The effects of microinjection of 5–10 μg of morphine into the midbrain periaqueductal gray (PAG) on the activity of neurons in the rostral ventral medulla (RVM) were studied in lightly anesthetized rats. Based on the relationship between changes in neuronal activity and the occurrence of the tail-flick reflex (TF), RVM neurons were divided into 3 groups: off-cells, on-cells and neutral cells. The off-cells exhibited an abrupt pause and the on-cells an acceleration beginning just prior to the occurrence of the TF. Neutral cell firing did not change at the time of the TF. Microinjections of morphine into the PAG which inhibited the TF had differential effects on the spontaneous activity of the 3 groups of neurons in RVM. Off-cells showed an increase and on-cells a decrease in spontaneous activity which preceded the inhibition of the TF. These microinjections also reduced the TF-related responses of off- and on-cells. The effects on cell activity were reversed by systemically administered naloxone and were not seen following microinjections which failed to block the TF. Neutral cell activity was unchanged following microinjection of morphine into the PAG. These results support the hypothesis that off- and on-cells in the RVM mediate the effects of microinjection of morphine into the PAG on spinal nociceptive reflexes.