Glutamate metabolism and transport in rat brain mitochondria

Dennis, Steven C.; Land, John M.; Clark, John B.

doi:10.1042/bj1560323

Cited by 56 publications

(40 citation statements)

References 30 publications

(34 reference statements)

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“…Furthermore, the extremely high specific activity of the aspartate formed from either IJC-or l3C-labelled or [l5N]glutamate [28,30,55,56] is consistent with the high affinity of astrocytic AAT for both gluta mate and the oxaloacetate formed from glutamate metabolism. In addition, these observations are consis tent with the important roles of intramitochondrial aspar tate in the transport of glutamate across the mitochon drial membrane [50] and in maintaining the activity of the malate-aspartate shuttle in astrocytes [5], Thus, it is difficult to argue against the strong evidence for an important role of AAT in glutamate metabolism in astro cytes. particularly when the concentration of glutamate is quite low.…”

Section: Discussionmentioning

confidence: 96%

New Insights into the Compartmentation of Glutamate and Glutamine in Cultured Rat Brain Astrocytes

Mc¹,

Jt²,

Jh³

et al. 1996

Dev Neurosci

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Studies from several groups have provided evidence that glutamate and glutamine are metabolized in different compartments in astrocytes. In the present study we measured the rates of ¹⁴C0₂ production from U-[¹⁴C]glutamate and U-[¹⁴C]glutamine, and utilized both substrate competition experiments and the transaminase inhibitor aminooxyacetic acid (AOAA) to obtain more information about the compartmentation of these substrates in cultured rat brain astrocytes. The rates of oxidation of 1 mM glutamine and glutamate were 26.4±1.4 and 63.0 ±7.4 nmol/h/mg protein, respectively. The addition of 1 mM glutamate decreased the rate of oxidation of glutamine to 26.3 % of the control rate, demonstrating that glutamate can effectively compete with the oxidation of glutamine by astrocytes. In contrast, the addition of 1 mM glutamine had little or no effect on the rate of oxidation of glutamate by astrocytes, demonstrating that the glutamate produced intracellularly from exogenous glutamine does not dilute the glutamate taken up from the media. The addition of 5 mM AOAA decreased the rate of ¹⁴C0₂ production from glutamine to 29.2% of the control rate, consistent with earlier studies by our group. The addition of 5 mM AOAA decreased the rate of oxidation of concentrations of glutamate ≤0.1 mM by ∼50%, but decreased the oxidation of 0.5–1 mM glutamate by only ∼20%, demonstrating that a substantial portion of glutamate enters the tricarboxylic acid (TCA) cycle via glutamate dehydrogenase (GDH) rather than transamination, and that as the concentration of glutamate increases the relative proportion entering the TCA cycle via GDH also increases. To determine if the presence of an amino group acceptor (i.e. a ketoacid) would increase the rate of metabolism of glutamate, pyruvate was added in some experiments. Addition of 1 mM pyruvate increased the rate of oxidation of glutamate, and the increase was inhibited by AOAA, consistent with enhanced entry of glutamate into the TCA cycle via transamination in the presence of pyruvate. Enzymatic studies showed that pyruvate increased the activity of mitochondrial aspartate aminotransferase (AAT). Overall, the data demonstrate that glutamate formed intracellularly from glutamine enters the TCA cycle primarily via transamination, but does not enter the same TCA cycle compartment as glutamate taken up from the extracellular milieu. In contrast, extracellular glutamate enters the TCA cycle in astrocytes via both transamination and GDH, and can compete with, or dilute, the oxidation of glutamate produced intracellularly from glutamine.

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Section: Discussionmentioning

confidence: 96%

New Insights into the Compartmentation of Glutamate and Glutamine in Cultured Rat Brain Astrocytes

Mc¹,

Jt²,

Jh³

et al. 1996

Dev Neurosci

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“…Consistent with the fact that glutamate also exists in nontransmitter pools, for example, as a participant in intermediary metabolism (Fonnum, 1984), glutamate labeling was also observed in several other subcellular compartments. For example, glutamate is transported into mitochondria (Dennis et al, 1976;Minn and Gayet, 1977), the primary site of glutamate synthesis by phosphate-activated glutaminase (Salganicoff and DeRobertis, 1965;Erecihska and Silver, 1990); accordingly, mitochondrial profiles were moderately to heavily labeled. Astrocytes, which actively accumulate glutamate (Drejer et al, 1982;Fonnum, 1984), possessed lower intensities of glutamate labeling than axon terminals forming asymmetric synapses (Ottersen et al, 1992 and references therein).…”

Section: Discussionmentioning

confidence: 99%

3-Hydroxyanthranilic acid oxygenase-containing astrocytic processes surround glutamate-containing axon terminals in the rat striatum

Roberts

McCarthy

et al. 1995

J. Neurosci.

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Glutamate, the major transmitter of the corticostriatal pathway, is present in abundance in the striatum. 3-Hydroxyanthranilic acid oxygenase (3HAO) is the biosynthetic enzyme for quinolinic acid, an endogenous agonist of the NMDA glutamate receptor subtype and a potent neurotoxin. In order to explore the anatomical basis of possible functional interactions between glutamate and quinolinic acid in the rat striatum, pre- and postembedding immunocytochemical methods were used to localize 3HAO immunoreactivity (-i) and glutamate-i at the electron microscopic level. In accordance with previous light microscopic and biochemical studies, 3HAO-i was detected exclusively in astrocytes throughout the striatum. Notably, 3HAO-i was present in fine- caliber glial processes that often surrounded or abutted synaptic profiles, both asymmetric and symmetric. Glutamate-i was heavily deposited (3–13-fold higher gold particle density than tissue average) in axon terminals forming asymmetric synapses with spines and, occasionally, dendrites. In contrast, terminals forming symmetric synapses, dendrites, neuronal somata, and glial cells contained significantly less labeling than terminals forming asymmetric synapses. In double-labeled material, 3HAO-i was observed in glial processes that partially surrounded or were adjacent to glutamate-labeled terminals forming asymmetric synapses. 3HAO-labeled glial processes were also adjacent to unlabeled terminals forming symmetric synapses. Since quinolinic acid is known to enter the extracellular compartment readily, these results suggest that astrocytic quinolinic acid may participate in the regulation of glutamatergic neurotransmission in the rat striatum.

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“…For example, the observa tion that aminooxyacetic acid (AOAA, an inhibitor of pyridoxal phosphate-utilizing enzymes) inhibits gluta mate-dependent oxygen uptake by brain mitochondria [63,64] provides some support for the hypothesis that MAS may be instrumental in coupling glycolytic and TCA cycle fluxes and prompts us to investigate the hypo thesis further employing primary cultures of neurons and astrocytes.…”

Section: Role O F Malate-aspartate Shuttle: Studies With Inhibitors Omentioning

confidence: 99%

“…There is increasing evidence suggesting that, in brain, the malate-aspartate shuttle (MAS) is perhaps the most important shuttle mechanism responsible for the transfer of reducing equivalents across the inner mitochondrial membrane [31,63,64,67,68], Thus, the elucidation of the functional roles of MAS will shed some light on the functional coupling between glycolytic and TCA cycle fluxes.…”

Section: Role O F Malate-aspartate Shuttle: Studies With Inhibitors Omentioning

confidence: 99%

“…For exam ple, since PDHC is localized on the matrix side of the inner mitochondrial membrane, pyruvate derived from glycolysis has to be imported into the matrix space for metabolism by PDHC. The pyruvate transporter is also found in brain mitochondria and imports pyruvate (and also (3-hydroxybutyrate) from the cytosol into the matrix [34,41,62], ' Another reasonably well characterized transporter important in metabolic control is the glutamate/aspartate antiporter which imports glutamate into mito chondrial matrix in exchange for aspartate output [34,41,63,64], Redox States (NADH/NAD) and Phosphate Poten tial (ATP/ADP). The redox and phosphate potentials are important in the control of substrate oxidation, res piratory chain activities, and oxidative phosphorylation in brain as well as peripheral tissue mitochondria al though their precise regulatory roles in brain mitochon dria have yet to be fully documented [34,35].…”

Section: Regulation O F Mitochondrial Intermediary Metabolismmentioning

confidence: 99%

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Glycolysis-Citric Acid Cycle Interrelation: A New Approach and Some Insights in Cellular and Subcellular Compartmentation

Lai

Behar

1993

Dev Neurosci

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In the development of an integrated approach to study metabolic compartmentation and regulation in brain, we have emphasized the importance, versatility, and need to exploit the recent methodological advances in (1) NMR spectroscopy, (2) primary cultures of neurons and glial cells, and (3) subcellular fractionation (especially brain mitochondrial isolation). The integrated approach has the advantage of being able to draw data and inferences based on some combination of results derived from in vivo, cellular, and subcellular studies. For example, some in vivo NMR data may suggest that an enzymatic step may be rate-limiting in a particular pathway. This information may be used to frame testable hypotheses and questions that can be investigated in experiments involving primary cultures of neural cells and subcellular fractions. Subsequently, the data from such in vitro studies could serve as the bases for constructing the hypothetical framework for predicting the regulatory role, in vivo, of the enzyme in the pathway. We have discussed the known as well as the as yet ill-defined facets of the cellular and subcellular aspects of the glycolysis-citric acid cycle interrelation and have attempted to illustrate how such an integrated approach could be applied to generate testable hypotheses for investigating the mechanisms concerned with metabolic compartmentation and regulation in brain. In the process of the illustration, we discuss some of the evidence in support of the general hypothesis that the transfer of reducing equivalents across the inner mitochondrial membrane plays a major role in mediating the coupling of the glycolytic flux to that of the citric acid cycle. We have given some indications as to how this hypothesis could be further investigated employing our approach. Moreover, we hope that other workers will find this integrated approach useful in designing multidisciplinary studies to investigate mechanistic issues related to this important theme.

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Glutamate metabolism and transport in rat brain mitochondria

Cited by 56 publications

References 30 publications

New Insights into the Compartmentation of Glutamate and Glutamine in Cultured Rat Brain Astrocytes

New Insights into the Compartmentation of Glutamate and Glutamine in Cultured Rat Brain Astrocytes

3-Hydroxyanthranilic acid oxygenase-containing astrocytic processes surround glutamate-containing axon terminals in the rat striatum

Glycolysis-Citric Acid Cycle Interrelation: A New Approach and Some Insights in Cellular and Subcellular Compartmentation

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