The signaling pathways involved in ischemic heart disease are not well characterized. In this study, the roles of Ras-GTPase, tyrosine kinases (TKs) and Ca2+/calmodulin-dependent protein kinase II (CaMKII) in global ischemia and reperfusion (I/R) in a perfused rat heart model were investigated and compared to beneficial effects produced by preconditioning (PC). A 40 min episode of global ischemia followed by a 30 min reperfusion in perfused rat hearts produced significantly impaired cardiac function, measured as left ventricular developed pressure (Pmax) and left ventricular end-diastolic pressure (LVEDP), and impaired coronary hemodynamics, measured as coronary flow (CF) and coronary vascular resistance (CVR). Hearts from male Wistar rats pre-treated with the tyrosine kinase inhibitor, genistein (1 mg/kg/day for 6 days), or the CaMKII inhibitor, KN-93 (578 ng/min for 6 days), produced detrimental effects on recovery of cardiac function and coronary hemodynamics. In contrast, pre-treatment with Ras-GTPase inhibitor FPT III (232 ng/min for 6 days) significantly enhanced cardiac recovery in terms of left ventricular contractility and coronary vascular hemodynamics. Treatment with FPT III also significantly reduced expression of the sodium-hydrogen exchanger-1 (NHE-1) which was elevated during I/R as detected by Western blotting. These data suggest that TKs and CaMKII are involved in signaling pathways leading to recovery from cardiac ischemia, whereas activation of Ras-GTPase signaling pathways are critical in the development of cardiac dysfunction due to I/R.