No description available.

Le kisspeptine est un peptide, encodé par le gène Kiss1, qui joue un rôle majeur dans le contrôle central de la fonction de reproduction en régulant la sécrétion du GnRH après la naissance. Les neurones exprimant Kiss1 apparaissent avant la naissance exclusivement dans le noyau arqué. Les objectifs de cette thèse étaient de décrire comment le système s‟organise avant la naissance en fonction du sexe à une période où il existe chez le mâle un pic prénatal de testostérone, puis de déterminer si cette organisation pouvait être influencée par les stéroïdes sexuels. Nos résultats ont permis 1) de caractériser précisément chez la souris la mise en place des cellules kisspeptine en fonction du sexe et d‟identifier un gradient de différenciation antéro-postérieur sexe-indépendant 2) de mettre en évidence une interaction réciproque avec les neurones à GnRH, chez les mâles et les femelles, suggérant ainsi une certaine maturité du système 3) de montrer la diversité des récepteurs aux stéroïdes sexuels déjà exprimés par ces cellules avant la naissance ainsi que l‟émergence d‟un dimorphisme sexuel 4) de démontrer la sensibilité et la réponse morphologique à l'oestradiol de ces cellules in vitro après la mise au point du premier modèle de culture primaire de cellules kisspeptine. / Kisspeptin neurons express the Kiss1 gene encoding kisspeptin, a potent neuropeptide secretagogue of gonadotropin releasing hormone (GnRH) that plays a fundamental role in regulation of reproductive life cycles after birth. Neurons expressing Kiss1 appear before birth only in the arcuate nucleus. The overall purpose of this study is to understand how kisspeptin neurons are set up during fetal development, if and how estrogen signaling in these cells could interfere with their development at a time of a prenatal testosterone peak in male. Our finding showed 1) precisely the placement of kisspeptin cells as a function of sex and to identify a sex-independent anteroposterior differentiation gradient 2) a reciprocal interaction with GnRH neurons, in males and females, thus suggesting a certain maturity of the system 3) the diversity of sexual steroids receptors already expressed by these cells before birth as well as the emergence of a sexual dimorphism 4) sensitivity and morphological response to estradiol of these cells in vitro after the development of the first primary culture model of kisspeptin cells.

Submitted by Ana Flávia Teresa Paiva (aflavia.paiva@gmail.com) on 2018-07-05T15:29:33Z No. of bitstreams: 1 Dissertação de mestrado.pdf: 699288 bytes, checksum: 3c4b62716218390065cac7fea5b0cb42 (MD5) / Rejected by Isabel Pereira de Matos (isabel@fmva.unesp.br), reason: Falta Ficha Catalográfica; Arrumar palavras chave on 2018-07-05T19:05:16Z (GMT) / Submitted by Ana Flávia Teresa Paiva (aflavia.paiva@gmail.com) on 2018-07-05T20:22:53Z No. of bitstreams: 1 Dissertação de mestrado.pdf: 790947 bytes, checksum: 15861ab0546530441a0c751b2150ab34 (MD5) / Approved for entry into archive by Ederson Vasconcelos Pereira null (edersonpereira@fmva.unesp.br) on 2018-07-06T14:09:52Z (GMT) No. of bitstreams: 1 paiva_aftp_me_araca_int.pdf: 790947 bytes, checksum: 15861ab0546530441a0c751b2150ab34 (MD5) / Made available in DSpace on 2018-07-06T14:09:52Z (GMT). No. of bitstreams: 1 paiva_aftp_me_araca_int.pdf: 790947 bytes, checksum: 15861ab0546530441a0c751b2150ab34 (MD5) Previous issue date: 2018-06-29 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Objetivou-se comparar a concentração de LH plasmático, em resposta à kisspeptina exógena, em novilhas Nelore, para avaliar a maturação sexual do Sistema Nervoso Central. As hipóteses testadas foram de que em resposta a kisspeptina, a secreção de LH aumenta com a idade e que em uma mesma idade, novilhas com maior peso à desmama secretam mais LH que as desmamadas mais leves. Utilizou-se 50 bezerras, segregadas em leves (174kg) e pesadas (232kg), desafiadas mensalmente dos 8 aos 16 meses, com 10 µg/kg (IM) de kisspeptina recombinante bovina, e após 20 min foi realizada a coleta de sangue. Aos 16 meses as novilhas foram expostas a touros por 60 dias e 30 dias depois a taxa de prenhez foi avaliada. A quantificação de LH, P4 e leptina foi feita por radioimunoensaio e IGF-I por ELISA. Houve aumento na concentração de LH em resposta a kisspeptina dos 8 aos 14 meses de idade. Comparando diferença na concentração de LH entre o 8º mês com meses os subsequentes, observou-se aumento no grupo prenhe no 11º mês, e no grupo não prenhe no 12º mês. A taxa de prenhez no grupo pesado foi de 92% enquanto que no grupo leve foi de 62,5% e o tempo gestacional estimado foi maior no grupo pesado. A concentração do IGF-I foi maior aos 12 meses no grupo leve e aos 15 meses no grupo não prenhe, enquanto que a concentração de leptina foi maior no grupo pesado aos 13, 15 e 16 meses, e prenhe aos 9 e 10 meses. A concentração de LH após a kisspeptina aumentou com a idade, mas não foi maior nos animais mais pesados à desmama, mas foi capaz de avaliar a maturação sexual do hipotálamo e futuramente pode ser usada para a triagem de novilhas Nelore com precocidade sexual. / The objective was to compare the plasma luteinizing hormone (LH) concentration in response to exogenous kisspeptin in Nellore heifers in order to evaluate the sexual maturation of the Central Nervous System. The tested hypotheses were: in response to kisspeptin the LH secretion increases with age and at the same age, heifers with higher weaning weight secrete more LH compared to lighter ones. Fifty heifers were used, secreted in light (174kg) and heavy (232kg), challenged monthly from 8-16 mo, with 10 μg / kg (IM) of recombinant bovine kisspeptin, and after 20 min blood sample was collected. At 16 months the heifers were exposed to bulls for 60 days, the pregnancy was evaluated 30 days later. The LH, P4 and leptin quantification were done by radioimmunoassay and IGF-I by ELISA. There was an increase in LH concentration in response to kisspeptin from 8th to 14th months of age. Comparing the difference on LH concentration between the 8th with the following months, the pregnant group presented an increase at 11th whereas the non-pregnant group it was observed at 12th month. The heavy heifers pregnancy rate was 92% while in the light group it was 62.5% and gestational time was also higher in the heavy group. IGF-1 levels was greater at 12th mo in the light group and at 15th mo in non-pregnant group. Leptin level was higher at 13th, 15th and 16th mo in the heavy group and at 9th and 10th mo in pregnant group. The LH concentration after kisspeptin injection increased according to age, was not higher in heavier animals at weaning, but was able to evaluate hypothalamus sexual maturation and in the future can be used as a screening tool for sexual precocity in Nellore heifers.

Orientador: Guilherme de Paula Nogueira / Banca: Claudia Maria Bertan Membrive / Banca: Rafael Silva Cipriano / Resumo: Objetivou-se comparar a concentração de LH plasmático, em resposta à kisspeptina exógena, em novilhas Nelore, para avaliar a maturação sexual do Sistema Nervoso Central. As hipóteses testadas foram de que em resposta a kisspeptina, a secreção de LH aumenta com a idade e que em uma mesma idade, novilhas com maior peso à desmama secretam mais LH que as desmamadas mais leves. Utilizou-se 50 bezerras, segregadas em leves (174kg) e pesadas (232kg), desafiadas mensalmente dos 8 aos 16 meses, com 10 µg/kg (IM) de kisspeptina recombinante bovina, e após 20 min foi realizada a coleta de sangue. Aos 16 meses as novilhas foram expostas a touros por 60 dias e 30 dias depois a taxa de prenhez foi avaliada. A quantificação de LH, P4 e leptina foi feita por radioimunoensaio e IGF-I por ELISA. Houve aumento na concentração de LH em resposta a kisspeptina dos 8 aos 14 meses de idade. Comparando diferença na concentração de LH entre o 8º mês com meses os subsequentes, observou-se aumento no grupo prenhe no 11º mês, e no grupo não prenhe no 12º mês. A taxa de prenhez no grupo pesado foi de 92% enquanto que no grupo leve foi de 62,5% e o tempo gestacional estimado foi maior no grupo pesado. A concentração do IGF-I foi maior aos 12 meses no grupo leve e aos 15 meses no grupo não prenhe, enquanto que a concentração de leptina foi maior no grupo pesado aos 13, 15 e 16 meses, e prenhe aos 9 e 10 meses. A concentração de LH após a kisspeptina aumentou com a idade, mas não foi maior nos animais mais p... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The objective was to compare the plasma luteinizing hormone (LH) concentration in response to exogenous kisspeptin in Nellore heifers in order to evaluate the sexual maturation of the Central Nervous System. The tested hypotheses were: in response to kisspeptin the LH secretion increases with age and at the same age, heifers with higher weaning weight secrete more LH compared to lighter ones. Fifty heifers were used, secreted in light (174kg) and heavy (232kg), challenged monthly from 8-16 mo, with 10 μg / kg (IM) of recombinant bovine kisspeptin, and after 20 min blood sample was collected. At 16 months the heifers were exposed to bulls for 60 days, the pregnancy was evaluated 30 days later. The LH, P4 and leptin quantification were done by radioimmunoassay and IGF-I by ELISA. There was an increase in LH concentration in response to kisspeptin from 8th to 14th months of age. Comparing the difference on LH concentration between the 8th with the following months, the pregnant group presented an increase at 11th whereas the non-pregnant group it was observed at 12th month. The heavy heifers pregnancy rate was 92% while in the light group it was 62.5% and gestational time was also higher in the heavy group. IGF-1 levels was greater at 12th mo in the light group and at 15th mo in non-pregnant group. Leptin level was higher at 13th, 15th and 16th mo in the heavy group and at 9th and 10th mo in pregnant group. The LH concentration after kisspeptin injection increased according to a... (Complete abstract click electronic access below) / Mestre

Les kisspeptides (KP), puissants sécrétagogues de la GnRH, agissent par l'intermédiaire du récepteur couplé aux protéines G, le GPR54. Une perfusion i.v. de KP est non seulement capable de synchroniser les ovulations chez la brebis cyclique, mais également d’induire un cycle chez la brebis en anoestrus. L'objectif de ma thèse a été d'étudier la neuroanatomie du système KP / GnRH, et de définir des applications potentielles du KP dans le contrôle du cycle chez la jument cyclique et en anoestrus. Nos résultats ont permis de montrer que 1) le KP interagit avec le système GnRH au niveau hypothalamique, 2) une perfusion i.v. de KP en début de phase folliculaire conduit à une stimulation - quoique transitoire – de la sécrétion des gonadotropines, 3) une capacité limitée pour reconstituer le stock en GnRH et / ou LH / FSH pourrait expliquer la nature transitoire de la stimulation par le KP, 4) une perfusion de KP pendant 3 jours, depuis le début de la phase folliculaire, tend à avancer le pic péri-ovulatoire de LH et l'ovulation, 5) une perfusion plus longue (soit plus de 3 jours) de KP, pendant toute la phase folliculaire, n’induit pas une ovulation plus précoce, 6) pendant la saison d'anoestrus, une perfusion de KP pendant 3 jours stimule la sécrétion de LH, avec une intensité néanmoins inférieure à celle observée avec une perfusion de GnRH. En conclusion, bien que l'utilisation potentielle du KP pour synchroniser ou induire l'ovulation pendant la saison de reproduction semble discutable, sa capacité à induire des cycles durant la période d'anoestrus présente un intérêt et mérite des études plus approfondies. / Kisspeptins (KP) are very potent secretagogues of GnRH which act through the G protein-coupled receptor GPR54. An i.v. infusion of KP is not only able to synchronize ovulations of cyclic ewes, but also to induce cycle in anoestrus ewes. The aim of my thesis was to investigate the neuroanatomy of the KP / GnRH system, and define potential applications for KP in the control of the estrus cycle in the cyclic and anoestrus mares. Our results showed that 1) KP interact with GnRH systems at the hypothalamic level, 2) an i.v. infusion of KP during early follicular phase leads to an enhanced - albeit transient - secretion of gonadotropins, 3) a limited ability to replenish GnRH and/or LH/FSH stores might explain the transient nature of the KP stimulation, 4) an infusion of KP for 3 days, since early follicular phase, modestly advances the periovulatory LH surge and the ovulation, 5) a longer (i.e. more than 3 days) infusion of KP during all the follicular phase does not lead to precocious ovulation, 6) during the anoestrus season, an infusion of KP for 3 days heightens LH secretion, with however a lower intensity to that observed with an infusion of GnRH. In conclusion, albeit the potential use of KP to synchronize or induce ovulation during the breeding season appears questionable, its ability to induce cycles during the anoestrus period is of interest and warrants further investigation.

Neurônios GnRH e kisspeptina representam as populações neuronais de maior importância no controle da reprodução. Estradiol liga-se ao seu receptor expresso pelos neurônios kisspeptina para regular a libertação de GnRH. No modelo animal OVX+E a atividade do neurônio GnRH e pico de LH é depende do estradiol e hora do dia. Nesse estudo, embora a taxa de disparo dos neurônios GnRH seja similar entre os grupos, o padrão dos potenciais revelou uma mudança para maior duração do estouro em camundongos no proestrous, além do fato de uma maior resposta da hipófise. A prolactina tem grande impacto na modulação do eixo HPG e kisspeptina são mediadores dos efeitos da prolactina sobre a reprodução. Uma pequena porcentagem de neurônios de kisspeptina do AVPV foi indiretamente despolarizada pela prolactina. Este efeito requeria a via de sinalização PI3K. Camundongos portadores de inativação de Stat5a/b em células kisspeptina exibiram um início precoce de ciclicidade estro, indicando que os fatores de transcrição STAT5 exercem um efeito inibitório sobre o momento da puberdade. / GnRH and kisspeptina neurons represent the most important neuronal populations in the control of reproduction. Estradiol binds to its receptor expressed by the kisspeptina neurons to regulate the release of GnRH. In the animal model OVX+E the activity of the GnRH neuron and LH surge is dependent of estradiol and time of day. In this study, although the firing rate of GnRH neurons was similar between groups, the pattern of potentials revealed a change to longer burst duration in mice in proestrous, and the pituitary response was greater in this group. Prolactin has impact on HPG axis modulation and kisspeptin is a mediator of the effects of prolactin on reproduction. A small percentage of AVPV kisspeptin neurons were indirectly depolarized by prolactin. This effect required the PI3K signaling pathway. Mice bearing Stat5a/b inactivation on kisspeptin cells exhibited an early onset of estrus cyclicity, indicating that STAT5 transcription factors exert an inhibitory effect on the time of puberty.

Les hypogonadismes hypogonadotropes congénitaux (CHH) sont des maladies héréditaires caractérisées par un déficit de sécrétion des gonadotrophines par l’hypophyse, à l’origine d’une infertilité ou d’une absence complète de puberté. On distingue les formes isolées avec olfaction normale (nCHH) et les formes syndromiques associant au déficit gonadotrope d’autres signes, tel qu’un défaut d’olfaction dans le cas du syndrome de Kallmann (SK), la forme plus fréquente de CHH. Les gènes identifiés dans le SK participent au développement embryonnaire et les gènes des nCHH sont impliqués dans la régulation de la sécrétion de la GnRH ou de son action. A ce stade, deux populations de neurones hypothalamiques gonadotropes sont connues, le neurone à GnRH et le neurone KNDy, sécrétant les Kisspeptines et la Neurokinine B. On estimait que l’ensemble des gènes identifiés couvraient moins de 20% des étiologies génétiques. L’objectif de ce doctorat était d’étudier prévalences et mécanismes physiopathologiques des gènes connus et d’identifier de nouvelles étiologies génétiques de CHH. Dans la première partie, nous avons caractérisé la fonctionnalité de tous les variants identifiés sur les gènes KISS1R, TACR3 et TAC3. Cela a permis de préciser les prévalences chez 600 patients, d’identifier un profil neuroendocrinien propre à l’altération de la signalisation Neurokinine B et de démontrer l’implication des Kisspeptines au cours de la vie embryonnaire. Enfin, nous proposons un modèle d’interaction entre le neurone à GnRH et le neurone KNDy. Dans la seconde partie, nous avons identifié deux nouveaux gènes, SEMA3A dans une forme familiale de SK et PNPLA6 dans une forme familiale rare de CHH syndromique. En conclusion, notre connaissance accrue des formes génétiques de CHH, a permis de développer un panel d’exome ciblé dédié au diagnostic par séquençage nouvelle génération permettant l’analyse simultanée de gènes candidats et de gènes connus. / Congenital hypogonadotropic hypogonadism (CHH) is characterized by deficient or absent pubertal development due to deficient or absent secretion of the pituitary gonadotropins. The many known genetic causes are generally classified into distinct nosological groups. One comprises abnormalities that affect the pre-natal development or migration of GnRH neurons, the paradigm of which is Kallmann syndrome. The other encompasses molecular abnormalities that only affect hypothalamic GnRH synthesis, GnRH release or GnRH signaling at pituitary level. At this stage, two populations of hypothalamic neurons implicated in a gonadotrop function are identified, GnRH neurons and KNDy neurons secreting kisspeptins and neurokinin B. All of the identified genes would represent less than 20% of genetic etiologies.The aim of this PhD was to study the prevalence and pathophysiology mechanisms of known genes and to identify new genetic etiologies of CHH.In the first part, we characterized the function of all molecular events identified on KISS1R, TACR3 and TAC3 genes. Prevalences were estimated in 600 patients. A particular neuroendocrine profile was identified in patients presenting an alteration of neurokinin B signaling. Importance of Kisspeptins during embryonic life was validated. According to these data, a model of interaction between GnRH and KNDy neurons was proposed.In the second part, we identified two new CHH genes using various molecular genetics approaches. SEMA3A was identified in a familial form of Kallmann syndrome and PNPLA6 in a rare familial form of CHH.Finally, our increased knowledge of the various genetic forms of CHH allows proposing a new genetic approach based on next generation sequencing to test together all known and several candidate genes.

No description available.

京都大学 / 0048 / 新制・課程博士 / 博士(薬科学) / 甲第18929号 / 薬科博第43号 / 新制||薬||5(附属図書館) / 31880 / 京都大学大学院薬学研究科医薬創成情報科学専攻 / (主査)教授 大野 浩章, 教授 高須 清誠, 教授 竹本 佳司 / 学位規則第4条第1項該当 / Doctor of Pharmaceutical Sciences / Kyoto University / DFAM

Le RFRP est une neuropeptide impliqué dans la régulation de l’axe reproducteur, mais ses effets varient en fonction du sexe et des espèces. Le but de cette étude était de décrire en détails l’organisation du système RFRP et de caractériser son rôle dans le contrôle circadien et saisonnier de l’axe reproducteur de hamsters femelles. Les résultats montrent que le système RFRP est régulé par la photopériode et que son niveau d’expression est plus élevé chez les femelles que chez les mâles. Cela se traduit par des actions spécifiques sur l’axe gonadotrope femelle. En effet, L’activité des neurones à RFRP est diminuée au moment du pic pré-ovulatoire de LH et des injections centrales de RFRP-3 dans l’heure qui précède le pic de LH induisent une diminution de l’amplitude de la sécrétion de LH, démontrant une implication du RFRP dans la régulation circadienne du pic pré-ovulatoire de LH. Par ailleurs, des infusions chroniques de RFRP-3 chez des hamsters femelles sexuellement inactifs sont capables de réactiver le fonctionnement de l‘axe reproducteur, ce qui montre que le RFRP a un également un rôle régulateur essentiel dans le contrôle saisonnier de la reproduction. / RFRP neurons regulate the reproductive axis, however, their effects depend on species and sex. Here, we aimed at providing a neuroanatomical description of the RFRP system in the Syrian hamster and at investigating the role of RFRP in the daily and seasonal control of female reproduction. We show that besides being regulated by annual changes in photoperiod, the RFRP system is more strongly expressed in females than in males. In line with this, we unveil that RFRP has multiple roles in regulating female reproduction. RFRP neuronal activity is specifically reduced at the time of the pre-ovulatory LH surge and central RFRP-3 administration prior to the surge decreases LH peak levels, altogether pointing towards a daily down-regulation of the inhibitory RFRP signal necessary for proper generation of the LH surge. Moreover, chronic RFRP-3 infusion in sexually inactive females, with endogenous low RFRP expression, completely reactivates the reproductive axis. Taken together, we demonstrate that RFRP is a key component in the seasonal control of reproduction while at the same time specifically regulating cyclic events controlling reproductive activity in females.