IL-12 is a crucial cytokine for the determination of a Th1/Th2 balance. It is important, therefore, to elucidate the mechanisms of IL-12R expression on Th cells. In this report, we present evidence to show that B7-2 costimulation plays a pivotal role in the expression of IL-12R on Th cells. A Th1 clone expressed a low density of IL-12R in a resting condition, the expression was enhanced by stimulation with specific Ag on splenic adherent cells and the enhancement was inhibited by anti-B7-2 or CTLA-4-Ig. When stimulated with anti-CD3 plus B7-2-transfected Chinese hamster ovary (CHO) cells, the clone strongly expressed IL-12R, although anti-CD3 by itself only weakly enhanced the expression. We obtained results that were similar to those in the Th1 clone in CD4+ CD45RB(low) memory T cells. In CD4+ CD44(low) naive T cells, costimulation with B7-2-CHO was found to play a more important role in IL-12R expression. The accumulation of both IL-12R beta 1 and -beta 2 chain mRNAs was detected in naive T cells only when they were costimulated with anti-CD3 and B7-2-CHO, but beta 2 mRNA was not expressed upon anti-CD3 stimulation alone. On the other hand, both Th1 clones and memory T cells expressed low amounts of these mRNA without any stimulation, and the expression was weakly enhanced by anti-CD3 stimulation alone. For the maximum expression of these mRNAs, however, these cells also required costimulation with anti-CD3 and B7-2-CHO.