scholarly journals Effect of Glutamine Synthetase Inhibition on Brain and Interorgan Ammonia Metabolism in Bile Duct Ligated Rats

2013 ◽  
Vol 34 (3) ◽  
pp. 460-466 ◽  
Author(s):  
Andreas W Fries ◽  
Sherry Dadsetan ◽  
Susanne Keiding ◽  
Lasse K Bak ◽  
Arne Schousboe ◽  
...  
Keyword(s):  
Amino Acids ◽  
Bile Duct ◽  
Glutamine Synthetase ◽  
Methionine Sulfoximine ◽  
Ammonia Metabolism ◽  
Liver And Kidney ◽  
Glutamine Synthesis ◽  
Alanine Synthesis ◽  
High Concentrations ◽  
The Brain

Ammonia has a key role in the development of hepatic encephalopathy (HE). In the brain, glutamine synthetase (GS) rapidly converts blood-borne ammonia into glutamine which in high concentrations may cause mitochondrial dysfunction and osmolytic brain edema. In astrocyte-neuron cocultures and brains of healthy rats, inhibition of GS by methionine sulfoximine (MSO) reduced glutamine synthesis and increased alanine synthesis. Here, we investigate effects of MSO on brain and interorgan ammonia metabolism in sham and bile duct ligated (BDL) rats. Concentrations of glutamine, glutamate, alanine, and aspartate and incorporation of 15NH4+ into these amino acids in brain, liver, muscle, kidney, and plasma were similar in sham and BDL rats treated with saline. Methionine sulfoximine reduced glutamine concentrations in liver, kidney, and plasma but not in brain and muscle; MSO reduced incorporation of 15NH4+ into glutamine in all tissues. It did not affect alanine concentrations in any of the tissues but plasma alanine concentration increased; incorporation of 15NH4+ into alanine was increased in brain in sham and BDL rats and in kidney in sham rats. It inhibited GS in all tissues examined but only in brain was an increased incorporation of 15N-ammonia into alanine observed. Liver and kidney were important for metabolizing blood-borne ammonia.

scholarly journals Methionine Sulfoximine, a Glutamine Synthetase Inhibitor, Attenuates Increased Extracellular Potassium Activity during Acute Hyperammonemia

1997 ◽  
Vol 17 (1) ◽  
pp. 44-49 ◽  
Author(s):  
Hideyoshi Sugimoto ◽  
Raymond C. Koehler ◽  
David A. Wilson ◽  
Saul W. Brusilow ◽  
Richard J. Traystman
Keyword(s):  
Glutamine Synthetase ◽  
Sodium Acetate ◽  
Ammonium Acetate ◽  
Ammonia Concentration ◽  
Methionine Sulfoximine ◽  
Extracellular Potassium ◽  
Edema Formation ◽  
Plasma Ammonia ◽  
Glutamine Synthesis ◽  
Ion Sensitive Microelectrodes

Hyperammonemia causes glutamine accumulation and astrocyte swelling. Inhibition of glutamine synthesis reduces ammonia-induced edema formation and watery swelling in astrocyte processes. Ordinarily, astrocytes tightly control extracellular K+ activity [K+]e. We tested the hypothesis that acute hyperammonemia interferes with this tight regulation such that [K+]e increases and that inhibition of glutamine synthetase reduces this increase in [K+]e. Ion-sensitive microelectrodes were used to measure [K+]e in parietal cortex continuously over a 6-h period in anesthetized rats. After i.v. sodium acetate infusion in eight control rats, plasma ammonia concentration was 33 ± 26 μmol/L (± SD) and [K+]e remained stable at 4.3 ± 1.6 mmol/L. During ammonium acetate infusion in nine rats, plasma ammonia increased to 594 ± 124 μmol/L at 2 h and to 628 ± 135 μmol/L at 6 h. There was a gradual increase in [K+]e from 3.9 ± 0.7 to 6.8 ± 2.7 mmol/L at 2 h and 11.8 ± 6.7 mmol/L at 6 h. In eight rats, L-methionine-D,L-sulfoximine (150 mg/kg) was infused 3 h before ammonium acetate infusion to inhibit glutamine synthetase. At 2 and 6 h of ammonium acetate infusion, plasma ammonia concentration was 727 ± 228 and 845 ± 326 μmol/L, and [K+]e was 4.5 ± 1.9 and 6.1 ± 3.8 mmol/L, respectively. The [K+]e value at 6 h was significantly less than that obtained with ammonium acetate infusion alone but was not different from that obtained with sodium acetate infusion. We conclude that acute hyperammonemia impairs astrocytic control of [K+]e and that this impairment is linked to glutamine accumulation rather than ammonium ions per se.

scholarly journals Hyperammonaemia does not impair brain function in the absence of net glutamine synthesis

1991 ◽  
Vol 277 (3) ◽  
pp. 697-703 ◽  
Author(s):  
R A Hawkins ◽  
J Jessy
Keyword(s):  
Amino Acids ◽  
Brain Function ◽  
Aromatic Amino Acids ◽  
Preceding Paper ◽  
Glucose Consumption ◽  
High Energy ◽  
High Energy Phosphates ◽  
Intermediary Metabolites ◽  
Glutamine Synthesis ◽  
The Brain

1. It has been established that chronic hyperammonaemia, whether caused by portacaval shunting or other means, leads to a variety of metabolic changes, including a depression in the cerebral metabolic rate of glucose (CMRGlc) increased permeability of the blood-brain barrier to neutral amino acids, and an increase in the brain content of aromatic amino acids. The preceding paper [Jessy, DeJoseph & Hawkins (1991) Biochem. J. 277, 693-696] showed that the depression in CMRGlc caused by hyperammonaemia correlated more closely with glutamine, a metabolite of ammonia, than with ammonia itself. This suggested that ammonia (NH3 and NH4+) was without effect. The present experiments address the question whether ammonia, in the absence of net glutamine synthesis, induces any of the metabolic symptoms of cerebral dysfunction associated with hyperammonaemia. 2. Small doses of methionine sulphoximine, an inhibitor of glutamine synthetase, were used to raise the plasma ammonia levels of normal rats without increasing the brain glutamine content. These hyperammonaemic rats, with plasma and brain ammonia levels equivalent to those known to depress brain function, behaved normally over 48 h. There was no depression of cerebral energy metabolism (i.e. the rate of glucose consumption). Contents of key intermediary metabolites and high-energy phosphates were normal. Neutral amino acid transport (tryptophan and leucine) and the brain contents of aromatic amino acids were unchanged. 3. The data suggest that ammonia is without effect at concentrations less than 1 mumol/ml if it is not converted into glutamine. The deleterious effect of chronic hyperammonaemia seems to begin with the synthesis of glutamine.

scholarly journals 2235 15N-Leucine transport across the blood brain barrier is significantly impaired in the glutamine synthetase-inhibited brain

2018 ◽  
Vol 2 (S1) ◽  
pp. 1-1
Author(s):  
Shaun E. Gruenbaum ◽  
Roni Dhaher ◽  
Kevin Behar ◽  
Hitten Zaveri ◽  
Mark Erfe ◽  
...  
Keyword(s):  
Amino Acid ◽  
Glutamine Synthetase ◽  
Dentate Gyrus ◽  
Extracellular Fluid ◽  
Ultra Performance Liquid Chromatography ◽  
Methionine Sulfoximine ◽  
Causative Role ◽  
Immune System Dysfunction ◽  
Tightly Coupled ◽  
The Brain

OBJECTIVES/SPECIFIC AIMS: Astroglial glutamine synthetase (GS), which metabolizes glutamate and ammonia to glutamine, is critical for the detoxification of brain ammonia, clearance of synaptic glutamate, and production of brain glutamine. Perturbations in the expression and activity of GS are thought to play a causative role in the pathogenesis of several conditions of abnormal neurotransmission. Although the long-term consequences of GS inhibition on amino acid homeostasis in the brain are unknown, it is thought that amino acid influx in the brain is tightly coupled with glutamine efflux via the L-type amino acid transporter. Both glutamine and leucine serve many critical functions in the brain including protein synthesis, gene expression, insulin regulation, and immune signaling. The objective of this study was to determine the effects of chronic GS inhibition with methionine sulfoximine (MSO) on glutamine and leucine homeostasis in the brain. METHODS/STUDY POPULATION: In total, 12 rats were surgically implanted with microdialysis guide cannulas in the bilateral dentate gyrus. Rats were randomly divided for surgical implantation of either a MSO (n=6) or phosphate buffer saline (PBS; n=6) pump in the right dentate gyrus. After 7 days, bilateral microdialysis probes were placed under brief isoflurane anesthesia, and microdialysis flow was established by infusing 0.5 µL/min of artificial extracellular fluid. Dialysate samples were collected every 30 minutes for the duration of the experiment. A 113 mM 15N-Leucine (3.6 mL/h) and 2 M 2–13C-sodium acetate (0.0633 μL/g/min for t=0–5 min, 0.0316 μL/g/min for t=5–10 min, and 0.0253 μL/g/min for t>10 min) solution was infused intravenously for 300 minutes. The EZ:Faast Free Amino Acid analysis kit and ultra-performance liquid chromatography/tandem mass spectrometry was used for quantification of amino acids in the dialysate fluid. RESULTS/ANTICIPATED RESULTS: At baseline (t=0 h), the concentrations of glutamine were significantly lower in MSO-treated rats (p<0.001) in the ipsilateral (GS-inhibited) hippocampus. There were no differences in glutamine concentrations between MSO and PBS-treated rats in the contralateral hippocampus. In PBS-treated rats, there was a significant increase in 15N-leucine between t=0 hour and t=5 hour in the contralateral (p<0.05) and ipsilateral (p<0.05) hippocampus. In MSO-treated rats, there was a significant increase in 15N-leucine between t=0 and t=5 hours in the contralateral (p<0.05) hippocampus, but not in the ipsilateral hippocampus (p=ns.). DISCUSSION/SIGNIFICANCE OF IMPACT: This study demonstrated for the first time that basal glutamine concentrations are low in areas of the brain where GS is acutely inhibited, and that leucine uptake in these brain areas are markedly decreased. Perturbations in glutamine and leucine homeostasis have been implicated in several disease processes including diabetes, obesity, liver disease, immune system dysfunction, epilepsy, and cancer, and the glutamine-dependent leucine influx in the brain may be a novel and important therapeutic target to treat these conditions.

scholarly journals Effects of inhibiting glutamine synthetase and blocking glutamate uptake on b-wave generation in the isolated rat retina

1999 ◽  
Vol 16 (2) ◽  
pp. 345-353 ◽  
Author(s):  
BARRY S. WINKLER ◽  
NATALIA KAPOUSTA-BRUNEAU ◽  
MATTHEW J. ARNOLD ◽  
DANIEL G. GREEN
Keyword(s):  
Amino Acids ◽  
Glutamine Synthetase ◽  
Glutamate Uptake ◽  
Müller Cells ◽  
Glutamate Transporters ◽  
Methionine Sulfoximine ◽  
Muller Cells ◽  
Rat Retina ◽  
Subsequent Conversion ◽  
Isolated Rat

The purpose of the present experiments was to evaluate the contribution of the glutamate-glutamine cycle in retinal glial (Müller) cells to photoreceptor cell synaptic transmission. Dark-adapted isolated rat retinas were superfused with oxygenated bicarbonate-buffered media. Recordings were made of the b-wave of the electroretinogram as a measure of light-induced photoreceptor to ON-bipolar neuron transmission. L-methionine sulfoximine (1–10 mM) was added to superfusion media to inhibit glutamine synthetase, a Müller cell specific enzyme, by more than 99% within 5–10 min, thereby disrupting the conversion of glutamate to glutamine in the Müller cells. Threo-hydroxyaspartic acid and D-aspartate were used to block glutamate transporters. The amplitude of the b-wave was well maintained for 1–2 h provided 0.25 mM glutamate or 0.25 mM glutamine was included in the media. Without exogenous glutamate or glutamine the amplitude of the b-wave declined by about 70% within 1 h. Inhibition of glutamate transporters led to a rapid (2–5 min) reversible loss of the b-wave in the presence and absence of the amino acids. In contrast, inhibition of glutamine synthetase did not alter significantly either the amplitude of the b-wave in the presence of glutamate or glutamine or the rate of decline of the b-wave found in the absence of these amino acids. Excellent recovery of the b-wave was found when 0.25 mM glutamate was resupplied to L-methionine sulfoximine–treated retinas. The results suggest that in the isolated rat retina uptake of released glutamate into photoreceptors plays a more important role in transmitter recycling than does uptake of glutamate into Müller cells and its subsequent conversion to glutamine.

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

2013 ◽  
Vol 33 (8) ◽  
pp. 1235-1241 ◽  
Author(s):  
Sherry Dadsetan ◽  
Eva Kukolj ◽  
Lasse K Bak ◽  
Michael SØrensen ◽  
Peter Ott ◽  
...  
Keyword(s):  
Hepatic Encephalopathy ◽  
Glutamine Synthetase ◽  
Drug Target ◽  
Methionine Sulfoximine ◽  
Concerted Action ◽  
N Incorporation ◽  
Dose Dependent ◽  
The Brain

Hyperammonemia is a major etiological toxic factor in the development of hepatic encephalopathy. Brain ammonia detoxification occurs primarily in astrocytes by glutamine synthetase (GS), and it has been proposed that elevated glutamine levels during hyperammonemia lead to astrocyte swelling and cerebral edema. However, ammonia may also be detoxified by the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT) leading to trapping of ammonia in alanine, which in vivo likely leaves the brain. Our aim was to investigate whether the GS inhibitor methionine sulfoximine (MSO) enhances incorporation of 15NH4+ in alanine during acute hyperammonemia. We observed a fourfold increased amount of 15NH4 incorporation in brain alanine in rats treated with MSO. Furthermore, co-cultures of neurons and astrocytes exposed to 15NH4Cl in the absence or presence of MSO demonstrated a dose-dependent incorporation of 15NH4 into alanine together with increased 15N incorporation in glutamate. These findings provide evidence that ammonia is detoxified by the concerted action of GDH and ALAT both in vivo and in vitro, a mechanism that is accelerated in the presence of MSO thereby reducing the glutamine level in brain. Thus, GS could be a potential drug target in the treatment of hyperammonemia in patients with hepatic encephalopathy.

Effects of Methionine Sulfoximine and Glycine on Nitrogen Metabolism of Maize Leaves in the Light

1983 ◽  
Vol 10 (2) ◽  
pp. 187 ◽  
Author(s):  
MG Berger ◽  
HP Fock
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Glutamine Synthetase ◽  
Nitrogen Cycling ◽  
Co2 Assimilation ◽  
Methionine Sulfoximine ◽  
Co2 Evolution ◽  
Ammonia Content ◽  
Ammonia Accumulation ◽  
Maize Leaves

Detached maize leaves with their cut bases in water or in solutions containing 15 mM [14C, 15N]glycine, 15 mM [14C]glutamate, 5 mM methionine sulfoximine (an inhibitor of glutamine synthetase) or the appropriate amino acid plus inhibitor, were incubated for up to 135 min in the light. The concentrations and the 15N content of ammonia and of amino acids involved in photorespiratory nitrogen cycling were determined. Incubation with methionine sulfoximine or glycine increased the ammonia content significantly, whereas glutamate showed no effect. The nitrogen of glycine was metabolized into serine and ammonia. Ammonia was first recycled into glutamine, and then into glutamate. The glycine carbon skeleton served as a precursor for serine. Based on the data for ammonia accumulation the minimum rate of photorespiratory CO2 evolution in maize leaves was estimated to be about 1% of the rate of CO2 assimilation.

Selected brain amino acids and ammonium during chronic hypercapnia in conscious rats

1978 ◽  
Vol 44 (3) ◽  
pp. 333-339 ◽  
Author(s):  
J. Weyne ◽  
F. Van Leuven ◽  
H. Kazemi ◽  
I. Leusen
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Glutamine Synthetase ◽  
Time Course ◽  
Aminobutyric Acid ◽  
Gamma Aminobutyric Acid ◽  
Ph Homeostasis ◽  
Glutamine Synthesis ◽  
Amino Acid Concentrations ◽  
Observation Period

Glutamic, aspartic, and gamma-aminobutyric acid (GABA), glutamine, and ammonium were measured in the brains of unanesthetized normocapnic and hypercapnic (10% CO2; 5 min to 3 wk) rats. Hypercapnia increased glutamine and GABA and decreased glutamic and aspartic acids. Changes occurred within 1 h and were maintained during the observation period of 3 wk. On return to normocapnia amino acid concentrations were almost normal after 1 h. Based on the time course it is concluded that intracerebral hypercapnia is more likely the stimulus for change than acidosis. Ammonium content was unchanged for at least 1 h after the onset of hypercapnia but increased thereafter. Experiments in which glutamine synthesis by brain was impeded by inhibiting the enzyme glutamine synthetase favor the hypothesis that the rise of ammonium content in hypercapnia is initially not seen because of increased glutamine synthesis. The changes observed may have a role in metabolic pH homeostasis of brain tissue and may also be relevant to the modified brain excitability in hypercapnia.

Native Ubiquitin Structural Changes Resulting from Complexation with β-methylamino-L-alanine (BMAA)

2020 ◽  
Author(s):  
Katie Mae Wilson ◽  
Aurora Burkus-Matesevac ◽  
Samuel Maddox ◽  
Christopher Chouinard
Keyword(s):  
Structural Changes ◽  
Noncovalent Complex ◽  
Unfolded Protein ◽  
Protein Size ◽  
High Concentrations ◽  
The Unfolded Protein Response ◽  
Critical Binding ◽  
Protein Response ◽  
The Brain ◽  
Lateral Sclerosis

β-methylamino-L-alanine (BMAA) has been linked to the development of neurodegenerative (ND) symptoms following chronic environmental exposure through water and dietary sources. The brains of those affected by this condition, often referred to as amyotrophic lateral sclerosis-parkinsonism-dementia complex (ALS-PDC), have exhibited the presence of plaques and neurofibrillary tangles (NFTs) from protein aggregation. Although numerous studies have sought to better understand the correlation between BMAA exposure and onset of ND symptoms, no definitive link has been identified. One prevailing hypothesis is that BMAA acts a small molecule ligand, complexing with critical proteins in the brain and reducing their function. The objective of this research was to investigate the effects of BMAA exposure on the native structure of ubiquitin. We hypothesized that formation of a Ubiquitin+BMAA noncovalent complex would alter the protein’s structure and folding and ultimately affect the ubiquitinproteasome system (UPS) and the unfolded protein response (UPR). Ion mobility-mass spectrometry revealed that at sufficiently high concentrations BMAA did in fact form a noncovalent complex with ubiquitin, however similar complexes were identified for a range of additional amino acids. Collision induced unfolding (CIU) was used to interrogate the unfolding dynamics of native ubiquitin and these Ubq-amino acid complexes and it was determined that complexation with BMAA led to a significant alteration in native protein size and conformation, and this complex required considerably more energy to unfold. This indicates that the complex remains more stable under native conditions and this may indicate that BMAA has attached to a critical binding location.

Native Ubiquitin Structural Changes Resulting from Complexation with β-methylamino-L-alanine (BMAA)

2020 ◽  
Author(s):  
Katie Mae Wilson ◽  
Aurora Burkus-Matesevac ◽  
Samuel Maddox ◽  
Christopher Chouinard
Keyword(s):  
Structural Changes ◽  
Noncovalent Complex ◽  
Unfolded Protein ◽  
Protein Size ◽  
High Concentrations ◽  
The Unfolded Protein Response ◽  
Critical Binding ◽  
Protein Response ◽  
The Brain ◽  
Lateral Sclerosis

β-methylamino-L-alanine (BMAA) has been linked to the development of neurodegenerative (ND) symptoms following chronic environmental exposure through water and dietary sources. The brains of those affected by this condition, often referred to as amyotrophic lateral sclerosis-parkinsonism-dementia complex (ALS-PDC), have exhibited the presence of plaques and neurofibrillary tangles (NFTs) from protein aggregation. Although numerous studies have sought to better understand the correlation between BMAA exposure and onset of ND symptoms, no definitive link has been identified. One prevailing hypothesis is that BMAA acts a small molecule ligand, complexing with critical proteins in the brain and reducing their function. The objective of this research was to investigate the effects of BMAA exposure on the native structure of ubiquitin. We hypothesized that formation of a Ubiquitin+BMAA noncovalent complex would alter the protein’s structure and folding and ultimately affect the ubiquitinproteasome system (UPS) and the unfolded protein response (UPR). Ion mobility-mass spectrometry revealed that at sufficiently high concentrations BMAA did in fact form a noncovalent complex with ubiquitin, however similar complexes were identified for a range of additional amino acids. Collision induced unfolding (CIU) was used to interrogate the unfolding dynamics of native ubiquitin and these Ubq-amino acid complexes and it was determined that complexation with BMAA led to a significant alteration in native protein size and conformation, and this complex required considerably more energy to unfold. This indicates that the complex remains more stable under native conditions and this may indicate that BMAA has attached to a critical binding location.

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