We investigated the effects of estrogen‐related compounds including xenoestrogens [17β‐estradiol (E2), 17α‐ethynylestradiol (EE), diethylstilbestrol (DES), p‐nonylphenol (PNP), bisphenol A (BPA) and 17α‐estradiol (17α)] on l‐glu uptake by cultured astrocytes via glutamate‐aspartate transporter (GLAST). After 24 h treatment, E2 inhibited the l‐glu uptake at 1 µm and higher concentrations. EE and DES also inhibited the l‐glu uptake at 1 nm and higher concentrations. The other four compounds had no effect. The effects of E2, EE and DES were completely blocked by 10 nm of ICI182 780 (ICI). β‐Estradiol 17‐hemisuccinate : bovine serum albumin (E2‐BSA), a membrane‐impermeable conjugate of E2, also elicited the inhibition of l‐glu uptake at 1 nm and higher concentrations, and the effect was blocked by ICI. 16α‐Iodo‐17β‐estradiol (16αIE2), an estrogen receptor α (ERα) selective ligand, revealed an inhibitory effect at 10 nm, while genistein, an ERβ selective ligand, failed to reveal such an effect at this concentration. Western blot analysis showed that the predominant ER of cultured astrocytes was ERα. The colocalization of ERα with GLAST on plasma membranes was immunohistochemically detected in these cells. From these results, we concluded that estrogens down‐regulate l‐glu uptake activity of astrocytes via membrane ERα.