It has been proposed that ischemic preconditioning involves the regulation of ATP-sensitive potassium (K ATP ) channels. The evidence is based largely on the ability of certain K ATP channel modulators to modify the protection in the various models of preconditioning. This study has investigated how two K ATP channel openers, pinacidil and nicorandil, affect both membrane currents and viability in isolated and ischemic rabbit cardiomyocytes. We used the whole-cell recording technique and in separate experiments viability was assessed by exposure to these drugs during ischemia. Pinacidil (50μmol/l) increased K ATP current approximately four-fold in isolated cardiomyocytes. This increase reversed rapidly after treatment with the K ATP channel blocker glibenclamide (200 nmol/l). After simulated ischemia, pinacidil protected cardiomyocytes (the area under cell-death curve was 29.5±1.1% · h) which was significantly less than that in control (46.9±2.0% · h). The protection from pinacidil could be completely eliminated by pretreatment with 10μmglibenclamide (46.9±2.0% · h). In contrast, nicorandil (1 mmol/l), which opens K ATP channels in some tissues, caused no detectable effect on the K ATP current. Similarly, nicorandil did not produce cardioprotection. These results indicate that pinacidil and nicorandil have very different effects on rabbit cardiomyocyte K ATP channels. Furthermore, because protection correlated with the ability of the agent to open the channel, they support a role for K ATP channels in preconditioning.