We studied the effects of supraspinally administered morphine on the expression of the hypothalamic pro-opiomelanocortin (POMC) gene and beta-endorphin. Mice were administered morphine intracerebroventricularly (i.c.v.) either once or 5 times for 5 days (once/day). A single morphine administration significantly increased the hypothalamic POMC gene and beta-endorphin expression at 2h after application in dose-dependent fashion; however, repeated morphine administration had no effect on the hypothalamic POMC gene and beta-endorphin expression. In the immunoblot and immunohistochemical study, the increase of beta-endorphin was observed in the arcuate nucleus of the hypothalamus. Moreover, the expressions of c-Fos, phosphorylated calcium/calmodulin-dependent protein kinase-IIalpha (pCaMK-IIalpha), and phosphorylated cAMP response element-binding protein (pCREB) were increased by a single i.c.v. morphine injection at various time points, but the expressions of phosphorylated extracellular signal-regulated protein kinase1/2 (pERK1/2) and phosphorylated IkappaB (pIkappaB) were not. We also found that the expressions of c-Fos, pCaMKIIalpha, and pCREB were co-localized with the POMC expression. Meanwhile, naloxone as well as muscimol and baclofen significantly attenuated the increases of the POMC gene expression induced by a single morphine administration. Furthermore, the pretreatment of muscimol and baclofen 10 min before morphine injection robustly attenuated the withdrawal behavior induced by a single morphine administration. These results imply that the hypothalamic POMC gene and beta-endorphin expression may play an important role in the development of an acute physical dependency of morphine. In that, GABAergic neurotransmission appear to be involved in the regulation of the hypothalamic POMC gene expression induced by supraspinal morphine administration.