Human leukocyte interferon (hIFN-α) preparations contain immunologically and biologically recognizable endorphin and corticotropin-like (ACTH-like) activities. The ACTH bioactivity was demonstrable only after pepsin or acid treatment. Highly purified hIFN-α was composed of two molecular species of interferon (18.500 and 23,000 daltons). Endorphin activity was associated with both of these molecules. Pepsin treatment of the 23,000-dalton but not the 18,500-dalton hIFN-α generated ACTH activity. In acid, the 23,000-dalton hIFN-α broke down into the 18,500-dalton form and ACTH (4500 daltons). The ACTH derived from hIFN-α by pepsin digestion comigrated with a purified ACTH standard in NaDodSO4/polyacrylamide gel electrophoresis. hIFN-α-producing lymphocytes showed positive immunofluorescence after staining with highly specific antisera to ACThα-(1-13) and γ-endorphin. Essentially 100% of the human peripheral lymphocytes were capable of producing both ACTH and γ-endorphin-related substances, presumably associated with hIFN-α. These results strongly suggest a circuit between the immune and neuroendocrine systems which involves neuroendocrine hormone-like substances, some of which are associated with hIFN-α.