There is substantial epidemiological evidence suggesting that alcohol consumption is associated with increased risk for breast cancer. However, possible biological mechanisms have not been clearly established. In the present studies, a direct effect of ethanol on the proliferation and intracellular content of cyclic AMP (cAMP) in two estrogen receptor-positive (ER+) and two estrogen receptor-negative (ER-) human breast cancer cell lines was examined. Treatment of ER+ human breast cancer cells (MCF-7 and ZR75.1) with ethanol at concentrations between 10 and 100 mM was associated with increased cell numbers compared to controls. The ERalpha content and the amount of intracellular cAMP also increased in ER+ cells exposed to ethanol, compared to controls. On the other hand, ethanol treatment did not increase cell proliferation or cAMP levels in the ER- (BT-20 and MDA-MB-231) human breast cancer cells. Therefore, ethanol added at physiologically relevant concentrations to ER+ human breast cancer cell cultures can enhance cell proliferation and increase the content of ERalpha.