Close Coupling between Astrocytic and Neuronal Metabolisms to Fulfill Anaplerotic and Energy Needs in the Rat Brain

Abstract

Carbon metabolism in the rat brain was studied in animals anesthetized with a light dose of pentobarbital and in awake animals under morphine, which were infused with either [1-13 C]glucose + acetate or glucose + [2-13 C]acetate for various periods of time. Brain amino-acid enrichments in tissue extracts were determined by nuclear magnetic resonance (NMR) spectroscopy and their time evolution was analyzed by automatic fitting. Acetyl-coenzyme A C2 enrichment and ratio between pyruvate carboxylase and pyruvate dehydrogenase activity (PC/PDH) were determined from glutamate and glutamine labeling. The following results were obtained: (i) amino-acid enrichment patterns implied metabolic compartmentation and occurrence of the glutamate-glutamine cycle; (ii) kinetics of aspartate, GABA, and glutamate labeling from [1-13 C]glucose and of glutamine labeling from [2-13 C]acetate indicated a twofold higher metabolic activity in awake than in anesthetized rat brain; (iii) evaluation of the contributions of the astrocytic and neuronal metabolisms to glutamine synthesis in both groups of rats indicated a coupling between neuronal tricarboxylic acid (TCA) cycle, glutamate-glutamine cycle and glial TCA cycle; and (iv) analyzing the extrapolations back to time zero and the steady-state values of PC/PDH indicated a close coupling between PC activity and both astrocytic and neuronal TCA cycles. All these results suggest a cooperative-like behavior of astrocytic and neuronal metabolisms to fulfill the anabolic and energy needs linked to brain activation.

Characterization of Cerebral Glutamine Uptake from Blood in the Mouse Brain: Implications for Metabolic Modeling of13C NMR Data

“…In addition, mice (n ¼ 4) were also infused for 60 minutes with rates adjusted such that infusion rates at tX8 minutes were 20, 37, 59, and 74 mmol/kg per minute, to evaluate the steady-state brain glutamine labeling at different levels of plasma glutamine. Blood was collected from the retro-orbital sinus just before the conclusion of infusion, centrifuged at high speed, and the supernatant (plasma) collected for the measurement of concentration and 13 C enrichment of glutamine in plasma. The head was frozen in situ with liquid nitrogen and stored at À 801C until further processing.…”

“…The metabolic pathways comprising these cycles have been revealed in vivo by 13 C Nuclear Magnetic Resonance (NMR) spectroscopy combined with infusions of 13 C-labeled substrates, 1,2 permitting detailed investigation of the metabolism of glucose and alternative substrates in the brain.…”

“…8,14,15 Models of brain metabolism used to estimate fluxes from 13 C time courses have dealt with this problem by either inclusion of a dilution flux (V dil(Gln) , V ex , or V dil ) at the level of glutamine 5,7,12,13,[16][17][18] or astroglial acetyl-CoA 6 derived from nonlabeled precursors. The presence of the glutamine-C4 dilution, and its incorporation into the metabolic model was recently shown to be critical for the accurate determination of the glutamate-glutamine cycle rate in NMR studies using [1][2][3][4][5][6][7][8][9][10][11][12][13] C]/ [1,6-13 C 2 ]glucose. 15 Thus, it is critical to better define the source of the astroglial glutamine dilution, allowing refinement of the metabolic models to estimate cerebral fluxes in 13 C-labeling studies, while exploring alternate 13 C-labeled substrates that might be exploited to study astrocytic and neuronal function in vivo.…”

“…The presence of the glutamine-C4 dilution, and its incorporation into the metabolic model was recently shown to be critical for the accurate determination of the glutamate-glutamine cycle rate in NMR studies using [1][2][3][4][5][6][7][8][9][10][11][12][13] C]/ [1,6-13 C 2 ]glucose. 15 Thus, it is critical to better define the source of the astroglial glutamine dilution, allowing refinement of the metabolic models to estimate cerebral fluxes in 13 C-labeling studies, while exploring alternate 13 C-labeled substrates that might be exploited to study astrocytic and neuronal function in vivo.…”

“…[8][9][10] In addition to the glutamate-C4 dilution (relative to glucose-C1), 13 C enrichment of glutamine-C4 is further B20% to 30% less than that of glutamate-C4 at isotopic steady state. 4,5,[11][12][13] As the majority of glutamine-C4 labeling in these experiments …”