REGULATION OF TESTICULAR GONOCYTE DEVELOPMENT

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The objectives of this application are to determine the mechanisms regulating neonatal testicular gonocyte proliferation. Anomalies of this process could jeopardize the entire spermatogenic process. We have shown that neonatal gonocytes specifically expressed the protein C zeta (ZetaPKC), and the phosphatidylinositol 3-kinase (PI 3-kinase), which may play a role in PDGF-activated cell proliferation. Thus, we examined the effect of PDGF on isolated gonocytes from 3-day-old rats. Since neonatal Sertoli cells secrete 17beta-estradiol, we also tested its effect. Both agents were found to stimulate gonocyte proliferation. These results constitute the basis of our hypothesis that the cross talk between the PDGF- and estradiol-activated signal transduction pathways drives gonocyte mitosis, development and differentiation into spermatogonia. Our first aim is to determine the components of the PDGF transduction pathway in gonocytes. The role of zetaPKC and PI 3-kinase in this pathway will be examined, by studying the expression, kinase activity, and substrates of each enzyme. The identification of two key elements of the cascade, the MEK and MAP-kinase, will also be pursued. These two experiments should reveal important components of PDGF pathway in gonocytes and confirm if zetaPKC and PI 3-kinase are part of this cascade. The second aim is to determine if there is a cross talk between PDGF and estradiol pathways in gonocytes. This will be examined by studying 1) the effects of estrogen antagonists of PDGF-dependent cell proliferation, kinase activities and protein phosphorylation patterns and 2) the effects of PDGF and PDGF-activated kinase inhibitors on estradiol-dependent proliferation. The third aim is to examine if perinatal exposure to xenoestrogens has detrimental effects on gonocyte functions. Pregnant rats will be treated with estrogens, and their functions of the gonocytes derived from their offspring will be examined, including the responses to PDGF and estradiol and percent of apoptotic cells. These experiments should reveal if estrogens play role in vivo in gonocyte maturation, and may provide clarifications regarding the effects of xenoestrogens on male reproductive functions.