1. Corticotropin (ACTH) was one of the first neuropeptides shown to bind to receptors on leukocytes and modulate immune responses. Generally ACTH inhibits immune responses, but certain functions can be enhanced. The present study was performed to determine the effects of ACTH on cytotoxic T-lymphocyte responses, the components, and the major phenotypes of the participating cells. 2. The action of ACTH on cytotoxicity was measured in vitro, in assays utilizing T-lymphocytes that had been previously sensitized in vivo. The cells were then cultured with ACTH and target cells bearing the appropriate stimulatory major histocompatiblity antigens. 3. ACTH did not significantly affect a primary mixed lymphocyte reaction whereas it enhanced a secondary (memory) cytotoxic response up to 100% following 2 days of ACTH treatment. The effect was a shift in the kinetics of effector cell generation so that ACTH-treated cultures demonstrated an augmented cytotoxic activity on day 2, that was not as pronounced on day 3 as cytotoxic activity in control cultures became maximal. ACTH also inhibited Concanavalin A-stimulated T-lymphocyte mitogenesis. Immature thymocyte mitogenesis was inhibited more than that of mature thymocytes. 4. The finding that IFN-gamma was elevated in the cultures suggested that ACTH may enhance memory cytotoxic responses through a combination of mechanisms such as direct cell alterations or synergy with regulatory cytokines. While corticosteroids are probably the most recognized neuroendocrine, stress hormone to affect immune functions, our study illustrates that other neuroendocrine factors such as ACTH, also directly affect immune functions.