Metabolic Effects of 2-deoxy-D-glucose in Isolated Fat Cells

The uptake and phosphorylation of 2-deoxy-D-glucose by isolated adipocytes of the rat was determined by a method of rapid flotation through oil coupled with separation of sugar from sugar phosphate by chromatography on Dowex-1-formate. Uptake of the sugar is rapid and linear over 5 min, with a gradual decline thereafter; by 1 h, no further uptake is observed. Initially only 2-deoxy-glucose phosphate is observed within the cells; by 1 h, however, free 2-deoxy-glucose accumulates to levels approximately those in the medium. Phosphorylation ceases when intracellular levels of 2-deoxyglucose phosphate are about 50 mM regardless of the medium concentration of 2-deoxyglucose; this does not represent feedback inhibition of hexokinase, since the enzyme in fat cell homogenates is not inhibited by 50 mM 2-deoxyglucose 6-phosphate. Accumulation of deoxyglucose 6-phosphate is associated with a marked decline in intracellular ATP levels. Fat cell respiration is also depressed by approximately 50 per cent after a 1 h preincubation with 10 or 20 mM 2-deoxyglucose. Intracellular ATP levels and O2 uptake are only partially corrected by the addition of pyruvate to the incubation medium. Since no glucose was present in the medium, and intracellular concentrations of glycogen are known to be small in adipose tissue, it is proposed that accumulation of 2-deoxyglucose 6-phosphate within fat cells has a direct inhibitory effect on cell respiration unrelated to inhibition of glycolysis. No increase in intracellular free fatty acids was observed to explain this, and under the conditions of the incubations it is unlikely that Pi availability was rate limiting. The exact locus of inhibition is unknown.