Intracellular calcium store depletion and acrosome reaction in human spermatozoa: role of calcium and plasma membrane potential

We evaluated the presence and role of internal calcium stores in human unca pacitated spermatozoa by determining the effects of two inhibitors of Ca2ATPase of the sarco-endoplasmic reticulum (SERCA-ATPase), thapsigargin and cyclopiazonic acid (CPA) on intracellular calcium concentrations, [Ca2+](i) , plasma membrane potential and acrosome reaction. Using a fluorescent conj ugate of thapsigargin, we localized internal Ca2+ stores on the acrosome, p ost-acrosomal region and sperm midpiece, SERCA-ATPase inhibitors induced a rise in [Ca2+](i) both in Ca2+ and Ca2+-free media but under these latter c onditions it was reduced with a progressive decline to baseline values; the re-addition of Ca2+-stimulated a rise in [Ca2+](i), This demonstrated that internal Ca2+ store depletion can evoke the opening of Ca2+-channels on sp erm plasma membrane, thus showing the existence of 'capacitative' Ca2+ entr y into these specialized cells. The addition of thapsigargin to human spema tozoa induced a dose-dependent increase in acrosome reaction percentages, b ut only when Ca2+ was present in the external medium. Plasma membrane poten tial monitoring showed that these inhibitors induced a depolarization depen dent on Ca2+ influx from external medium and that this was preceded by a tr ansient hyperpolarization caused by activation of Ca2+-dependent K+ channel s. When K+-dependent plasma membrane hyperpolarization was inhibited, the t hapsigargin- and CPA-stimulated rise in [Ca2+](i) plasma membrane depolariz ation and acrosome reaction were abolished, In conclusion, the present stud y demonstrates that human spermatozoa possess internal Ca2+ stores and that the capacitative Ca2+ entry pathway present in these cells regulates impor tant biological processes that are fundamental for the acrosome reaction.