This study investigated the effects of long-term treatment with glibenclamide (GLIB) on the susceptibility of streptozotocin (STZ)-induced diabetic heart to ischemia/reperfusion insults. Starting 4 weeks after the injection of STZ, rats were treated with GLIB (0.1 mg/kg, ip, three times a week, STZ-GLIB group) or vehicle (STZ-VEH group) for 8 weeks. The recovery of cardiac performance, released creatine kinase (CK), and incidence of ventricular arrhythmias were studied during the reperfusion period in isolated hearts from rats in STZ-GLIB (n = 14) and in STZ-VEH groups (n = 13) and from age-matched control rats (CNT group, n = 14). Each heart was subjected to 5 min of global low-flow ischemia followed by 25 min of no-flow ischemia, with a subsequent 30 min of reperfusion. Plasma glucose level was not significantly different between the STZ-GLIB and STZ-VEH groups. The recovery of cardiac performance and the released CK during reperfusion period were significantly lower in both STZ-VEH and STZ-GLIB groups than in the CNT group (P < 0.01 and P < 0.05, respectively). Reperfusion resulted in an incidence of ventricular fibrillation in 23% and 21% in STZ-VEH and STZ-GLIB groups, respectively (P = ns). These values were significantly lower than that of the CNT group (100%, P < 0.001 for both). More importantly, the incidence of ventricular tachycardia in the STZ-GLIB group was significantly higher than that in the STZ-VEH group (93% vs 54%, P < 0.05) and was not significantly different from that in the CNT group (93% vs 100%, P = ns). The results suggest that STZ-induced protection against reperfusion-induced ventricular arrhythmias in diabetic heart may be partially abrogated by long-term treatment with GLIB.