The physiological roles of aldose reductase [alditol:NAD(P)+1-oxidoreductase] have not been fully elucidated yet, although it has been implicated in the pathogenesis of diabetic complications. In the rat ovary we found remarkable changes in the enzyme level during the 4-day estrous cycle. After diestrus, the activity and protein content of aldose reductase increased to the maximum level on proestrous morning and rapidly fell off to the lowest level on the early morning of estrus. At this time its mRNA level in the ovary was significantly decreased compared with that on the morning of proestrus. Immunohistochemical staining of the diestrous ovary demonstrated localization of the enzyme protein in the granulosa cells and in the oocytes. At the end of proestrus when its level was low, immunoreactive aldose reductase in the granulosa cells was localized preferentially to the antrum side, with lesser staining in the cells lining the follicles. Administration of chlorpromazine to the rats on proestrus significantly restored the enzyme level on the following morning of the expected estrus. This effect of chlorpromazine was abolished when human chorionic gonadotropin was administered to the chlorpromazine-treated rats. When chlorpromazine was administered to the rats treated with bromocriptine, an inhibitor of pituitary prolactin secretion, aldose reductase activity in the ovary was significantly elevated compared with that in the rats treated with chlorpromazine alone. These findings suggest that in the rat ovary it is under hormonal regulation during the estrous cycle. The enzyme may possess a new functional role in the reproductive system of the female rat, which can be disordered under diabetic conditions.