The possible modulatory role of microtubules or closely related colchicine-sensitive structures in the response of human lymphocytes to mitogenic lectins was investigated. Colchicine (0.1 to 10 muM) AND VINBLASTINE (0.1 TO 10 MUM) inhibited early [14C]-aminoisobutyric acid and late [3H]-thymidine uptake in phytohemagglutinin-and concanavalin A-stimulated human lymphocytes but failed to alter 45Ca uptake. Lumicolchicine, an inactive congener of colchicine, was ineffective in all three systems. Both microtubular agents accentuated and prolonged the early cyclic AMP response to lectin. Little or no alteration in cyclic AMP levels was seen with colchicine or vinblastine alone or in combination with PGE (10MUM) or epinephrine (1muM) suggesting that the effect on cyclic AMP metabolism is largely selective for lectin stimulation. Neither microtunular agent altered 125I-concanacalin A binding. Since the inhibition of DNA synthesis was throughout the culture period and early aminoisobutyric acid uptake is affected, it appears that these agents are acting on an early event, or events, in the activation sequence. Although the mechanism of the inhibition is not known, the effect of colchicine and vinblastine in prolonging the cyclic AMP response to lectin may be involved. Alternatively, alterations in microtubules assembly may exert effects on membrane architecture interfering with propagation of the stimulus from the membrane to the cell interior.