D-Glucose uptake into jejunal brush-border membrane vesicles was studied in suckling (2-wk-old), weanling (3-wk-old), and adolescent (6-wk-old) rats. The purity of the membrane vesicles from all age groups was validated by the finding that the specific activity of brush-border enzyme markers was severalfold greater in membrane vesicles compared with corresponding values in mucosal homogenate. D-Glucose uptake was inversely related to increasing medium osmolality, indicating that uptake of D-glucose was into the intravesicular space rather than binding. D-Glucose uptake was sodium dependent at all age groups; however, the initial uptake at 20 s was significantly greater in adolescent rats compared with suckling rats. The addition of valinomycin to KCl-preincubated vesicles in the presence of Na+ gradient resulted in a severalfold increase in D-glucose initial uptake over Na+ gradient alone, indicating that D-glucose uptake was electrogenic at all age groups. To delineate the mechanism for the decrease in the initial uptake in suckling rats, two experiments were performed: 1) an exchange tracer study that indicated the activity of D-glucose-Na+ transporters was similar in suckling and adolescent rats, and 2) a study that indicated 22Na uptake, as an indicator for Na+ permeability, was significantly greater in suckling rats compared with adolescent rats. These findings suggest that a Na+-dependent, electrogenic D-glucose uptake is already developed in the suckling period; however, because of the increased permeability to Na+, the Na+ gradient dissipates faster, resulting in a decrease in initial uptake.