Studies of inwardly rectifying K^+ channels and an approach to develop new methods for assaying K^+ channel activity.

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Inwardly rectifying K^+ channels (Kir channels) consist of more than 15 members and they are highy selective for K^+. They have, been estimated to possess two transmembrane domains and one pore-forming (H5) region. Amino acid sequence of H5 region of Kir channels has high similarity to each other and contains K^+ channel signature sequence of Gly-Tyr(Phe)-Gly. Each Kir channel consists of four subunits. Functions of Kir channels have been considered stabilizing and decreasing membrane potential, inhibiting excitability of cell, transport of K^+, and secretion of insulin. To evaluate Kir channel activity, patch-clamp method and measurement of influx of radioactive Rb have been used. But these methods have some defects. We planed to develop a new method to assay Kir channels, and at the same time, we planed to further study expression, distribution and function of Kir channels. Furthermore, we started to determine the chromosomal localization of some Kir channels to know the interaction of Kir channels and diseases. Our achievements are as follows. (1) Kir4.l/K_<AB>-2is expressed in glial cells and gastric parietal cells, the function of which is transport of K4. (2) new sulfonylurea receptor subunit (SUR2B) was cloned and identified as smooth vascular K_<ATP> channel subunit. (3) Kir3.2b, Kir1.lf, and cTBAK-1 were cloned. The chromosomal localization of Kir4.1/K_<Ab>-2, Kir5.1, Kir6.1, SUR2 and cTBAK-1 were identified. (4) We tried to develop measuring K^+ channel activity using PBFI.