We tested the hypothesis that the previously observed loss of thymic lymphocytes in mice after treatment with time-release morphine pellets was occurring through the process of apoptosis. Apoptosis is a form of cell death, distinct from necrosis, which involves a specific endonuclease that fragments the cell's own DNA. Forty-eight hours after implantation of a time-release morphine pellet in B6C3F1 mice, thymus weight and cellularity was reduced to 30% of that observed in placebo-treated mice. Thymocytes from morphine pellet-treated mice were found to have a significantly greater percentage of their DNA fragmented than did thymocytes from either placebo pellet-implanted or naive control mice. The peak level of DNA fragmentation was found to occur approximately 12 hr postpellet implant. When separated on agarose gels, the sizes of the DNA fragments observed corresponded to the multiples of 180 base pairs which are characteristic of apoptosis. In vivo, the use of either the opiate receptor antagonist naloxone, or the glucocorticoid receptor antagonist RU-38486, was able to block completely the morphine mediated increase in thymocyte apoptosis. In vitro experiments in which thymocytes were cultured with morphine concentrations as high as 10(-4) M showed no evidence of an increased rate of DNA fragmentation. These data indicate that both opiate and glucocorticoid receptors are involved in morphine-induced apoptosis and that the opiate receptor responsible is not located on the thymic lymphocytes.