published_or_final_version / Zoology / Master / Master of Philosophy

Thesis (M.S.)--West Virginia University, 2004. / Title from document title page. Document formatted into pages; contains iv, 124 p. : ill. (some col.). Includes abstract. Includes bibliographical references (p. 82-124).

Call number: LD2668 .T4 1984 S585 / Master of Science

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.

Although much more work needs to be done to elucidate the functional roles of FSH and LH in fish reproduction, the preset study provides a relatively comprehensive study for us to understand the potential roles of FSH and LH during ovarian development in fish, especially the importance of FSH. / At the same time, functional studies were carried out to examine and compare bioactivities of the CHO-derived zfFSH and zfLH in zebrafish ovary, which is the major part of the present project. The following aspects were covered to investigate the actions of zfFSH and zfLH: steroidogenesis and folliculogenesis. / Both recombinant zfGTHs stimulated activin betaA expression but slightly suppressed activin betaB expression. During short-term treatment, zfFSH and zfLH exhibited similar stimulatory effects on activin betaA expression; the effect of zfLH became more prominent after 24 h treatment while zfFSH had little effect. / Previously, our laboratory had established two stable Chinese hamster ovary (CHO) cell lines expressing recombinant zebrafish FSH (zfFSH) and LH (zfLH). However, the production yields are very low. Therefore, the present study tried to adopt the yeast Pichia pastoris as another bioreactor to produce recombinant zfFSH and zfLH. Two different forms of expression vectors for a native form and a fusion form carrying a His-tag, respectively, were constructed for each hormone. Their bioactivities were monitored and confirmed by receptor-based reporter gene assays as well as ovarian fragment incubation. As expected, the native form exhibited much higher activities than the fusion form. / The pituitary gonadotropins (GTHs), follicle-stimulating hormone (FSH) and luteinizing hormone (LH), are the key hormones controlling vertebrate reproduction. Although the two gonadotropins have been characterized in numerous teleost species, our understanding of their biological functions remains rather limited. This is largely due to the lack of pure form of homologous gonadotropins and inadequate understanding of gonadal physiology in most species studied as well as species variation of hormone actions. The present study aims at systematically investigating the functional roles of FSH and LH in the ovary using zebrafish as the model. Zebrafish is becoming more and more popular as the model of reproductive and developmental studies due to several advantages. First, though its body size is small, its ovary is relatively large and available all the year around. Second, zebrafish spawns everyday and its development is fast. Last but not least, its bioinformatics information is tremendous compared to other fish models. / We investigated the effects of zfFSH and zfLH on steroidogenesis by examining the regulation of aromatase by these two hormones. Aromatase catalyzes the conversion of androgens into estrogens during steroidogenesis. Both recombinant zfGTHs stimulated the aromatase expression during short-term treatment (8 h) in ovarian fragment culture, with zfFSH much more potent than zfLH. However, zfFSH continued to exhibit powerful effect on aromatase expression after 24 h treatment while zfLH had little effect at all. The stimulatory effect of zfFSH on aromatase expression was time-, dose- and stage-dependent and was also confirmed by in vivo study. Furthermore, it was also zfFSH but not zfLH that significantly stimulated StAR protein expression during short-term treatment. StAR protein is critical to steroidogenesis by facilitating the movement of cholesterol across the mitochondrial membrane. / zfLH was found to be able to induce GVBD in zebrafish, as demonstrated in other fish species. However, our preliminary data showed that zfFSH was also involved in this process. To our knowledge, this is the first time to demonstrate that homologous FSH induces GVBD in teleosts. / Yu, Xiaobin. / Adviser: Wei Ge. / Source: Dissertation Abstracts International, Volume: 70-09, Section: B, page: . / Thesis submitted in: December 2008. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 152-181). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

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.

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