Quantitative Assessment of Neurometabolism by Deuterium and Proton NMR Spectroscopy Using 6,6′- 2 H 2 Glucose

Neurometabolic analysis is routinely performed using 13 C NMR/MRS in conjunction with the administration of 13 C-labeled tracers. Despite its widespread use, the low sensitivity of 13 C NMR necessitates long acquisition times. To address this limitation, the present study introduces an indirect approach that uses a single 1 H NMR acquisition to measure deuterium labeling by exploiting labeling at specific sites, thereby providing a novel and more efficient strategy for assessing neurometabolism. In this study, male SD rats were infused with 6,6’- 2 H 2 glucose for 10–90 min, and the 2 H labeling of cortical metabolites was measured ex vivo using 2 H NMR spectroscopy. After 90 min of 6,6′- 2 H 2 glucose infusion, resonances corresponding to 3- 2 H 2 lactate, 4- 2 H x glutamate, 4- 2 H x glutamine, and 3- 2 Haspartate (∼2.60 ppm) were detected. In contrast, signals corresponding to 2- 2 Haspartate (∼3.90 ppm), 2- 2 Hglutamate (∼3.76 ppm), and 3- 2 Hglutamate (∼2.09 ppm) were not detected, even after prolonged infusion, suggesting complete loss of the 2 H label during the second turn of the tricarboxylic acid cycle. The cerebral metabolic rate of glucose oxidation, estimated by fitting an exponentially saturating curve to the pooled labeling of 4- 2 H x glutamate, 4- 2 H x glutamine, and 3- 2 Haspartate, was 0.389 ± 0.031 μmol/g/min. The 4- 2 H x glutamate levels were also derived by subtracting the 4- 1 H 2 glutamate resonance intensity from total 4- 1 H 2 glutamate intensity estimated using the 3- 1 H 2 glutamate intensity in unedited 1 H NMR spectra. The concentrations of 4- 2 H x glutamate determined from 1 H NMR spectra were comparable to those determined from 2 H NMR. Moreover, when applied to 5xFAD mice (2.53 ± 0.43 μmol/g, n = 7), a model of Alzheimer’s disease, this indirect approach revealed significantly reduced ( p = 0.028) 4- 2 H x glutamate levels compared with controls (2.99 ± 0.15 μmol/g, n = 7). This decrease was comparable to that observed with the direct 2 H NMR measurements and was consistent with findings from 13 C tracer-based studies.