Development of ovarian granulosa cells is controlled by peptide and steroid hormones, neurotransmitters and growth factors. We have shown that these cells undergo hormone-dependent differentiation, luteinization and new hormone receptor acquisition in a serum-free culture system. Our present aims are: (1) To continue studies on the auto- and hetero-regulation of FSH, LH and prolactin receptors in vitro, accompanied by studies on the gonadotropin regulation of phosphodiesterase activity and cAMP production. Since growth hormone (GH) and insulin-like growth factors (IGFs) enhance FSH action, we will test the modulatory role of GH, IGF-I, IGF-II and insulin on steroidogenesis and gondatropin receptor induction in vitro. The hypothesis that GH may induce granulosa cells to release IGFs to exert autocrine action will be tested by elucidating the GH regulation of ovarian content of IGFs. We will characterize granulosa cell receptors for GH and IGFs, and study their modulation by gonadotropins. (2) To elucidate intra-ovarian roles of steroids. We will study the inhibitory effect of androgens as well as the augmenting action of estrogens on gonadotropin-stimulated LH and prolactin receptor induction. Using high-titer estrogen antiserum, we will block the action of endogenously produced steroids to test the role of intrafollicular steroids in autocrine control mechanisms. Triphenylethylene """"""""antiestrogens"""""""" exert disparate actions on granulosa cell estrogen and progestin biosynthesis, while a new """"""""anti-estrogen"""""""" (LY-117018) may be anti-gonadotropic. The presence of specific """"""""anti-estrogen"""""""" receptors and the actions of these pharmacologic agents will be tested in vitro and in vivo. (3) To study the direct modulatory roles of cholinergic and GABAnergic agents on granulosa cell steroidogenesis and gonadotropin receptor induction. Since vasoactive intestinal peptide (VIP) induces granulosa cell steroidogenesis, the ovarian presence and mechanism of action of this putative neuropeptide will be studied. (4) To separate and characterize subpopulations of granulosa cells based on a density gradient centrifugation method. The differential distribution of various hormone receptors as well as differentiated functions will be studied to understand the heterogeneity of the follicular cells. The present proposal will enhance our understanding of the hormonal control processes involved in follicular maturation and luteinization.
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