Objective: Prolonged donor heart preservation is important in cardiovascular surgery. This study examined the effect of pinacidil, a nonselective adenosine triphosphate-sensitive potassium channel opener, on donor heart preservation when added to histidine tryptophan ketoglutarate solution and the role of mitochondria in this protection. Methods: Sprague-Dawley rat hearts received one of 5 preservation solutions in the Langendorff perfusion apparatus (24 per group): (1) histidine tryptophan ketoglutarate solution; (2) histidine tryptophan ketoglutarate solution containing pinacidil; (3) histidine tryptophan ketoglutarate solution containing pinacidil and 5-hydroxydecanote, a mitochondrial adenosine triphosphate-sensitive potassium channel blocker; (4) histidine tryptophan ketoglutarate solution containing pinacidil and Hoechst-Marion-Roussel 1098, a sarcolemmal adenosine triphosphate-sensitive potassium channel blocker; and (5) histidine tryptophan ketoglutarate solution containing pinacidil, 5-hydroxydecanote, and Hoechst-Marion-Roussel 1098. After a 10-minute equilibration period, all the hearts in the different preservation solutions were placed in cold storage for 8 hours, followed by 60 minutes of reperfusion. Hemodynamics, mitochondrial respiratory function, adenosine triphosphate level, cardiac troponin I release, and ultrastructure were examined. Results: Histidine tryptophan ketoglutarate solution containing 0.5 mmol/L pinicidal significantly improved heart function, coronary flow, myocardial ultrastructure, and cardiac troponin I release after reperfusion (P < .01 or P < .05). In the pinacidil group at the end of storage and the end of reperfusion, mitochondrial respiratory function and myocardial adenosine triphosphate levels were superior when compared with other groups (P < .01 or P < .05). These beneficial effects of pinacidil were blocked by 100 μmol/L 5-hydroxydecanote. Conclusion: Histidine tryptophan ketoglutarate solution containing pinacidil provides better cardioprotection with preservation of mitochondrial energy. This effect of pinacidil appears to depend on both mitochondrial and sarcolemmal adenosine triphosphate sensitive potassium channel. © 2010 The American Association for Thoracic Surgery.