Loss of libido is a major side effect that reduces the quality of life of prostate cancer patients on androgen-deprivation therapy. Estrogen restores sexual interest to some extent in castrated male mammals; however, the beneficial effects of estrogen vary greatly among different studies. We investigated whether the timing of estrogen treatment after castration affected its ability to restore sexual interest in male rats. For each rat, sexual behavior was tested with receptive female rats before castration, and after 2 weeks of either oil alone (as a control) or oil plus estradiol (E2) treatment administered via Silastic tubes implanted immediately, at 1 month (Short-Term), or at 3 months (Long-Term) after castration. Intromission frequency decreased and genital sniffing frequency increased significantly after castration compared to pre-castration levels, regardless of the testing time post-castration. E2 treatment at any time point did not reverse these changes. However, more E2-treated than control rats exhibited mounting behavior, with a significant difference between the Long-Term groups. Mounting frequency did not differ from pre-castration levels for either E2 or control rats under the Immediate condition, but declined significantly in rats treated with oil only under both the Short- and Long-Term conditions. In contrast, E2 treatment elevated mounting frequency above the castrate levels to a similar extent in both the Short and Long-term groups. In conclusion, E2 administration partially restores sexual interest in castrated male rats, and the length of post-castration delay in E2 administration does not affect the ability of E2 to restore mounting behavior.