Lim I, Yun J, Kim S, Lee C, Seo S, Kim T, Bang H
Department of Physiology, College of Medicine, Chung-Ang University, Seoul, Korea.
Skin Pharmacol Physiol. 2005 Nov-Dec;18(6):279-87. doi: 10.1159/000088013. Epub 2005 Sep 5.
In order to investigate the large-conductance Ca(2+)-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.
为了研究大电导钙激活钾（BK(Ca)）通道，并确定一氧化氮（NO）对人皮肤成纤维细胞中该通道的影响，我们对人生殖器皮肤培养物的第5代细胞进行了电生理膜片钳记录。全细胞外向钾电流随去极化增加，并被证明对NS1619（一种选择性BK(Ca)通道激活剂）和iberiotoxin（一种特异性BK(Ca)通道抑制剂）敏感。单通道电流在对称钾溶液中显示平均电导为226 pS。硝普钠（SNP；一种NO供体）在全细胞模式下显著增加钾电流幅度，在细胞贴附模式下增加通道开放概率（NPo），但在向外膜片模式下无此作用。S-亚硝基-N-乙酰青霉胺（一种NO供体）和8-溴-cGMP（一种可透过膜的cGMP类似物）也增加BK(Ca)通道活性。SNP对BK(Ca)通道的刺激作用被用1H-[1,2,4]-恶二唑并[4,3-a]喹喔啉-1-酮（一种可溶性鸟苷酸环化酶抑制剂）或KT5823[一种特异性蛋白激酶G（PKG）抑制剂]预处理所抑制。细胞质PKG也增加向外膜片的通道活性。总之，目前的数据表明BK(Ca)通道构成人皮肤成纤维细胞中钾电流的很大一部分，并且NO增加BK(Ca)通道的NPo，这是通过cGMP/PKG途径介导的，而对通道无直接影响。
