In the present study, we investigated immunohistochemically the time-course alterations in estrogen receptor alpha (ER) expression and cell proliferating activity in the developing uteri of Donryu rats exposed neonatally to a high dose p-tert-octylphenol (OP), an endocrine disrupting chemical (EDC). OP-treatment (sc injections of 100 mg/kg, every other day from postnatal days 1 to 15) induced an early and enhanced ER expression in the luminal epithelium compared with age-matched controls from postnatal day (PND) 10, and increased proliferating cell nuclear antigen (PCNA) positive cells up to PND21. At PND28, ER expression in the luminal epithelium of the OP-treated group was decreased, in association with decline in the luminal epithelial areas. PND14, the second week of life, is coincident with the normal time for differentiation when the luminal epithelium invaginates into the stroma to form uterine glands. OP-treatment, however, delayed and inhibited gland-formation, and suppressed ER expression in the invaginated-luminal and glandular epithelium at this time. These results indicate that ER expression in these sites is strongly linked with cell proliferating activity. In stromal cells, ER was expressed from PND6 in both groups without any PCNA positive cells, but significantly lower values were noted in the OP-treated group up to PND10. Our immunohistochemical investigation did not reveal any abnormalities in expression of the proto-oncogene c-fos, mitotic inhibitor p21, or epidermal growth factor antigen, although the apoptotic index in the luminal epithelium was slightly increased in the OP-treated group. These results demonstrate neonatal effects of a high dose of OP, already detectable at PND10, with early and enhanced ER expression, resulting in increase of cell proliferative activity in the luminal epithelium, though expression in the glandular epithelium was suppressed in relation to inhibited gland-genesis. The present study thus suggests that neonatal exposure to high doses of EDCs with estrogenic activity can induce abnormal differentiation in the developing rat uteri via abnormal ER expression and subsequent alteration of cell proliferating activity.