Balthasar, Nina; (2001) Transgenic studies of growth hormone secretagogue physiology. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The synthetic growth hormone secretagogues (GHS) exert their growth hormone (GH) releasing effects by activating their receptor (GHS-R) in hypothalamic neurones and pituitary GH cells. The aim of this study was to investigate hypothalamic and pituitary GHS physiology by in vitro studies and transgenesis, over-expressing the GHS-R in hypothalamic GH releasing hormone (GHRH) neurones or pituitary GH cells. Initially, to analyse transgene expression driven by two targeting cosmids - a rat GHRH or a human GH cosmid - transgenic mice expressing enhanced green fluorescent protein (eGFP) were generated. In rGHRH-eGFP transgenic mice eGFP protein was visible in brightly fluorescing GHRH neurone cell bodies and median eminence nerve terminals. hGH-eGFP transgenic mice showed high levels of eGFP fluorescence limited to GH expressing pituitary cells. These mice provide useful new tools to study GHRH and GH cell physiology. In vitro studies with a stably GHS-R transfected GH cell line showed that processing of the GHS-R cDNA resulted in a functionally active receptor. In transgenic mice overexpression of the GHS-R in GHRH neurones caused alteration of the GH axis with increased pituitary GH content and body weight. GH responses to a GHS injection seemed enhanced. However, prolonged GHS exposure did not cause enhanced body weight gain compared to WT littermates. These studies suggested that enhancement of GHS-R abundance in GHRH neurones caused alteration of the GH and GHS axis, implicating GHRH neurones as an important GHS target in GH release. In contrast, transgenic mice with 50-fold GHS-R over-expression in GH cells showed reduced pituitary GH content, body weight and GH responses to GHRH. These data suggest that somatotroph function might be compromised by high levels of GHS-R over-expression. Crossing the hGHS-R over-expressing mice with their eGFP counterpart mice will facilitate the further investigation of their physiological phenotype by enabling in vivo analysis of GHS effects directly in identified cells.

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