31P and 13C NMR spectroscopic study of wild type and multidrug resistant Ehrlich ascites tumor cells

Steady-state 31P NMR spectra of wild type EHR2 Ehrlich ascites tumor cells and multidrug resistant EHR2/DNR+ cells, immobilized in agarose threads, and continuously perfused with medium, showed temperature-dependent differences in the levels of intracellular phosphate metabolites. At 37 degrees C, the EHR2/DNR+ cells contained four times more phosphocreatine (PCr) than the EHR2 cells. At 20 degrees C, the EHR2 cells contained 80% more of phosphodiesters (PDE), the levels of PCr being equal. The quantitative metabolite level data are based on T1 relaxation times data and are normalized for the protein content of the cells. Perfusion of the cells with azide, an inhibitor of mitochondrial respiration, had no effect on the ATP level, and caused no changes in glucose consumption and lactate production. Azide perfusion, combined with glucose depletion, caused rapid drop in the ATP content, which was reestablished after renewed perfusion with glucose. Similarly, perfusion with 2,4-dinitrophenol, an uncoupler of the respiration chain, had no effect on the phosphate metabolites. These results demonstrate that aerobic glycolysis is the main route by which glucose is metabolised under the conditions used (glucose concentration in medium 2 g/L). Rates of uptake and phosphorylation of 2-deoxy-D-glucose were measured by following the formation of intracellular [6-13C]2-deoxy-D-glucose-6-phosphate by 13C NMR; at 37 degrees C the observed rates for EHR2 and EHR2/DNR+ cells were equal, about 10 nmol/(min x mg protein), whereas at 20 degrees C the wild type cells produced the 6-phosphate at an approximately twice the rate found for the resistant cells [about 4 and 2 nmol/(min x mg protein), respectively].(ABSTRACT TRUNCATED AT 250 WORDS)