INDUCTION OF GRANULOSA-CELL DIFFERENTIATION BY FORSKOLIN - STIMULATION OF ADENOSINE-3',5'-MONOPHOSPHATE PRODUCTION, PROGESTERONE SYNTHESIS, AND LUTEINIZING-HORMONE RECEPTOR EXPRESSION

The effects of forskolin on the acquisition of differentiated functions in cultured ovarian granulosa cells were compared with the actions of FSH and prostaglandin E2 (PGE2). In 48 h granulosa cell cultures from immature diethylstilbestrol-treated rats, 100 .mu.M forskolin caused a 45-fold increase in cAMP accumulation and stimulated progesterone production from undetectable levels (< 0.02 ng/ml) to 80 ng/ml. The forskolin-induced increase in cAMP was similar to the maximum response to FSH, and progesterone production was .apprx. 50% of that elicited by FSH. PGE2 also enhanced cAMP and progesterone production in a concentration-dependent manner, with a maximum 8-fold increase in cAMP accumulation and an increase in progesterone to 5.6 ng/ml when the PGE2 concentration was 10 .mu.g/ml. The time course of forskolin-stimulated cAMP production was notable for its rapid rise to the maximum level during the first 24 h of culture, followed by a plateau for up to 72 h. This contrasted with FSH-stimulated cAMP production, which increased progressively for up to 72 h when measured at 24 h intervals. LH [luteinizing hormone] receptor levels were low in untreated cells and after exposure to the various stimuli for 24 h, but increased 9- to 11-fold after culture with FSH or forskolin for 48-72 h. PGE2-induced LH receptor formation was .apprx. 20% of that seen after FSH stimulation. Forskolin enhanced cAMP and progesterone production in response to FSH and choleragen, but impaired the effects of these ligands on LH receptor formation. Exposure of the cultured cells to a potent GnRH agonist inhibited forskolin-induced progesterone and LH receptor synthesis, but did not influence forskolin-stimulated cAMP production. The ability of forskolin to serve as a nonhormonal stimulator of granulosa cell differentiation is demonstrated and the importance of cAMP in this process is indicated as well as the ability of GnRH[gonadotropin-releasing hormone] agonists to exert inhibitory effects on post-cAMP steps in cellular maturation.