The role of membrane potential changes in T cell activation was studied on human peripheral blood lymphocytes stimulated with phytohemagglutinin. Addition of bretylium tosylate, a sodium channels opener, to PHA treated lymphocytes modified the membrane potential and consequently blocked cell activation in a dose-dependent fashion. BT was non-toxic even in long-term (72 hr) incubations. It was reversibly removable, and the removal restored the stimulatory effect of PHA. 3H-thymidine incorporation was blocked if BT was present during the first 20-24 hr of the mitogenic activation. The later BT was added after PHA, the less inhibition of proliferation was observed. BT hyperpolarized the lymphocytes also in the presence of PHA. BT hindered the depolarizing effect of high extracellular potassium concns. The sustained polarized state of the lymphocytes did not influence the intracellular calcium increase upon PHA treatment. IL-2 and transferrin receptor expression was not hindered by BT during PHA stimulation of lymphocytes. Addition of rIL-2 did not abolish the inhibitory effect of BT. According to cell-cycle analysis BT arrested the majority of the cells in G1 phase. It is suggested that cell activation demands the flexible maintenance of a relatively narrow membrane potential "window". Any sustained and significant hyper-, or depolarization, may dramatically decrease the effectivity of transmembrane signalling.