Hypoxia is a well-known signal for vessel formation, and a common cause of perinatal morbidity and mortality. The ability to adapt to hypoxic state depends on cardiovascular, respiratory, and renal responses. Although much is known about the role of aldosterone in the myocardium, the relationships between the role of aldosterone, myocardial hypoxia, and hypoxia inducible gene expression are unknown, especially in the neonates. Previous our studies demonstrated that aldosterone-blockade impairs cell proliferation and apoptosis in the neonatal rat heart. In the present study, we hypothesized that aldosterone-blockade affects myocardial hypoxia and hypoxia-inducible gene expression. Newborn rat pups were treated with spironolactone (200 mg/kg/d) or olive oil for 7 days. Spironolactone treatment resulted in a 50% mortality, reduced body weight, decreased heart weight, and decreased heart weight/body weight ratio (P<0.05). Immunoblot for the expression of hypoxia-inducible factor (HIF)-1 alpha, and vascular endothelial growth factor (VEGF), and immunohistochemical staining for hypoxia marker, pimonidazole were performed. In the spironolactone group, VEGF protein expression was significantly increased (P<0.05), whereas HIF-1 alpha protein expression was not changed, compared with control group. In immunostaining, hypoxia marker immunoreactivity was more detected in the spironolactone group. These results indicate that aldosterone inhibition in the developing rat heart induces hypoxic responses, and upregulates hypoxia-inducible gene VEGF .