Objective: To investigate the effects of aquaporin-4 (AQP4) on female fertility. Design: Prospective, randomized animal study. Setting: Medical university. Animal(s): AQP4^+/+ and AQP4^-/- female mice. Intervention(s): Eight- to 10-week-old virgin female AQP4^+/+ and AQP4^-/- mice were subjected to a mating study. Superovulation was carried out on 4-week-old AQP4^+/+ and AQP4^-/- mice. The protocol consisted of a single intraperitoneal (IP) injection with 5 units of pregnant mare serum gonadotropin (PMSG) for 48 hours, followed by 5 units of human chorionic gonadotropin (hCG) for an additional 14 or 48 hours. Main Outcome Measure(s): Pregnancy success, oocyte count, morphological changes in ovaries and uteri, and alterations of estrous cycle. Result(s): AQP4-null mice displayed subfertility as evidenced by a lower rate of pregnancy and decreased litter size. Further studies revealed that in AQP4-null mice, there were fewer numbers of antral follicles and corpora lutea in the ovaries and that uteri presented as hyporeactive to gonadotropins. In addition, in AQP4-null mice, superovulation induced by hCG and PMSG yielded fewer numbers of antral follicles, atretic follicles, corpora lutea, and oocytes in ovaries, and uteri exhibited a decreased response to exogenous gonadotropins. Conclusion(s): AQP4 deficiency induces subfertility in female mice. © 2009 American Society for Reproductive Medicine.