published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

published_or_final_version / Zoology / Master / Master of Philosophy

The ovarian steroids, progesterone and estradiol, regulate luteinizing hormone synthesis and secretion during the estrous cycle of mature ewes. During the luteal phase of the cycle the ovarian steroids inhibit luteinizing hormone secretion. Luteinizing hormone is secreted from the pituitary when stimulated by the hypothalamic neuropeptide, gonadotropin-releasing hormone. Ovarian steroids can inhibit luteinizing hormone secretion indirectly, by decreasing the secretion of gonadotropin-releasing hormone or directly, by modulating the response of the pituitary to gonadotropin-releasing hormone. These studies have examined the direct control of pituitary luteinizing hormone secretion by using an in vivo model in which endogenous gonadotropin-releasing hormone (GnRH) release has been ablated and replaced with exogenous GnRH release at a constant frequency. Progesterone directly inhibited pituitary LH secretion in an estradiol-dependent manner and this may not require inhibition of pituitary LH synthesis. Progesterone inhibition of pituitary luteinizing hormone secretion is associated with enhanced progesterone binding by the pituitary.

Hui Sau Lei Raydi. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (leaves 71-82). / Abstracts in English and Chinese. / ABSTRACT (English and Chinese) --- p.i / ACKNOWLEDGMENT --- p.1 / LIST OF FIGURES --- p.2 / LIST OF TABLES --- p.3 / LIST OF ABBREVIATIONS --- p.4 / Chapter I. --- Introduction and Objectives --- p.5-8 / Chapter II. --- Literature review --- p.9 / Chapter II.A --- Corticotrophin-releasing hormone --- p.9 / Chapter II.A.1. --- Structure of Corticotrophin-releasing hormone --- p.9-10 / Chapter II.A.2. --- Corticotrophin releasing hormone and normal Physiology --- p.11 / Chapter II.A.2.a. --- Pituitary-adrenal axis --- p.11-12 / Chapter II.A.2.b. --- Role of Pituitary-adrenal axis --- p.12 / Chapter II.A.3. --- Placental Corticotrophin releasing hormone --- p.13 / Chapter II.A.3.a. --- Origin --- p.13 / Chapter II.A.3.b. --- Physiology --- p.14 / Chapter II.A.3.C. --- Normal pregnancy --- p.15 / Chapter II.A.3.d. --- Association with human parturition --- p.16 / Chapter II.A.3.e. --- Association with preterm delivery and other abnormal pregnancy outcomes --- p.17-18 / Chapter II.B. --- Alpha-fetoprotein --- p.19 / Chapter II.B.1. --- Physiology --- p.19-20 / Chapter II.B.2. --- Maternal alpha-fetoprotein levels in the second trimester --- p.21-22 / Chapter III : --- Materials & Method --- p.23 / Chapter III.A. --- Study population --- p.23-24 / Chapter III.B. --- Sample collection and Analysis --- p.25 / Chapter III.C. --- Corticotrophin releasing hormone radioimmunoassay --- p.26 / Chapter III.C.l.a. --- Theoretical basis for radioimmunoassay --- p.26-27 / Chapter III.C.l.b. --- Vycor extraction of maternal plasma samples --- p.28-30 / Chapter III.C.l.c. --- Standard curve --- p.31-32 / Chapter III.C.l.d. --- Antisera --- p.33 / Chapter III.C.1.e. --- Tracer --- p.34-35 / Chapter III.C.l.f. --- HPLC Tracer Purification --- p.36-37 / Chapter III.C.l.g. --- Separation of bound from unbound Cortico- trophin-releasing hormone: second antibody --- p.38 / Chapter III.C.1.h. --- Corticotrophin-releasing hormone radio- immunoassay procedure --- p.39 / Chapter III.C.2. --- Corticotrophin-releasing hormone reagents --- p.40-41 / Chapter III.C.3. --- Estimation of Corticotrophin-releasing hormone extraction recovery --- p.42 / Chapter III.C.4. --- Sample dilution --- p.43 / Chapter III.D. --- Alpha-fetoprotein: microparticle enzyme immunoassay --- p.44 / Chapter III.D.1. --- Principles --- p.44 / Chapter III.D.2. --- Reaction Process --- p.45-46 / Chapter III.D.3. --- MEIA Optical Assembly --- p.47 / Chapter III.D.4. --- Operation --- p.47 / Chapter III.D.5. --- Alpha-fetoprotein reagents --- p.48 / Chapter III.D.6. --- Sample Dilution --- p.49 / Chapter III.D.7. --- Inter-assay and Intra-assay Variation --- p.50-52 / Chapter III.E. --- Data handling --- p.53 / Chapter III.F. --- Statistical Analysis --- p.53 / Chapter IV: --- Results --- p.54 / Chapter IV.A. --- Demographic Data --- p.55-57 / Chapter IV.B. --- Corticotrophin-releasing hormone levels --- p.58 / Chapter IV.B.1. --- Corticotrophin-releasing hormone levels increases as gestation advances --- p.58 / Chapter IV.B.2 --- The association between the plasma Corticotrophin releasing hormone levels and the time to delivery --- p.59 / Chapter IV.B.3 --- Elevated Corticotrophin-releasing hormone levels among the preterm group --- p.60 / Chapter IV.B.4. --- Corticotrophin releasing hormone levels and history of threatened abortion --- p.61 / Chapter IV.C. --- Alpha-fetoprotein levels --- p.62 / Chapter IV.C.1. --- Alpha-fetoprotein levels and gestational age --- p.62 / Chapter IV.C.2. --- Alpha-fetoprotein levels and preterm labour --- p.63 / Chapter V: --- Discussion --- p.64-65 / Chapter V.A. --- Importance of dating --- p.64 / Chapter V.B. --- Diagnosis of preterm labour --- p.64-65 / Chapter V.C.1. --- Corticotrophin-releasing hormone and labour --- p.65-66 / Chapter V.C.2. --- Corticotrophin-releasing hormone and infection --- p.66-67 / Chapter V.C.3. --- Diurnal rhythm of Corticotrophin-releasing hormone --- p.67 / Chapter V.C.4. --- Laboratory assays of Corticotrophin-releasing hormone --- p.68 / Chapter V.D. --- Alpha-fetoprotein and labour --- p.69-70 / Chapter VI. --- Reference --- p.71-82

Orientador: Rita Cássia Menegati Dornelles / Co-orientador: Janete Aparecida Anselmo-Franci / Banca: Maristela de Oliveira Poletini / Banca: Jacqueline Nelisis Zanoni / Resumo: As alterações nos componentes reprodutivos do eixo hipotálamo-hipófise-gônadas em muitas fêmeas de mamíferos determinam a transição gradual de ciclos reprodutivos regulares para ciclos irregulares, com perda de fertilidade. A interação dos neurônios do hormônio liberador de gonadotrofinas (GnRH) e esteróides gonadais representa função chave na neurobiologia do envelhecimento, pois a sobreposição temporal da senescência endócrina e neural está mecanicamente interligada pelas alças de retroalimentação. Estímulos do locus coeruleus (LC) para a área pré-óptica (APO) e eminência mediana são essenciais para a liberação das gonadotrofinas e seus neurônios apresentam receptores para estrógeno e progesterona, sugerindo controle dos esteróides ovarianos. Neste estudo foi avaliado a atividade de células neuronais localizadas em áreas e núcleos envolvidos com o controle de ação dos neurônios GnRH de ratas Wistar no período de transição para a aciclicidade. Para este trabalho foram utilizadas fêmeas Wistar cíclicas (4 meses) e acíclicas (18-20 meses) submetidas à decapitação ou perfusão às 10, 14 e 18 h na fase do diestro. Após serem retirados, os cérebros dos animais decapitados foram congelados e armazenados para posterior determinação do conteúdo de GnRH hipotalâmico e do conteúdo de noradrenalina e dopamina na APO. Os cérebros perfundidos foram cortados seriadamente em secções coronais de 30 μm para a APO e o LC e... / Abstract: Changes in reproductive components of the hypothalamic-pituitary-gonadal axis in many female mammals determine the gradual transition from regular reproductive cycles to irregular cycles, with loss of fertility. The interaction of neurons of gonadotropin-releasing hormone (GnRH) and gonadal steroids represents key role in the neurobiology of aging, because the temporal overlap of endocrine and neural senescence is mechanically interconnected by feedback loops. Stimulation of the locus coeruleus (LC) for the preoptic area (POA) and median eminence are essential for the release of gonadotropins and their neurons have receptors for estrogen and progesterone, suggesting control of ovarian steroids. Therefore, in this study we evaluated the activity of neuronal cells located in areas and nuclei involved in the control of action of GnRH neurons of female rats during the transition to acyclicity. For this study, we used cyclic female (4 months) and acyclic (18-20 months) rats underwent perfusion or decapitation at 10, 14 and 18 h of diestrus day. The brains from decapitated animals, after removed, were frozen and stored for subsequent determination of the hypothalamic GnRH content and the noradrenaline and dopamine content in the POA. The perfused brains were serially cut into coronal sections of 30 μm to POA and LC and subsequently submitted to immunohistochemical labeling for Fos (FRA) and FRA / TH, respectively. For quantitative analysis of the POA were considered plates containing AVPe being the counting of neurons FRA-ir performed from the insertion of the box with... / Mestre

The gonadotropin-releasing hormone (GnRH) neurons are the final output neurons of a complex neuronal network that controls fertility in all mammals. The GnRH neurons reside in a scattered continuum throughout the anterior hypothalamus. The majority of GnRH neurons project an axon to the median eminence where GnRH is secreted into the hypophyseal-pituitary portal vessels from whence it travels to the anterior pituitary gland. GnRH acts on the gonadotrophs of the anterior pituitary gland to cause the secretion of luteinising hormone (LH) and follicle stimulating hormone (FSH) into the peripheral circulation. LH and FSH act on the gonads to control gametogenesis and steroidogenesis. This thesis focuses on two unanswered questions in reproductive neurobiology that are fundamental to fertility 1) how the GnRH neurons become activated at puberty to produce patterned GnRH secretion and 2) the nature of the positive feedback mechanism that drives the preovulatory GnRH and LH surges. Recently, a novel neuropeptide called kisspeptin and its G-protein coupled receptor GPR-54 were found to be essential for pubertal activation of GnRH neurons, with GPR-54 mutation or deletion resulting in failed puberty and infertility in humans and mice. In addition, kisspeptin administration potently stimulates GnRH neuron-mediated gonadotropin secretion and advances the onset of pubertal maturation suggesting an important role for kisspeptin in the activation and perhaps post-pubertal modulation of GnRH neurons. In this thesis I have used immunocytochemical, whole animal manipulations and knockout mouse approaches to investigate the role of kisspeptin in both the activation of GnRH neurons at puberty and in the estrogen positive feedback mechanism in the mouse. I have demonstrated that kisspeptin neurons are located principally in the rostral periventricular area of the third ventricle (RP3V) and the arcuate nucleus (ARN), which are both known to be important areas for the modulation of GnRH neuronal activity. Kisspeptin fibres are found in abundance throughout the hypothalamus, but of particular interest are the kisspeptin fibres found in close apposition with a subset of GnRH neurons in the rostral preoptic area (rPOA). The kisspeptin neurons in the RP3V are sexually dimorphic with up to ten times more neurons in the female than the male. The number of kisspeptin neurons in the RP3V increases throughout pubertal development reaching adult levels at the time of puberty in both males and females. In concert with the increase in the number of kisspeptin neurons in the RP3V there is an increase in the percentage of GnRH neurons in the rPOA which exhibited a close apposition with a kisspeptin fibre indicating that kisspeptin neurons may target GnRH neurons to activate them at puberty. Additionally, I demonstrate that the increase in the number of neurons in the RP3V of the female mouse approaching puberty is driven by estrogen secreted from the ovary. A significant number of kisspeptin neurons in the RP3V were shown to express tyrosine hydroxylase (TH). The number and percentage of kisspeptin cells colocalised with TH cells in the RP3V did not change throughout the estrous cycle. Some colocalisation of kisspeptin and TH was observed at terminal appositions with GnRH neurons in the rPOA, though the magnitude of colocalisation also did not change throughout the estrous cycle. I demonstrate that RP3V kisspeptin neurons are a critical part of the estrogen positive feedback mechanism which drives the preovulatory GnRH and LH surges. Kisspeptin neurons in the RP3V express steroid receptors and are activated by estrogen positive feedback. Loss of kisspeptin-GPR-54 signalling prevents the GnRH neurons from being activated by estrogen positive feedback indicating that the RP3V kisspeptin neurons not only contribute to the estrogen positive feedback mechanism, but are a critical component of the mechanism. The results of these studies demonstrate that kisspeptin is an integral component in both the activation of GnRH neurons at puberty and in the estrogen positive feedback mechanism which drives the preovulatory GnRH and LH surges. Therefore, kisspeptin plays an important role in the neuroendocrine control of reproduction in the mouse.

Thesis (Ph. D.)--Georgia State University, 2007. / Title from file title page. Gabor Patonay, committee chair; A.L. Baumstark, G. Davon Kennedy, Gregg Pratt, committee members. Electronic text (170 p. : ill. (some col.)) : digital, PDF file. Description based on contents viewed Dec. 10, 2007. Includes bibliographical references.

Graduation date: 2002

To determine whether membrane-related events may be involved in attenuated luteal function after gonadotropin-releasing hormone (GnRH) administration, corpora lutea (CL) were removed from 10 beef heifers on day 7 of the estrous cycle after i.v. injection of GnRH or saline on day 2 of the cycle. Luteal slices were incubated with saline (control), luteinizing hormone (LH) or 8-bromo-cAMP for 2 h. In vivo administration of GnRH reduced LH and cAMP-stimulated progesterone production by tissue (p<0.01), but basal progesterone production was not affected (p>0.05). Luteal adenylyl cyclase activity did not differ between saline and GnRH-treated animals (p>0.05). Results of this experiment suggested that GnRH-induced alteration of bovine luteal function may be due to an effect distal to the point of cAMP accumulation. To explore further the effect of GnRH on luteal cell function, 10 heifers were injected with saline or GnRH and CL removed as above. Dissociated (mixed) and small luteal cells (SC) were cultured overnight, then incubated for 2 h with medium alone (control), LH or cAMP. In vitro treatment with LH and cAMP increased progesterone in the medium relative to controls (p<0.01), however, there was no effect of GnRH injection on progesterone production (p>0.05) nor in the percentage of large cells (LC) present in the mixed cell cultures (p=0.95). It has been previously found that the ratio of LC to SC increases in GnRH-treated animals. Many LC can be ruptured during dissociation of the CL, and it is possible that this procedure altered the number of LC, such that any differences that may have existed between the saline and GnRH-exposed CL were minimized. These data suggest that differences in the LC to SC ratio may indeed account for attenuated luteal function after exposure to GnRH. To examine if early administration of GnRH alters response of the CL to prostaglandin (PG) Fav beef heifers were injected with saline or GnRH on day 2 of the cycle (n=4/group), then injected with PGF[subscript 2��], on day 8 and the CL removed 60 min later. Blood samples were collected for oxytocin (OT) analysis at frequent intervals after PGF[subscript 2��], injection and for progesterone at 0 and 60 min. Induction of the early response gene c-jun or release of OT by PGF[subscript 2��], was not altered by GnRH injection (p>0.05). Injection of PGF[subscript 2��], decreased serum progesterone by 60 min post-injection (p<0.05), but was also unaffected by GnRH (p>0.05). These data support the hypotheses that c-jun expression and OT release are involved in PGF[subscript 2��]-induced luteolysis, but early administration of GnRH did not affect these processes. / Graduation date: 1996

Graduation date: 1995