To determine the relationship between cerebral Glc metabolism and glutamatergic neuronal function, we used 13 C NMR spectroscopy to measure, simultaneously, the rates of the tricarboxylic acid cycle and Gln synthesis in the rat cortex in vivo. From these measurements, we calculated the rates of oxidative Glc metabolism and glutamate-neurotransmitter cycling between neurons and astrocytes (a quantitative measure of glutamatergic neuronal activity). By measuring the rates of the tricarboxylic acid cycle and Gln synthesis over a range of synaptic activity, we have determined the stoichiometry between oxidative Glc metabolism and glutamateneurotransmitter cycling in the cortex to be close to 1:1. This finding indicates that the majority of cortical energy production supports functional (synaptic) glutamatergic neuronal activity. Another implication of this result is that brain activation studies, which map cortical oxidative Glc metabolism, provide a quantitative measure of synaptic glutamate release.Glc metabolism is the major pathway of energy production in the mature brain (1). During brain activation, increases in Glc metabolism directly form the basis of brain functional mapping by using both 2-deoxyglucose autoradiography (2, 3) and positron-emission tomography (4) and indirectly influence signal changes observed with functional MRI (5). Despite the extensive use of these methods for mapping brain function, the mechanism linking Glc metabolism and functional neuronal activity and the fraction of cerebral energy production that supports neuronal function are still unknown.Glutamate is the major excitatory neurotransmitter in the brain (6), and a high percentage of cortical neurons are glutamatergic (7). It has been proposed that a neuronalastrocytic neurotransmitter cycle exists in the brain in which glutamate from the neuronal pool is released into the synaptic cleft as a neurotransmitter, taken up by astrocytes, converted to Gln, and returned to the neuron in this synaptically inactive form where it is converted back to glutamate (6). The development of in vivo 13 C NMR spectroscopy has enabled the direct investigation of cerebral glutamate metabolism (8, 9). We recently have shown that the rate of glutamateneurotransmitter cycling between neurons and astrocytes can be calculated by using the flux of the 13 C label from glutamate to Gln in the rat brain in vivo during a [1-13 C]Glc infusion (10). Thus, we can obtain an in vivo measure of glutamatergic neuronal activity. In the same experiment, the flux of the 13 C label from [1-13 C]Glc into glutamate yields a simultaneous in vivo measurement of the cerebral tricarboxylic acid (TCA) cycle rate, from which oxidative Glc consumption can be derived (11,12). Therefore, by using the combined measurement of these two fluxes, we can determine quantitatively the stoichiometry between cerebral Glc metabolism and glutamatergic-synaptic activity in vivo.In the present study, we have used direct 13 C NMR spectroscopy to determine the cerebral (primarily cortical)...

The study replicates the findings of decreased GABA concentrations in the occipital cortex of subjects with MDD. It also demonstrates that there is a change in the ratio of excitatory-inhibitory neurotransmitter levels in the cortex of depressed subjects that may be related to altered brain function. Last, the combined data set suggests that magnetic resonance spectroscopy GABA measures may serve as a biological marker for a subtype of MDD.

Recent 13 C NMR studies in rat models have shown that the glutamate͞glutamine cycle is highly active in the cerebral cortex and is coupled to incremental glucose oxidation in an Ϸ1:1 stoichiometry. To determine whether a high level of glutamatergic activity is present in human cortex, the rates of the tricarboxylic acid cycle, glutamine synthesis, and the glutamate͞glutamine cycle were determined in the human occipital͞parietal lobe at rest. During an infusion of [1-13 C]-glucose, in vivo 13 C NMR spectra were obtained of the time courses of label incorporation into [4-13 C]-glutamate and [4-13 C]-glutamine. Using a metabolic model we have validated in the rat, we calculated a total tricarboxylic acid cycle rate of 0.77 ؎ 0.07 mol͞min͞g (mean ؎ SD, n ‫؍‬ 6), a glucose oxidation rate of 0.39 ؎ 0.04 mol͞min͞g, and a glutamate͞ glutamine cycle rate of 0.32 ؎ 0.05 mol͞min͞g (mean ؎ SD, n ‫؍‬ 6). In agreement with studies in rat cerebral cortex, the glutamate͞glutamine cycle is a major metabolic f lux in the resting human brain with a rate Ϸ80% of glucose oxidation.The regulation of the release and re-uptake of the excitatory neurotransmitter glutamate is critical for mammalian brain function (1, 2). Glutamate released from the neuron may be cleared from the synaptic cleft through uptake by neuronal or glial glutamate transporters (3, 4). Recent studies have supported the glia as the major pathway of glutamate clearance (3). Neurons lack the enzymes necessary to perform net glutamate synthesis and depend on the glia to supply precursors. One of the pathways proposed for neuronal glutamate repletion is the glutamate͞glutamine cycle (5-8). In this pathway, glutamate taken up by the glia is converted to glutamine by glutamine synthetase (9-11). Glutamine then is released to the extracellular fluid, where it is taken up by neurons and is converted back to glutamate by the action of phosphate-activated glutaminase (12).The rate of the glutamate͞glutamine cycle has been controversial because of difficulties in performing measurements in the living brain. The prevailing belief has been that the glutamate͞glutamine cycle is a minor metabolic flux relative to total cellular glutamate metabolism. This view is largely based on the small size of the vesicular glutamate pool compared with other cellular glutamate pools (13,14). Additional evidence comes from the low flux of isotope from [1-13 C] glucose into glutamine in studies of brain slices (15).We have demonstrated that in vivo 13 C NMR may be used to measure the rate of glutamine labeling (16, 17) from [1-13 C] glucose in human occipital͞parietal cortex. These and subsequent studies (18) demonstrated that, in contrast with results from nonactivated brain slices (15), glutamine labeling is rapid. However, the rate of the glutamate͞glutamine cycle was not uniquely determined from these first experiments because of the inability to distinguish the glutamate͞glutamine cycle from other sources of glutamine labeling. The major alternate pathway of brain glutamine metaboli...

This study provides the first evidence of abnormally low cortical GABA concentrations in the brains of depressed patients.

Increasing evidence supports a crucial role for glial metabolism in maintaining proper synaptic function and in the etiology of neurological disease. However, the study of glial metabolism in humans has been hampered by the lack of noninvasive methods. To specifically measure the contribution of astroglia to brain energy metabolism in humans, we used a novel noninvasive nuclear magnetic resonance spectroscopic approach. We measured carbon 13 incorporation into brain glutamate and glutamine in eight volunteers during an intravenous infusion of [2-13C] acetate, which has been shown in animal models to be metabolized specifically in astroglia. Mathematical modeling of the three established pathways for neurotransmitter glutamate repletion indicates that the glutamate/glutamine neurotransmitter cycle between astroglia and neurons (0.32 +/- 0.07 micromol x gm(-1) x min(-1)) is the major pathway for neuronal glutamate repletion and that the astroglial TCA cycle flux (0.14 +/- 0.06 micromol x gm(-1) x min(-1)) accounts for approximately 14% of brain oxygen consumption. Up to 30% of the glutamine transferred to the neurons by the cycle may derive from replacement of oxidized glutamate by anaplerosis. The further application of this approach could potentially enlighten the role of astroglia in supporting brain glutamatergic activity and in neurological and psychiatric disease.

Previous studies have shown that the glutamate͞glutamine (Glu͞ Gln) neurotransmitter cycle and neuronal glucose oxidation are proportional (1:1), with increasing neuronal activity above isoelectricity. GABA, a product of Glu metabolism, is synthesized from astroglial Gln and contributes to total Glu͞Gln neurotransmitter cycling, although the fraction contributed by GABA is unknown. In the present study, we used 13 C NMR spectroscopy together with i.v. infusions of [1,6-13 C2]glucose and [2-13 C]acetate to separately determine rates of Glu͞Gln and GABA͞Gln cycling and their respective tricarboxylic acid cycles in the rat cortex under conditions of halothane anesthesia and pentobarbital-induced isoelectricity. Under 1% halothane anesthesia, GABA͞Gln cycle flux comprised 23% of total (Glu plus GABA) neurotransmitter cycling and 18% of total neuronal tricarboxylic acid cycle flux. In isoelectric cortex, glucose oxidation was reduced >3-fold in glutamatergic and GABAergic neurons, and neurotransmitter cycling was below detection. Hence, in both cell types, the primary energetic costs are associated with neurotransmission, which increase together as cortical activity is increased. The contribution of GABAergic neurons and inhibition to cortical energy metabolism has broad implications for the interpretation of functional imaging signals.GABAergic neurons ͉ glutamatergic neurons ͉ magnetic resonance spectroscopy ͉ neuronal-glial interactions G lutamate (Glu) and GABA are the major excitatory and inhibitory neurotransmitters in the mature cerebral cortex and together account for Ϸ90% of total cortical synapses (1). Excitatory synapses outnumber inhibitory synapses Ϸ5:1 (2), suggesting that excitation plays an energetically dominant role in the cortex. The energetic cost of GABAergic neurotransmission remains an open question, however, because current methods used to assess cortical activity are based on changes in local blood f low or metabolism (e.g., glucose or oxygen consumption), which cannot differentiate glutamatergic from GABAergic neurons. Thus, the interpretation of changes in cortical activity in terms of the energetics of excitation and inhibition requires other methods that are sensitive to the synthesis of these neurotransmitters.Glutamatergic neurotransmitter cycling flux and energy consumption have been reported for rat and human cortex by using 13 C NMR spectroscopy during the i.v. infusion of 13 C-labeled glucose (3-10). These studies show that the Glu͞glutamine (Gln) cycling flux (V cyc ) is substantial, from Ϸ30% to 42% of neuronal tricarboxylic acid (TCA) cycle flux in anesthetized rats (V TCAn ) (4, 5, 8) to Ϸ38-50% of V TCAn in resting awake rat and human cortex (7, 9, 10). In the cortex of anesthetized rats, changes in V cyc and V TCAn are proportional (Ϸ1:1) over a large cortical activity range above isoelectricity (4,8).The determination of V cyc from Gln and Glu 13 C turnover, using [1-13 C]glucose as tracer, includes contributions from GABA as well as Glu (4,5,11). This is because Gln is a com...

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 Ϸ0.05 mM to Ϸ0.35 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 f lux from glutamate to glutamine through the glutamine synthetase pathway (V gln ). Under hyperammonemia the value of V TCA was 0.57 ؎ 0.16 mol͞min 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 mol͞min 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 mol͞min 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 mol͞ min per g, n ‫؍‬ 5). We propose that the majority of the glutamine synthetase f lux 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 eff lux 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.Glutamine synthetase in the brain is predominantly an astrocytic enzyme that catalyzes the formation of glutamine from glutamate and ammonia. It has been suggested that this pathway forms part of a neurotransmitter cycle between neurons and glia (1). In this cycle, astrocytes take up neurotransmitter glutamate from the synaptic cleft, convert it to glutamine via glutamine synthetase, and release this glutamine into the extracellular space for uptake by neurons, where it is converted back to glutamate primarily by glutaminase. ␥-Aminobutyric acid (GABA) released from inhibitory neurons also may be taken up by astrocytes, converted to glutamate, and recycled to neruons in the same way. The function of such a cycle would be to remove the synaptically active metabolite from the synaptic cleft and to recycle the carbon skeletons back to the neuronal pool by a synaptically inactive form.While ammonia is a normal constituent of tissue, elevated concentrations of blood and brain ammonia have been found to interfere with cerebral energy metabolism and may redu...

Summary: 13C isotopic tracer data previously obtained by 13C nuclear magnetic resonance in the human brain in vivo were analyzed using a mathematical model to deter mine metabolic rates in a region of the human neocortex. The tricarboxylic acid (TCA) cycle rate was 0.73 ± 0.19 fLmol min -I g-I (mean ± SD; n = 4). The standard deviation reflects primarily intersubject variation, since individual uncertainties were low. The rate of a-ketoglu tarate/glutamate exchange was 57 ± 26 fLmol min -1 g -1 (n = 3), which is much greater than the TCA cycle rate; the high rate indicates that a-ketoglutarate and glutamate are in rapid exchange and can be treated as a single com bined kinetic pool. The rate of synthesis of glutamine from glutamate was 0.47 fLmol min -I g-I (n = 4), withThe stable isotope 13C combined with nuclear magnetic resonance (NMR) spectroscopy allows