Existence and importance of Na+K+-ATPase in the plasma membrane of boar spermatozoa

Sodium-potassium-ATPase (Na+K+-ATPase), a target to treat congestive heart failure, is the only known receptor for cardiac glycosides implicated in intracellular signaling and additionally functions enzymatically in ion transport. Spermatozoa need transmembrane ion transport and signaling to fertilize, and Na+K+-ATPase is identified here for the first time in boar spermatozoa. Head plasma membrane (HPM) isolated from boar spermatozoa was confirmed pure by marker enzymes acid and alkaline phosphatase (218 +/- 23% and 245 +/- 38% enrichment, respectively, versus whole spermatozoa). Western immunoblotting detected alpha and 6 subunits (isoforms alpha 1, alpha 3, 61, 62, and 63) in different concentrations in whole spermatozoa and HPM. Immunofluorescence of intact sperm only detected alpha 3 on the post-equatorial exterior membrane; methanol-permeabilized sperm also had alpha 3 post-equatorially and other isoforms on the acrosomal ridge and cap. Mass spectrometry confirmed the presence of all isoforms in HPM. Incubating boar sperm in capacitating media to induce the physiological changes preceding fertilization significantly increased the percentage of capacitated sperm compared to 0 h control (33.0 +/- 2.6% vs. 19.2 +/- 2.6% capacitated sperm, respectively; p = 0.014) and altered the 62 immunofluorescence pattern. These results demonstrate the presence of Na+K+-ATPase in boar sperm HPM and that it changes during capacitation.