Depletion of red cell ATP by incubation with 2-deoxyglucose

In order to devise a more physiologic system for measuring depletion of red cell ATP levels, the effect of incubating human erythrocytes with 2-deoxyglucose has been investigated. ATP depletion proceeds very slowly at a 20 mM concentration of 2-deoxyglucose, a level which exceeds the Km of hexokinase for this substrate by more than 10-fold. However, at 160 mM concentration of 2-deoxyglucose, ATP depletion proceeds sufficiently rapidly that nearly 90% of ATP has disappeared from the red cell after 2 1/2 hr of incubation. To explain this observation, a number of additional studies were carried out. It was found that 2-deoxyglucose penetrated rapidly into red cells. Phosphorylation of 2-deoxyglucose in red cells was inhibited by both products of the 2-deoxyglucose-phosphorylating reaction, namely, 2-deoxyglucose-6-phosphate and ADP. Inhibition of 2-deoxyglucose phosphorylation was diminished at higher-than-physiologic pH levels. Red cells may be relatively rapidly depleted of ATP by incubation with 100 mM 2-deoxyglucose in a saline-phosphate-buffered medium, pH 7.8. In such rapidly depleted cells, the morphologic changes which formerly were attributed to ATP depletion do not occur.