Using pharmacologic agents as probes, three aspects of the mitogen-induced cellular cytotoxicity reaction (MICC) were evaluated. These included the requirement for a concomitant lymphoid DNA synthetic response, the role of microtubules and microfilaments in effecting target cell lysis, and the potential inhibitory activity of hydrocortisone. Results, using both PHA and Con A stimulated guinea pig lymph node cells, indicate that this form of cytotoxicity occurs as a phenomenon independent of 3H-Tdr incorporation. Animals treated with cyclophosphamide showed cytotoxic reactions equivalent to untreated controls whereas mitogen-induced proliferative responses were greatly impaired. Further studies indicated that the in vitro exposure of lymph node cells to either mitomycin or cytosine arabinoside markedly reduced DNA synthesis but did not depress the ability of mitogen-stimulated lymphocytes to effect target cell lysis. The microtubular antagonists, vincristine, vinblastine and colchicine, inhibited cytotoxic reactions if the agents were present at the time that lymphocyte-target cell interaction was established. However, the antimicrofilament agent cytochalasin B was not inhibitory; this contrasts to this agent's ability to suppress antigen-induced cell-mediated cytotoxic reactions. In a third series of experiments, hydrocortisone showed a dose-related inhibition of MICC responses. The action of the steroid was apparent only if the drug was present at the onset of the culture; no suppression was observed if it was added after an initial one day incubation.