The long term goal of this research is to understand the molecular basis of cardiac excitability. In this proposal the investigators will study molecular determinants of voltage-gated K+ channel block by antiarrhythmic drugs and local anesthetics. Voltage-gated K+ channels play a crucial role in controlling cardiac excitability and repolarization and are the molecular target for many new antiarrhythmic agents. The investigators will utilize cloned subunits that are considered to contribute to native cardiac currents (Kv1.5, Kv4.2, HERG, KvLQT1). Increasing evidence indicates that the molecular architecture of the channel protein complex includes function-altering accessory subunits (beta subunits, minK) which may impact on drug binding. The hypotheses to be tested include that specific pore residues in the S6 segment are involved in binding of open channel blocking drugs. Furthermore, the investigators will test whether hydrophobic interactions determine affinity and stereoselectivity of drug block. In addition, they will test whether amino acid differences in the pore lining segments explain isoform-specific affinities for antiarrhythmic drugs. They will address whether antiarrhythmic drugs utilize a conserved receptor site for inactivating beta-subunits by testing for mutual interactions in the inner mouth of the pore. Finally, they will test whether the intrinsic pharmacology of KcLQT1 is altered when it co-assembles with the minK subunit to reconstitute the native current Iks. In these studies they will use contemporary techniques of molecular biology to modify channel structure and a variety of patch clamp techniques to test for specific functional changes. The results will be interpreted in terms of mathematical and thermodynamic models for channel gating and drug action. These studies will address molecular determinants of drug block and drug interactions with activation and inactivation gates . The information gained from this project will expand our knowledge of the molecular pharmacology of these important channels, which may ultimately result in improved understanding of mechanisms of arrhythmias and the development of better antiarrhythmic treatments.
