Automated patch clamp analysis of heterologously expressed Kir6.2/SUR1 and Kir6.1/SUR2B K_ATP currents

ATP-sensitive potassium (K_ATP) channels are therapeutic targets for numerous metabolic, cardiovascular, and neurological disorders. Drug development for K_ATP channels requires electrophysiology assays for detailed compound characterization. Parallel automated patch clamp (APC) techniques offer considerable advantages over low-throughput manual patch clamp electrophysiology. Here, we characterized the functional properties and pharmacological sensitivity of heterologously expressed Kir6.2/SUR1 and Kir6.1/SUR2B using a SyncroPatch 384PE APC instrument. Ruptured-membrane and perforated-patch whole cell recordings in potassium fluoride and fluoride-free assay buffers and electrophysiology chips were evaluated for both subtypes. Effects of internal ATP and ADP, and magnesium (Mg²⁺) addition were also assessed. Kir6.2/SUR1 currents were constitutively active in all potassium fluoride-based recordings, insensitive to activation by the SUR1 agonist, VU0071063, and variably inhibited by glibenclamide. Success rates, current rundown, and glibenclamide sensitivity were associated with internal buffer composition. Recordings in fluoride-free buffers revealed a minor population of constitutively active Kir6.2/SUR1 currents and a larger population of currents exhibiting low basal activity and activation by VU0071063. Success rate and stability were associated with internal buffer composition. Kir6.1/SUR2B currents, which were most readily assayed in ruptured-membrane and potassium fluoride-based conditions, were stable, activatable with pinacidil, and inhibited by glibenclamide. Our study sheds new light on the behavior of Kir6.2/SUR1 and Kir6.1/SUR2B currents under available APC conditions and represents an important step toward developing truly high-throughput APC techniques for K_ATP.NEW & NOTEWORTHY Highly parallel automated patch clamp (APC) methods have revolutionized the way electrophysiology is performed in the pharmaceutical and biotechnology industries and increasingly in academic laboratories. Here, we characterized the functional and pharmacological properties of heterologously expressed Kir6.2/SUR1 and Kir6.1/SUR2B using a SyncroPatch 384PE APC instrument. The results of our studies highlight heretofore unappreciated effects of fluoride-base internal solutions on Kir6.2/SUR1 and provide foundational support for developing truly high-throughput electrophysiology methods for both drug targets.