Opioids and volatile anesthetics produce marked cardioprotective effects against myocardial infarction via the activation of ATP-sensitive potassium (K(ATP)) channels, however, the effect of combined treatment with both drugs is unknown. We examined the hypothesis that opioids and volatile anesthetics potentiate cardiac K(ATP) channel opening, thereby enhancing cardioprotection. Rats were treated with the delta opioid agonists, TAN-67 or BW373U86, or isoflurane, together or alone with and without diazoxide, a mitochondrial K(ATP) channel opener. Glibenclamide, a non-selective K(ATP) channel blocker, was used to further characterize the signaling mechanism involved. Myocardial infarct size (IS) was determined by tetrazolium staining and was expressed as a percent of the area at risk (AAR). High doses of TAN-67 (10 mg/kg), diazoxide (10 mg/kg), and isoflurane (1 MAC) produced a significant reduction in IS compared with the control group (30+/-3%, 36+/-5%, and 42+/-2 vs. 58+/-2%, respectively), whereas lower doses of the drugs had no effect except for the low dose of isoflurane (0.5 MAC). The combination of TAN-67 and diazoxide or isoflurane and diazoxide resulted in a marked reduction in IS compared with controls in the presence of high (9+/-3% and 14+/-3%) and low (17+/-7% and 31+/-7%) dose combinations, respectively. The combination of TAN-67 or BW373U86 and isoflurane also caused a striking reduction in IS/AAR (16+/-7% and 7+/-2%, respectively). To date, this is the first demonstration that opioids and volatile anesthetics work in conjunction to confer protection against myocardial infarction through potentiation of cardiac K(ATP) channel opening.