ACUTE EFFECTS OF PROSTAGLANDIN-F(2-ALPHA), LUTEINIZING-HORMONE, AND ESTRADIOL ON 2ND MESSENGER SYSTEMS AND ON THE SECRETION OF OXYTOCIN AND PROGESTERONE FROM GRANULOSA AND EARLY LUTEAL CELLS OF THE EWE

Previous reports have suggested that gonadotropins, estradiol, and prostaglandin F2alpha (PGF2alpha) have varying effects on progesterone and oxytocin synthesis or secretion in cultured granulosa and luteal cells collected at different stages of the estrous cycle. The experiments reported here were designed to investigate whether effects of these agonists on secretion of hormones and their coupling to second messenger systems changed around the time of ovulation. Granulosa cells and Day 2 luteal cells of the ewe were cultured for three days and then treated for 30 min with varying doses of PGF2alpha, LH, or estradiol. LH increased intracellular cAMP at both stages, but granulosa cells were more responsive in terms of both minimum effective dose (10 compared with 100 ng/ml) and degree of stimulation. LH caused no change in intracellular inositol phosphate levels. Both granulosa and early luteal cells responded to LH treatment by an increase in progesterone output in a dose-responsive fashion. PGF2alpha increased inositol phosphate accumulation in cells collected at both stages of the cycle. All doses tested (10(-6)-10(-8) M) stimulated the release of oxytocin into the culture medium from both granulosa and luteal cells. Progesterone secretion was also increased, but only at the highest dose (10(-6) M). Estradiol treatment (10(-6) M) did not affect either the inositol phosphate or cAMP second messenger systems, but it did inhibit the secretion of oxytocin from granulosa cells. It is concluded that PGF2alpha stimulates oxytocin secretion in vitro via activation of the inositol phosphate/Ca2+ second messenger pathway, and that LH increases progesterone synthesis acting via cAMP in both luteinized granulosa and early luteal cells, whereas estradiol does not work through either of these pathways. These data suggest that previously reported changes in the chronic effects of gonadotropins, PGF2alpha, and estradiol on oxytocin synthesis during the periovulatory period cannot be explained in terms of activation of alternative signalling pathways. The short-term stimulatory effects of PGF2alpha on progesterone output differ from its reported inhibitory effect in the mature ovine CL and support the idea that granulosa/luteal cell responsiveness to the same agonists alters during luteal development.