Call number: LD2668 .T4 1985 L825 / Master of Science

This study was designed to evaluate the possible existence of masked gonadotropin binding sites in the corpus luteum of the rhesus monkey. Pretreatment of macaque luteal particulates and cells with neuraminidase increased LH binding. In vitro exposure to alcohols also enhanced LH binding to these preparations. Ethanol modulation of LH binding was a time- and temperature-dependent process. The optimal concentration of ethanol for enhancing LH uptake was inversely proportional to the incubation temperature. Longer straight-chain alcohols were more potent than ethanol in increasing LH binding. Ethanol and neuraminidase increased the number of binding sites with no affect on affinity. Moreover, the effects of ethanol and NA were additive. Since alcohols and temperature are modulators of membrane fluidity, we examined the hypothesis that the unmasking of gonadotropin binding sites may be related to changes in the fluid state of the lipid bilayer of the luteal membrane. First, membrane fluidity was estimated from the fluorescence polarization of the membrane probe diphenylhexatriene. Conditions which resulted in enhanced gonadotropin binding (1-8% ethanol, increased temperature), increased the fluidity of luteal membranes. Moreover, changes in gonadotropin binding were highly correlated (r = -0.97) with changes in membrane fluidity under these conditions. Pretreatment of luteal particulates with neuraminidase had no apparent effect on membrane fluidity. Second, gonadotropin receptors were removed from the luteal membrane by detergent solubilization, and the effects of ethanol on soluble receptors were compared to those on receptors associated with the lipid bilayer. Solubilization resulted in the recovery of 50% more gonadotropin binding sites than are available in particulate preparations of the corpus luteum; these sites displayed lower affinity for gonadotropin. Moreover, conditions which increase LH binding to luteal particulates (1-8% ethanol at 25C) decreased LH uptake by soluble receptors. The data suggest that two populations of LH binding sites are masked within the membranes of the monkey corpus luteum. The ability of two markedly different agents, alcohol and neuraminidase, to increase LH binding indicates the diverse mechanisms may modulate the masking/unmasking of gonadotropin receptors in target cell membranes. As such, changes in membrane fluidity may play an important role in this response.

No description available.

Kwok Hin-Fai. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 84-100). / Abstracts in English and Chinese. / Abstract (in English) --- p.i / Abstract (in Chinese) --- p.iv / Acknowledgement --- p.vi / Table of contents --- p.vii / List of figures --- p.xi / List of tables --- p.xiv / Symbols and abbreviations --- p.xv / List of fish names mentioned in the thesis --- p.xviii / Chapter Chapter 1 --- General Introduction / Chapter 1.1 --- Gonadotropins / Chapter 1.1.1 --- Structure --- p.1 / Chapter 1.1.2 --- Function --- p.4 / Chapter 1.2 --- Gonadotropin receptor / Chapter 1.2.1 --- Structure --- p.5 / Chapter 1.2.2 --- Expression --- p.7 / Chapter 1.2.3 --- Signaling / Chapter 1.2.3.1 --- cAMP-mediated pathway --- p.7 / Chapter 1.2.3.2 --- Phospholipase C-mediated pathway --- p.9 / Chapter 1.2.4 --- Regulation of expression --- p.12 / Chapter 1.2.5 --- Desensitization of receptors / Chapter 1.2.5.1 --- Uncoupling --- p.13 / Chapter 1.2.5.2 --- Internalization --- p.13 / Chapter 1.3 --- Structure of ovarian follicles --- p.14 / Chapter 1.4 --- The project objectives and long-term significance --- p.16 / Chapter Chapter 2 --- Cloning and Characterization of Zebrafish Follicle-stimulating Hormone (FSH) and Luteinizing Hormone (LH) Receptors ´ؤ Evidence for Distinct Functions of FSH and LH in Follicle Development / Chapter 2.1 --- Introduction --- p.19 / Chapter 2.2 --- Materials and Methods / Chapter 2.2.1 --- Animals and chemicals --- p.22 / Chapter 2.2.2 --- Isolation of total RNA --- p.22 / Chapter 2.2.3 --- Cloning of zebrafish FSHR (zfFSHR) and LHR (zfLHR) cDNA fragments from the zebrafish ovary --- p.23 / Chapter 2.2.4 --- Rapid amplification of 5´ةcDNA ends (5'-RACE) and full-length cDNA --- p.24 / Chapter 2.2.5 --- Isolation of ovarian follicles --- p.25 / Chapter 2.2.6 --- Sampling of the ovaries from sexually immature zebrafish --- p.25 / Chapter 2.2.7 --- Reverse transcription-polymerase chain reaction (RT-PCR) --- p.25 / Chapter 2.2.8 --- Construction of expression plasmids --- p.26 / Chapter 2.2.9 --- Transient transfection and reporter gene assay --- p.27 / Chapter 2.2.10 --- Establishment and characterization of stable zfFSHR or zfLHR-expressing cell lines --- p.28 / Chapter 2.3 --- Results / Chapter 2.3.1 --- Cloning of FSHR and LHR cDNA from the zebrafish ovary --- p.29 / Chapter 2.3.2 --- Functional characterization of zfFSHR and zfLHR --- p.30 / Chapter 2.3.3 --- Expression of zfFSHR and zfLHR during sexual maturation --- p.31 / Chapter 2.3.4 --- Stage-dependent expression of zfFSHR and zfLHR in the ovarian follicles --- p.32 / Chapter 2.4 --- Discussion --- p.33 / Chapter Chapter 3 --- Down-regulation of FSHR and LHR Expression in the Zebrafish Follicle Ceils by Gonadotropin (hCG) and Its Sigaling Mechanism / Chapter 3.1 --- Introduction --- p.51 / Chapter 3.2 --- Materials and Methods / Chapter 3.2.1 --- Animals --- p.54 / Chapter 3.2.2 --- Chemicals and hormones --- p.54 / Chapter 3.2.3 --- Primary follicle cell culture --- p.55 / Chapter 3.2.4 --- Total RNA isolation --- p.55 / Chapter 3.2.5 --- "Validation of semi-quantitative RT-PCR assays for FSHR, LHR and GAPDH" --- p.56 / Chapter 3.2.6 --- Data analysis --- p.57 / Chapter 3.3 --- Results / Chapter 3.3.1 --- Validation of semi-quantitative RT-PCR assays --- p.57 / Chapter 3.3.2 --- Gonadotropin regulation of FSHR and LHR expression in cultured zebrafish ovarian follicle cells --- p.58 / Chapter 3.3.3 --- Effect of db-cAMP and forskolin on FSHR and LHR expression --- p.59 / Chapter 3.3.4 --- Effects of H89 on hCG-induced suppression of FSHR and LHR expression --- p.60 / Chapter 3.4 --- Discussion --- p.60 / Chapter Chapter 4 --- General Discussion --- p.75 / Chapter 4.1 --- Cloning of zebrafish FSHR and LHR cDNAs and demonstration of receptor specificity --- p.77 / Chapter 4.2 --- Evidence for the differential expression of FSHR and LHR in the zebrafish ovarian and follicle development --- p.78 / Chapter 4.3 --- Down-regulation of FSHR and LHR expression in the zebrafish follicle cells by gonadotropin (hCG) --- p.79 / Chapter 4.4 --- Future research direction --- p.80 / References --- p.84

So Wai-Kin. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 100-127). / Abstracts in English and Chinese. / Acknowledgement --- p.I / Abstract (in English) --- p.II / Abstract (in Chinese) --- p.IV / Table of contents --- p.VI / List of Figures --- p.X / Symbols and Abbreviations --- p.XII / List of fish names mentioned in the thesis --- p.XIV / Chapter Chapter 1 --- General Introduction / Chapter 1.1 --- Pituitary --- p.1 / Chapter 1.2 --- Gonadotropins --- p.1 / Chapter 1.2.1 --- Structure --- p.2 / Chapter 1.2.2 --- Signaling --- p.3 / Chapter 1.2.3 --- Expression --- p.5 / Chapter 1.2.4 --- Functions --- p.7 / Chapter 1.2.4.1 --- Gonadotropin actions on gametogenesis --- p.7 / Chapter 1.2.4.2 --- Gonadotropin actions on steroidogenesis --- p.8 / Chapter 1.2.5 --- Regulation --- p.9 / Chapter 1.2.5.1 --- Neuroendocrine control --- p.10 / Chapter 1.2.5.1.1 --- Gonadotropin-releasing hormone (GnRH) --- p.10 / Chapter 1.2.5.1.2 --- Dopamine (DA) --- p.12 / Chapter 1.2.5.2 --- Gonadal steroid feedback --- p.12 / Chapter 1.2.5.2.1 --- Positive feedback --- p.13 / Chapter 1.2.5.2.2 --- Negative feedback --- p.14 / Chapter 1.2.5.3 --- Paracrine regulators within pituitary --- p.15 / Chapter 1.3 --- Objectives of the present study --- p.16 / Chapter Chapter 2 --- "Molecular Cloning and Functional Characterization of Zebrafish FSHβ, LHβ and GTHα subunits" / Chapter 2.1 --- Introduction --- p.19 / Chapter 2.2 --- Materials and methods --- p.21 / Chapter 2.2.1 --- Chemicals --- p.21 / Chapter 2.2.2 --- Animals --- p.21 / Chapter 2.2.3 --- Genomic DNA isolation --- p.22 / Chapter 2.2.4 --- Total RNA isolation --- p.22 / Chapter 2.2.5 --- Cloning of zebrafish FSHp，LHβ and GTHa fragments --- p.23 / Chapter 2.2.5.1 --- LHβ and GTHα --- p.23 / Chapter 2.2.5.2 --- FSHβ --- p.23 / Chapter 2.2.6 --- "5'- and 3'-RACE of zebrafish FSHp, LHβ and GTHα subunits" --- p.24 / Chapter 2.2.7 --- Construction of expression constructs --- p.25 / Chapter 2.2.8 --- Cell culture and transfection of Flp-In´ёØ CHO cell --- p.26 / Chapter 2.2.9 --- Recombinant production of zebrafish FSH and LH --- p.27 / Chapter 2.2.10 --- Reverse transcription-polymerase chain reaction (RT-PCR) analysis --- p.27 / Chapter 2.2.11 --- Northern blot hybridization --- p.28 / Chapter 2.2.12 --- SEAP reporter gene assay --- p.28 / Chapter 2.2.13 --- Data analysis --- p.29 / Chapter 2.3 --- Results --- p.30 / Chapter 2.3.1 --- "Cloning of zebrafish FSHβ, LHβ and GTHα subunits" --- p.30 / Chapter 2.3.2 --- "Expression of zebrafish FSHp, LHβ and GTHα in the zebrafish pituitary" --- p.31 / Chapter 2.3.3 --- Recombinant production of zebrafish FSH and LH --- p.32 / Chapter 2.3.4 --- Functional analysis of zebrafish FSH and LH --- p.33 / Chapter 2.4 --- Discussion --- p.34 / Chapter Chapter 3 --- "Spatial Expression Patterns of Zebrafish FSHβ, LHβ and GTHα Subunits in the Pituitary and Their Temporal Expression Profiles during Sexual Maturation and Ovulatory Cycle" / Chapter 3.1 --- Introduction --- p.58 / Chapter 3.2 --- Materials and methods --- p.61 / Chapter 3.2.1 --- Chemicals --- p.61 / Chapter 3.2.2. --- Animals --- p.62 / Chapter 3.2.3 --- Total RNA isolation from zebrafish pituitaries and reverse transcription --- p.62 / Chapter 3.2.4 --- Validation of RT-PCR on single pituitary --- p.63 / Chapter 3.2.5 --- Real-time PCR --- p.64 / Chapter 3.2.6 --- Tissue preparation for in situ hybridization --- p.64 / Chapter 3.2.7 --- In situ hybridization --- p.65 / Chapter 3.2.8 --- Data analysis --- p.66 / Chapter 3.3 --- Results --- p.66 / Chapter 3.3.1 --- "PCR amplification of FSHβ, LHβ and GTHα and GAPDH in single zebrafish pituitary" --- p.67 / Chapter 3.3.2 --- "Establishement of real-time RT-PCR for zebrafish FSHβ, LHβ and GTHa and GAPDH" --- p.67 / Chapter 3.3.3 --- "Temporal expression profiles of zebrafish FSHβ, LHβ and GTHα subunits during sexual maturation" --- p.67 / Chapter 3.3.4 --- "Temporal expression profiles of zebrafish FSHp, LHβ and GTHα subunits during ovulatory cycle" --- p.68 / Chapter 3.3.5 --- "In situ hybridization of zebrafish FSHβ, LHβ and GTHα" --- p.69 / Chapter 3.4 --- Discussion --- p.70 / Chapter Chapter 4 --- General Discussion / Chapter 4.1 --- Cloning of zebrafish gonadotropin subunit cDNAs --- p.91 / Chapter 4.2 --- Bioactivity and receptor specificity of recombinant zebrafish FSH and LH --- p.91 / Chapter 4.3 --- Expression of gonadotropin subunits during zebrafish sexual maturation and ovulatory cycle --- p.92 / Chapter 4.4 --- "Localization of FSHβ, LHβ and GTHα subunits in zebrafish pituitary" --- p.93 / Chapter 4.5 --- Contributions of the present study --- p.94 / Chapter 4.6 --- Future prospects --- p.95 / References --- p.100

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

No description available.

Thesis (Ph. D.)--Washington State University, December 2009. / Title from PDF title page (viewed on Dec. 16, 2009). "School of Molecular Biosciences." Includes bibliographical references.

An experiment was conducted with Brangus cows to evaluate the effect of loss of body weight and condition on pituitary responsiveness to gonadotropin releasing hormone (GnRH) stimulation during late lactation. The treatment groups were lactating intact (LI), lactating ovariectomized (LO), nonlactating intact (NLI), and nonlactating ovariectomized (NLO). The study was carried out in two separate blocks, each one consisting of 3 periods. During period 1 the cows were fed a ration that supplied 90% and 88% of the NRC recommendations for TDN in lactating and nonlactating cows, respectively. This period lasted 170 in block 1 and 130 days in block 2. During period 2 the TDN was reduced to 55% or 52% for lactating and nonlactating cows, respectively. Period 2 lasted 100 days for cows in block 1 and 63 days for cows in block 2. At the beginning of period 3 TDN was further reduced to 25% or 27% for the lactating and nonlactating cows, respectively. Cows in block 1 were challenged with GnRH 40 days after the beginning of the 1st energy reduction, 30 days later and 7 days after the 2nd energy reduction. The cows in block 2 were challenged with GnRH 30 days after the 1st energy reduction, 30 days later and 25 days after the 2nd energy reduction. At the end of the study body composition parameters and organ gland weights were determined. No significant differences in the weights of the cows among the treatment groups were found. All cows were, however, losing weight through the course of this study. The nonlactating cows maintained higher body condition (P < .05) than lactating cows from 31 days after ovariectomies were performed until the end of the study. The pituitary glands were significantly heavier in the lactating ovariectomized (2.3 g vs. 1.7 g, P < .05) than the nonlactating intact cows. The weight of the adrenals per unit of body weight of LO cows was significantly higher (.057 g/kg vs. .040 g, P < .05) than among NLO cows. The percent of carcass lipid was significantly higher (P < .05) in nonlactating as compared to lactating cows. Percent moisture and protein were higher (P < .05) in lactating cows. Amount of LH released after GnRH stimulation tended to be higher in lactating than nonlactating cows. The magnitude of the LH peak did not differ significantly among the treatment groups at each of the dates GnRN was injected. Ovariectomized cows (LO and NLO) responded more rapidly (P < .05) to GnRH stimulation than intact cows (LI and NLI). Time on reduced TDN did not affect cow's response pattern after GnRH injection.

The present investigation was designed to further our understanding of the interaction of pituitary (luteinizing hormone, LH) and placental (chorionic gonadotropin, CG) gonadotropins with the primate corpus luteum. Studies were performed (1) to characterize the LH/CG receptor population in the