Regulation of Prostaglandin Endoperoxide Synthase by Cyclic Adenosine 3′,5′-Monophosphate in the in Vitro-Perfused Rat Ovary1

The preovulatory regulation of two enzymes in the prostaglandin biosynthetic pathway, prostaglandin endoperoxide synthase (PGS) and prostacyclin synthase (ISN), was examined in granulosa cells and residual tissue of rat ovaries perfused in vitro. Ovaries from rats primed with pregnant mare’s serum gonadotropin (20 IU) were perfused for up to 20 h starting the morning of induced proestrus. The amounts of PGS and ISN present were analyzed with immunoblotting techniques. Soluble extracts from granulosa cells and residual ovarian tissues were obtained at different times (0 h, 3 h, 7 h, 12 h) after treatment in vitro with luteinizing hormone (LH, 0.1 μg/ml) and 3-isobutyl-1-methylxanthine (IBMX; 0.2 mM) and at 7 h in untreated control ovaries or after treatment with forskolin (30 μM) or LH (0.1 μg/ml). The levels in the perfusion medium of cyclic adenosine 3′,5′-monophosphate (cAMP), progesterone, testosterone, and estradiol were measured and the number of ovulations were examined. The levels of PGS after treatment with LH + IBMX increased up 1o7 hand remained high at 12 h, a time that is close to the rime of ovulation. The increase was more pronounced in the granulosa cells than in the residual tissue. Treatment with forsko!in induced synthesis of PGS in granulosa cells, and the levels at 7 h were similar to those after stimulation with LH + IBMX. The levels of PGS were lower in granulosa cells of the group stimulated with LH alone than in granulosa cells from ovaries stimulated with LH + IBMX orforskolin. There were significantly (p <O.O1) more ovulations in the groups treated with LH + IBMX (242 ± 5.1) orforskolin (212 ± 4.8) than in the group treated with LH alone (3.8 ± 1.4). The release of cAMP was greatest in the group treated with forskolin, followed by the group treated with LH + IBMX and the group treated with LH alone. Progesterone remained highest in the forskolin-ereared group,followed by the group treated with LH + IBMX and LH alone. The levels of testosterone and estradiol were similar in all treatment groups. In conclusion, the induction of PGS but not of ISN was stimulated by LH and mediated by cAMP. Moreover, the total ovarian levels of PGS were related to the nwnber of ovulating follicles, unplying that a rise in PGS levels is one mechanism by which the ovulatory rise infollicularprostaglandin levels is mediated.