A kinetic analysis of D-glucose transport by adipocyte plasma membranes

The measurement of the initial rate of glucose uptake is of critical importance for the kinetic analysis of the effects of temperature, ions, or hormones on the uptake process. Presented here is a highly reproducible double isotope assay system which for the first time allowed the measurement of the initial rates of glucose uptake into isolated adipocyte plasma membrane vesicles by having time points as early as 1/2 s. Measurement of the initial rates of uptake at 1 s allowed the calculation of a Km of 9.0 mM and a Vmax of 3209 pmol/mg of protein/s for specific D-glucose transport (D-glucose minus L-glucose). The time course of specific D-glucose uptake is extremely rapid, reaching an equilibrium value in approximately 20 s with a half-time of 4 s. This uptake process is markedly inhibited by cytochalasin B. The similarity of these findings to those for intact adipocytes indicates that the plasma membrane preparation and the assay conditions used have retained the structural and functional integrity of the glucose transport system from the intact cell.