The negative affective aspects of nicotine withdrawal have been hypothesized to contribute to tobacco dependence. In the present studies in rats, brain stimulation reward thresholds, conditioned place aversions, and somatic signs of withdrawal were used to investigate the role of central and peripheral nicotinic acetylcholine and opioid receptors in nicotine withdrawal. Rats prepared with s.c. osmotic mini-pumps delivering 9.0 mg/kg/day nicotine hydrogen tartrate or saline were administered various doses of the nicotinic antagonists mecamylamine (s.c.), chlorisondamine (s. c. or i.c.v.), dihydro-beta-erythroidine (s.c.), or the opiate antagonist naloxone (s.c.). Nicotine-treated rats receiving mecamylamine or i.c.v. chlorisondamine exhibited elevated thresholds and more somatic signs than saline-treated rats. Nicotine-treated rats receiving s.c. chlorisondamine, at doses that do not readily cross the blood-brain barrier, exhibited more somatic signs than saline-treated rats with no threshold elevations. Naloxone administration produced threshold elevations and somatic signs only at high doses that induced similar magnitude effects in both nicotine- and saline-treated subjects. Mecamylamine or dihydro-beta-erythroidine administration induced conditioned place aversions in nicotine-treated rats but required higher doses than those needed to precipitate threshold elevations. In contrast, naloxone administration induced conditioned place aversions at lower doses than those required to precipitate threshold elevations and somatic signs. These data provide evidence for a dissociation between centrally mediated elevations in reward thresholds and somatic signs that are both centrally and peripherally mediated. Furthermore, threshold elevations and somatic signs of withdrawal appear to be mediated by cholinergic neurotransmission, whereas conditioned place aversions appear to be primarily mediated by the opioid system.