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

Abstract

13 C Nuclear Magnetic Resonance (NMR) studies of rodent and human brain using [1-13 C]/[1,6-13 C 2 ]glucose as labeled substrate have consistently found a lower enrichment (B25% to 30%) of glutamine-C4 compared with glutamate-C4 at isotopic steady state. The source of this isotope dilution has not been established experimentally but may potentially arise either from blood/brain exchange of glutamine or from metabolism of unlabeled substrates in astrocytes, where glutamine synthesis occurs. In this study, the contribution of the former was evaluated ex vivo using 1 H-[ 13 C]-NMR spectroscopy together with intravenous infusion of [U-13 C 5 ]glutamine for 3, 15, 30, and 60 minutes in mice. 13 C labeling of brain glutamine was found to be saturated at plasma glutamine levels 41.0 mmol/L. Fitting a blood-astrocyte-neuron metabolic model to the 13 C enrichment time courses of glutamate and glutamine yielded the value of glutamine influx, V Gln(in) , 0.036±0.002 mmol/g per minute for plasma glutamine of 1.8 mmol/L. For physiologic plasma glutamine level (B0.6 mmol/L), V Gln(in) would be B0.010 mmol/g per minute, which corresponds to B6% of the glutamine synthesis rate and rises to B11% for saturating blood glutamine concentrations. Thus, glutamine influx from blood contributes at most B20% to the dilution of astroglial glutamine-C4 consistently seen in metabolic studies using [1-13 C]glucose. Keywords: glutamate; neurotransmitter; nuclear magnetic resonance spectroscopy INTRODUCTION Glucose, the primary source of energy in the mature brain, is mainly oxidized in neurons to support neuronal firing, and the release and recycling of neurotransmitters, glutamate, and GABA, through the neuron-astrocyte glutamate-glutamine and GABAglutamine cycles. 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.A consistent finding in NMR studies of rodent and human brain metabolism using [1-13 C] or [1,6-13 C 2 ]glucose as precursor is the lower fractional 13 C enrichment of brain glutamate and glutamine at isotopic steady state compared with blood glucose, 3-5 revealing the presence of constant 'dilutional' flows of unlabeled substrates into the respective tricarboxylic acid (TCA) cycles of neurons and astroglia. For glutamate, this dilution amounts to 25% to 30% in rodents 5,6 (B14% in human brain 7 ) of the theoretical maximum expected from the 13 C-labeled glucose, and is thought to arise within neurons mainly from metabolism of blood-borne substrates such as lactate or ketone bodies and within the brain from glutamine produced in astrocytes through the glutamate-glutamine cycle. [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...

Fluorination at the 4 position alters the substrate behavior of l-glutamine and l-glutamate: Implications for positron emission tomography of neoplasias

“…These factors, together with the relatively small amounts of unlabeled [2 S ,4 S ]-4-FGlu [2 S ,4 R ]-4-FGlu expected to be present in the blood after injection of [ 18 F][2 S ,4 S ]-4-FGln and [ 18 F][2 S ,4 R ]-4-FGln [15–17], suggest that uptake of [2 S ,4 S ]-4-FGlu and [2 S ,4 R ]-4-FGlu into the brain will be mimimal. On the other hand, circulating glutamine has been shown to enter the mouse and human brain [52]. However, this uptake contributes maximally 20% to the brain astrocytic glutamine pool [52].…”

Brain interstitial fluid glutamine homeostasis is controlled by blood–brain barrier SLC7A5/LAT1 amino acid transporter

“…13 C5]-Gln infusion did not increase total cortical Gln and Gln-C4 13 C enrichment was &2%, 38 while it cannot be ruled out that hGln is efficiently transendothelially transported and subsequently rapidly sequestered in brain cells (e.g. astrocytes) and therefore not detected in ISF.…”

How to measure metabolic fluxes: a taxonomic guide for 13 C fluxomics

“…Recently, the understanding of the distribution and exchange of tracer between glial cells and neurons (GABAergic, glutamatergic) has been significantly advanced [46,47,48 ] (expertly reviewed in [49]). The INST models are focused on glycolysis, citric acid cycle, glutamine/glutamate metabolism and the intercellular nutrient exchange.…”

“…INST [48] INST [46] PWA [59] PWA [60] IST [56] INST [62] IST [64] INST [63] [9] FR [50] Single Cell FR/KFP [68] [53]…”