The c-fos gene is expressed in the central nervous system (CNS) in response to neuronal stimuli. Induction of c-fos in certain CNS regions occurs following naltrexone precipitated withdrawal in morphine dependent rats. Non-competitive (MK801) and competitive (LY274614) NMDA receptor antagonists and clonidine, an alpha2 partial agonist, attenuate the intensity of naltrexone precipitated withdrawal. We determined the levels of c-fos mRNA by solution hybridization in several brain regions in control and morphine dependent rats following pretreatment with saline, MK801 (1 mg/kg, s.c.), LY274614 (100 mg/kg, i.p.), or clonidine (1.5 mg/kg, i.p.). Morphine treatment increased c-fos mRNA levels in striatum (STR) and amygdala (AMY). Naltrexone did not alter c-fos mRNA levels in placebo-treated rats. However, naltrexone increased c-fos mRNA levels in morphine dependent rats in the nucleus accumbens (NA), frontal cortex (FC), AMY, and hippocampus (HIP) but not in STR or spinal cord. Pretreatment with MK801 blocked this effect of naltrexone in AMY but not in NA, FC, or HIP, while pretreatment with LY274614 or clonidine blocked this effect of naltrexone in AMY and NA but not in FC or HIP. These results further delineate both the neuroanatomical pathways involved in morphine withdrawal and the locus of action of compounds that reduce morphine-withdrawal symptoms.