[1-C-13] GLUCOSE-METABOLISM IN RAT CEREBELLAR GRANULE CELLS AND ASTROCYTES IN PRIMARY CULTURE - EVALUATION OF FLUX PARAMETERS BY C-13-NMR AND H-1-NMR SPECTROSCOPY

The metabolism of [1-C-13]glucose in rat cerebellum astrocytes and granule cells was investigated using C-13- and H-1-NMR spectroscopy. Near homogeneous primary cultures of each cell type were incubated with [1-C-13]glucose, under the same conditions. Analysing the relative C-13 enrichments of metabolites in spectra of cell perchloric acid extracts, on the one hand, the C-13-H-1 spin-coupling patterns in H-1-NMR spectra of cell medium lactate and the C-13-C-13 spin-coupling patterns in C-13-NMR spectra of purified cell glutamate, on the other hand, showed significant differences, between the two cell types, in the activity of various metabolic ways. First, the carbon flux through the oxidative branch of the hexose monophosphate shunt, which leads to unenriched lactate, was found higher in granule cells than in astrocytes. Second, although the specific C-13 enrichment of lactate was higher in astrocytes than in granule cells, the fraction of C-13-enriched acetyl-CoA entering the citric acid cycle was more than twice as high in granule cells as in astrocytes. Lactate C3 and acetyl-CoA C2 enrichments were very similar in granule cells, whereas acetyl-CoA C2 enrichment was 60% lower than that of lactate C3 in astrocytes. These results can be explained by the fact that granule cells used almost exclusively the exogenous glucose to fuel the citric acid cycle, whereas astrocytes used concomitantly glucose and other carbon sources. Last, in the case of granule cells, glutamate C2 and C3 enrichments were equivalent; the carbon flux through the pyruvate carboxylase route was evaluated to be around 15% of the carbon flux through the citrate synthetase route. In astrocytes, glutamate C2 enrichment was higher than that of C3, which could be explained by a pyruvate carboxylase activity much more active in these cells than in granule cells.