Atrial natriuretic peptide-induced stimulation of cyclic GMP formation by isolated bovine luteal cells.

Incubation of bovine luteal cells with an atrial natriuretic peptide (rat atriopeptin II, rAP-II) did not affect hCG-stimulated or basal cyclic AMP accumulation and progesterone production, but cyclic GMP formation was stimulated by rAP-II in a dose-dependent manner, being maximally stimulatory in the nanomolar range. This stimulatory influence of rAP-II on cyclic GMP formation results from a specific stimulation of particulate guanylate cyclase. We suggest that, although rAP-II mediated cyclic GMP formation can be demonstrated in bovine luteal cells, there appear to be no acute effects of the atrial peptide on the regulation of progesterone production by these cells.

Renin and prorenin-like activities in bovine ovarian follicles.

Recent reports emerging from several laboratories have strengthened the concept of a physiological intrinsic ovarian renin-angiotensin system. Encouraged by these studies carried out mostly so far in human subjects, we decided to investigate if prorenin- and renin-like activities could be demonstrated in the follicular fluid obtained from bovine ovaries. The results obtained in this study show that in a total of 58 follicles examined, significant quantities of both prorenin- and renin-like activities could be demonstrated. The prorenin-like activity measured was invariably 30-40 times greater than the renin-like activity in the follicular fluid samples. There was an inverse relationship between the ratio of estradiol to progesterone concentrations and prorenin-like activity in the follicular fluid. Similarly, a significant negative correlation was seen between the estradiol concentration and prorenin-like activity in follicular fluid. On the other hand, there was no discernible relationship between androstendione concentrations and prorenin-like activity. With respect to progesterone, in large follicles with high (greater than 50 ng/ml) progesterone content, the prorenin-like activity was 3 times as high as that in the low (less than 50 ng/ml) progesterone group. There was, however, no difference in the levels of prorenin-like activity between the high and low progesterone groups in the case of small follicles. Furthermore, analysis of the contents of prorenin- and renin-like activities in extract prepared from granulosa and thecal cells revealed thecal cells of bovine ovarian follicles to be a major source of the enzyme activities. The ratio of prorenin to renin activity in thecal cell extract was close to 1. By analyzing the relationship between various steroid concentrations and prorenin-like activity in follicular fluid, it appears that the atretic follicles are likely to have significantly higher levels of prorenin-like activity in their fluid.

Atrial natriuretic factor stimulates luteal guanylate cyclase.

Effect of a synthetic atrial natriuretic peptide, rat atriopeptin II (rAP-II) on the formation of cyclic nucleotides and progesterone production in Percoll-purified rat luteal cells was investigated. Incubation of luteal cells with varying concentrations of rAP-II resulted in a dose-related stimulation of intracellular cyclic GMP content; maximum stimulation being achieved with 10 nM rAP-II. The increase in cyclic GMP formation was extremely rapid and a 12-fold increase in the cyclic GMP content over basal level was attained within 5 min of incubation of the cells with 10 nM rAP-II. In the presence of phosphodiesterase inhibitor, 3-isobutyl-1-methyl-xanthine, both basal and rAP-II-stimulated levels of cyclic GMP were increased approximately 10 times, but the magnitude of stimulation remained similar in the presence or absence of the inhibitor. The atrial peptide at the concentration of 1-100 nM, however, had no effect on either basal or gonadotropin-stimulated progesterone production and cyclic AMP formation by the luteal cells. Furthermore, the increase in the level of cellular cyclic GMP content of rAP-II was demonstrated to result from a selective activation of particulate guanylate cyclase.

Phorbol ester stimulates progesterone production by isolated bovine luteal cells.

The tumor promoter, phorbol 12-myristate-13-acetate (PMA), is known to modulate the response of several steroidogenic tissues presumably by activating a Ca++- and phospholipid-dependent protein kinase (protein kinase C). The presence of this kinase has been demonstrated in bovine corpus luteum, although its role in steroidogenesis by these cells is unknown. We report here the effects of PMA on progesterone production by the enzymically dispersed bovine luteal cells in vitro. PMA (1-50 nM) produced a dose- and time-related increase in progesterone production by the luteal cells. The maximum stimulation was achieved with 10 nM PMA. Higher concentrations of PMA led to a decline of steroidogenesis close to the basal level. A nonpromoting derivative, 4 alpha-phorbol 12,13-didecanoate had no effect. The PMA-induced stimulation of progesterone production was not associated with a change in the cAMP level. PMA added together with suboptimal doses of human CG, 8Br-cAMP, cholera toxin, or forskolin significantly increased the amount of progesterone produced. PMA as well as human CG-induced steroidogenesis was sensitive to cycloheximide inhibition. The conversion of exogenous pregnenolone or 25-hydroxycholesterol to progesterone was not altered by PMA. We conclude that PMA at nanomolar concentrations is able to stimulate progesterone production by bovine luteal cells and that the site of action of PMA is distal to the formation of cAMP but before the formation of pregnenolone. The observed effects of PMA in luteal cells are probably linked to its ability to activate protein kinase C, since a diacylglycerol could mimic the steroidogenic action of PMA.