Reconstituted Human Cardiac K _ATP Channels

Abstract

Abstract-ATP-sensitive potassium (K ATP ) channels in striated myocytes are heteromultimers of K IR 6.2, a weak potassium inward rectifier, plus SUR2A, a low-affinity sulfonylurea receptor. We have cloned human K IR 6.2 (huK IR 6.2) and a huSUR2A that corresponds to the major, full-length splice variant identified by polymerase chain reaction analysis of human cardiac poly A ϩ mRNA. ATP-and glibenclamide-sensitive K ϩ channels were produced when both subunits were coexpressed in COSm6 and Chinese hamster ovary cells lacking endogenous K ATP channels, but not when huSUR2A or huK IR 6.2 were transfected alone. Recombinant channels activated by metabolic inhibition in cell-attached configuration or in inside-out patches with ATP-free internal solution were compared with sarcolemmal K ATP channels in human ventricular cells. The single-channel conductance of Ϸ80 pS measured at Ϫ40 mV in quasi-symmetrical Ϸ150 mmol/L K ϩ solutions, the intraburst kinetics that were dependent on K ϩ driving force, and the weak inward rectification were indistinguishable for both channels. Similar to the native channels, huSUR2A/huK IR 6.2 recombinant channels were inhibited by ATP at quasi-physiological free Mg 2ϩ (Ϸ0.7 mmol/L) or in the absence of Mg 2ϩ , with an apparent IC 50 of Ϸ20 mol/L and a pseudo-Hill coefficient of Ϸ1. They were "refreshed" by MgATP and stimulated by ADP in the presence of Mg 2ϩ when inhibited by ATP. The huSUR2A/huK IR 6.2 channels were stimulated by cromakalim and pinacidil in the presence of ATP and Mg 2ϩ but were insensitive to diazoxide. The results suggest that reconstituted huSUR2A/huK IR 6.2 channels represent K ATP channels in sarcolemma of human cardiomyocytes and are an adequate experimental model with which to examine structure-function relationships, molecular physiology, and pharmacology of these channels from human heart. (Circ Res. 1998;83:1132-1143.)

A Conserved Inhibitory and Differential Stimulatory Action of Nucleotides on KIR6.0/SUR Complexes Is Essential for Excitation-Metabolism Coupling by KATP Channels

“…Electrophysiology and Single-channel Kinetic Analysis-Patchclamp recordings were done as described previously (23,24,28 (Fig. 1).…”

The ATP-sensitive potassium (K ATP ) channel-encoded dSUR gene is required for Drosophila heart function and is regulated by tinman

“…P was calculated by a 2 ϫ 2 2 test. (B and C) Electrical pacing-induced heart failure in flies treated with drugs that affect the mammalian sulfonylurea receptors (46). (B) One-week-old yw flies fed with SUR blocker tolbutamide showed a higher heart failure rate than DMSO control-treated yw flies (45%, n ϭ 114; 25%, n ϭ 100, respectively; P Ͻ 0.01).…”

Identification of the Potassium Channel Opener Site on Sulfonylurea Receptors

“…E-mail: m.schwanstecher@tu-bs.de. 1 The abbreviations used are: KCO, potassium channel opener; K ATP , ATP-sensitive K ϩ channel; KCO I and KCO II, potassium channel opener binding regions; K IR , inwardly rectifying K ϩ channel; NBF, nucleotide binding fold; SU, sulfonylurea; SUBR, sulfonylurea binding region; SUR, sulfonylurea receptor; TMD, transmembrane domain; TMDI or TMDII, first (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11) or second (12)(13)(14)(15)(16)(17) set of transmembrane domains (see Fig. 1, A or F); DMEM, Dulbecco's modified Eagle's medium.…”

“…SUR1/ K IR 6.2 have been proposed to reconstitute the neuronal/pancreatic ␤-cell (5), SUR2A/K IR 6.2, the cardiac (6,13,14), and SUR2B/K IR 6.1 (or K IR 6.2), the vascular smooth muscle-type K ATP channels (8,11,15,16).…”