A proposed membrane-mediated mechanism of rapid nongenomic response to estrogen has been the intense focus of recent research. Estren, a synthetic steroid, is reported to act selectively through a rapid membrane-mediated pathway, rather than through the classical nuclear estrogen receptor (ER)-mediated pathway, to maintain bone density in ovariectomized mice without uterotropic effects. To evaluate the mechanism and physiological effects of estren, we studied responses in adult ovariectomized mice. In a 3-d uterine bioassay, we found that 300 μg estren significantly increased uterine weight; in comparison, a more maximal response was seen with 1 μg estradiol (E2). The estren response was partly ERα independent, because ERα knockout (αERKO) uteri also exhibited a more moderate weight increase. Estren induced epithelial cell proliferation in wild-type, but not αERKO, mice, indicating ERα dependence of the epithelial growth response. Examination of estren-regulated uterine genes by microarray indicated that early (2 h) changes in gene expression are similar to the early responses to E2. These gene responses are ERα dependent, because they are not seen in αERKO mice. Later estren-induced changes in gene expression (24 h) are blunted compared with those seen 24 h after E2. In contrast to early genes, these later estren responses are independent of ERα, because the αERKO shows a similar response to estren at 24 h. We found that E2 or estren treatments lead to depletion of ERα in the uterine cytosol fraction and accumulation in the nuclear fraction within 30-60 min, consistent with the ability of estren to regulate genes through a nuclear ERα rather than a nongenomic mechanism. Interestingly, estren, but not E2, induces accumulation of androgen receptor (AR) in the nuclear fraction of both wild-type and αERKO samples, suggesting that AR might be involved in the later ERα-independent genomic responses to estren. In conclusion, our studies suggest that estren is weakly estrogenic in the mouse uterus and might induce nuclear ERα- and AR-mediated responses. Given its activity in our uterine model, the use of estren as a bone-selective clinical compound needs to be reconsidered.