Pretreatment of rat thymus lymphocytes with N-ethyl-maleimide prevents the stimulation of 3-O-methyl-glucose transport by concanavalin A or ionophore A23187 but does not affect the ability of concanavalin A to induce a rapid increase in cellular Ca2+take. N-ethyl-maleikide added after concanavalin A amplifies rather than prevents the subsequent stimulation of 3-O-methyl-glucose transport. Incubation of thymocytes with concanavalin A produces a decrease of 43% in the apparent Ki for phloretin, a competitive inhibitor of 3-O-methyl-glucose transport, without affecting the apparent Km for the substrate. Similarly, very low concentrations of cytochalasin B inhibit concanavalin A-stimulated glucose transport preferentially, without markedly affecting the unstimulated transport rates. The similarity between concanavalin A-stimulated 3-O-methyl-glucose transport in thymocytes and insulin-stimulated glucose transport in adipose tissue, with particular emphasis on the effects of the modifying agents described in this paper, is discussed.