Role for Ca2+ channels in the signal transduction pathway leading to acrosomal exocytosis in human spermatozoa

Progesterone interaction with human spermatozoa promotes a rise in intracellular Ca²⁺ and can trigger acrosomal exocytosis in capacitated cells. We have used nifedipine, a 1,4-dihydropyridine Ca²⁺ channel antagonist, to investigate the possibility that Ca²⁺ channels play a role in the progesterone-stimulated exocytotic response. Cells were assessed biochemically for the generation of diacylglycerol (DAG) and microscopically for acrosome loss using chlortetracycline fluorescence. When motile cells were preincubated for 5 hr using culture conditions similar to those used for successful human in vitro fertilization, a short exposure to progesterone significantly stimulated DAG formation and acrosomal exocytosis. The addition of nifedipine (10 and 100 nM), either at time 0 or just prior to progesterone introduction, significantly inhibited both DAG formation and exocytosis, suggesting that Ca²⁺ channels are involved in the responses observed. Treatment of capacitated cells with a synthetic permeant DAG stimulated exocytosis irrespective of whether nifedipine was present, indicating that Ca²⁺ channels function prior to DAG generation. The possibility that an influx of Na⁺, as well as Ca²⁺, might be involved in the exocytotic pathway was investigated using the monovalent cation ionophores monensin and nigericin. Both significantly stimulated DAG generation and acrosome loss, but the prior inclusion of nifedipine significantly inhibited all responses. These results strongly suggest that the entry of Ca²⁺ through Ca²⁺ channels, with characteristics similar to those of L-type, voltage-sensitive Ca²⁺ channels found in cardiac and skeletal muscle, is a crucial step in the sequence of events leading to exocytosis in progesterone-stimulated human spermatozoa. An influx of Na⁺ also may play a role, but at a point prior to the opening of Ca²⁺ channels. © 1996 Wiley-Liss, Inc.