We have found that the induction of suppressor T cells (Ts) function in pokeweed mitogen (PWM)-stimulated cultures of peripheral blood mononuclear cells (PBMC) depends on the concentration of monocytes in inductive cultures. When cultured for 7 to 8 days in a physiologic concentration of monocytes (10 to 15%) suppressor cells differentiated with the surface phenotype CD8+ DR+ Leu-7- Leu-8+ Tp44-. Suppressor function, as assessed in secondary cultures of fresh PWM-stimulated CD4+ cells, was partially radiosensitive, and its induction could be partially inhibited by treatment with indomethacin. Culture supernatants of PWM-pulsed CD4+ cells could induce CD8+ suppressor cells only when monocytes were included in cultures for supernatant production. In marked contrast, if the monocyte concentration of PWM-stimulated T cells was reduced to < 5%, CD8+ cells harvested from such cultures substantially augmented proliferation of PWM-stimulated CD4+ cells in assay cultures. Amplifier cells from monocyte-depleted cultures, were also DR+Tp44-. Suppressive activity of the CD8+ subset was restored simply by addition of monocytes to purified T cells, reconstituting the concentration present among unfractionated PBMC. Addition of IL 1 could not replace the requirement of monocytes for suppressor cell induction either in PWM-stimulated primary cultures or in culture supernatants. The data thus demonstrate that the regulatory activity of CD8+ lymphocytes is critically dependent on the conditions of cellular activation; at concentrations of monocytes normally present in peripheral blood, PWM stimulation induced potent suppressor activity, whereas under conditions of moderate monocyte depletion the regulatory activity of the same phenotypic subset of CD8+ cells was reversed. The experiments thus suggest that the functional consequences of an activating stimulus may depend on regulatory effects of monocytes in CD8+ suppressor cell induction.