Brain Alanine Formation as an Ammonia-Scavenging Pathway during Hyperammonemia: Effects of Glutamine Synthetase Inhibition in Rats and Astrocyte—Neuron Co-Cultures

Dadsetan, Sherry; Kukolj, Eva; Bak, Lasse K.; Sørensen, Michael; Ott, Peter; Vilstrup, Hendrik; Schousboe, Arne; Keiding, Susanne; Waagepetersen, Helle S.

doi:10.1038/jcbfm.2013.73

Cited by 44 publications

(34 citation statements)

References 38 publications

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“…Instead, our results indicate that glutamine synthetase dampens the harmful effects of acute NH 4 + /NH 3 intoxication by reducing its effective concentration. In addition, NH 4 + /NH 3 might be metabolized not only via glutamine synthetase, but by the action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT) which results in alanine formation [59]. These enzymes, might, therefore, additionally protect the tissue from harmful effects of hyperammonemia.…”

Section: Discussionmentioning

confidence: 99%

Dysbalance of Astrocyte Calcium under Hyperammonemic Conditions

2014

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Increased brain ammonium (NH4 +/NH3) plays a central role in the manifestation of hepatic encephalopathy (HE), a complex syndrome associated with neurological and psychiatric alterations, which is primarily a disorder of astrocytes. Here, we analysed the influence of NH4 +/NH3 on the calcium concentration of astrocytes in situ and studied the underlying mechanisms of NH4 +/NH3-evoked calcium changes, employing fluorescence imaging with Fura-2 in acute tissue slices derived from different regions of the mouse brain. In the hippocampal stratum radiatum, perfusion with 5 mM NH4 +/NH3 for 30 minutes caused a transient calcium increase in about 40% of astrocytes lasting about 10 minutes. Furthermore, the vast majority of astrocytes (∼90%) experienced a persistent calcium increase by ∼50 nM. This persistent increase was already evoked at concentrations of 1–2 mM NH4 +/NH3, developed within 10–20 minutes and was maintained as long as the NH4 +/NH3 was present. Qualitatively similar changes were observed in astrocytes of different neocortical regions as well as in cerebellar Bergmann glia. Inhibition of glutamine synthetase resulted in significantly larger calcium increases in response to NH4 +/NH3, indicating that glutamine accumulation was not a primary cause. Calcium increases were not mimicked by changes in intracellular pH. Pharmacological inhibition of voltage-gated sodium channels, sodium-potassium-chloride-cotransporters (NKCC), the reverse mode of sodium/calcium exchange (NCX), AMPA- or mGluR5-receptors did not dampen NH4 +/NH3-induced calcium increases. They were, however, significantly reduced by inhibition of NMDA receptors and depletion of intracellular calcium stores. Taken together, our measurements show that sustained exposure to NH4 +/NH3 causes a sustained increase in intracellular calcium in astrocytes in situ, which is partly dependent on NMDA receptor activation and on release of calcium from intracellular stores. Our study furthermore suggests that dysbalance of astrocyte calcium homeostasis under hyperammonemic conditions is a widespread phenomenon, which might contribute to the disturbance of neurotransmission during HE.

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Section: Discussionmentioning

confidence: 99%

Dysbalance of Astrocyte Calcium under Hyperammonemic Conditions

2014

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“…The ALAT reaction in concert with glutamate dehydrogenase (GDH) may be a particularly important mechanism for fixation of ammonia during hyperammonemic conditions when the normal activity of glutamine synthetase (see below) is inhibited (Dadsetan et al 2011, 2013). This aspect will be discussed in further detail below.…”

Section: Enzymatic Reactions Involving Glutamate As Substrate or Productmentioning

confidence: 99%

“…As pointed out above, de novo synthesis of glutamine requires CO 2 fixation by pyruvate carboxylase to form oxaloacetate; interestingly, hyperammonemic conditions have been shown to stimulate this pathway (Leke et al 2011). On the other hand, when GS was inhibited and ammonia was fixed in alanine, the glycolytic pathway was accelerated while the anaplerotic pathway did not increase (Dadsetan et al 2011, 2013). This shows that astrocytes are able to switch back and forth between anaplerosis and glycolysis depending on whether pyruvate or a TCA cycle intermediate is needed.…”

Section: Glutamate and Ammonia Homeostasis Under Normal And Hyperammomentioning

confidence: 99%

Glutamate Metabolism in the Brain Focusing on Astrocytes

Schousboe

Scafidi

Bak

et al. 2014

Advances in Neurobiology

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301

217

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Metabolism of glutamate, the main excitatory neurotransmitter and precursor of GABA, is exceedingly complex and highly compartmentalized in brain. Maintenance of these neurotransmitter pools is strictly dependent on the de novo synthesis of glutamine in astrocytes which requires both the anaplerotic enzyme pyruvate carboxylase and glutamine synthetase. Glutamate is formed directly from glutamine by deamidation via phosphate activated glutaminase a reaction that also yields ammonia. Glutamate plays key roles linking carbohydrate and amino acid metabolism via the tricarboxylic acid (TCA) cycle, as well as in nitrogen trafficking and ammonia homeostasis in brain. The anatomical specialization of astrocytic endfeet enables these cells to rapidly and efficiently remove neurotransmitters from the synaptic cleft to maintain homeostasis, and to provide glutamine to replenish neurotransmitter pools in both glutamatergic and GABAergic neurons. Since the glutamate-glutamine cycle is an open cycle that actively interfaces with other pathways, the de novo synthesis of glutamine in astrocytes helps to maintain the operation of this cycle. The fine-tuned biochemical specialization of astrocytes allows these cells to respond to subtle changes in neurotransmission by dynamically adjusting their anaplerotic and glycolytic activities, and adjusting the amount of glutamate oxidized for energy relative to direct formation of glutamine, to meet the demands for maintaining neurotransmission. This chapter summarizes the evidence that astrocytes are essential and dynamic partners in both glutamatergic and GABAergic neurotransmission in brain.

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“…The main mechanism of osmolyte accumulation is fixation of ammonia and production of glutamine by the astrocytic ATP-dependent glutamine synthetase [33, 144]. The osmotic burden due to the enhanced activity of glutamine synthetase is further accentuated by accumulation of brain alanine via the work of the alternative detoxification mechanism incorporating glutamate dehydrogenase and alanine aminotransferase [41, 201]. Actions of astrocytic glutamine are not restricted to its direct osmotic effects; since this amino acid is also thought to induce cellular pathology indirectly, via oxidative and nitrosative stress and impaired mitochondrial function (the "Trojan horse" hypothesis [8]).…”

Section: Common and Unique Roles Of Vrac Within The Brainmentioning

confidence: 99%

Volume-regulated anion channel—a frenemy within the brain

Mongin

2015

Pflugers Arch - Eur J Physiol

115

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Volume regulated anion channel (VRAC) is a ubiquitously expressed yet highly enigmatic member of the superfamily of chloride/anion channels. It is activated by cellular swelling and mediates regulatory cell volume decrease in a majority of vertebrate cells, including those in the central nervous system (CNS). In the brain, besides its crucial role in cellular volume regulation, VRAC is thought to play a part in cell proliferation, apoptosis, migration, and release of physiologically active molecules. Although these roles are not exclusive to the CNS, the relative significance of VRAC in the brain is amplified by several unique aspects of its physiology. One important example is contribution of VRAC to release of the excitatory amino acid neurotransmitters, glutamate and aspartate. This latter process is thought to have impact on both normal brain functioning (such as astrocyte-neuron signaling) and neuropathology (via promoting the excitotoxic death of neuronal cells in stroke and traumatic brain injury). In spite of much work in the field the molecular nature of VRAC remained unknown until less than two years ago. Two pioneer publications identified VRAC as the hetero-hexamer formed by the leucine-rich repeat-containing 8 (LRRC8) proteins. These findings galvanized the field and are likely to result in dramatic revisions to our understanding of the place and role of VRAC in the brain, as well as other organs and tissues. The present review briefly recapitulates critical findings in the CNS, and focuses on anticipated impact on the LRRC8 discovery on further progress in neuroscience research.

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Brain Alanine Formation as an Ammonia-Scavenging Pathway during Hyperammonemia: Effects of Glutamine Synthetase Inhibition in Rats and Astrocyte—Neuron Co-Cultures

Cited by 44 publications

References 38 publications

Dysbalance of Astrocyte Calcium under Hyperammonemic Conditions

Dysbalance of Astrocyte Calcium under Hyperammonemic Conditions

Glutamate Metabolism in the Brain Focusing on Astrocytes

Volume-regulated anion channel—a frenemy within the brain

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