To monitor the possible effect of morphine on sigma sites, binding characteristics of [3H](+)-pentazocine and [3H]1,3-di-(2-tolyl)guanidine (DTG) to brain and spinal cord membranes of morphine-treated and control mice were compared. For morphine treatment, a single injection (100 mg/kg, s.c.) of morphine was followed 4 h later by pellet implantation (75 mg morphine free base). Animals were sacrificed 24, 72 h or 7 days later. The equilibrium dissociation value (Kd) and the density (Bmax) of [3H](+)-pentazocine binding remained unaffected by morphine treatment. Also, no change was found in Kd and Bmax values of [3H]DTG labeled sigma2 subtypes after any morphine treatment schedule when measured in the presence of 100 nM (+)-pentazocine. However, the Bmax of [3H]DTG binding in the spinal cord in the absence of 100 nM (+)-pentazocine, was significantly elevated 72 h after implantation of the morphine pellet and recovered by 7 days, a time when the antinociceptive effect produced by the morphine pellet had dissipated. These data suggest that one population of sigma sites, that has a high affinity for [3H]DTG, but is not equivalent with the [3H](+)-pentozocine labeled sigma1 subtype or the [3H]DTG labeled sigma2 subtype, is upregulated by morphine and, therefore, may play a role in the development of tolerance to or dependence on the effects of morphine.