Quantitative assessment of brain glucose metabolic rates using in vivo deuterium magnetic resonance spectroscopy

Abstract

Quantitative assessment of cerebral glucose consumption rate (CMR_glc) and tricarboxylic acid cycle flux (V_TCA) is crucial for understanding neuroenergetics under physiopathological conditions. In this study, we report a novel in vivo Deuterium (²H) MRS (DMRS) approach for simultaneously measuring and quantifying CMR_glc and V_TCA in rat brains at 16.4 Tesla. Following a brief infusion of deuterated glucose, dynamic changes of isotope-labeled glucose, glutamate/glutamine (Glx) and water contents in the brain can be robustly monitored from their well-resolved ²H resonances. Dynamic DMRS glucose and Glx data were employed to determine CMR_glc and V_TCA concurrently. To test the sensitivity of this method in response to altered glucose metabolism, two brain conditions with different anesthetics were investigated. Increased CMR_glc (0.46 vs. 0.28 µmol/g/min) and V_TCA (0.96 vs. 0.6 µmol/g/min) were found in rats under morphine as compared to deeper anesthesia using 2% isoflurane. This study demonstrates the feasibility and new utility of the in vivo DMRS approach to assess cerebral glucose metabolic rates at high/ultrahigh field. It provides an alternative MRS tool for in vivo study of metabolic coupling relationship between aerobic and anaerobic glucose metabolisms in brain under physiopathological states.

Keywords: Brain glucose metabolisms; TCA cycle; cerebral metabolic rate of glucose; deuterium magnetic resonance spectroscopy (2H MRS); glycolysis.

Figure 1.

The ²H-labeling scheme in the brain tissue for dynamic DMRS application using D-Glucose-6,6-d₂ (d66) as an isotopic tracer. Labeling firstly incorporates into pyruvate pool to form [3,3-d₂] Pyruvate through glycolysis, which is then converted to [3,3-d₂] lactate catalyzed by lactate dehydrogenase (LDH). [3,3-d₂] Pyruvate can also be transported into mitochondria and transformed into [2,2-d₂] Acetyl-CoA via pyruvate decarboxylation by pyruvate dehydrogenase (PDH). By entering the TCA cycle, intermediates of [4-d] or [4,4-d₂] Citrate and [4-d] or [4,4-d₂] α-ketoglutarate will be produced, which could exchange with glutamate to generate [4-d] or [4,4-d₂] glutamate. During the following steps of TCA cycle, ²H-labeling may depart from the cycle and exchange with the proton(s) in water molecule to become deuterated water. ‘*’: Pools labeled with ²H; black square boxes: pools to be detected by in vivo DMRS.

Figure 2.

Simplified kinetic modeling. Symbols: Glc: glucose; Gly: glycogen; L: combined pool for Pyr (pyruvate) and Lac (lactate); K: α-KG, α-ketoglutarate; Glx: combined pool for Glu (glutamate) and Gln (glutamine). V_x stands for the α-KG/Glx exchange rate. V_y is the glycogen synthetic rate. V_out represents an efflux of lactate. ‘*’: ²H-labeled metabolites.

Figure 3.

Representative original (black trace in upper row and grey trace in bottom row) and fitted (red trace in bottom row) DMR spectra obtained from deuterated glucose (d66) phantom solution (a) and a rat brain under constant morphine sulfate infusion pre- (b), 5 min (c), 30 min (d), 60 min (e) and 120 min (f) post-deuterated glucose (d66) infusion. Each in vivo spectrum displayed in this figure was summed from 1 min of data acquisitions (four spectra). ²H resonance assignments: (1) water (4.8 ppm, set as a chemical shift reference); (2) glucose (3.8 ppm); (3) Glx (2.4 ppm); and (4) lactate (1.4 ppm).

Figure 4.

Dynamic changes and time courses of deuterated brain glucose (d66) and labeled Glx concentrations during sequential DMRS acquisitions (15 s temporal resolution) in two representative rat brains under 2% isoflurane anesthesia vs. constant morphine sulfate infusion, respectively. Time = 0, starting time point for model fitting.

Figure 5.

Fitting of both glucose (d66, blue circles) and Glx (green circles) labeling curves with kinetic modeling for representative rats under 2% isoflurane anesthesia (a) vs. constant morphine sulfate infusion (b). Solid lines are the model fittings of labeled glucose (red) and Glx (black) changes. Starting time point (0 min) for model fitting corresponds to 5 min after ending of the d66 infusion.