CatSper channels are regulated by protein kinase A

Mammalian sperm must undergo capacitation as a preparation for entering into hyperactivated motility, undergoing the acrosome reaction, and acquiring fertilizing ability. One of the initial capacitation events occurs when sperm encounter an elevated HCO³_- concentration. This anion activates the atypical adenylyl cyclase Adcy10, increases intracellular cAMP, and stimulates protein kinase A (PKA). Moreover, an increase in intracellular Ca²⁺ concentration ([Ca²⁺]i) is essential for sperm capacitation. Although a cross-talk between cAMP-dependent pathways and Ca²⁺ clearly plays an essential role in sperm capacitation, the connection between these signaling events is incompletely understood. Here, using three different approaches, we found that CatSper, the main sperm Ca²⁺ channel characterized to date, is up-regulated by a cAMP-dependent activation of PKA in mouse sperm. First, HCO³_- and the PKA-activating permeable compound 8-Br-cAMP induced an increase in [Ca²⁺]i, which was blocked by the PKA peptide inhibitor PKI, and H89, another PKA inhibitor, also abrogated the 8-Br-cAMP response. Second, HCO³_- increased the membrane depolarization induced upon divalent cation removal by promoting influx of monovalent cations through CatSper channels, which was inhibited by PKI, H89, and the CatSper blocker HC-056456. Third, electrophysiological patch clamp, wholecell recordings revealed that CatSper activity is up-regulated by HCO³_- and by direct cAMP injection through the patch-clamp pipette. The activation by HCO³_- and cAMP was also blocked by PKI, H89, Rp-cAMPS, and HC-056456, and electrophysiological recordings in sperm from CatSper-KO mice confirmed CatSper's role in these activation modes. Our results strongly suggest that PKA-dependent phosphorylation regulates [Ca²⁺]i homeostasis by activating CatSper channel complexes.