The purpose of this experiment is to determine if immature female white mice will react to the gonadotropic hormone from the anterior pituitary of several species of fish. If they can be shown to do so, a second purpose is to establish an assay unit for this fish gonadotropin in order that it may be used with predictable results in inducing extra-seasonal spawning for fresh-water conservation practices.

Protocolos de sucesso para a maturação in vitro de oócitos de peixe são importantes, uma vez que é necessário para garantir uma fertilização bem sucedida, formação do zigoto, crescimento do embrião e seu completo desenvolvimento. Em algumas espécies, a eficiência deste processo ainda é muito baixa ou restrita a poucas substâncias que podem ser utilizadas. Assim, pesquisou-se a utilização de hormônios alternativos ao protocolo já existente para maturação in vitro de ovócitos de zebrafish. O objetivo foi avaliar a eficiência do extrato de hipófise de carpa (EHC), dos hormônios folículo estimulante (FSH) e luteinizante (LH) para fazer a maturação dos ovócitos estádio III de zebrafish. Os oócitos estádio III foram colocados em meio de cultivo Leibovitz modificado, suplementado com soro fetal bovino e adicionado o hormônio correspondente a seu tratamento (T1-controle; T2-16 μg/ml de EHC; T3- 32 μg/ml de EHC; T4- 48 μg/ml de EHC; T5- 64 μg/ml de EHC; T6- 80 μg/ml de EHC; T7- 0,5 μg/ml de FSH; T8- 0,5 μg/ml de LH e T9- 0,5 μg/ml de FSH e 0,5 μg/ml de LH). A taxa de maturação foi avaliada através da visualização da quebra da vesícula germinal (GVBD). Em todos os tratamentos houve maturação, embora o EHC tenha demonstrado taxas de maturação muito baixas (T2= 12,8%; T3=24,8%; T4=27%; T5=22,7%; T6=9,7%) e inferiores em relação a maior eficiência dos hormônios gonadotrópicos (T7=16%; T8=35%; T9=50%). Além disso foi possível verificar a viabilidade dos oócito através da fertilização in vitro do melhor tratamento (T9) com uma taxa de eclosão e desenvolvimento em larva de 60%. Os resultados da maturação in vitro utilizando estes indutores hormonais em oócitos estádio III de zebrafish mostraram-se promissores, e reforçam as perspectivas para o aprimoramento e uso desta técnica para produção in vitro de embriões viáveis. / Successful protocols for maturation of oocytes are important, as it is necessary for ensuring successful fertilization, zygote formation, embryo growth and full development. In some species the efficiency of in vitro maturation is still very low or is still restricted to a little amount of substances which can be used for the matter. Thus, we studied the use of alternative hormones to the existing protocol for in vitro maturation of zebrafish oocytes. The aim of this study was to evaluate the efficiency of the use of carp pituitary extract (CPE), the follicle stimulating hormone (FSH) and luteinizing hormone (LH) to oocyte maturation stage III of zebrafish. Oocytes stage III were placed in modified Leibovitz culture medium, suplemented with fetal bovine serum and added to the correnponding hormone treatment (T1-control; T2-16 g / ml of CHE; T3 32 g / ml of CHE, T4 - 48 g / ml of CHE; T5- 64 g / ml of CHE; T6- 80 g / ml of CHE; T7- 0.5 g / ml of FSH, T8 0.5 mg / ml of LH and T9- 0.5 g / ml of FSH and 0.5 mg / ml LH). The maturation rate was assessed by the germinal vesicle break down (GVBD). In all cases there was maturation, though the EHC has demonstrated fairly low maturation rate (T2= 12,8%; T3=24,8%; T4=27%; T5=22,7%; T6=9,7%) and lower in relation of the high efficiency presented by the gonadotropic hormones (T7=16%; T8=35%; T9=50%). In addition it was possible to verify the viability of the oocyte through IVF of the best treatment (T9) with a result of 60% of hatching and larvae development rate. The results of maturation in turn using this hormones in stage III oocytes of zebrafish proved promising, and enhance the prospects for improvement and use of this technique for in vitro production of viable embryos.

Protocolos de sucesso para a maturação in vitro de oócitos de peixe são importantes, uma vez que é necessário para garantir uma fertilização bem sucedida, formação do zigoto, crescimento do embrião e seu completo desenvolvimento. Em algumas espécies, a eficiência deste processo ainda é muito baixa ou restrita a poucas substâncias que podem ser utilizadas. Assim, pesquisou-se a utilização de hormônios alternativos ao protocolo já existente para maturação in vitro de ovócitos de zebrafish. O objetivo foi avaliar a eficiência do extrato de hipófise de carpa (EHC), dos hormônios folículo estimulante (FSH) e luteinizante (LH) para fazer a maturação dos ovócitos estádio III de zebrafish. Os oócitos estádio III foram colocados em meio de cultivo Leibovitz modificado, suplementado com soro fetal bovino e adicionado o hormônio correspondente a seu tratamento (T1-controle; T2-16 μg/ml de EHC; T3- 32 μg/ml de EHC; T4- 48 μg/ml de EHC; T5- 64 μg/ml de EHC; T6- 80 μg/ml de EHC; T7- 0,5 μg/ml de FSH; T8- 0,5 μg/ml de LH e T9- 0,5 μg/ml de FSH e 0,5 μg/ml de LH). A taxa de maturação foi avaliada através da visualização da quebra da vesícula germinal (GVBD). Em todos os tratamentos houve maturação, embora o EHC tenha demonstrado taxas de maturação muito baixas (T2= 12,8%; T3=24,8%; T4=27%; T5=22,7%; T6=9,7%) e inferiores em relação a maior eficiência dos hormônios gonadotrópicos (T7=16%; T8=35%; T9=50%). Além disso foi possível verificar a viabilidade dos oócito através da fertilização in vitro do melhor tratamento (T9) com uma taxa de eclosão e desenvolvimento em larva de 60%. Os resultados da maturação in vitro utilizando estes indutores hormonais em oócitos estádio III de zebrafish mostraram-se promissores, e reforçam as perspectivas para o aprimoramento e uso desta técnica para produção in vitro de embriões viáveis. / Successful protocols for maturation of oocytes are important, as it is necessary for ensuring successful fertilization, zygote formation, embryo growth and full development. In some species the efficiency of in vitro maturation is still very low or is still restricted to a little amount of substances which can be used for the matter. Thus, we studied the use of alternative hormones to the existing protocol for in vitro maturation of zebrafish oocytes. The aim of this study was to evaluate the efficiency of the use of carp pituitary extract (CPE), the follicle stimulating hormone (FSH) and luteinizing hormone (LH) to oocyte maturation stage III of zebrafish. Oocytes stage III were placed in modified Leibovitz culture medium, suplemented with fetal bovine serum and added to the correnponding hormone treatment (T1-control; T2-16 g / ml of CHE; T3 32 g / ml of CHE, T4 - 48 g / ml of CHE; T5- 64 g / ml of CHE; T6- 80 g / ml of CHE; T7- 0.5 g / ml of FSH, T8 0.5 mg / ml of LH and T9- 0.5 g / ml of FSH and 0.5 mg / ml LH). The maturation rate was assessed by the germinal vesicle break down (GVBD). In all cases there was maturation, though the EHC has demonstrated fairly low maturation rate (T2= 12,8%; T3=24,8%; T4=27%; T5=22,7%; T6=9,7%) and lower in relation of the high efficiency presented by the gonadotropic hormones (T7=16%; T8=35%; T9=50%). In addition it was possible to verify the viability of the oocyte through IVF of the best treatment (T9) with a result of 60% of hatching and larvae development rate. The results of maturation in turn using this hormones in stage III oocytes of zebrafish proved promising, and enhance the prospects for improvement and use of this technique for in vitro production of viable embryos.

Protocolos de sucesso para a maturação in vitro de oócitos de peixe são importantes, uma vez que é necessário para garantir uma fertilização bem sucedida, formação do zigoto, crescimento do embrião e seu completo desenvolvimento. Em algumas espécies, a eficiência deste processo ainda é muito baixa ou restrita a poucas substâncias que podem ser utilizadas. Assim, pesquisou-se a utilização de hormônios alternativos ao protocolo já existente para maturação in vitro de ovócitos de zebrafish. O objetivo foi avaliar a eficiência do extrato de hipófise de carpa (EHC), dos hormônios folículo estimulante (FSH) e luteinizante (LH) para fazer a maturação dos ovócitos estádio III de zebrafish. Os oócitos estádio III foram colocados em meio de cultivo Leibovitz modificado, suplementado com soro fetal bovino e adicionado o hormônio correspondente a seu tratamento (T1-controle; T2-16 μg/ml de EHC; T3- 32 μg/ml de EHC; T4- 48 μg/ml de EHC; T5- 64 μg/ml de EHC; T6- 80 μg/ml de EHC; T7- 0,5 μg/ml de FSH; T8- 0,5 μg/ml de LH e T9- 0,5 μg/ml de FSH e 0,5 μg/ml de LH). A taxa de maturação foi avaliada através da visualização da quebra da vesícula germinal (GVBD). Em todos os tratamentos houve maturação, embora o EHC tenha demonstrado taxas de maturação muito baixas (T2= 12,8%; T3=24,8%; T4=27%; T5=22,7%; T6=9,7%) e inferiores em relação a maior eficiência dos hormônios gonadotrópicos (T7=16%; T8=35%; T9=50%). Além disso foi possível verificar a viabilidade dos oócito através da fertilização in vitro do melhor tratamento (T9) com uma taxa de eclosão e desenvolvimento em larva de 60%. Os resultados da maturação in vitro utilizando estes indutores hormonais em oócitos estádio III de zebrafish mostraram-se promissores, e reforçam as perspectivas para o aprimoramento e uso desta técnica para produção in vitro de embriões viáveis. / Successful protocols for maturation of oocytes are important, as it is necessary for ensuring successful fertilization, zygote formation, embryo growth and full development. In some species the efficiency of in vitro maturation is still very low or is still restricted to a little amount of substances which can be used for the matter. Thus, we studied the use of alternative hormones to the existing protocol for in vitro maturation of zebrafish oocytes. The aim of this study was to evaluate the efficiency of the use of carp pituitary extract (CPE), the follicle stimulating hormone (FSH) and luteinizing hormone (LH) to oocyte maturation stage III of zebrafish. Oocytes stage III were placed in modified Leibovitz culture medium, suplemented with fetal bovine serum and added to the correnponding hormone treatment (T1-control; T2-16 g / ml of CHE; T3 32 g / ml of CHE, T4 - 48 g / ml of CHE; T5- 64 g / ml of CHE; T6- 80 g / ml of CHE; T7- 0.5 g / ml of FSH, T8 0.5 mg / ml of LH and T9- 0.5 g / ml of FSH and 0.5 mg / ml LH). The maturation rate was assessed by the germinal vesicle break down (GVBD). In all cases there was maturation, though the EHC has demonstrated fairly low maturation rate (T2= 12,8%; T3=24,8%; T4=27%; T5=22,7%; T6=9,7%) and lower in relation of the high efficiency presented by the gonadotropic hormones (T7=16%; T8=35%; T9=50%). In addition it was possible to verify the viability of the oocyte through IVF of the best treatment (T9) with a result of 60% of hatching and larvae development rate. The results of maturation in turn using this hormones in stage III oocytes of zebrafish proved promising, and enhance the prospects for improvement and use of this technique for in vitro production of viable embryos.

The purpose of this research is, first, to determine by laboratory methods, that species specificity does not exist in closely allied taxonomic animals; second, to determine a unit of activity for the gonadotropic hormone. For this purpose a quantitative method for determining potency is necessary to ascertain the seasonal production of the gonadotropic factor in fish. A further aim in this investigation is to demonstrate that the diversity of the gonadotropic factor, in relation to phylogenetic variations, is not as ineffectual as previously reported.

Les processus physiopathologiques qui mènent à la dégénérescence neuronale ainsi qu’aux symptômes de la maladie de Huntington (MH) demeurent non identifiés et les hypothèses actuelles ne permettent pas d’expliquer l’hétérogénéité intra et interindividuelle de l’évolution de ces symptômes. Ainsi, la progression de la maladie reste donc difficile voire impossible à prédire. Dans ce contexte, il est important d’explorer d’autres facteurs qui semblent être impliqués dans le processus pathogène de la maladie mais qui pourraient également influencer l’évolution de ces symptômes et ainsi prédire la progression de la maladie. Plusieurs éléments de preuve renforcent l’hypothèse de l’existence de troubles hormonaux dans la MH tels que l’atteinte de l’hypothalamus et la perte de poids. Cependant, en raison du peu d’études, de leur qualité et de la discordance de leurs résultats, l’existence des modifications hormonales dans la maladie de Huntington et plus particulièrement leur lien avec la progression de la maladie reste controversée. L’objectif de ce travail est de décrire le profil hormonal de l’axe hypothalamohypophysaire dans la MH afin de mieux comprendre le rôle de ces hormones sur la progression et éventuellement sur la physiopathologie de la maladie. Dans notre étude transversale, nous avons mis en évidence une activation de l’axe somatotrope (Growth Hormone/Insulin Growth Factor 1), une inhibition en fonction de la sévérité de la maladie de deux axes : gonadotrope (Testostérone) et thyréotrope (Thyroid Stimulating Hormone et triiodothyronine) mais aucune modification des hormones de l’axe corticotrope ni de la prolactine. De plus, la modification hormonale de l’axe somatotrope était non pathologique et précoce alors qu’elle était tardive pour les deux autres axes. Pour expliquer le lien entre ces modifications et la progression de la maladie une étude longitudinale a été mise en place. Les résultats de cette étude montre que seule l’élévation plasmatique d’IGF1 était prédictive de la détérioration cognitive. L’ensemble de nos résultats apporte une meilleure description et compréhension du profil de l’axe hypothalamo-hypophysaire dans la maladie de Huntington. Les axes pituitaires ne sont pas tous atteints et leur atteinte n’est pas dans le même sens. La relation inverse entre l’activation de l’axe somatotrope et la détérioration cognitive renforce l’hypothèse d’une résistance à l’effet de l’IGF1 dans la maladie de Huntington comme pour la maladie d’alzheimer. En conclusion, compte tenu de l’implication de l’IGF1 dans la prédiction de la progression cognitive dans la maladie de Huntington, il serait intéressant de détecter si les modifications biologiques de l’IGF1 existent dès la phase asymptomatique cognitive afin d’envisager d’utiliser l’IGF1 comme biomarqueur de l’apparition ou de l’évolution des symptômes cognitives. D’un autre côté, il serait important d’étendre les recherches sur les mécanismes responsables des modifications hormonales dans la maladie de Huntington afin de mieux comprendre l’effet de cause à effet s’il existe entre ces modifications et les symptômes de la maladie / The pathophysiological processes leading to neurodegeneration and the symptoms of Huntington's disease (HD) remain unidentified and current hypothesis do not explain the intra and interindividual heterogeneity of the evolution of these symptoms. Thus, the progression of the disease remains difficult or impossible to predict. In this context, it is important to explore other factors that appear to be involved in the pathogenic process of the disease but could also influence the evolution of these symptoms and predict disease progression. Several evidences reinforce the hypothesis of the existence of hormonal disorders in HD such as the atrophy of the hypothalamus and weight loss. Because of few studies, their quality and the discrepancies of their results, the existence of hormonal changes in Huntington's disease and particularly their relationship to disease progression remains controversial. The objective of this work is to describe the hormonal profile of the hypothalamicpituitary axis in HD in order to better understand the role of these hormones on the progression and on the pathophysiology of the disease. In our cross-sectional study, we identified an activation of the somatotropic axis (Growth Hormone / Insulin Growth Factor 1), an inhibition according to the severity of the disease in two axes: gonadotrope (Testosterone) and thyréotrope (Thyroid Stimulating hormone and triiodothyronine) but no change in hormones of corticotropic axis and prolactin. In addition, the somatotropic axis is overactive even in patients with early disease. To explain the link between these changes and the progression of the disease, a longitudinal study was done. The results of this study showed that only the elevated plasma IGF1 was predictive of cognitive impairment. All of our results provide a better description and understanding of the profile of the hypothalamic-pituitary axis in Huntington's disease. Pituitary axes are not all disturbed. The inverse relationship between activation of the somatotropic axis and cognitive impairment strengthens the hypothesis of a resistance to the effect of IGF1 in Huntington's disease like in Alzheimer's disease. In conclusion, given the involvement of IGF1 in the prediction of cognitive progression in Huntington's disease, it would be interesting to detect whether the biological changes of IGF1 are already present at the asymptomatic cognitive stage in order to use IGF1 as a biomarker of the onset or changes in cognitive symptoms. On the other hand, , it would be important to extend research on the mechanisms responsible for hormonal changes in Huntington's disease to better understand the link between these changes and symptoms of the disease