We have examined the role of interleukin 2, interferon-gamma and interferon-alpha in the generation of natural cytotoxic (NC) activity and cytotoxic T-lymphocyte (CTL) activity in peripheral blood lymphocyte cultures stimulated with influenza virus, using the immunosuppressive effects of prednisolone. In addition to an inhibitory effect on the generation of CTL activity, prednisolone also inhibited the generation of NC activity in a similar dose-and time-dependent manner. Prednisolone suppressed the production of interferon-gamma when it was added on the first day of culture of PBL with influenza virus. Levels of interferon-alpha were not affected. The effects of prednisolone on the generation of NC activity and CTL activity in kinetic terms were not paralleled by the effects on interferon-alpha and interferon-gamma production. The diminished generation of NC activity could be reversed by the addition of interleukin 2 (IL-2), but interferon-gamma had little if any restorative effects. Interferon-alpha had no effect. These findings support the hypothesis that IL-2 is the major inducer of NC activity in CTL generation cultures. The inhibitory effect on CTL generation could only be reversed by IL-2 and not by interferon-alpha- and interferon-gamma. Thus, in the absence of IL-2, interferon-alpha and interferon-gamma cannot support the generation of CTL activity or the concomitantly induced NC activity.