ATP-sensitive K + (K A T P ) channels couple metabolic changes to membrane excitability in vascular smooth muscle cells (SMCs). While the electrophysiological properties of K A T P channels have been examined, little is known about the molecular basis of K A T P complex in vascular SMCs. We identified and cloned four K A T P subunit genes from rat mesenteric artery, namely rvKir6.1, rvKir6.2, rvKirSUR1, and rvSUR2B. These clones showed over 99.6% amino acid sequence identity with other previously reported isoforms. The mRNA expression patterns of the K A T P subunits varied among rat aorta, mesenteric artery, pulmonary artery, tail artery, hepatic artery, and portal vein. Heterologous co-expression of rvKir6.1 and rvSUR2B yielded functional K A T P channels that were inhibited by glibenclamide, and opened by pinacidil. Our results for the first time reported the expression of four K A T P subunits in same vascular tissues, unmasking the diversity of native K A T P channels in vascular SMCs.