In vitro capacitation of bovine spermatozoa: role of intracellular calcium.

The development of successful methods of in vitro fertilization for bovine oocytes has advanced the bovine as a model for reproductive technology. The discovery of heparin as a capacitating agent has made it possible for investigators to have an inexpensive, readily available supply of bovine gametes for experimentation in reproductive biotechnologies such as gene transfer and cloning. The central event that mammalian sperm must undergo before being able to fertilize an oocyte is capacitation. Although we have methods which lead to efficient in vitro fertilization, we still lack understanding about the molecular mechanisms of capacitation. While numerous events occur during capacitation, it appears that regulation of intracellular Ca2+ (Ca(i)) is one of the most important. We found that the influx of Ca2+ into sperm during the first 2 hours of incubation is critical to heparin-induced capacitation. This is a period during capacitation when Ca(i) has not yet increased. We propose that during capacitation, the initial influx of Ca2+ into sperm is used to fill an intracellular Ca2+ store located in the acrosome. We found that thapsigargin, an inhibitor of an acrosomal Ca2+-ATPase, can stimulate capacitated sperm to acrosome react, trigger the opening of a store-operated calcium channel in the plasma membrane and has greater effects on capacitated sperm compared to noncapacitated sperm. An increase in intracellular Ca2+ was also detected in the anterior sperm head during capacitation, suggesting the loading of the acrosome with Ca2+. These observations may be important in the development of new methods for capacitation and understanding the death of sperm after cryopreservation.