Introduction: Sex-dependent changes of behavioral responses in humans and animals has been widely investigated. There are also controversial reports regarding the role of sex hormones in memory and learning. The present study carried out to compare spatial learning of male, female and ovariectomized (OVX) rats in Morris water maze.  Materials and Methods: Twenty four rats were divided into 3 groups: 1) male, 2) female, 3) OVX. The animals were ovariectomized under ketamine anesthesia (150 mg/kg, I.P.). The same procedure as OVX rats was performed on the female and male rats except the wound was closed without removing the ovaries.8 weeks later, the animals of all groups were tested in Morris Water Maze. The escape latency, traveled path lengh and swimming speed to reach the platform were compared between groups. Results: Time latency in both OVX and male groups was significantly higher than female group . However, path length in OVX group was higher than female group, there was no significant difference in path length between male and female or between male and OVX groups. The swimming speed in male group was lower than female group. Conclusion: It might be concluded that endogenous estrogen has an important role in sex dependent differences of spatial learning.