Wild-type and mu-opioid receptor knockout (MORKO) mice were used to investigate the role of corticosterone (CORT) and the mu-opioid receptor (MOR) in chronic morphine-mediated immunosuppression. We found that although plasma CORT concentrations in CORT infusion (10 mg/kg/day) and morphine-pellet implantation (75 mg) mice were similar (400-450 ng/ml), chronic morphine treatment resulted in a significantly higher (two- to threefold) inhibition of thymic, splenic, and lymph node cellularity; inhibition of thymic-lymphocyte proliferation; inhibition of IL-2 synthesis; and activation of macrophage nitric oxide (NO) production when compared with CORT infusion. In addition, results show that the inhibition of IFN-gamma synthesis and splenic- and lymph node-lymphocyte proliferation and activation of macrophage TNF-alpha and IL-1beta synthesis occurred only with chronic morphine treatment but not with CORT infusion. These morphine effects were abolished in MORKO mice. The role of the sympathetic nervous system on morphine-mediated effects was investigated by using the ganglionic blocker chlorisondamine. Our results show that chlorisondamine was able to only partially reverse morphine's inhibitory effects. The results clearly show that morphine-induced immunosuppression is mediated by the MOR and that although some functions are amplified in the presence of CORT or sympathetic activation, the inhibition of IFN-gamma synthesis and activation of macrophage-cytokine synthesis is CORT-independent and only partially dependent on sympathetic activation.