The objective of this proposal is to pursue studies on the genesis of the resting membrane potential in visceral smooth muscle. The resting potential is one of the major determinants of smooth muscle excitability and is largely governed by the permeability to K+ ions. Preliminary studies demonstrate that the resting potential of several gastrointestinal smooth muscle is, in part, controlled by a unique human ether-a-go-go (HERG)-like K+ channel. This K+ current demonstrates inward rectification and appears to be a potential target of the commonly used prokinetic agent, cisapride. The first specific aim is to characterize the detailed biophysical properties of the HERG-like K+ current in gastrointestinal smooth muscle cells. The voltage- dependent kinetics of this current in single cells will be determined using the patch clamp technique and its physiological role in resting potential and repolarization of the action potential will be characterized. The second specific aim is to identify the pharmacological regulation of the smooth muscle HERG-like K+ currents. in these studies, the effects of class III antiarrhythmic compounds which are known HERG channel blockers will be determined, and the mechanism of action of the prokinetic agent, cisapride on the HERG currents will be investigated. Preliminary data show that cisapride attenuates the HERG currents in single gastrointestinal smooth muscle cells and depolarizes muscle strips. The effects of the excitatory neurotransmitter, acetylcholine on these K currents will be determined and the involvement of the second messenger regulation by protein kinase C will be evaluated. In the third specific aim, the protein expression of HERG-like channels will be localized by immunofluorescence techniques and by Western blotting. The fourth specific aim is to define the relationship of the HERG-like conductance with those of the other inwardly rectifying currents in smooth muscle. In these studies the role of IKi, ATP- sensitive K+ channel and the hyperpolarization-activated cation currents will be examined. Inhibitory modulators of the HERG-ike K+ channels in smooth muscle may provide new approaches to treatment of disorders that involve smooth muscle hyperexcitability.
