Stimulation of Ca2+-dependent exocytosis of the sperm acrosome by cAMP acting downstream of phospholipase A(2)

Spermatozoa undergo exocytosis in response to agonists that induce Ca2+ inf lux and, in turn, activation of phosphoinositidase C, phospholipase C, phos pholipase A(2), and cAMP formation. Since the role of cAMP downstream of Ca 2+ influx is unknown, this study investigated whether cAMP modulates phosph olipase C or phospholipase A(2) using a ram sperm model stimulated with A23 187 and Ca2+. Exposure to dibutyryl-cAMP, phosphodiesterase inhibitors or f orskolin resulted in enhancement of exocytosis. However, the effect was not due to stimulation of phospholipase C or phospholipase A(2): in spermatozo a prelabelled with [H-3]palmitic acid or [C-14]arachidonic acid, these reag ents did not enhance [H-3]diacylglycerol formation or [C-14]arachidonic aci d release. Spermatozoa were treated with the phospholipase A(2) inhibitor a ristolochic acid, and dibutyryl-cAMP to test whether cAMP acts downstream o f phospholipase A(2). Under these conditions, exocytosis did not occur in r esponse to A23187 and Ca2+. However, inclusion of dibutyryl-cAMP and the ph ospholipase A(2) metabolite lysophosphatidylcholine did result in exocytosi s (at an extent similar to that seen when cells were treated with A23187/Ca 2+ and without the inhibitor). Inclusion of lysophosphatidylcholine alone, without dibutyryl-cAMP, enhanced exocytosis to a lesser extent, demonstrati ng that cAMP requires a phospholipase A(2) metabolite to stimulate the fina l stages of exocytosis. These results indicate that cAMP may act downstream of phospholipase A(2), exerting a regulatory role in the exocytosis trigge red by physiological agonists.