Peripheral blood V gamma 9/V delta 2 T cells proliferate vigorously in response to heat-killed Mycobacterium tuberculosis (Mtb) and Daudi Burkitt's lymphoma cells. We have analyzed the respective roles of IL-10 and IL-12 in gamma delta T cell activation, using the selective expansion of V gamma 9 cells in Mtb- or Daudi-stimulated PBMC as a readout. IL-10 inhibited in a dose-dependent fashion the V gamma 9 responsiveness to Mtb. In contrast, IL-10 did not impair reactivity toward Daudi cells, even when added at 10 ng/ml. The inhibitory action of IL-10 could be overcome by the exogenous supply of IL-2 or IL-12. Blockade of endogenous IL-10 by neutralizing mAb increased responsiveness to Mtb but not to Daudi cells. Low concentrations of exogenous IL-12 increased the V gamma 9 responsiveness to Mtb in 6 of 11 healthy donors. In contrast, IL-12 inhibited V gamma 9 cell expansion in response to Daudi stimulation in all of the tested individuals. Neutralizing anti-IL-12 Ab significantly suppressed V gamma 9 responsiveness to Mtb in IL-12 nonresponders but only slightly inhibited the latter responsiveness in IL-12 responders. The effect of anti-IL-12 Ab correlated with the level of Mtb-stimulated endogenous IL-12 production. In purified gamma delta T cells, IL-12 synergized with IL-2 to stimulate cellular expansion in response to Daudi cells. This effect was reduced when T cell-depleted APC were added back. Finally, neutralization of endogenous IFN-gamma by Ab abrogated V gamma 9 cell responsiveness to Mtb but exerted only a minor inhibition of the reactivity toward Daudi cells. Taken together, these results reveal significant differences in the cytokine requirement of gamma delta T cell activation by Mtb or Daudi lymphoma cells, respectively.