Estrogen receptor (ER)-beta is a member of the nuclear receptor superfamily and mediates various estrogenic actions. Changes in ER-alpha mRNA expression induced by estrogen have been well documented, whereas those with regard to ER-beta have only been reported for a part of the hypothalamus. In the present study, we examined the effect of estrogen on ER-beta mRNA expression in the female rat brain. Detection of ER-beta mRNA using the in situ hybridization method with a digoxigenin-labeled RNA probe was performed in two groups of female rats: ovariectomized (OVX) and estrogen (E2)-treated. A wide distribution of ER-beta mRNA-containing cells was demonstrated in both groups. In the E2-treated group compared with the OVX group, the number of ER-beta mRNA-containing cells was significantly reduced in the external plexiform layer of the olfactory bulb, entorhinal cortex, intermediate part of the lateral septal nucleus, nucleus of the horizontal limb of the diagonal band, amygdala (lateral, medial and basolateral part), thalamus (anteroventral, laterodorsal and lateral posterior part), medial geniculate nucleus, suprachiasmatic nucleus and Purkinje cells in the cerebellum. These results reveal that ER-beta mRNA-containing cells were decreased by estrogen in several brain regions in the female rat brain, suggesting that ER-beta mRNA is downregulated by the physiological level of estrogen in a region-specific manner.