KATP channels in mouse spermatogenic cells and sperm, and their role in capacitation

Abstract

Mammalian sperm must undergo a series of physiological changes after leaving the testis to become competent for fertilization. These changes, collectively known as capacitation, occur in the female reproductive tract where the sperm plasma membrane is modified in terms of its components and ionic permeability. Among other events, mouse sperm capacitation leads to an increase in the intracellular Ca(2+) and pH as well as to a hyperpolarization of the membrane potential. It is well known that ion channels play a crucial role in these events, though the molecular identity of the particular channels involved in capacitation is poorly defined. In the present work, we report the identification and potential functional role of K(ATP) channels in mouse spermatogenic cells and sperm. By using whole-cell patch clamp recordings in mouse spermatogenic cells, we found K(+) inwardly rectifying (K(ir)) currents that are sensitive to Ba(2+), glucose and the sulfonylureas (tolbutamide and glibenclamide) that block K(ATP) channels. The presence of these channels was confirmed using inhibitors of the ATP synthesis and K(ATP) channel activators. Furthermore, RT-PCR assays allowed us to detect transcripts for the K(ATP) subunits SUR1, SUR2, K(ir)6.1 and K(ir)6.2 in total RNA from elongated spermatids. In addition, immunoconfocal microscopy revealed the presence of these K(ATP) subunits in mouse spermatogenic cells and sperm. Notably, incubation of sperm with tolbutamide during capacitation abolished hyperpolarization and significantly decreased the percentage of AR in a dose-dependent fashion. Together, our results provide evidence for the presence of K(ATP) channels in mouse spermatogenic cells and sperm and disclose the contribution of these channels to the capacitation-associated hyperpolarization.

Calcium Channels in the Development, Maturation, and Function of Spermatozoa

“…This structure has been detected in spermatozoa from several mammals and in sea urchin (210), and it is now proposed that it functions as a Ca 2ϩ store. In support of this proposal, inositol trisphosphate (IP 3 ) receptors (IP 3 Rs) have been detected in the RNE (261). The second structure found in mammalian sperm is the cytoplasmic droplet ( see FIG.…”

“…IP 3 Rs and RyRs are the major intracellular Ca 2ϩ channels located primarily in ER-type intracellular stores. In addition, two-pore channels (TPCs) were recently described as intracellular Ca 2ϩ channels located in acidic organelles such as lysosomes (199,585,586).…”

“…There are three genes coding for IP 3 Rs with differential tissue expression and distinct properties. The generation of IP 3 together with DAG occurs via stimulation of G proteincoupled receptors (GPCR) or tyrosine kinase-coupled receptors which activate PLCs that in turn cleave PIP 2 to produce IP 3 and DAG.…”

Involvement of Cystic Fibrosis Transmembrane Conductance Regulator in Mouse Sperm Capacitation

“…Studies from our group have demonstrated that inwardly rectifying K ϩ channels contribute to this hyperpolarization (5,6). In addition, we documented that an electrogenic Na ϩ /HCO 3 Ϫ cotransporter hyperpolarizes mouse sperm when external HCO 3 Ϫ is elevated (7).…”

Sperm-Surface ATP in Boar Spermatozoa is Required for Fertilization: Relevance to Sperm Proteasomal Function

“…There are at least three different sources of ATP generation in the spermatozoa [RodriguezMiranda et al 2008], including oxidative phosphorylation in the sperm mitochondria [Halangk et al 1985], glycolysis in the fibrous sheath of the sperm tail principal piece [Krisfalusi et al 2006], and the conversion of ADP to ATP by adenylate kinase [Schoff et al 1989;Foresta et al 1992]. Of particular interest to the present study is the detection of adenylate kinase-dependent KATP-potassium channels in the sperm acrosome [Acevedo et al 2006].…”