Exposure to microgravity results in postflight cardiovascular deconditioning in astronauts. Vascular oxidative stress injury and mitochondrial dysfunction have been reported during this process. To elucidate the mechanism for this condition, we investigated whether mitochondrial oxidative stress regulates calcium homeostasis and vasoconstriction in hindlimb unweighted (HU) rat cerebral arteries.Three-week HU was used to simulate microgravity in rats. The contractile responses to vasoconstrictors, mitochondrial fission/fusion, CaAn increase of cytoplasmic CaThe present results suggest that mitochondrial oxidative stress enhances cerebral vasoconstriction by regulating calcium homeostasis during simulated microgravity.