal4 gene trap line to investigate sclerotome ablation. I characterised the chordacentra and chordoblasts in our model system and verified the specificity of a promoter as a chordoblast marker. With this promoter, I established a method to target the chordoblasts for KillerRed-induced phototoxicity. I demonstrated that intact chordoblasts are necessary for chordacentrum formation, but that vertebral arches are unaffected. Fused perichordal centra are still able to form, but the underlying sheath has a very different structure. This supports the ‘duality’ hypothesis that in teleosts the role of the sclerotome in vertebra formation is limited to the arches and perichordal centra, whereas the chordoblasts are responsible for the chordacentra.
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Sullivan, Matthew J. "The Zebrafish as a Model for Cystic Fibrosis." Fogler Library, University of Maine, 2008. http://www.library.umaine.edu/theses/pdf/SullivanMJ2008.pdf.
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Raghupathy, Rakesh Kotapati. "Zebrafish as a model for Leber Congenital Amaurosis." Thesis, Glasgow Caledonian University, 2015. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.700985.
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