Biphasic role of calcium in mouse sperm capacitation signaling pathways

Mammalian sperm acquire fertilizing ability in the female tract in a process known as capacitation. At the molecular level, capacitation is associated with up-regulation of a cAMP-dependent pathway, changes in intracellular pH, intracellular Ca(2+), and an increase in tyrosine phosphorylation. How these signaling systems interact during capacitation is not well understood. Results presented in this study indicate that Ca(2+) ions have a biphasic role in the regulation of cAMP-dependent signaling. Media without added Ca(2+) salts (nominal zero Ca(2+)) still contain micromolar concentrations of this ion. Sperm incubated in this medium did not undergo PKA activation or the increase in tyrosine phosphorylation suggesting that these phosphorylation pathways require Ca(2+). However, chelation of the extracellular Ca(2+) traces by EGTA induced both cAMP-dependent phosphorylation and the increase in tyrosine phosphorylation. The EGTA effect in nominal zero Ca(2+) media was mimicked by two calmodulin antagonists, W7 and calmidazolium, and by the calcineurin inhibitor cyclosporine A. These results suggest that Ca(2+) ions regulate sperm cAMP and tyrosine phosphorylation pathways in a biphasic manner and that some of its effects are mediated by calmodulin. Interestingly, contrary to wild-type mouse sperm, sperm from CatSper1 KO mice underwent PKA activation and an increase in tyrosine phosphorylation upon incubation in nominal zero Ca(2+) media. Therefore, sperm lacking Catsper Ca(2+) channels behave as wild-type sperm incubated in the presence of EGTA. This latter result suggests that Catsper transports the Ca(2+) involved in the regulation of cAMP-dependent and tyrosine phosphorylation pathways required for sperm capacitation.