Growth hormone (GH) is secreted by the pituitary in a pulsatile pattern that acts on tissues throughout the body both directly and through the stimulation of insulin-like growth factor 1 (IGF-1). Treatments that increase GH have been shown to be effective in the treatment of GH deficient children and adults, and also have wide ranging potential applications toward the relative GH deficiency associated with aging and obesity, the weight loss and muscle wasting of AIDS, and the agricultural production of milk and lean meat. Growth hormone-releasing hormone (GHRH) is a hypothalamic neuropeptide that acts as the major stimulus for GH synthesis and release. GHRH mediates its actions by binding to pituitary GHRH receptors (GHRH-R) and activating an intracellular signal transduction pathway. An improved understanding of GHRH receptor structure, function, physiology and regulation will reveal the information needed to facilitate clinical and pharmacological manipulations of the GH axis. Toward this goal we propose the following specific aims: 1. Which domains of the GHRH-R are in close proximity to bound GHRH? A panel of five GHRH photoaffinity crosslinking probes and three receptor-directed antisera will be used to identify receptor domains in close contact with specific residues of bound hormone. These results will then be used to establish structure/function relationships and suggest structural requirements for receptor activation. 2. What mechanisms are involved in GHRH-R down regulation? We will examine how the GHRH induced receptor modification we report here relates to receptor down regulation and internalization in stable cell lines, primary cultured rat pituitary, and in naturally truncated ovine receptor. The roles of PKC, PKA and betaARK kinases will be tested. Mutant receptors will be prepared to identify the receptor domains involved. 3. Is GHRH the primary regulator of GHRH-R and is GHRH-R expression directly correlates to hypothalamic GHRH drive? A rat model with GH suppression of hypothalamic drive to the somototroph will be examined. This suppression will be challenged by either elevating endogenous GHRH with intracerebroventricular injection of GH antagonist or by iv infusion of exogenous GHRH.
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