Emerging Concepts in Ion Channel Biophysics

Emerging Concepts in Ion Channel Biophysics

Poster Abstracts

56-POS Board 56 CatSper Channels Are Activated by PKA in Mouse Sperm

Gerardo Orta 1 , José L. De la Vega-Beltran 1 , David Martín-Hidalgo 2 , Pablo E. Visconti 2 , Alberto Darszon 1 . 1 Instituto de Biotecnología-Universidad Nacional Autónoma de México, Cuernavaca, Morelos, Mexico, 2 University of Massachusetts, Amherst, MA, USA. In mammals, sperm become motile during ejaculation and swim up the female reproductive tract where they undergo physiological changes named capacitation before being capable of fertilizing the egg. One of the first signaling events is an intracellular cAMP elevation triggered by HCO3- influx which activates the soluble adenylyl cyclase (sAC)1, leading to PKA-dependent protein phosphorylation2. CatSper, a sperm exclusive, alkali- and voltage-gated Ca2+ channel, has been proposed to be responsible for the Ca2+ influx that accompanies and is necessary for capacitation. The key role of cAMP in capacitation and fertilization was demonstrated using pharmacological tools for gain and loss of function of PKA-dependent pathways3 and confirmed with sAC null mouse models whose sperm are sterile4. We hypothesized that CatSper is the molecular candidate in this mechanistic pathway. We tested this possibility using western blot analysis, Em and [Ca2+]i measurements, and CatSper whole-cell patch-clamp current recordings. Our results show a direct regulation of CatSper by PKA when stimulated by HCO3- influx. Notably, in whole-cell patch-clamp recordings, cAMP in the pipette solution activates CatSper currents strongly and this effect is blocked by PKA inhibitors H89 and PKI. Pharmacological inhibition of the PKA-pathway and CatSper blockade annul the effect of HCO3- on: [Ca2+]i levels, Em and CatSper currents. Additionally, our experiments with sperm from CatSper KO mice confirm that HCO3- does not alter the ionic currents obtained under CatSper channel recording conditions. All together our results indicate that CatSper channels are activated by PKA directly, having a major role in the Ca2+ influx occurring during capacitation. These findings have significant implications for our understanding of fertilization. Support acknowledged from CONACyT Fronteras 71, PAPIIT/UNAM IN205516 and NIH RO1 HD038082-13 to Darszon A. NIH RO1 HD038082-13 to Visconti PE.

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