Only a subpopulation of mouse sperm displays a rapid increase in intracellular calcium during capacitation.

Mammalian sperm must undergo a functionally defined process called capacitation to be able to fertilize oocytes. They become capacitated in vivo by interacting with the female reproductive tract or in vitro in a defined capacitation medium that contains bovine serum albumin, calcium (Ca ), and bicarbonate (HCO ). In this work, sperm were double stained with propidium iodide and the Ca dye Fluo-4 AM and analyzed by flow cytometry to determine changes in intracellular Ca concentration ([Ca ] ) in individual live sperm. An increase in [Ca ] was observed in a subpopulation of capacitated live sperm when compared with noncapacitated ones. Sperm exposed to the capacitating medium displayed a rapid increase in [Ca ] within 1 min of incubation, which remained sustained for 90 min. These rise in [Ca ] after 90 min of incubation in the capacitating medium was evidenced by an increase in the normalized median fluorescence intensity. This increase was dependent on the presence of extracellular Ca and, at least in part, reflected the contribution of a new subpopulation of sperm with higher [Ca ] . In addition, it was determined that the capacitation-associated [Ca ] increase was dependent of CatSper channels, as sperm derived from CatSper knockout (CatSper KO) or incubated in the presence of CatSper inhibitors failed to increase [Ca ] . Surprisingly, a minimum increase in [Ca ] was also observed in CatSper KO sperm suggesting the existence of other Ca transport systems. Altogether, these results indicate that a subpopulation of sperm increases [Ca ] very rapidly during capacitation mainly due to a CatSper-mediated influx of extracellular Ca .