Intracellular calcium store depletion and acrosome reaction in human spermatozoa: role of calcium and plasma membrane potential.
ABSTRACT We evaluated the presence and role of internal calcium stores in human uncapacitated spermatozoa by determining the effects of two inhibitors of Ca2+ ATPase 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 conjugate of thapsigargin, we localized internal Ca2+ stores on the acrosome, post-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 conditions 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 sperm plasma membrane, thus showing the existence of "capacitative" Ca2+ entry into these specialized cells. The addition of thapsigargin to human spematozoa induced a dose-dependent increase in acrosome reaction percentages, but only when Ca2+ was present in the external medium. Plasma membrane potential monitoring showed that these inhibitors induced a depolarization dependent on Ca2+ influx from external medium and that this was preceded by a transient hyperpolarization caused by activation of Ca2+-dependent K+ channels. When K+-dependent plasma membrane hyperpolarization was inhibited, the thapsigargin- and CPA-stimulated rise in [Ca2+](i) plasma membrane depolarization and acrosome reaction were abolished. In conclusion, the present study demonstrates that human spermatozoa possess internal Ca2+ stores and that the capacitative Ca2+ entry pathway present in these cells regulates important biological processes that are fundamental for the acrosome reaction.
Article: Estradiol inhibits the effects of extracellular ATP in human sperm by a non genomic mechanism of action.[show abstract] [hide abstract]
ABSTRACT: Steroid hormones, beside their classical genomic mechanism of action, exert rapid, non genomic effects in different cell types. These effects are mediated by still poorly characterized plasma membrane receptors that appear to be distinct from the classic intracellular receptors. In the present study we evaluated the non genomic effects of estradiol (17betaE(2)) in human sperm and its effects on sperm stimulation by extracellular ATP, a potent activator of sperm acrosome reaction. In human sperm 17betaE(2) induced a rapid increase of intracellular calcium (Ca(2+)) concentrations dependent on an influx of Ca(2+) from the extracellular medium. The monitoring of the plasma membrane potential variations induced by 17betaE(2) showed that this steroid induces a rapid plasma membrane hyperpolarization that was dependent on the presence of Ca(2+) in the extracellular medium since it was absent in Ca(2+) free-medium. When sperm were pre-incubated in the presence of the K(+) channel inhibitor tetra-ethylammonium, the 17betaE(2) induced plasma membrane hyperpolarization was blunted suggesting the involvement of K(+) channels in the hyperpolarizing effects of 17betaE(2). Extracellular ATP induced a rapid plasma membrane depolarization followed by acrosome reaction. Sperm pre-incubation with 17betaE(2) inhibited the effects of extracellular ATP on sperm plasma membrane potential variations and acrosome reaction. The effects of 17betaE(2) were specific since its inactive steroisomer 17alphaE(2) was inactive. Furthermore the effects of 17betaE(2) were not inhibited by tamoxifen, an antagonist of the classic 17betaE(2) intracellular receptor.Purinergic Signalling 01/2006; 1(4):369-75. · 3.16 Impact Factor