The potential contribution of oxidative stress to cardioprotection in infants induced by adaptation to chronic hypoxia and by ischemic preconditioning is poorly understood. Under conditions of oxidative stress, reactive oxygen species and reactive nitrogen species may contribute to phenotypic changes in hearts adapted to chronic hypoxia and to the pathogenesis of myocardial injury during both ischemia/reperfusion and hypoxia/reoxygenation. Hearts from infant rabbits normoxic from birth can be preconditioned by brief periods of ischemia. In contrast, hearts from infant rabbits adapted to hypoxia from birth appear resistant to ischemic preconditioning. Chronically hypoxic infant rabbit hearts are already resistant to ischemia compared with age-matched normoxic controls, and thus additional cardioprotection by ischemic preconditioning may not be possible. Endothelial nitric oxide synthase (NOS3) protein and its product nitric oxide are increased, but not NOS3 message, in chronically hypoxic infant hearts to protect against ischemia. Chronic hypoxia from birth also increases cardioprotection of infant hearts by increasing association of heat shock protein 90 with NOS3. Normoxic infant hearts also generate more superoxide by an N(omega)-nitro-L-arginine methyl ester-inhibitable mechanism than chronically hypoxic hearts. Thus, NOS3 appears to be critically important in adaptation of infant hearts to chronic hypoxia and in resistance to subsequent ischemia by regulating the production of reactive oxygen and nitrogen species.