REGULATION OF LH RECEPTORS AND PROGESTERONE SECRETION

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The goal of this research is to obtain a better understanding of the factors which regulate secretion of progesterone from the corpus luteum in order to develop better contraceptives and treat infertility. The proposed research has five specific aims. 1) To determine if mLH and mCG have different receptor-mediated actions in primate luteal cells, rhodamine-labeled hormones will be used to measure the lateral mobility of the receptor for LH by laser photobleach recovery. If mCG, but not mLH, immobilizes the receptor for LH in primate luteal cells, it will be determined if mCG also causes a greater increase in secretion of progesterone than mLH. The results of this study have very important implications for treatment of infertility. 2) To determine the follicular source of origin of the two types of steroidogenic luteal cells, lipophilic dyes will be used to specifically label thecal or granulosal cells in the preovulatory follicle or the corpus hemorrhagicum. Luteal tissue will be collected during the early, mid and late luteal phases of the estrous cycle and the origins of large and small luteal cells determined based on the presence of the fluorescent, lipophilic dyes. 3) To elucidate the role of growth factors in controlling the division, differentiation and function of luteal cells, the number of receptors for IGF-I, IGF-II, EGF and FGF will be determined at different stages of the estrous cycle. In situ hybridization of cRNA probes will be used to localize mRNA for the growth factors to determine which cell types synthesize these factors. Subsequent studies will evaluate the actions of growth factors and control of their secretion in purified preparations of luteal cells. 4) To study control of steroidogenesis in the two types of steroidogenic luteal cells, initial studies will evaluate the lipoprotein pathway. The role of calcium will be studied using "patch clamp" procedures to evaluate the number and activity of calcium channels and fluorescent dyes will be used to monitor intracellular calcium shifts. Inhibitors and stimulators of calmodulin and protein kinase C activity will be used to evaluate the activity of these second messengers. 5) Finally, numbers of luteal cells, their receptor content and responsiveness to various hormones will be determined in the two steroidogenic luteal cell types to define at the cellular level changes occurring during transition of the corpus luteum of the estrous cycle to the corpus luteum of pregnancy.