Expression and differential cell distribution of low-threshold Ca 2+ channels in mammalian male germ cells and sperm

Abstract

Numerous sperm functions including the acrosome reaction (AR) are associated with Ca 2+ influx through voltage-gated Ca 2+ (Ca V) channels. Although the electrophysiological characterization of Ca 2+ currents in mature sperm has proven difficult, functional studies have revealed the presence of low-threshold (Ca V3) channels in spermatogenic cells. However, the molecular identity of these proteins remains undefined. Here, we identified by reverse transcription polymerase chain reaction the expression of Ca V3.3 mRNA in mouse male germ cells, an isoform not previously described in these cells. Immunoconfocal microscopy revealed the presence of the three Ca V3 channel isoforms in mouse spermatogenic cells. In mature mouse sperm only Ca V3.1 and Ca V3.2 were detected in the head, suggesting its participation in the AR. Ca V3.1 and Ca V3.3 were found in the principal and the midpiece of the flagella. All Ca V3 channels are also present in human sperm, but only to a minor extent in the head. These findings were corroborated by immunogold transmission electron microscopy. Tail localization of Ca V3 channels suggested they may participate in motility, however, mibefradil and gossypol concentrations that inhibit Ca V3 channels did not significantly affect human sperm motility. Only higher mibefradil doses that can block high-threshold (HVA) Ca V channels caused small but significant motility alterations. Antibodies to HVA channels detected Ca V1.3 and Ca V2.3 in human sperm flagella.