Neurokinin B (NKB) gene expression is elevated in the infundibular (arcuate) nucleus of the hypothalamus in postmenopausal women. Estrogen replacement decreases both the number of NKB mRNA-expressing neurons and the level of expression within individual cells. Similarly, NKB gene expression is elevated in ovariectomized rats and reduced after estrogen treatment. The actions of estrogen in the brain can be mediated via either estrogen receptor alpha (ERalpha) or estrogen receptor beta (ERbeta). In the rodent arcuate nucleus (ARC), more ERalpha- than ERbeta-containing cells are present, suggesting that ERalpha might be directly responsible for estrogen regulation of NKB gene expression. However, an indirect effect via ERbeta could not be ruled out. Here we used ERalpha knockout and ERbeta knockout mice to identify the type of ER responsible for mediating estrogen action on NKB gene expression in the ARC. Using in situ hybridization histochemistry, we have found that estrogen treatment significantly reduced NKB gene expression in the ARC of ovariectomized ERbeta knockout mice, but had no effect on NKB mRNA levels in ERalpha knockout mice. These data indicate that ERalpha mediates the increase in NKB gene expression associated with ovariectomy in rodents and might also be responsible for the increase in NKB in postmenopausal women.