Tolerance to the analgesic effects of morphine results in part from the development of a compensatory response in neurons that express the opioid receptor or of neural circuits in which those neurons participate. According to this formulation, withdrawal of morphine results in an overshoot of several neuronal properties because of the unopposed action of the compensatory response system. To identify the population of spinal cord neurons that underlies this state, we monitored expression of Fos-like immunoreactivity, after naltrexone-precipitated abstinence in normal and morphine-tolerant rats. After daily (five days) implantation of morphine or placebo pellets, the rats received an injection of saline or naltrexone and behavior was monitored for 1 h. The rats were then killed, their spinal cords removed and 50-microns transverse sections of the lumbar cord were immunostained with a rabbit polyclonal antiserum directed against Fos. Naltrexone injection in the placebo group did not increase spinal cord Fos expression. Naltrexone-precipitated abstinence resulted in an increase in Fos expression at all levels of the spinal cord; the greatest increase and densest staining was in laminae I through VI. Importantly, when withdrawal was precipitated in anesthetized rats, we recorded a significant reduction in Fos expression, particularly in laminae III through VI, but there was persistent expression in the superficial dorsal horn, particularly in lamina I. These results suggest that spinal cord nociresponsive neurons are sensitized during the development of tolerance. This sensitization is unmasked by the administration of naltrexone and is manifested by fos induction in laminae I/II in awake or anesthetized withdrawing animals. The underlying mechanisms of tolerance development may be similar to those that underlie injury-induced central sensitization and hyperalgesia.