Apical SK potassium channels and Ca2+-dependent anion secretion in endometrial epithelial cells.
Apical uridine triphosphate (UTP) stimulation was shown to increase short circuit current (I(sc)) in immortalized porcine endometrial gland epithelial monolayers. Pretreatment with the bee venom toxin apamin enhanced this response. Voltage-clamp experiments using amphotericin B-permeablized monolayers revealed that the apamin-sensitive current increased immediately after UTP stimulation and was K(+) dependent. The current-voltage relationship was slightly inwardly rectifying with a reversal potential of -52 +/- 2 mV, and the P(K)/P(Na) ratio was 14, indicating high selectivity for K(+). Concentration-response relationships for apamin and dequalinium had IC(50) values of 0.5 nm and 1.8 microm, respectively, consistent with data previously reported for SK3 channels in excitable cells and hepatocytes. Treatment of monolayers with 50 microm BAPTA-AM completely blocked the effects of UTP on K(+) channel activation, indicating that the apamin-sensitive current was also Ca(2+) dependent. Moreover, channel activation was blocked by calmidazolium (IC(50) = 5 microm), suggesting a role for calmodulin in Ca(2+)-dependent regulation of channel activity. RT-PCR experiments demonstrated expression of mRNA for the SK1 and SK3 channels, but not SK2 channels. Treatment of monolayers with 20 nm oestradiol-17beta produced a 2-fold increase in SK3 mRNA, a 2-fold decrease in SK1 mRNA, but no change in GAPDH mRNA expression. This result correlated with a 2.5-fold increase in apamin-sensitive K(+) channel activity in the apical membrane. We speculate that SK channels provide a mechanism for rapidly sensing changes in intracellular Ca(2+) near the apical membrane, evoking immediate hyperpolarization necessary for increasing the driving force for anion efflux following P2Y receptor activation.
Department of Physiology, 495 Animal Science/Veterinary Medicine Bldg, University of Minnesota, 1988 Fitch Ave., St Paul, MN 55108, USA.
Melissa L
ML
Palmer
Katherine R
KR
Schiller
Scott M
SM
O'Grady
D000595:Amino Acid Sequence; D000818:Animals; D000838:Anions / Q000378:metabolism*; D001030:Apamin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D004717:Endometrium / Q000166:cytology / Q000378:metabolism*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D004958:Estradiol / Q000494:pharmacology; D004967:Estrogens / Q000494:pharmacology; D005260:Female; D008969:Molecular Sequence Data; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051640:Small-Conductance Calcium-Activated Potassium Channels / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D013552:Swine; D014544:Uridine Triphosphate / Q000494:pharmacology
D000838:Anions; D004967:Estrogens; D012333:RNA, Messenger; D051640:Small-Conductance Calcium-Activated Potassium Channels; D001030:Apamin; D004958:Estradiol; D011188:Potassium; D002118:Calcium; D014544:Uridine Triphosphate