Maize (Zea mays L.) endosperm suspension cultures are a useful model system for studying biochemical and physiological events in developing maize endosperm. In this report, sugar uptake by the cultures is characterized. Uptake of '4C-labeled fructose and L-glucose was linear with time, while the rate of uptake of radioactivity from sucrose increased over a 120 min period. Both saturable and linear components of uptake were observed for fructose, glucose, sucrose, 1'-deoxy-1'-fluorosucrose, and maltose. Uptake of mannitol, sorbitol, and L-glucose took place at lower rates and was linear with concentration. Rates of incorporation of radioactivity from fructose and glucose exceeded that of sucrose at all concentrations tested. Kinetics of 1'-deoxy-1'-fluorosucrose uptake indicated that '4C from sucrose can be taken up by a saturable carrier of intact sucrose as well as by invertase hydrolysis and subsequent uptake of hexoses. Cell wall invertase was demonstrated histochemically. Further study of fructose uptake at a concentration at which the saturable component predominated revealed sensitivity to metabolic inhibitors, respiratory uncouplers, the nonpermeant sulfhydryl reagent p-chloromercuribenzenesulfonic acid, and nigericin. Uptake was not affected by valinomycin plus K' and was stimulated by fusicoccin. Fructose and glucose uptake was not pH-sensitive below pH 7.0, whereas uptake of radioactivity from sucrose and 1 '-deoxy-1 '-fluorosucrose declined as the pH was increased above 5.0. Fructose uptake was not completely inhibited by glucose and vice versa, suggesting the presence of specific carriers. These results indicate that maize endosperm suspension cultures (a) absorb fructose via a typical, energy-requiring, carrier-mediated proton cotransport system; (b) possess saturable carriers for glucose and sucrose; and (c) also absorb sucrose via hexose uptake after sucrose hydrolysis by extracellular invertase.