STUDIES ON FUNCTIONAL AND METABOLIC ROLE OF UREA CYCLE INTERMEDIATES IN BRAIN

Abstract— The distribution of argininosuccinate synthetase, argininosuccinase and arginase, and the synthesis of urea in cerebullum. cerebral cortex and brain stem have been studied. Cerebral cortex had high levels of argininosuccinate synthetase and argininosuccinase. and a high ability to synthesize urea from aspartic acid and citrulline. Of the three regions, cerebullum had the highest arginase activity. The activities of the enzymes transamidinase and ornithine aminotransferase in the metabolism of arginine and ornithine in pathways other than urea formation have been studied in the three regions of the rat brain. The activity of creatine phosphokinase in all regions was the same: carbamylphosphatase activity was highest in cerebullum. Cerebral cortex had a high activity of aspartic acid transcarba-mylase. The brain stem, among the three regions, had the lowest activities of glutamine synthetase and glutaminase. The activities of these enzymes in the different regions are discussed in relation to urea production and the utilization of the urea cycle intermediates.
Intraperitoneal injection of high amounts of citrulline brought about a rise in the glutamine synthetase activity of cerebellum and brain stem and a rise in ornithine aminotransferase in cerebral cortex and liver. These results are discussed in relation to the mechanism of action of citrulline in alleviating the toxicity in hyperammonaemic states.     

Effect of arginine,ornithine and citrulline supplementation upon performance and metabolism of trained rats

During intense exercise there is an augmented production of ammonia and IMP in the exercised muscle that could be related to the establishment of peripheral fatigue. In order to prevent this accumulation, the urea cycle in the liver eliminates ammonia in the form of urea and the skeletal muscle buffers the increase of ammonia via transamination reactions. In the present study we evaluated the effect of arginine, citrulline and ornithine supplementation, intermediates of the urea cycle, on the performance of sedentary and swimming-trained rats submitted to a single bout of exhaustive exercise. We also measured the glycogen content of the soleus and gastrocnemius muscles and of the liver, as well as the plasma concentrations of ammonia, urea, glutamine, glucose and lactate. The results indicate that arginine, citrulline and ornithine supplementation increased the flux of substrate through the reaction catalysed by glutamine synthetase, leading to increased glutamine production after an exhaustive bout of exercise, and of the mechanism involved in ammonia buffering.     

Enteral arginase II provides ornithine for citrulline synthesis

The synthesis of citrulline from arginine in the small intestine depends on the provision of ornithine. To test the hypothesis that arginase II plays a central role in the supply of ornithine for citrulline synthesis, the contribution of dietary arginine, glutamine, and proline was determined by utilizing multitracer stable isotope protocols in arginase II knockout (AII(-/-)) and wild-type (WT) mice. The lack of arginase II resulted in a lower citrulline rate of appearance (121 vs. 137 μmol·kg(-1)·h(-1)) due to a reduced availability of ornithine; ornithine supplementation was able to restore the rate of citrulline production in AII(-/-) to levels comparable with WT mice. There were significant differences in the utilization of dietary citrulline precursors. The contribution of dietary arginine to the synthesis of citrulline was reduced from 45 to 10 μmol·kg(-1)·h(-1) due to the lack of arginase II. No enteral utilization of arginine was observed in AII(-/-) mice (WT = 25 μmol·kg(-1)·h(-1)), and the contribution of dietary arginine through plasma ornithine was reduced in the transgenic mice (20 vs. 13 μmol·kg(-1)·h(-1)). Dietary glutamine and proline utilization were greater in AII(-/-) than in WT mice (20 vs. 13 and 1.4 vs. 3.7 μmol·kg(-1)·h(-1), respectively). Most of the contribution of glutamine and proline was enteral rather than through plasma ornithine. The arginase isoform present in the small intestinal mucosa has the role of providing ornithine for citrulline synthesis. The lack of arginase II results in a greater contribution of plasma ornithine and dietary glutamine and proline to the synthesis of citrulline.     

DL-7-azatryptophan and citrulline metabolism in the cyanobacterium Anabaena sp. strain 1F.

An alternative route for the primary assimilation of ammonia proceeds via glutamine synthetase-carbamyl phosphate synthetase and its inherent glutaminase activity in Anabaena sp. strain 1F, a marine filamentous, heterocystous cyanobacterium. Evidence for the presence of this possible alternative route to glutamate was provided by the use of amino acid analogs as specific enzyme inhibitors, enzymological studies, and radioistopic labeling experiments. The amino acid pool patterns of continuous cultures of Anabaena sp. strain 1F were markedly influenced by the nitrogen source. A relatively high concentration of glutamate was maintained in the amino acid pools of all cultures irrespective of the nitrogen source, reflecting the central role of glutamate in nitrogen metabolism. The addition of 1.0 microM azaserine increased the intracellular pools of glutamate and glutamine. All attempts to detect any enzymatic activity for glutamate synthase by measuring the formation of L-[14C]glutamate from 2-keto-[1-14C]glutarate and glutamine failed. The addition of 10 microM DL-7-azatryptophan caused a transient accumulation of intracellular citrulline and alanine which was not affected by the presence of chloramphenicol. The in vitro activity of carbamyl phosphate synthetase and glutaminase increased severalfold in the presence of azatryptophan. Results from radioisotopic labeling experiments with [14C]bicarbonate and L-[1-14C]ornithine also indicated that citrulline was formed via carbamyl phosphate synthetase and ornithine transcarbamylase. In addition to its effects on nitrogen metabolism, azatryptophan also affected carbon metabolism by inhibiting photosynthetic carbon assimilation and photosynthetic oxygen evolution.     

Effect of hyperammonemia on brain amino acids in young and adult ferrets

Summary Effects of arginine deficiency and hyperammonemia on the brain concentrations of amino acids and urea cycle enzyme activities in young and adult ferrets were investigated. Only young ferrets developed hyperammonemia and encephalopathy immediately after consuming the arginine-free diet. Brain ornithine and citrulline concentrations in young ferrets fed arginine containing diet were significantly lower than those in adult ferrets. Compared to rats and other animals, young and adult ferrets had lower concentrations of brain glutamic acid and glutamine. Unlike in other species, brain glutamine was not elevated in young, hyperammonemic ferrets. Brain arginase and glutamate dehydrogenase activities were significantly increased in young ferrets fed arginine-free diet. Young ferrets provide a useful animal model for investigating the neurotoxicity of acute hyperammonemia.Abbreviations ACD Arginine-containing diet - AFD Arginine-free diet This work was presented, in part, at the annual meeting of the Midwest Society for Pediatric Research, Chicago, IL, 1991.     

Changes in urea cycle-related metabolites in the mouse after combined administration of valproic acid and an amino acid load

Increased blood ammonia was induced in fasting mice by ip administration of 200 mg/kg Na-valproate followed 1 h later by 13 and 4 mmol/kg alanine and ornithine, respectively. When valproate was not used blood or liver ammonia was not increased, but increases were observed in liver glutamate (5-fold), glutamine (2-fold), aspartate (5-fold), acetylglutamate (15-fold), citrulline (35-fold), argininosuccinate (11-fold), arginine (11-fold), and urea (3-fold). The level of carbamoyl phosphate (less than 2 nmol/g) was, by far, the lowest of all urea cycle intermediates. The large increase in citrulline indicates that argininosuccinate synthesis was limiting, and that the increase in acetylglutamate induced a considerable activation of carbamoyl phosphate synthetase, which agrees with theoretical expectations, irrespective of the actual KD value for acetylglutamate. Pretreatment with valproate resulted in lower hepatic levels of glutamate, glutamine, aspartate, acetyl-CoA, and acetylglutamate. At the level found of acetylglutamate the activation of carbamoyl phosphate synthetase would be expected to be similar to that without valproate. Indeed, the levels of citrulline were similar with or without valproate. Argininosuccinate, arginine, and urea levels exhibited little if any change. Although the model used may not replicate exactly the situation in patients, from our results it appears that changes in citrullinogenesis or in other steps of the urea cycle do not account for the increase in blood ammonia induced by valproate, and it is proposed that valproate may alter glutamine metabolism.     

Serum amino acid profile in patients with acute pancreatitis

Patients in the early phase of acute pancreatitis (AP) have reduced serum levels of arginine and citrulline. This may be of patho-biological importance, since arginine is the substrate for nitric oxide, which in turn is involved in normal pancreatic physiology and in the inflammatory process. Serum amino acid spectrum was measured daily for five days and after recovery six weeks later in 19 patients admitted to the hospital for acute pancreatitis. These patients had abnormal levels of most amino acids including arginine, citrulline, glutamine and glutamate. Phenylalanine and glutamate were increased, while arginine, citrulline, ornithine and glutamine were decreased compared to levels after recovery. NO(2)/NO(3) concentration in the urine, but not serum arginase activity, was significantly increased day 1 compared to day 5 after admission. Acute pancreatitis causes a disturbance of the serum amino acid spectrum, with possible implications for the inflammatory process and organ function both in the pancreas and the gut. Supplementation of selected amino acids could possibly be of value in this severe condition.     

Changes of free amino acids in plasma of healthy subjects induced by physical exercise.

In eight healthy men a 20-min load of 1.5 W/kg body weight on a bicycle ergometer led to a significant increase of alanine and decline of leucine. Exhausting exercise caused in the same subjects a highly significant increase of alanine and decline of isoleucine, threonine, ornithine, leucine, serine, glycine, and asparasine and glutamine. The methionine and citrulline level declines also significantly. The total amino acids practically did not change. Physical exercise led furthermore to a marked increase of serum ammonia and uric acid. Urea nitrogen changed only little and on average had rather a declining tendency. The rise of alanine suggests the existence of a glucose-alanine cycle, the drop of ornithine and citrulline is most probably associated with the inhibition of ureogenesis in the liver. The reduction of leucine and isoleucine is probably the result of the entry of these amino acids into muscle and their deamination.     

Regulation of growth and polyamine biosynthesis of the ciliated protozoan Tetrahymena thermophila. Effects of L-arginine metabolites and polyamines

Growth of Tetrahymena thermophila in a synthetic nutrient medium with or without the essential amino acid L-arginine was studied in the presence or absence of the arginine metabolites L-citrulline and L-ornithine and the polyamines putrescine, spermidine, and spermine. The effects of the growth conditions on the stimulations of the enzymes of the arginine metabolic and polyamine biosynthetic pathway, arginine deiminase (ADI), citrulline hydrolase (CH), ornithine decarboxylase (ODC), and ornithine-oxo-acid aminotransferase were determined. Tetrahymena cells were unable to grow in the absence of L-arginine and the amino-acid utilization was greatly impaired. None of the metabolites or polyamines was able to substitute for arginine. In the presence of arginine, Tetrahymena cultures grew well and citrulline and ornithine did not alter the growth behaviour in any way. In the presence of putrescine, the lag period was decreased from 3 h to 2 h. Spermidine and spermine acted similar to putrescine but less pronounced. The stimulation of the activity of ADI, the key enzyme of arginine degradation, was absolutely dependent upon the presence of arginine in the medium: in the absence of arginine, the low ADI activity which was present in the cells before inoculation was decreased to zero levels within 30 min. In the presence of arginine, the stimulation of ADI was not altered by citrulline and ornithine but putrescine, spermidine, and spermine decreased ADI-stimulation to half of the control values. The stimulation of CH activity in the presence of arginine was not altered by any added metabolite or polyamine. In the media without arginine, stimulation of CH was greatly reduced, in the presence of ornithine more than in its absence, and even more in the presence of putrescine and spermidine. Stimulation of ODC activity in the presence of arginine was not affected by citrulline and ornithine but in the presence of polyamines it was rapidly decreased to unstimulated levels after an initial ca. 10-fold increase. The "hyperstimulation" of ODC in the absence of free arginine was reduced to normal in the presence of citrulline, the stimulation was decreased even below normal levels in the presence of ornithine and polyamines decreased ODC activity to zero levels. O delta T activity was stimulated more in the presence of arginine than in its absence. In both cases the stimulation was enhanced in the presence of polyamines and only in the absence of arginine--by ornithine.(ABSTRACT TRUNCATED AT 400 WORDS)     

Arginine catabolism in the cyanobacterium Synechocystis sp. Strain PCC 6803 involves the urea cycle and arginase pathway

Cells of the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 supplemented with micromolar concentrations of L-[(14)C]arginine took up, concentrated, and catabolized this amino acid. Metabolism of L-[(14)C]arginine generated a set of labeled amino acids that included argininosuccinate, citrulline, glutamate, glutamine, ornithine, and proline. Production of [(14)C]ornithine preceded that of [(14)C]citrulline, and the patterns of labeled amino acids were similar in cells incubated with L-[(14)C]ornithine, suggesting that the reaction of arginase, rendering ornithine and urea, is the main initial step in arginine catabolism. Ornithine followed two metabolic pathways: (i) conversion into citrulline, catalyzed by ornithine carbamoyltransferase, and then, with incorporation of aspartate, conversion into argininosuccinate, in a sort of urea cycle, and (ii) a sort of arginase pathway rendering glutamate (and glutamine) via Delta(1)pyrroline-5-carboxylate and proline. Consistently with the proposed metabolic scheme (i) an argF (ornithine carbamoyltransferase) insertional mutant was impaired in the production of [(14)C]citrulline from [(14)C]arginine; (ii) a proC (Delta(1)pyrroline-5-carboxylate reductase) insertional mutant was impaired in the production of [(14)C]proline, [(14)C]glutamate, and [(14)C]glutamine from [(14)C]arginine or [(14)C]ornithine; and (iii) a putA (proline oxidase) insertional mutant did not produce [(14)C]glutamate from L-[(14)C]arginine, L-[(14)C]ornithine, or L-[(14)C]proline. Mutation of two open reading frames (sll0228 and sll1077) putatively encoding proteins homologous to arginase indicated, however, that none of these proteins was responsible for the arginase activity detected in this cyanobacterium, and mutation of argD (N-acetylornithine aminotransferase) suggested that this transaminase is not important in the production of Delta(1)pyrroline-5-carboxylate from ornithine. The metabolic pathways proposed to explain [(14)C]arginine catabolism also provide a rationale for understanding how nitrogen is made available to the cell after mobilization of cyanophycin [multi-L-arginyl-poly(L-aspartic acid)], a reserve material unique to cyanobacteria.     

Submitochondrial localization of arginase and other enzymes associated with urea synthesis and nitrogen metabolism, in liver of Squalus acanthias

The submitochondrial localization of the four mitochondrial enzymes associated with urea synthesis in liver of Squalus acanthias (spiny dogfish), a representative elasmobranch, was determined. Glutamine- and acetylglutamate-dependent carbamoyl-phosphate synthetase, ornithine carbamoyltransferase, glutamine synthetase, and arginase were all localized within the matrix of liver mitochondria. The subcellular and submitochondrial localization and activities of several related enzymes involved in nitrogen metabolism and gluconeogenesis in liver and dogfish are also reported. Pyruvate carboxylase and phosphoenolpyruvate carboxykinase were localized in the mitochondrial matrix. Synthesis of citrulline by isolated mitochondria from ornithine proceeds at a near optimal rate at ornithine concentrations as low as 0.08 mM. The same stoichiometry and rates of citrulline synthesis are observed when ornithine is replaced by arginine. The mitochondrial location of arginase does not appear to reflect a mechanism for regulating ornithine availability.     

Analysis of regulatory factors for urea synthesis by isolated perfused rat liver. I. Urea synthesis with ammonia and glutamine as nitrogen sources.

Urea synthesis was studied using the isolated liver perfusion with ammonium cholride and glutamine as nitrogen sources. The rate of urea formation increases with ammonium cholorde concentration up to 5mM, and the rate remained constant in the range between 5 and 20mM of ammonium chloride as the substrate. The concentration of ammonia in the medium to support the half-maximum velocity of urea formation was 0.7mM. The rate of urea formation was stimulated by the addition of 2.5mM ornithine, and the greater part of the ornithine which was taken up into the liver was accumulated as citrulline in the presence of ammonia. A considerable accelerating effect of N-acetylglutamate on the synthetic rate was observed, but a rather high concentration of N-acetylglutamate was required in order to obtain the maximum effect possibly, because its permeability into liver cells may be limited. A marked additive effect on the rate of urea formation was observed with the combined addition of ornithine and N-acetylglutamate. The metabolic conversion of glutamine nitrogen to urea in the perfused rat liver and the effect of several compounds which stimulated urea synthesis with ammonia were further examined. The process of conversion of glutamine nitrogen to urea might be composed of the following three steps. In the first lag phase, a small amount of glutamine was removed from the medium. In the second stage, the glutamine level decreased rapidly and ammonia was accumulated in the perfusate. The third stage was a period in which glutamine concentration remained at a constant low level, and the accumulated ammonia was rapidly conversed to urea. The rate of urea formation in this third stage was found to be much higher than that with ammonia as the substrate. The maximum rate of glutamine removal was obtained at pH 7.7 of the perfusate and at a concentration of 10mM glutamine. Urea formation with glutamine was also stimulated by the addition of ornithine, malate, or N-acetylglutamate, which had accelerating effects on the urea synthesis with ammonia. This stimulation was due to an effective conversion of ammonia to urea, but no change in the rate of removal glutamine was obtained.     

Analysis of ven3 and ven6 reticulate mutants reveals the importance of arginine biosynthesis in Arabidopsis leaf development

Arabidopsis thaliana reticulate mutants exhibit differential pigmentation of the veinal and interveinal leaf regions, a visible phenotype that often indicates impaired mesophyll development. We performed a metabolomic analysis of one ven6 (venosa6) and three ven3 reticulate mutants that revealed altered levels of arginine precursors, namely increased ornithine and reduced citrulline levels. In addition, the mutants were more sensitive than the wild-type to exogenous ornithine, and leaf reticulation and mesophyll defects of these mutants were completely rescued by exogenous citrulline. Taken together, these results indicate that ven3 and ven6 mutants experience a blockage of the conversion of ornithine into citrulline in the arginine pathway. Consistent with the participation of VEN3 and VEN6 in the same pathway, the morphological phenotype of ven3 ven6 double mutants was synergistic. Map-based cloning showed that the VEN3 and VEN6 genes encode subunits of Arabidopsis carbamoyl phosphate synthetase (CPS), which is assumed to be required for the conversion of ornithine into citrulline in arginine biosynthesis. Heterologous expression of the Arabidopsis VEN3 and VEN6 genes in a CPS-deficient Escherichia coli strain fully restored bacterial growth in minimal medium, demonstrating the enzymatic activity of the VEN3 and VEN6 proteins, and indicating a conserved role for CPS in these distinct and distant species. Detailed study of the reticulate leaf phenotype in the ven3 and ven6 mutants revealed that mesophyll development is highly sensitive to impaired arginine biosynthesis.     

Chemical modification of the mitochondrial ornithine/citrulline carrier by SH reagents: effects on the transport activity and transition from carrier to pore-like function

The transport activity of the purified and reconstituted ornithine/citrulline carrier from rat liver mitochondria was correlated to modification of its sulfhydryl groups by various reagents. Both the ornithine/ornithine (antiport) and the ornithine/H(+) (unidirectional) transport modes catalysed by the ornithine/citrulline carrier were inhibited by methanethiosulfonates, mercurial reagents, N-ethylmaleimide (NEM) and 5,5'-dithiobis(2-nitrobenzoate) (DTNB). The treatment of the ornithine/citrulline carrier with mercurial reagents, at concentrations above 5 microM, caused the induction of an additional (pore-like) transport mode, characterized by loss of substrate specificity and a transport activity higher than that of the unmodified carrier. The S-S forming reagent Cu(2+)-phenanthroline inhibited the transport catalysed by the carrier, indicating the presence of close sulfhydryl groups. The effect of consecutive addition of the various reagents revealed a peculiar aspect of the ornithine/citrulline carrier, i.e. the presence of three distinct populations of sulfhydryl groups. The first was responsible for the inhibition of the physiological transport modes by methanethiosulfonates, NEM and DTNB and low concentrations (<5 microM) of mercurials; the second population was responsible for the transition to the pore-like activity induced by higher concentrations (>5 microM) of mercurials; the third population was involved in S-S bridge formation.     

Functional significance of the activities of glutaminase and ornithine-ω-aminotransferase in rat brain

The activities of glutaminase, glutamine synthetase (GS), arginase and ornithine amino transferase (orn-T) were studied in three regions of rat brain in heightened neuronal activity by producing convulsions by leptazol. These enzymes were studied in preconvulsive, convulsive and postconvulsive phases. Glutaminase activity was found to increase in all the three regions in the preconvulsive and convulsive phases. GS activity decreased in the preconvulsive phase but rose gradually to the control level when the postconvulsive phase was reached. The activity of arginase decreased in the cerebellum in preconvulsive and convulsive phases. However, in the cerebral cortex there was a decrease in the activity of this enzyme only in the convulsive phase. The results suggest that glutamine acts more likely as a precursor for the neurotransmitter pool of glutamate, while ornithine serves more as a precursor for the neurotransmitter pool of GABA.     

Regulation of ornithine aminotransferase gene expression and activity by all-transretinoic acid in Caco-2 intestinal epithelial cells

Ornithine aminotransferase (OAT) is a crucial enzyme in the synthesis of citrulline and arginine from glutamine/glutamate and proline by enterocytes of the small intestine. However, a role for OAT in intestinal polyamine synthesis and cell growth is not known. All-transretinoic acid (RA), an active metabolite of vitamin A, regulates the activity of several metabolic enzymes related to OAT, including ornithine decarboxylase and arginase, which may influence the function of OAT through effects on substrate (ornithine) availability. The objective of the present study was to test the hypothesis that RA regulates OAT mRNA expression and enzymatic activity in intestinal epithelial cells. Caco-2 cells were cultured for 12-72 h in the presence of 0, 0.01 and 1 microM RA and then used for measurements of OAT mRNA levels and enzyme activity as well as ornithine and polyamines. Treatment with RA induced increases in OAT gene expression and enzymatic activity, which resulted in decreased intracellular concentrations of ornithine and polyamines (putrescine, spermidine and spermine) in a dose-dependent manner. These changes occurred concomitantly with a decrease in the total number of cells, and the increase in OAT activity was due to increased OAT mRNA expression. In cells treated with 1 microM RA, addition of 10 microM putrescine to culture medium restored both cellular levels of polyamines and cell numbers to the values for the control group (without addition of RA). We conclude that exposure of Caco-2 cells to RA induces OAT expression for increasing ornithine catabolism. This leads to a reduced availability of intracellular ornithine for polyamine synthesis, thereby decreasing cell proliferation. These novel findings indicate a functional role for OAT in regulating intestinal polyamine synthesis and growth.     

The production of extracellular and intracellular free amino acids during aerated fermentation of glucose by Baker’s yeast (Saccharomyces cerevisiae)

During a study of the effects of a high level of NaCl on the content of free intracellular amino acids in baker’s yeast grown in aerated fermentation of glucose it was found (Malaneyet al. 1988, 1989; Malaney and Tanner 1988) that 0.6 mol/L exogenous NaCl significantly increased the content of free intracellular citrulline, glutamine, ornithine, arginine and lysine (all basic amino acids) over that observed at zero mol/L exogenous NaCl. (Exogenous is defined as salt added beyond that present in the mineral salts in the culture medium.) This paper describes the production and relative relationships of both extracellular and free intracellular amino acids byS. cerevisiae under conditions of high NaCl content in the growth medium at pH 5 and 32 °C. For early culture times (6 h), the production of glutamine, citrulline, valine, isoleucine, ornithine, lysine and histidine were all enhanced by the addition of NaCl. For late times (24 h), except for ornithine, the early-time-enhanced amino acids continued to be enhanced by the addition of NaCl. In addition, the yields of several other amino acids also were increased by exogenous salt at this late time. These include aspartic acid, threonine, glutamic acid, cystine, methionine, tyrosine, phenylalanine and arginine. Deceased, August 12, 1988. This study was supported by theNational Science Foundation (Grant No. CPE8209945) in conjunction with the NSF United States — Taiwan Cooperative Science Program.     

Partial characterization of ornithine carbamoyltransferase in three microalgae : anabolic role only

Although the existence of isozymes of ornithine carbamoyltransferase (carbamoylphosphate:l-ornithine carbamoyltransferase, EC 2.1.3.3) in higher plants has been reported, and the possibility exists that one or more of these operates catabolically to produce ornithine and carbamoylphosphate from citrulline and inorganic phosphate, no proof has been forthcoming. In view of the fact that many unicellular algae degrade arginine via arginine deiminase to citrulline and ammonium, and that the pathway of utilization of citrulline is unknown, we decided to investigate the possibility of the presence of a catabolic form of ornithine carbamoyltransferase in three microalgae known to have arginine deiminase activity. These were Chlorella autotrophica, Chlorella saccharophila, and Dunaliella tertiolecta. Our results show that the properties of OCT from these three algae are similar to OCTs from many higher plants with respect to general kinetics (K_m values for ornithine and carbamoylphosphate), substrate inhibition by ornithine at high pHs, apparent sequential ordered kinetic mechanisms and paucity of apparent regulatory properties. Our data indicate an exclusively anabolic role of ornithine carbamoyltransferase in these algae.     

Decreased glutamate, glutamine and citrulline concentrations in plasma and muscle in endotoxemia cannot be reversed by glutamate or glutamine supplementation: a primary intestinal defect?

Endotoxemia affects intestinal physiology. A decrease of circulating citrulline concentration is considered as a reflection of the intestinal function. Citrulline can be produced in enterocytes notably from glutamate and glutamine. The aim of this work was to determine if glutamate, glutamine and citrulline concentrations in blood, intestine and muscle are decreased by endotoxemia, and if supplementation with glutamate or glutamine can restore normal concentrations. We induced endotoxemia in rats by an intraperitoneal injection of 0.3?mg?kg^?1 lipopolysaccharide (LPS). This led to a rapid anorexia, negative nitrogen balance and a transient increase of the circulating level of IL-6 and TNF-α. When compared with the values measured in pair fed (PF) animals, almost all circulating amino acids (AA) including citrulline decreased, suggesting a decrease of intestinal function. However, at D2 after LPS injection, most circulating AA concentrations were closed to the values recorded in the PF group. At that time, among AA, only glutamate, glutamine and citrulline were decreased in gastrocnemius muscle without change in intestinal mucosa. A supplementation with 4% monosodium glutamate (MSG) or an isomolar amount of glutamine failed to restore glutamate, glutamine and citrulline concentrations in plasma and muscle. However, MSG supplementation led to an accumulation of glutamate in the intestinal mucosa. In conclusion, endotoxemia rapidly but transiently decreased the circulating concentrations of almost all AA and more durably of glutamate, glutamine and citrulline in muscle. Supplementation with glutamate or glutamine failed to restore glutamate, glutamine and citrulline concentrations in plasma and muscles. The implication of a loss of the intestinal capacity for AA absorption and/or metabolism in endotoxemia (as judged from decreased citrulline plasma concentration) for explaining such results are discussed.     

Effect of cholera toxin on glutamine metabolism and transport in rabbit ileum

The aim of the present study was to evaluate the effect of cholera toxin on energy balance from intestinal glutamine metabolism and oxidation, glutamine-dependent sodium absorption, and cholera toxin-dependent ion flux. Cholera toxin-stimulated sodium and L-glutamine ileal transport and metabolism were studied in Ussing chambers. Glutamine (10 mM) transport and metabolism were simultaneously studied using (14)C flux and HPLC. In the same tissues, the flux of each amino acid was studied by HPLC, and glutamine metabolism and oxidation were studied by the determination of amino acid specific activity and (14)CO(2) production. In control tissues, glutamine stimulated sodium absorption and was mainly oxidized. The transepithelial flux of intact glutamine represented 45% of glutamine flux across the luminal membrane. The other metabolites were glutamate and, to a lesser degree, citrulline, ornithine, and proline. Cholera toxin did not alter glutamine-stimulated sodium absorption, glutamine oxidation, transport, and metabolism. In conclusion, the present results indicate that cholera toxin does not alter glutamine intestinal function and metabolism. In addition, approximately 95% of the energy provided by glutamine oxidation remains available to the enterocyte.