Calcium-dependent, cyclic nucleotide-independent steroidogenesis in the bovine placenta

Dispersed bovine placental cells (fetal cotyledon and maternal caruncle) were shown to synthesize progesterone. To determine if their steroidogenic activity could be modulated by a cyclic nucleotide-mediated process, we added luteinizing hormone, 8-bromoadenosine 3',5'-monophosphate, 8-bromoguanosine 3',5'-monophosphate, adenosine, or cholera toxin to dispersed cells from placentomes of 100-283 days gestational age and examined progesterone synthesis during 3-to 16-hr incubation periods. Net progesterone production, defined as the amount of progesterone released in excess of the zero-time cellular progesterone content, was determined by using a specific RIA. None of these agents significantly affected progesterone synthesis. In contrast, the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (MIX; 0.2-0.5 mM) caused a marked increase in progesterone formation. In time course studies it was found that MIX produced a 5-fold increase in progesterone production in 16 hr, with steroid production increasing linearly during this time. MIX also increased the conversion of exogenous pregnenolone to progesterone by placental cells. In view of the failure of cyclic nucleotide analogues and activators of adenylate cyclase to stimulate steroidogenesis, it was necessary to consider other modes of action of MIX. Since MIX is known to affect intracellular calcium translocation, we examined the effects of the calcium ionophore A23187 on progesterone formation. This drug enhanced progesterone formation and augmented the stimulatory effects of MIX. The stimulatory action of A23187 was not affected by cyclic nucleotide analogues. Our data suggest that progesterone synthesis in the bovine placentome is calcium dependent and cyclic nucleotide independent.