Activity of the Na,K-ATPase α4 isoform is important for membrane potential, intracellular Ca2+, and pH to maintain motility in rat spermatozoa

While the function of the ubiquitous Na,K-ATPase alpha 1 subunit has been well documented, the role of the sperm-specific alpha 4 isoform of this ion transporter is less known. We have explored the importance of a4 in rat sperm physiology by taking advantage of the high sensitivity of this isoform for the inhibitor ouabain. Using concentrations that selectively block alpha 4 activity, we found ouabain to reduce not only sperm total motility, but also multiple parameters of sperm movement, including progressive motility, straight line, curvilinear, and average path velocities, lateral head displacement, beat cross frequency, and linearity. According to a direct role of alpha 4 in Na C transport, ouabain inhibition of alpha 4 increased [Na+](i) in the male gametes. In addition, interference of alpha 4 activity with ouabain produced cell membrane depolarization, diminished pH, and increased [Ca2+](i) in spermatozoa. Inhibition of alpha 4 was sufficient to cause all these effects and additional blockage of alpha 1, the other Na, K-ATPase a isoform expressed in sperm, and higher doses of ouabain did not result in further changes in the cell parameters studied. These results show that alpha 4 is the Na,K-ATPase isoform primarily involved in controlling the transmembrane Na+ gradient in sperm, and that alpha 4 activity is necessary for maintaining membrane potential, [Ca2+](i), and [H+](i) in the cells. The high dependence of sperm motility on membrane excitability, [Ca2+](i), and acid-base balance suggests that their regulation is the mechanism by which alpha 4 maintains motility of the male gametes. Reproduction (2010) 139 835-845