Characterization of the ATP-sensitive potassium channels (K-ATP) expressedin guinea pig bladder smooth muscle cells

ATP-sensitive K+ (K-ATP) channels play an important role in the regulation of smooth muscle membrane potential. To investigate the properties of K-ATP channels in guinea pig urinary bladder smooth muscle cells, fluorescence-b ased assays were carried out with the membrane potential-sensitive probe bi s-(1,3-dibutylbarbituric acid)trimethine oxonol [DiBAC(4)(3)]. The prototyp ical channel openers, including pinacidil, (-)-cromakalim, and diazoxide, e licited concentration-dependent decreases in membrane potential that were a ttenuated by glyburide. Similar responses were evoked by a reduction in int racellular ATP levels by metabolic inhibition. The observed rank order pote ncy (EC50) for evoking membrane potential changes by potassium channel open ers, P1075 (53 nM) similar to Bay X 9228 > (-)-cromakalim similar to ZD6169 similar to pinacidil > Bay X 9227 similar to ZM244085 > diazoxide (59 mu M ), showed a good correlation with that of bladder smooth muscle relaxation, as assessed by isolated tissue bath studies. The maximal efficacies of (-) -cromakalim, pinacidil, Bay X 9228, and ZD6169 were comparable with the res ponse achieved by the reference activator P1075. Whole cell currents in bla dder smooth muscle cells were increased in both inward and outward directio ns by P1075 and were reversed by glyburide to control levels. The molecular composition assessed by reverse transcriptase-polymerase chain reaction an alysis using subunit-specific primers revealed the presence of mRNA for inw ard rectifying potassium channel (K(IR)6.2) and sulfonylurea receptors (SUR )2B and SUR1. The subunit profile together with pharmacological properties suggests that the K-ATP channel in bladder smooth muscle cells could be com posed of SUR2B associated with a single inward rectifier, K(IR)6.2. In summ ary, these studies have characterized the pharmacological profile using flu orescent imaging plate reader-based membrane potential techniques and provi de evidence for the molecular identity of K-ATP channels expressed in guine a pig bladder smooth muscle cells.