Heat stress (HS) induced-cardioprotection is associated with activation of focal adhesion kinase (FAK) and protein kinase B (Akt) in neonatal rat ventricular myocytes (NRVM) suggesting that stress-induced activation of survival pathways may be important in protecting intact hearts from irreversible injury. Aims: The purposes of this study were: 1) to examine the subcellular signaling pathways activated by heat stress (HS) and ischemic preconditioning (IP) in intact hearts; 2) to determine if HS and IP activate an integrated survival pathway similar to that activated by HS in cultured NRVM; and 3) to determine if HS and IP reduce lethal cell injury in perfused intact hearts. Methods: Adult rat hearts perfused in the Langendorff mode were subjected to 25 minutes of global ischemia and 30 minutes of reperfusion (IR) either 24 hours after whole animal heat shock (HS) or following a standard IP protocol. Myocardial signaling was analyzed using Western blot techniques while cell death was assayed by measuring lactate dehydrogenase (LDH) release into the perfusate and confirmed by light microscopy. Results: Similar to NRVM, HS performed in the whole animal 24 hours prior to IR increased phosphorylation of focal adhesion kinase (FAK) at tyrosine 397 and protein kinase B (Akt) and resulted in protection from cell death. Using IP as a myocardial stress also resulted in increased phosphorylation/activation of both FAK and Akt and resulted in reduced cell death in adult perfused rat hearts subjected to IR. Conclusions: 1) Myocardial stress caused by whole animal HS activates cytoskeletal-based survival signaling pathways in whole heart tissue and reduces lethal IR injury; 2) IP activates the same stress-induced survival pathway and the activation correlates with the well known cardioprotective effect of IP on lethal IR injury.