We have previously reported that cromakalim and U-89232 reduce infarct size in a rabbit model of myocardial ischemia. Because U-89232 appeared to lack activity in the vasculature, we tested its reversibility with glibenclamide. Twenty-eight ketamine-xylazine anesthetized open-chest, New Zealand White rabbits were instrumented for regional coronary occlusion and reperfusion. Study animals received either cromakalim, U-89232 or vehicle. In some animals, glibenclamide was administered. All animals were then subjected to ischemia (30 min) and reperfusion (120 min), and necrosis was determined using tetrazolium. With comparable hemodynamics and myocardium at risk, infarct size in control animals was 35.5 +/- 4.6% of risk region, and was not different from glibenclamide-treated animals (37.7 +/- 5.8%). Cromakalim alone has been shown to be protective, however when combined with glibenclamide necrosis amounted to 35.1 +/- 3.8% of the risk region (p = NS vs. control). In contrast, U-89232 was protective in the presence of glibenclamide (17.2 +/- 4.9% of the risk region). We conclude that U-89232 produces myoprotection independent of K-ATP channel inhibition, indicating that this compound possesses novel anti-ischemic characteristics.