Prolactin modulates steroidogenesis by rat granulosa cells: II. Effects on estradiol

Abstract

The effects of prolactin on secretion of estradiol by rat granulosa cells obtained at two stages of differentiation and cultured under various conditions were determined. Relatively undifferentiated granulosa cells were obtained from immature, diethylstilbestrol-treated, hypophysectomized (HPX) rats and cultured in serum-free medium or with 10% serum. More highly differentiated granulosa cells were obtained on the morning of proestrous from the preovulatory follicles of immature rats in which an estrous cycle had been induced with 4 IU pregnant mare's serum gonadotropin; these cells were cultured in medium containing 10% serum. Cells were cultured for 3 days with graded doses of prolactin (0, 0.02, 0.2, 2, or 10 μg/ml) alone or in combination with follicle-stimulating hormone (FSH; 300 ng/ml), testosterone (0.5 μM), or FSH + testosterone. In control cultures (no prolactin) the relatively undifferentiated granulosa cells from HPX rats secreted negligible quantities of estradiol except when both FSH and testosterone were supplied. Prolactin alone or in combination with FSH or testosterone had no effect on estradiol secretion, but prolactin in combination with FSH + testosterone significantly decreased secretion in a dose-dependent fashion. This set of prolactin treatments was applied in both serum-free medium and medium containing 10% serum, with similar results under both culture conditions. The inhibitory effects of prolactin appeared to be reversible if cells were cultured with prolactin for only 1 day, but were not reversed if cells were cultured with prolactin for 2 or 3 days. Prolactin inhibited secretion of estradiol by cultures that had been exposed initially to FSH + testosterone (to induce aromatase), but the prior exposure to FSH increased the lag time before the effects of prolactin were evident. Cells from proestrous rats secreted only small amounts of estradiol in medium alone or with FSH. Either FSH or FSH + testosterone greatly increased estradiol secretion on the first 2 days of culture, but only FSH + testosterone maintained aromatase activity on the third day of culture. Prolactin alone or in combination with FSH had no effect on estradiol secretion. On the thrid day of culture, prolactin significantly increased estradiol secretion in the presence of testosterone alone, but decreased secretion when both FSH and testosterone were present. In summary, under various culture conditions and at two different stages of differentiation, prolactin inhibited FSH-stimulated secretion of estradiol. The data suggest that prolactin inhibits secretion of estradiol by rat granulosa cells by inhibiting the induction of aromatase by FSH.