We explored whether hypoxic preconditioning minimizes oxidative injury induced by overdistension/emptying in the rat bladder. For hypoxic preconditioning, female Wistar rats were placed in a hypobaric chamber (380 Torr) 15 h/day for 28 days. Overdistension was induced by infusion of two times the threshold volume of saline into the bladder and was maintained for 1 or 2 h, followed by drainage/emptying. During overdistension (ischemia) and emptying (reperfusion) periods, a bursting increase of reactive oxygen species (ROS) from the bladder was originated from the large numbers of infiltrating leukocytes and scattered resident cells, including urothelial, submucosal, and smooth muscle cells. ROS impaired the voiding function by a reduction of bladder afferent and efferent nerve activity and acetylcholine- or ATP-induced detrusor contraction. ROS enhanced pro-apoptotic mechanisms, including increases in the Bax/Bcl-2 ratio, CPP32 expression, and PARP fragments with subsequent apoptotic cell formation in the insulted bladders. Hypoxia preconditioning upregulated Bcl-2 expression in the bladder and significantly reduced the levels of ROS and apoptosis detected in the overdistension/emptying bladders and preserved partial voiding function. In conclusion, Bcl-2 upregulation by hypoxia preconditioning contributes protection against overdistension/emptying-induced oxidative stress and injury in the bladder.