Suppression of two cloned smooth muscle-derived delayed rectifier potassium channels by cholinergic agonists and phorbol esters.
Functional coupling between muscarinic (m3) receptors and two voltage-gated K+ (Kv) channels (Kv1.2 and Kv1.5) cloned originally from canine colonic smooth muscle was studied using the Xenopus oocytes expression system and a mammalian cell line (COS cells). Oocytes were coinjected with cRNAs encoding the human m3 receptor and the Kv channel clones. COS cells were stably transfected with the hm3 cDNA and the cDNA encoding Kv1.5 channels. In oocytes coexpressing hm3 receptors and Kv channels, acetylcholine (ACh, 100 microM) decreased the whole-oocyte Kv channel current (IKv) by 72% over 20 min. ACh was equally effective at suppressing IKv1.2 as IKv1.5. In oocytes expressing only Kv channels phorbol esters (phorboldibutyrate) and phorbol dideconoate (10-30 nM) mimicked the action of ACh on IKv in oocytes coexpressing hm3 receptors. At the single-channel level, both ACh and phorbol dibutyrate applied to the extra-patch membrane reduced the open probability of Kv channels in the cell-attached patches without affecting single-channel conductance. In cotransfected COS cells, over a similar time course as in oocytes ACh suppressed whole-cell IKv1.5, but only by 30% and the effect was not reversible. These data indicate that stimulation of m3 receptors in cells that express Kv1.2 and Kv1.5 channels causes a poorly reversible decrease in the open probability of these channels.
Vogalis
F
F
Department of Physiology, University of Nevada, School of Medicine, Reno 89557-0046, USA.
Ward
M
M
Horowitz
B
B
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D018679:Cholinergic Agonists / Q000494:pharmacology*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004285:Dogs; D004789:Enzyme Activation; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism / Q000502:physiology; D009130:Muscle, Smooth / Q000737:chemistry* / Q000187:drug effects / Q000502:physiology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology*; D010703:Phorbol Esters / Q000494:pharmacology*; D015221:Potassium Channels / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D011493:Protein Kinase C / Q000378:metabolism / Q000502:physiology; D011976:Receptors, Muscarinic / Q000235:genetics / Q000502:physiology; D015290:Second Messenger Systems / Q000187:drug effects / Q000502:physiology; D014162:Transfection; D014982:Xenopus laevis
D018679:Cholinergic Agonists; D018076:DNA, Complementary; D010703:Phorbol Esters; D015221:Potassium Channels; D011976:Receptors, Muscarinic; C008876:phorbol-12,13-didecanoate; D015240:Phorbol 12,13-Dibutyrate; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D000109:Acetylcholine; D002118:Calcium