In order to investigate the large-conductance Ca²⁺-activated K⁺ (BK_Ca) channel and determine the effects of nitric oxide (NO) on the channel in human skin fibroblasts, we performed electrophysiological patch clamp recordings on 5th-passage cells of human genital skin cultures. The whole-cell outward K⁺ current was increased with depolarization, and proved to be sensitive to NS1619 (a selective BK_Ca channel activator) and iberiotoxin (a specific BK_Ca channel inhibitor). The single-channel currents showed 226 pS of mean conductance in symmetrical K⁺. Sodium nitroprusside (SNP; an NO donor) significantly increased the K⁺ current amplitude in the whole-cell mode, and open probability of the channel (NPo) in the cell-attached mode, but not in the inside-out mode. S-nitroso-N-acetylpenicillamine (an NO donor) and 8-Br-cGMP (a membrane-permeant cGMP analogue) also increased the BK_Ca channel activity. The stimulatory effect of SNP on BK_Ca channels was inhibited by pretreatment with 1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-1-one (a soluble guanylyl cyclase inhibitor), or KT5823 [a specific protein kinase G (PKG) inhibitor]. Cytoplasmic PKG also increased the channel activity in inside-out patches. In conclusion, the present data indicate that BK_Ca channels constitute a significant fraction of K⁺ current in human skin fibroblasts, and that NO increases NPo of BK_Ca channels, which are mediated via the cGMP/PKG pathway, without direct effects on the channel.

porc / maturation des ovocytes / protéine kinase C / protéine kinase AMP! dépendante / calmoduline