Mechanisms by which calcium ionophore and phorbol ester modulate steroid production by goldfish preovulatory ovarian follicles

Abstract Recent studies have shown that agents that increase intracellular calcium content (calcium ionophore A23187) and activate protein kinase C (phorbol 12‐myristate 13‐acetate; PMA) modulate basal and cAMP‐stimulated steroid production by goldfish preovulatory ovarian follicles. This report describes studies evaluating sites in the steroidogenic cascade (pre‐ and postadenylyl cyclase) influenced by calcium and protein kinase C. PMA (12.5–400 nM) inhibits hCG stimulated cAMP production, whereas phorbol 12, 13‐dideconate, an inactive phorbol ester that does not activate protein kinase C, was without effect. Earlier studies suggested that PMA also acts post cAMP formation, but this action was shown to be exerted prior to cholesterol side‐chain cleavage as metabolism of 25‐hydroxycholesterol to testosterone was not inhibited by PMA. A high dosage of A23187 (4000 nM) inhibited the stimulatory action of hCG on cAMP and testosterone production; lower dosages of A23187 were without effect. In contrast, a wide range of A23187 dosages (250–4,000 nM) enhanced conversion of 25‐hydroxycholesterol to testosterone. This effect appears to be exerted at the level of the cholesterol side‐chain cleavage enzyme as A23187 did not affect conversion of exogenous pregnenolone to testosterone. These data demonstrate specific regulatory actions of calcium and protein kinase C at multiple sites in the steroidogenic cascade both proximal and distal to cAMP generation. © 1992 Wiley‐Liss, Inc.