ATP-sensitive K⁺ channels (K_ATP channels) of arterial smooth muscle are critical regulators of arterial tone, and so blood flow, in response to vasoactive transmitters [1]. Recent biochemical evidence suggests that these channels localise to cholesterol and sphingolipid-enriched invaginations of the smooth muscle surface membrane termed caveolae [2]. These specialised lipid microdomains are thought to assist in the spatial organisation of signal transduction pathways by aggregating interacting proteins into functional signalling compartments [3]. Here we investigate the potential role of the caveolae-associated scaffold protein caveolin in regulating K_ATP channel activity. Immunogold electron microscopy of rat aortic smooth muscle plasma membrane sheets confirmed the presence of Kir6.1, the pore-forming inwardly rectifying subunit of the vascular K_ATP channel, in morphologically identifiable caveolin-rich regions of the membrane. Antibodies directed against vascular K_ATP channel subunits (Kir6.1 and sulphonylurea receptor (SUR2B)) co-immunoprecipitated caveolin-1 from rat aortic homogenates, suggesting that caveolin-1 interacts with the channel protein. To investigate whether this interaction has any functional effect on K_ATP channel activity pinacidil-evoked recombinant whole-cell K_ATP currents were recorded in HEK293 cells. Whole-cell K_ATP currents recorded at -60 mV in HEK293 cells stably expressing caveolin-1 were significantly smaller than currents recorded in wild-type HEK293 where caveolin is absent (69.6 ± 8.3 pA/pF, n=8; 179.7 ± 35.9 pA/pF, n=6, respectively; mean ± SEM; p<0.05, Student’s t test). In cell-attached patch clamp recordings, the presence of caveolin-1 had no significant effect upon pinacidil-evoked single K_ATP channel chord conductance (39.4 ± 2.5 pS, n=5; 30.6 ± 4.2 pS, n=5, respectively; membrane potential -60 mV; p>0.05, Student’s t test). However, analysis of open and closed time distributions revealed that in the presence of caveolin-1 the channel spends significantly more time in longer-lived closed states (p<0.05, Student’s t test). Together these findings suggest that interaction with caveolin-1 has an inhibitory effect on vascular K_ATP channel activity that may be important in the physiological control of channel function.