Progesterone primes zona pellucida‐induced activation of phospholipase A 2 during acrosomal exocytosis in guinea pig spermatozoa

We investigated, using guinea‐pig spermatozoa as a model, whether phospholipase A 2 (PLA 2 ) is involved in progesterone or zona pellucida (ZP)‐stimulated acrosomal exocytosis, if progesterone enhances ZP‐induced activation of PLA 2 , and mechanisms underlying PLA 2 regulation. Spermatozoa were capacitated and labeled in low Ca 2+ medium with [ 14 C]choline chloride or [ 14 C]arachidonic acid, washed, and then exposed to millimolar Ca 2+ and progesterone and/or ZP. Each agonist stimulated decrease of phosphatidylcholine (PC) and release of arachidonic acid and lysoPC, indicative of PLA 2 activation. Aristolochic acid (a PLA 2 inhibitor) abrogated lipid changes and exocytosis, indicating that these lipid changes are essential for exocytosis. Exposure of spermatozoa to submaximal concentrations of both progesterone and ZP resulted in a synergistic increase of arachidonic acid and lysoPC releases, and exocytosis, suggesting that, under natural conditions, both agonists interact to bring about acrosomal exocytosis. Progesterone‐induced PLA 2 activation appears to be mediated by a GABA A ‐like receptor, because bicuculline (a GABA A receptor antagonist) blocked arachidonic acid release and exocytosis. In agreement with this, GABA mimicked progesterone actions. ZP‐induced activation of PLA 2 seemed to be transduced via G i proteins because pertussis toxin blocked arachidonic acid release and acrosomal exocytosis. PLA 2 may be regulated by PKC because progesterone‐ or ZP‐induced release of arachidonic acid was blocked by the PKC inhibitors staurosporine or chelerythrine chloride. PLA 2 could also be regulated by the cAMP‐PKA pathway; inclusion of the PKA inhibitor 14–22 amide or H‐89 led to a reduction in arachidonic acid release or exocytosis after progesterone or ZP. Taken together, these results suggest that PLA 2 plays an essential role in progesterone or ZP‐stimulated exocytosis with progesterone priming ZP action. © 2005 Wiley‐Liss, Inc.