In vivo (13)C NMR measurements of cerebral glutamine synthesis as evidence for glutamate–glutamine cycling

The cerebral tricarboxylic acid (TCA) cycle rate and the rate of glutamine synthesis were measured in rats in vivo under normal physiological and hyperammonemic conditions using (13)C NMR spectroscopy. In the hyperammonemic animals, blood ammonia levels were raised from control values of ≈0.05 mM to ≈0.35 mM by an intravenous ammonium acetate infusion. Once a steady-state of cerebral metabolites was established, a [1-(13)C]glucose infusion was initiated, and (13)C NMR spectra acquired continuously on a 7- tesla spectrometer to monitor (13)C labeling of cerebral metabolites. The time courses of glutamate and glutamine C-4 labeling were fitted to a mathematical model to yield TCA cycle rate (V(TCA)) and the flux from glutamate to glutamine through the glutamine synthetase pathway (V(gln)). Under hyperammonemia the value of V(TCA) was 0.57 ± 0.16 μmol/min per g (mean ± SD, n = 6) and was not significantly different (unpaired t test; P > 0.10) from that measured in the control animals (0.46 ± 0.12 μmol/min per g, n = 5). Therefore, the TCA cycle rate was not significantly altered by hyperammonemia. The measured rate of glutamine synthesis under hyperammonemia was 0.43 ± 0.14 μmol/min per g (mean ± SD, n = 6), which was significantly higher (unpaired t test; P < 0.01) than that measured in the control group (0.21 ± 0.04 μmol/min per g, n = 5). We propose that the majority of the glutamine synthetase flux under normal physiological conditions results from neurotransmitter substrate cycling between neurons and glia. Under hyperammonemia the observed increase in glutamine synthesis is comparable to the expected increase in ammonia transport into the brain and reported measurements of glutamine efflux under such conditions. Thus, under conditions of elevated plasma ammonia an increase in the rate of glutamine synthesis occurs as a means of ammonia detoxification, and this is superimposed on the constant rate of neurotransmitter cycling through glutamine synthetase.