Morphine has been reported to possess immunosuppressive actions in both in vitro as well as in vivo assays of immune function. Our work in female B6C3F1 mice, surgically implanted with a 75-mg time release morphine pellet, has confirmed previous reports of a rapid loss in the cellularity of the spleen and thymus. To evaluate the effect of morphine on the subpopulations of cells in the thymus, two color fluorescence flow cytometry studies were performed. Fluorescently conjugated monoclonal antibodies specific for the murine cell surface CD4 and CD8 markers were used to identify the four major subpopulations of thymocytes. These studies indicated that morphine pellet-implanted mice suffered a loss in each of the four thymocyte subpopulations in comparison to placebo-implanted mice. However, the loss (> 90%) in the important CD4+/CD8+ subpopulation of immature thymocytes greatly exceeded that which was observed for any other subpopulation. Kinetic studies of morphine's effect on the thymocyte subpopulations revealed that the maximal depletion of the CD4+/CD8+ cells occurs approximately 4 days after pellet implantation. Thymocyte cell populations recovered by 14 days, with an increase above placebo for the double positive cells. Naltrexone administration blocked thymic alterations, suggesting that these immunologic consequences of morphine may be mediated through an opiate receptor. Measurements in thymocytes from morphine pellet-implanted mice showed an increased level of DNA fragmentation, whereas in vitro exposure to morphine (1-100 microM) produced no such increases. This suggests morphine may be working indirectly to induce apoptosis of immature thymocytes.