Abstract
Activation of the sperm-specific Ca(2+) channel CatSper by progesterone evokes rapid changes in intracellular Ca(2+) in human sperm that are required for fertilization. However, the mechanisms regulating the progesterone-induced Ca(2+) signals have remained elusive. Here, we used quantitative kinetic fluorimetry with fast voltage-sensitive fluorescent indicators to investigate how progesterone affects the membrane potential (V(m)) of human sperm. Additionally, we employed the FAST(M) technique to simultaneously record at millisecond time resolution changes in both V(m) and intracellular Ca(2+). We show that progesterone evokes a rapid pulse-like depolarization and repolarization. The depolarization is caused by Ca(2+) influx through CatSper, which pulls V(m) away from a resting membrane potential (V(rest)) of -65 mV set by the sperm-specific K(+) channel Slo3. We further show that V(m)- and Ca(2+)-dependent mechanisms limit the CatSper-mediated Ca(2+) influx, thereby promoting repolarization and enabling K(+) efflux through Slo3 channels to restore V(rest). Our findings demonstrate that non-genomic progesterone signaling in human sperm is regulated by negative feedback on CatSper and involves a dynamic interplay between CatSper and Slo3 in controlling V(m). We anticipate that our novel kinetic, quantitative V(m) recording and V(m)/Ca(2+)-multiplexing techniques will reveal additional molecular mechanisms underlying CatSper-mediated Ca(2+) signaling in human sperm both in health and disease.