Glutamine synthetase deficiency in murine astrocytes results in neonatal death

He, Youji; Hakvoort, Theodorus B. M.; Vermeulen, Jacqueline L.M.; Labruyère, Wil T.; Waart, Dirk R. de; Hel, W. Saskia van der; Uylings, H.B.M.; Lamers, Wouter H.

doi:10.1002/glia.20960

Cited by 49 publications

(65 citation statements)

References 60 publications

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“…We, therefore, also challenged mice systemically by infusing incremental doses of NH 4 HCO 3 into the jugular vein. Because constitutive GS deficiency is lethal, (2) some control mice were . At the same portal vein ammonia concentration, that in the hepatic vein was 2.3-fold higher in GS-deficient mice (P < 0.001) and the ammonia-detoxifying capacity of the liver was 50% of controls (P < 0.001).…”

Section: Ammonia-detoxifying Capacity Upon Administration Of Incremenmentioning

confidence: 99%

“…GS deficiency causes only moderate hyperammonemia, but affected humans and mice suffer from encephalopathy and die neonatally. (2,3) GS is predominantly expressed in the nervous system, kidney, and liver, that is, in established glutamine-consuming organs, (4)(5)(6) whereas skeletal muscle, with a much lower expression but large mass, is considered the main net producer of glutamine. (7) The nervous system synthesizes glutamine to scavenge /Gs fl/fl mice; GS-KO/0.5, Gs fl/LacZ mice; K 0.5 , substrate concentration at which process proceeds at 50% of maximum velocity; KO, knockout; MSO, methionine sulfoximine.…”

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confidence: 99%

“…(9) To assess the ammonia-detoxifying function of GS in an in vivo setting, we generated mice with the proteincoding region of the GS gene (Glul) flanked by loxP sequences. (2,10) We used genetic (muscle-specific (10) and liver-specific [this study] GS-knockout [KO] mice) or pharmacological methods to modulate GS activity and stepwise increments of enterally or intravenously administered NH 4 HCO 3 to challenge ammonia detoxification (see Supporting Fig. S1).…”

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Pivotal role of glutamine synthetase in ammonia detoxification

et al. 2016

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Glutamine synthetase (GS) catalyzes condensation of ammonia with glutamate to glutamine. Glutamine serves, with alanine, as a major nontoxic interorgan ammonia carrier. Elimination of hepatic GS expression in mice causes only mild hyperammonemia and hypoglutaminemia but a pronounced decrease in the whole-body muscle-to-fat ratio with increased myostatin expression in muscle. Using GS-knockout/liver and control mice and stepwise increments of enterally infused ammonia, we show that 35% of this ammonia is detoxified by hepatic GS and 35% by urea-cycle enzymes, while 30% is not cleared by the liver, independent of portal ammonia concentrations £ 2 mmol/L. Using both genetic (GSknockout/liver and GS-knockout/muscle) and pharmacological (methionine sulfoximine and dexamethasone) approaches to modulate GS activity, we further show that detoxification of stepwise increments of intravenously (jugular vein) infused ammonia is almost totally dependent on GS activity. Maximal ammonia-detoxifying capacity through either the enteral or the intravenous route is 160 lmol/hour in control mice. Using stable isotopes, we show that disposal of glutaminebound ammonia to urea (through mitochondrial glutaminase and carbamoylphosphate synthetase) depends on the rate of glutamine synthesis and increases from 7% in methionine sulfoximine-treated mice to 500% in dexamethasone-treated mice (control mice, 100%), without difference in total urea synthesis. Conclusions: Hepatic GS contributes to both enteral and systemic ammonia detoxification. Glutamine synthesis in the periphery (including that in pericentral hepatocytes) and glutamine catabolism in (periportal) hepatocytes represents the high-affinity ammonia-detoxifying system of the body. The dependence of glutamine-bound ammonia disposal to urea on the rate of glutamine synthesis suggests that enhancing peripheral glutamine synthesis is a promising strategy to treat hyperammonemia. Because total urea synthesis does not depend on glutamine synthesis, we hypothesize that glutamate dehydrogenase complements mitochondrial ammonia production. (HEPATOLOGY 2017;65:281-293). G lutamine synthetase (GS) catalyzes condensation of ammonia with glutamate to glutamine. (Note: NH 3 is protonated for 98% to NH 4 1 at physiological pH. We use the term "ammonia" to refer to the sum of NH 3 and NH 4 1 unless specified as either "NH 3 " or "NH 41 .") Glutamine serves, with alanine, as a major nontoxic interorgan ammonia shuttle in the body (1) and as an aminomoiety donor for the synthesis of nucleotides, amino acids, amino-sugars, and oxidized nicotinamide adenine dinucleotide. Its intracellular turnover rate exceeds that of all other amino acids. GS deficiency causes only moderate hyperammonemia, but affected humans and mice suffer from encephalopathy and die neonatally. (2,3) GS is predominantly expressed in the nervous system, kidney, and liver, that is, in established glutamine-consuming organs, (4)(5)(6) whereas skeletal muscle, with a much lower expression but large mass, is considered the main net ...

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Section: Ammonia-detoxifying Capacity Upon Administration Of Incremenmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Pivotal role of glutamine synthetase in ammonia detoxification

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“…18 and 19, respectively. The floxed GS allele is equally active as the wild-type allele (19). Accordingly, both GS fl/LacZ and GS fl/fl mice are phenotypically normal and fertile.…”

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Glutamine Synthetase in Muscle Is Required for Glutamine Production during Fasting and Extrahepatic Ammonia Detoxification

Hakvoort

Köhler³

et al. 2010

Journal of Biological Chemistry

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The main endogenous source of glutamine is de novo synthesis in striated muscle via the enzyme glutamine synthetase (GS). The mice in which GS is selectively but completely eliminated from striated muscle with the Cre-loxP strategy (GS-KO/M mice) are, nevertheless, healthy and fertile. Compared with controls, the circulating concentration and net production of glutamine across the hindquarter were not different in fed GS-KO/M mice. Only a ϳ3-fold higher escape of ammonia revealed the absence of GS in muscle. However, after 20 h of fasting, GS-KO/M mice were not able to mount the ϳ4-fold increase in glutamine production across the hindquarter that was observed in control mice. Instead, muscle ammonia production was ϳ5-fold higher than in control mice. The fasting-induced metabolic changes were transient and had returned to fed levels at 36 h of fasting. Glucose consumption and lactate and ketone-body production were similar in GS-KO/M and control mice. Challenging GS-KO/M and control mice with intravenous ammonia in stepwise increments revealed that normal muscle can detoxify ϳ2.5 mol ammonia/g muscle⅐h in a muscle GS-dependent manner, with simultaneous accumulation of urea, whereas GS-KO/M mice responded with accumulation of glutamine and other amino acids but not urea. These findings demonstrate that GS in muscle is dispensable in fed mice but plays a key role in mounting the adaptive response to fasting by transiently facilitating the production of glutamine. Furthermore, muscle GS contributes to ammonia detoxification and urea synthesis. These functions are apparently not vital as long as other organs function normally.Glutamine is among the most abundant free amino acids in mammals. Almost 90% of the daily glutamine production originates from endogenous sources, because 30 -35% of all nitrogen derived from protein catabolism is transported in the form of glutamine (1, 2). This glutamine can serve, after transport via the vasculature, as an oxidative fuel for enterocytes and immune cells, a precursor for purine and pyrimidine synthesis, a modulator of protein turnover, or an intermediate for gluconeogenesis and acid-base balance. The only enzyme capable of glutamine synthesis is glutamine synthetase (L-glutamate: ammonia ligase (ADP); EC 6.3.1.2). Because of the prominent role of glutamine in the interorgan transport of carbon and nitrogen, the plasma glutamine pool is turning over very rapidly (3). Glutamine tracer kinetic studies in humans have shown that ϳ50% of plasma glutamine is oxidized and that of the remainder, 10 -20% is used for gluconeogenesis, and most of the rest is used for protein synthesis and incorporation into macromolecules (2).Thus far, the irreversible GS 5 inhibitor methionine sulfoximine (MSO) was the tool of choice to interfere with cellular glutamine synthesis. Administration of MSO for 4 -6 days results in a 40 -50% decrease in plasma glutamine levels, a 55-65% decrease in intracellular muscle glutamine, and a 50% increase in muscle ammonia levels (4 -6). An inherent drawback of the...

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“…Immunohistochemical analyses were performed as described previously (He et al 2010). Briefly, tissues were fixed in 4% paraformaldehyde in phosphate-buffered saline (PBS) overnight at 4ºC, embedded in paraffin, and sectioned at 7 μm.…”

Section: Immunohistochemical Analysesmentioning

confidence: 99%

Cellular Localization and Biochemical Analysis of Mammalian CDC50A, a Glycosylated β-subunit for P4 ATPases

Folmer

Mok

Wee

et al. 2012

J Histochem Cytochem.

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CDC50 proteins are β-subunits for P4 ATPases, which upon heterodimerization form a functional phospholipid translocation complex. Emerging evidence in mouse models and men links mutations in P4 ATPase genes with human disease. This study analyzed the tissue distribution and cellular localization of CDC50A, the most abundant and ubiquitously expressed CDC50 homologue in the mouse. The authors have raised antibodies that detect mouse and human CDC50A and studied CDC50A localization and glycosylation status in mouse liver cells. CDC50A is a terminal-glycosylated glycoprotein and is expressed in hepatocytes and liver sinusoidal endothelial cells, where it resides in detergent-resistant membranes. In pancreas and stomach, CDC50A localized to secretory vesicles, whereas in the kidney, CDC50A localized to the apical region of proximal convoluted tubules of the cortex. In WIF-B9 cells, CDC50A partially costains with the trans-Golgi network. Data suggest that CDC50A is present as a fully glycosylated protein in vivo, which presumes interaction with distinct P4 ATPases.

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Glutamine synthetase deficiency in murine astrocytes results in neonatal death

Cited by 49 publications

References 60 publications

Pivotal role of glutamine synthetase in ammonia detoxification

Pivotal role of glutamine synthetase in ammonia detoxification

Glutamine Synthetase in Muscle Is Required for Glutamine Production during Fasting and Extrahepatic Ammonia Detoxification

Cellular Localization and Biochemical Analysis of Mammalian CDC50A, a Glycosylated β-subunit for P4 ATPases

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