Stress is known to either up or down regulate immunity. In this study, mice were subjected to handling combined with rectal temperature measurement (RTM) stress or handling only stress. We investigated whether there were any significant differences in the effect of handling combined with RTM and handling only on NK cell activity, serum cytokine (IL-1β, IL-6, and TNF-α) and ACTH and β-endorphin levels, and splenic cytokine (IL-1β, IL-6, TNF-α, IFN-α, and IFN-β) levels. Circulating cytokines and hormones and splenic cytokine mRNA levels were measured in individual mice. NK cell activity was significantly increased in both stress groups when compared to the control group. Handling combined with RTM produced significantly increased serum levels of IL-1β, IL-6, and β-endorphin. Serum IL-1β, ACTH, and β-endorphin were elevated significantly in the handling only group. Splenic TNFα mRNA in both of the stress groups and IL-6 mRNA in handling only group decreased significantly. Our observations are supported by existing literature demonstrating that various stressors have differential effects on immune functions and the neuroendocrine hormones and cytokines, which regulate them. © 2002 Elsevier Science Inc. All rights reserved.