Effects of insulin and diabetes on 2-deoxy-D-glucose uptake by the isolated perfused rat lung.

2-Deoxy-D-glucose uptake was studied in the normal, insulin-treated, and streptozotocin-induced diabetic rat lung. Using an isolated perfused lung preparation, 2deoxy-o-glucose uptake was enhanced by insulin and depressed by diabetes. The uptake of 2-deoxy-o-glucose was inhibited by both phloridzin and cytochalasin B. Further, the uptake process was found to be stereospecific with L-glucose uptake being 45 times lower than that of 2-deoxy-D-glucose. D-Glucose also markedly inhibited 2-deoxy-o-glucose uptake. The uptake of 2-deoxy-D-glucose was saturable at high media substrate concentration and followed Michaelis-Menten type kinetics. The apparent kinetic constants for 2-deoxyglucose uptake were K,,, values of 1.55, 2.87, and 3.57 mM and V,, values (micromoles/g dry weight/min) of 0.94, 2.02, and 0.53 for control, insulin-treated, and diabetic rat lungs, respectively. The lungs accumulated considerable amounts of free, unphosphorylated 2-deoxy-D-glucose. This would suggest that the perfused lung has either decreased sugarphosphorylating capacity or a nonspecific phosphatase capable of breaking down 2-deoxy-D-glucose-6-P. In a cell-free assay system, the kinetic constants for hexokinase activity were an apparent K,,, of 0.36 XIIM and an apparent V,, 0 f 1.21 pmol/g dry weight/min. The hexokinase activity was found to be essentially the same when measured in the presence and absence of 10 mM 2-deoxy-D-glucose-&P, indicating that the intracellular accumulation of the sugar phosphate probably did not inhibit the rate of phosphorylation in the perfused system. It was further found that ATP levels in lungs perfused with 2-deoxy-D-glucose, plus lactate as a utilizable substrate, were only slightly decreased over glucose-perfused controls. These findings indicate that, in the lungs perfused with 2-deoxy-D-glucose, the hexokinase activity is not decreased significantly enough to account for the observed intracellular sugar distribution. The rat lung was found to have a nonspecific phosphatase capable of breaking down 2-deoxy-o-glucose6-P to the free sugar. Using glucose-6-P as substrate, the lung phosphatase showed two peaks of enzymatic activity, a broad peak at pH 5.5 and a sharp peak at 9.5. The activity was lowest in the neutral pH range, but at pH 7.35, there was still enough activity to account