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Randomized clinical outcome study of critically ill patients given glutamine-supplemented enteral nutrition.

Glutamine is normally an abundant amino acid in the body. It has many important metabolic roles, which may protect or promote tissue integrity and enhance the immune system. Low plasma and tissue levels of glutamine in the critically ill suggest that demand may exceed endogenous supply. A relative deficiency of glutamine could compromise recovery, resulting in prolonged illness and an increase in late mortality, morbidity, and consequently hospital costs. Using a prospective block-randomized, double-blind treatment study design, we tested whether a glutamine-containing enteral feed compared with an isonitrogenous, isoenergetic control feed would influence outcome. The study endpoints were morbidity, mortality, and hospital cost at 6 mo postintervention. In one general intensive care unit (ICU), to ensure consistency of management policies, 78 critically ill adult patients with Acute Physiological and Chronic Health Evaluation (APACHE) II score of 11 and greater and who were considered able to tolerate introduction of enteral nutrition were studied. Fifty patients successfully received enteral nutrition (26 glutamine, 24 control). There was no mortality difference between those patients receiving glutamine-containing enteral feed and the controls. However, there was a significant reduction in the median postintervention ICU and hospital patient costs in the glutamine recipients $23,000 versus $30,900 in the control patients (P = 0.036). For patients given glutamine there was a reduced cost per survivor of 30%. We conclude that in critically ill ICU patients enteral feeds containing glutamine have significant hospital cost benefits.

Adult↗

Epidermal growth factor activation of intestinal glutamine transport is mediated by mitogen-activated protein kinases.

Glutamine is an essential nutrient for gut functions, but the regulation of its uptake by intestinal mucosal cells is poorly understood. Given the pivotal role of epidermal growth factor (EGF) in regulating gut metabolism, growth, and differentiation, this in vitro study was designed to investigate the intracellular signaling pathways involved in the regulation of EGF-mediated intestinal glutamine transport in intestinal epithelia. Continuous incubation with EGF (>30 hours, 100 ng/ml) stimulated glutamine transport activity across intestinal epithelial Caco-2 cell apical membrane. Exposure to EGF for 48 hours resulted in an increase in transport activity (50%) and glutamine transport system B gene ATB(0) mRNA levels (ninefold). EGF stimulated glutamine transport activity by increasing the glutamine transporter maximal velocity (V(max)) without altering the transporter apparent affinity (K(m)). Furthermore, EGF stimulated both intracellular protein kinase C and mitogen-activated protein kinase MEK1/2 activities. The EGF-stimulated glutamine transport activity was attenuated individually by the specific protein kinase C inhibitor chelerythrine chloride and the mitogen-activated protein kinase MEK1 inhibitor PD 98059. These data suggest that EGF activates glutamine transport activity across intestinal epithelial membrane via a signaling mechanism that involves activation of protein kinase C and the mitogen-activated protein kinase MEK1/2 cascade. EGF activates glutamine transport via alterations in transporter mRNA levels and the number of functional copies of transporter units.

Amino Acid Transport System ASC↗

Kinetic analysis of L-glutamine transport into porcine jejunal enterocyte brush-border membrane vesicles.

L-Glutamine transport into porcine jejunal enterocyte brush border membrane vesicles was studied. Uptake was mediated by a Na(+)-dependent and a Na(+)-independent pathway as well as by diffusion. The initial rates of glutamine uptake over a range of concentrations is both Na(+)-gradient and Na(+)-free conditions were analyzed and kinetic parameters were obtained. Na(+)-dependent glutamine transport had a K(m) of 0.77 +/- 0.16 mM and a Jmax of 70.7 +/- 5.8 pmol mg protein-1 s-1; Na(+)-independent glutamine transport had a K(m) of 3.55 +/- 0.78 mM and a Jmax of 55.1 +/- 6.6 pmol mg protein-1 s-1. The non-saturable component measured with HgCl2-poisoned brush border membrane vesicles in the Na(+)-free condition contained passive diffusion and non-specific membrane binding and was defined to be apparent glutamine diffusion and the glutamine permeability coefficient (Kdiff) was estimated to be Kdiff = 3.78 +/- 0.06 pmol 1 mg protein-1 mmol-1 s-1. Results of inhibition experiments showed that Na(+)-dependent glutamine uptake occurred primarily through the brush border system-B degree transporters, whereas Na(+)-independent glutamine uptake occurred via the system-L transporters. Furthermore, the kinetics of L-leucine and L-cysteine inhibition of L-glutamine uptake demonstrated that neutral amino acids sharing the same brush border transporters can effectively inhibit each other in their transport.

Animals↗

Structures and hydration enthalpies of cationized glutamine and structural analogues in the gas phase.

The structures of lithiated and sodiated glutamine, both with and without a water molecule, are investigated using experiment and theory. Loss of water from these complexes and from lithiated and sodiated complexes of asparagine methyl ester, asparagine ethyl ester, and glutamine methyl ester is probed with blackbody infrared radiative dissociation experiments performed over a wide temperature range. Threshold dissociation energies, E(o), for loss of a water molecule from these complexes are obtained from master equation modeling of these data. The values of E(o) are 63 +/- 1 and 53 +/- 1 kJ/mol for the lithiated and sodiated glutamine complexes, respectively. These values are similar to those for the nonzwitterionic model complexes and are in excellent agreement with calculated values. In contrast, water binding to the zwitterionic form is calculated to be significantly higher. These results indicate that glutamine in these lithiated and sodiated complexes with a water molecule are nonzwitterionic. Complexes with the asparagine side chain have slightly higher E(o) values than those with the glutamine side chain, a result consistent with more effective solvation of the metal ion due to the slightly longer side chain of glutamine. Calculations indicate that lithiated and sodiated glutamine are nonzwitterionic, with the metal ion interacting with the amine nitrogen and carbonyl oxygen from the amino acid backbone and the amide oxygen of the side chain. Addition of a water molecule does not affect the lowest-energy structure of lithiated glutamine, whereas, for sodiated glutamine, the lowest-energy zwitterionic and nonzwitterionic structures are essentially isoenergetic.

Cations↗

Effects of potassium and glutamine on metabolism of glucose in astrocytes.

The metabolic effects of extracellular glutamine (2.5 mM) or high potassium (25 mM) on glucose metabolism were studied in cultured cerebellar astrocytes. High potassium caused an increased glycolytic flux and an increase in glutamine release. Exposure to glutamine increased glycolytic flux and alanine formation, indicating that glutamine uptake is an energy requiring process. The effects of glutamine and high potassium on glycolytic flux were additive. Formation of metabolites from [1-13C]glucose and [2-13C]acetate confirmed the effects of glutamine and high potassium on glycolytic metabolism. In the presence of extracellular glutamine, analysis of the 13C labeling patterns of citrate and glutamine indicated a decrease in the cycling ratio and/or pyruvate carboxylation and glutamine synthesis from [1-13C]glucose did occur, but was decreased. Exposure to high potassium led to extracellular accumulation of acetate, presumably through non-enzymatic decarboxylation of pyruvate.

Acetates↗

Glutamine transport in rat brain synaptic and non-synaptic mitochondria.

Glutamine transport into rat brain mitochondria (synaptic and non-synaptic) was monitored by the uptake of [3H]glutamine as well as by mitochondrial swelling. The uptake is inversely correlated to medium osmolarity, temperature-dependent, saturable and inhibited by mersalyl, and glutamine is upconcentrated in the mitochondria. These results indicate that glutamine is transported into an osmotically active space by a protein catalyzed mechanism. The uptake is slightly higher in synaptic mitochondria than in non-synaptic ones. It is inhibited both by rotenone and the protonophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone, the latter at pH 6.5, showing that the transport is activated by an electrochemical proton gradient. The K+/H+ ionophore nigericin also inhibits the uptake at pH 6.5 in the presence of external K+, which indicates that glutamine, at least in part, is taken up by a proton symport transporter. In addition, glutamine uptake as measured by the swelling technique revealed an additional glutamine transport activity with at least 10 times higher Km value. This uptake is inhibited by valinomycin in the presence of K+ and is thus also activated by the membrane potential. Otherwise, the two methods show similar results. These data indicate that glutamine transport in brain mitochondria cannot be described by merely a simple electroneutral uniport mechanism, but are consistent with the uptake of both the anionic and the zwitterionic glutamine.

Animals↗

Effect of blood ammonia elevation following oral glutamine load on the psychometric performance of cirrhotic patients.

Oral glutamine challenge is a method to increase blood ammonia and may be used to study the ammonia lowering effect of drugs potentially useful in hepatic encephalopathy (HE). We tested its influence on the psychometric performance of 18 cirrhotic patients without HE. Twelve nonencephalopatic cirrhotic patients were studied before and after glutamine load (20 g in 100 mL tap water) and six patients before and after placebo (100 mL tap water) by using the Number Connection Test (NCT), the Covert Visual Attention Orienting Test (CVAOT), and the Scan Test (SCT). Blood ammonia increased significantly after glutamine (from 79 +/- 34 to 211 +/- 66 microg/dL) but not after placebo (from 94 +/- 41 to 88 +/- 26). No difference in the NCT was found before and after glutamine load or placebo. The CVAOT was similar after glutamine challenge and placebo, nor any interaction between Loads (glutamine or placebo) x Cue position was found, suggesting that glutamine load did not influence attention-orienting. SCT results were also similar after glutamine and placebo, suggesting a lack of influence on the working memory. Glutamine challenge is a safe method to induce hyperammonemia in nonencephalopatic cirrhotic patients and, therefore, to study the efficacy of ammonia lowering treatments.

Administration, Oral↗

Role of manganese accumulation in increased brain glutamine of the cirrhotic rat.

Cirrhosis promotes increases of both manganese and glutamine in brain. Manganese is a modulator and glutamine is the product of glutamine synthetase. This work studies the relationship between manganese and glutamine synthetase in a model of cirrhosis in the rat. We administered manganese (1 g/L) in the drinking water of sham-operated and bile-duct obstructed rats. We evaluated the manganese and glutamine accumulation and the glutamine synthetase activity in frontal cortex, striatum, and pallidum after 2, 4, and 6 weeks of biliary obstruction or sham surgery. Cirrhotic rats receiving manganese increased their brain content of metal about 400%-600% after 4 weeks of treatment (P < .05) and also remarkably accumulated glutamine through time in the three regions studied (P < .05 at week 6). Interestingly, bile-duct obstructed rats treated with manganese showed no effect on glutamine synthetase activity. Results from this study suggest that manganese induces increases of brain glutamine independently of its synthesis.

Animals↗

Glutamine distribution in mitochondria from normal and acidotic rat kidneys.

Mitochondria from control and acidotic rat renal cortex were incubated with labeled compounds to determine the distribution of L-glutamine. The mitochondrial preparations lacked detectable phosphate-independent glutaminase activity. In control mitochondria, the distribution of glutamine exceeded that of mannitol by 8% without an added energy source in the medium and by 9.9% when tetramethyl-p-phenylenediamine (TMPD) and ascorbate were present. The distribution of D-ribose (MW 150) was similar to that of L-glutamine (MW 146), indicating that these molecules penetrate into a slightly larger space than D-mannitol (MW 182). Inhibition of phosphate-dependent glutaminase activity with 5 mM glutamate did not alter the volume of glutamine distribution. In mitochondria from chronically acidotic rats, the glutamine/mannitol ratio was 1.03 in the absence of an energy source. With an energy source this ratio fell to 0.88 due to metabolism of glutamine during the separation process. Inhibition of glutamine metabolism with glutamate or alpha-ketoglutarate resulted in distribution ratios of 1.07 and 0.97, respectively. It is concluded that, although glutamine/mannitol ratios greater than 1.00 can be demonstrated in rat kidney mitochondria, such a finding may arise from causes other than the presence of free glutamine in the matrix space. Thus, it has yet to be established whether the inner membrane carrier for glutamine releases glutamine or glutamate into the matrix space.

Acidosis, Renal Tubular↗

Hormonal regulation of glutamine metabolism by OK cells.

The precise mechanism(s) of action of PTH, insulin or glucagon in the regulation of renal glutamine and ammonia metabolism is unknown. Our aim was to delineate the effects and the site(s) of action of these hormones on renal glutamine metabolism. Experiments were carried out using OK cells as a model system. Cell cultures were incubated for three hours in a bicarbonate buffer of pH 7.4 supplemented with either 1 mM [2-15N] or [5-15N] glutamine and 10(-7) M PTH, insulin or glucagon. Comparative studies were performed at pH 6.8, 7.4 or 7.6 without hormone. PTH and acute acidosis significantly stimulated glutamine metabolism via both the phosphate-dependent glutaminase (PDG) and glutamate dehydrogenase (GLDH) pathways. The opposite was observed at pH 7.6. Insulin augmented flux via PDG with little effect on the GLDH pathway. Glucagon had insignificant effects on either PDG or GLDH pathways. Intracellular [15N] glutamate formed from [2-15N] glutamine was removed partially by transamination to alanine, aspartate and serine and partially by translocation to an extracellular compartment. Acidosis, PTH and insulin enhanced the formation of [15N] alanine with little effect on [15N] aspartate. PTH, insulin and glucagon significantly stimulated the production of [15N]serine, whereas acidosis had little effect. The translocation of intracellular glutamate was significantly increased by acidosis, PTH and insulin and decreased by acute alkalosis. The data indicate that: (a) PTH mimicks the effect of acute acidosis on renal glutamine metabolism, that is, augmented glutamine metabolism through both PDG and GLDH pathways and stimulated the output of intracellular glutamate. This effect might be mediated via decreased activity of the Na(+)-H+ exchanger associated with cellular acidification and/or through a second messenger; (b) insulin, but not glucagon, increased glutamine uptake and metabolism, and simultaneously enhanced output of intracellular glutamate sufficiently to stimulate the PDG pathway; and (c) overall, glucagon had little effect on glutamine metabolism by OK cells compared with either PTH or insulin.

Adenosine Triphosphate↗

Effect of low-dose oral glutamine on painful stomatitis during bone marrow transplantation.

Painful oral mucositis is a common complication after bone marrow transplantation (BMT). Glutamine is a nutrient for rapidly dividing cells and the major energy source for intestinal epithelium. This study tested whether an oral glutamine preparation could decrease the severity of oral mucositis in patients undergoing BMT. Glutamine or a placebo (glycine) were administered from admission until day +28 in 193 BMT patients in a randomized, double-blind, placebo-controlled study at a dose of 1.0 g amino acid/m2/dose swish and swallow four times a day. In autologous BMT patients (n = 87) glutamine was associated with significantly less mouth pain by self report and by opiate use (5.0+/-6.2 days of morphine for glutamine vs 10.3+/-9.8 days for placebo; P= 0.005). Matched sibling BMT patients had no effect by self report and an increased duration of opiate use (23.2+/-5.7 days for glutamine vs 16.3+/-8.3 days for placebo) (P = 0.002). However, day 28 survival of allogeneic patients was improved by glutamine. No significant differences in TPN use, rate of relapse or progression of malignancy, parenteral antibiotic use, acute or chronic GVHD, or days of hospitalization were observed in either autologous or allogeneic recipients. No toxicity of glutamine was observed. We conclude that oral glutamine can decrease the severity and duration of oropharyngeal mucositis in autologous BMT patients but not in allogeneic BMT patients, possibly due to interaction with methotrexate.

Administration, Oral↗

Glutamate is preferred over glutamine for intermediary metabolism in cultured cerebellar neurons.

The glutamate-glutamine cycle is thought to be of paramount importance in the mature brain; however, its significance is likely to vary with regional differences in distance between astrocyte and synapse. The present study is aimed at evaluating the role of this cycle in cultures of cerebellar neurons, mainly consisting of glutamatergic granule cells. Cells were incubated in medium containing [U-13C]glutamate or [U-13C]glutamine in the presence and absence of unlabeled glutamine and glutamate, respectively. Cell extracts and media were analyzed using high-performance liquid chromatography (HPLC) and gas chromatography combined with mass spectrometry (GC/MS). Both [U-13C]glutamate and [U-13C]glutamine were shown to be excellent precursors for synthesis of neuroactive amino acids and tricarboxylic acid (TCA) cycle intermediates. Labeling from [U-13C]glutamate was higher than that from [U-13C]glutamine in all metabolites measured. The presence of [U-13C]glutamate plus unlabeled glutamine in the experimental medium led to labeling very similar to that from [U-13C]glutamate alone. However, incubation in medium containing [U-13C]glutamine in the presence of unlabeled glutamate almost abolished labeling of metabolites. Thus, it could be shown that glutamate is the preferred substrate for intermediary metabolism in cerebellar neurons. Label distribution indicating TCA cycle activity showed more prominent cycling from [U-13C]glutamine than from [U-13C]glutamate. Labeling of succinate was lower than that of the other TCA cycle intermediates, indicating an active role of the gamma-amino butyric acid shunt in these cultures. It can be concluded that the cerebellar neurons rely more on reuptake of glutamate than supply of glutamine from astrocytes for glutamate homeostasis.

Amino Acids↗

Glutamine metabolism in the kidney during induction of, and recovery from, metabolic acidosis in the rat.

Experiments were carried out on rats to evaluate the possible regulatory roles of renal glutaminase activity, mitochondrial permeability to glutamine, phosphoenolpyruvate carboxykinase activity and systemic acid-base changes in the control of renal ammonia (NH(3) plus NH(4) (+)) production. Acidosis was induced by drinking NH(4)Cl solution ad libitum. A pronounced metabolic acidosis without respiratory compensation [pH=7.25; HCO(3) (-)=16.9mequiv./litre; pCO(2)=40.7mmHg (5.41kPa)] was evident for the first 2 days, but thereafter acid-base status returned towards normal. This improvement in acid-base status was accompanied by the attainment of maximal rates of ammonia excretion (onset phase) after about 2 days. A steady rate of ammonia excretion was then maintained (plateau phase) until the rats were supplied with tap water in place of the NH(4)Cl solution, whereupon pCO(2) and HCO(3) (-) became elevated [55.4mmHg (7.37kPa) and 35.5mequiv./litre] and renal ammonia excretion returned to control values within 1 day (recovery phase). Renal arteriovenous differences for glutamine always paralleled rates of ammonia excretion. Phosphate-dependent glutaminase and phosphoenolpyruvate carboxykinase activities and the rate of glutamine metabolism (NH(3) production and O(2) consumption) by isolated kidney mitochondria all increased during the onset phase. The increases in glutaminase and in mitochondrial metabolism continued into the plateau phase, whereas the increase in the carboxykinase reached a plateau at the same time as did ammonia excretion. During the recovery phase a rapid decrease in carboxykinase activity accompanied the decrease in ammonia excretion, whereas glutaminase and mitochondrial glutamine metabolism in vitro remained elevated. The metabolism of glutamine by kidney-cortex slices (ammonia, glutamate and glucose production) paralleled the metabolism of glutamine in vivo during recovery, i.e. it returned to control values. The results indicate that the adaptations in mitochondrial glutamine metabolism must be regulated by extra-mitochondrial factors, since glutamine metabolism in vivo and in slices returns to control values during recovery, whereas the mitochondrial metabolism of glutamine remains elevated.

Acid-Base Equilibrium↗

Bidirectional transport of glutamine across the cell membrane in rat liver.

Hepatocytes isolated from fed rats were used to investigate glutamine transport. Glutamine transport appears as a composite process involving at least two saturable components. The Na+-dependent component probably represents the entry through the N system. The Na+-independent component was also inhibited by histidine and exhibited trans-stimulation, suggestive of a facilitated diffusion process. Kinetic parameters for both systems suggest that facilitated diffusion only plays a minor role in glutamine influx. In contrast, the Km for glutamine efflux was consistent with a physiological role of the facilitated-diffusion component in glutamine release. In Na+ medium, relatively constant distribution ratios (about 8) between intra- and extra-cellular concentrations were observed, with external glutamine ranging from 0.5 to 5 mM. The present observations suggest that glutamine influx might largely be mediated by the N system, whereas facilitated diffusion allows hepatocytes to release glutamine when intracellular concentrations are elevated. The physiological consequences of this bidirectional transfer of glutamine across the liver cell membrane is discussed.

Amino Acids↗

Glucose and glutamine metabolism in rat thymocytes.

The metabolism of glucose and glutamine in freshly prepared resting and concanavalin A-stimulated rat thymocytes was studied. Concanavalin A addition enhanced uptake of both glucose and glutamine and led to an increase in oxidative degradation of both substrates to CO2. With variously labelled [14C]glucose, it was shown that the pathways of glucose dissimilation were equally stimulated by the mitogen. A disproportionately large percentage of the extra glucose taken up was converted into lactate, but concanavalin A also caused an increase in the oxidation of pyruvate as judged by the enhanced release of 14CO2 from [2-14C]-, [3,4-14C]- and [6-14C]-glucose. Addition of glutamine did not affect glucose metabolism. The major end products of glutamine metabolism by resting and mitogen-stimulated rat thymocytes were glutamate, aspartate, CO2 and NH3. Virtually no lactate was formed from glutamine. Concanavalin A enhanced the formation of all end products except glutamate, indicating that more glutamine was metabolized beyond the stage of glutamate in the mitogen-activated cells. Addition of glucose caused a significant decrease in the rates of glutamine utilization and conversion into aspartate and CO2 in the absence and in the presence of concanavalin A. In the presence of glucose, almost all nitrogen of the metabolized glutamine was accounted for as NH3 released via the glutaminase and/or glutamate dehydrogenase reactions. In the absence of glucose, part (18%) of the glutamine nitrogen was metabolized by the resting and to a larger extent (38%) by the mitogen-stimulated thymocytes via a transaminase or amidotransferase reaction.

Animals↗

Glutamine metabolism in isolated incubated adipocytes of the rat.

1. Phosphate-dependent glutaminase activity in the epididymal fat-pad was 15.1 nmol/min per mg of protein. Glutaminase activity demonstrated differences with respect to adipose-tissue sites. Considerable variation was found in different sites of adipose tissue from lean control and Zucker obese animals. 2. Adipocytes incubated in the presence of 2 mM-glutamine utilized glutamine at a rate of 1.8 mumol/h per g dry wt., and glutamate, ammonia, lactate and alanine were produced. Addition of glucose plus insulin increased the rates of glutamine utilization and glutamate, ammonia, lactate and alanine production. Isoprenaline alone or plus glucose further stimulated the rate of glutamine utilization and formation of end products. 3. The rate of incorporation of 14C from glutamine into CO2 was similar to that of glucose, but the rate of incorporation into triacylglycerol was much less. Addition of unlabelled glucose or glucose plus insulin stimulated the rate of incorporation of [14C]glutamine into triacylglycerol, but had no effect on that of 14CO2 formation. Isoprenaline plus glucose increased the rate of incorporation of [14C]glutamine into CO2, but decreased the rate of incorporation into triacylglycerol. 4. In the absence of insulin, the rate of [14C]glutamine incorporation into triacylglycerol was related to the glucose concentration (0-10 mM). However, in the presence of insulin, the rate of incorporation of [14C]glutamine was maximal at 1 mM-glucose.

Adipose Tissue↗

Glutamine-dependent carbamoyl-phosphate synthetase and other enzyme activities related to the pyrimidine pathway in spleen of Squalus acanthias (spiny dogfish).

The first two steps of urea synthesis in liver of marine elasmobranchs involve formation of glutamine from ammonia and of carbamoyl phosphate from glutamine, catalysed by glutamine synthetase and carbamoyl-phosphate synthetase, respectively [Anderson & Casey (1984) J. Biol. Chem. 259, 456-462]; both of these enzymes are localized exclusively in the mitochondrial matrix. The objective of this study was to establish the enzymology of carbamoyl phosphate formation and utilization for pyrimidine nucleotide biosynthesis in Squalus acanthias (spiny dogfish), a representative elasmobranch. Aspartate carbamoyltransferase could not be detected in liver of dogfish. Spleen extracts, however, had glutamine-dependent carbamoyl-phosphate synthetase, aspartate carbamoyltransferase, dihydro-orotase, and glutamine synthetase activities, all localized in the cytosol; dihydro-orotate dehydrogenase, orotate phosphoribosyltransferase, and orotidine-5'-decarboxylase activities were also present. Except for glutamine synthetase, the levels of all activities were very low. The carbamoyl-phosphate synthetase activity is inhibited by UTP and is activated by 5-phosphoribosyl 1-pyrophosphate. The first three enzyme activities of the pyrimidine pathway were eluted in distinctly different positions during gel filtration chromatography under a number of different conditions; although complete proteolysis of inter-domain regions of a multifunctional complex during extraction cannot be excluded, the evidence suggests that in dogfish, in contrast to mammalian species, these three enzymes of the pyrimidine pathway exist as individual polypeptide chains. These results: (1) establish that dogfish express two different glutamine-dependent carbamoyl-phosphate synthetase activities, (2) confirm the report [Smith, Ritter & Campbell (1987) J. Biol. Chem. 262, 198-202] that dogfish express two different glutamine synthetases, and (3) provide indirect evidence that glutamine may not be available in liver for biosynthetic reactions other than urea formation.

Animals↗

Glutamine transport by basolateral plasma-membrane vesicles prepared from rabbit intestine.

L-Glutamine, a major energy substrate for intestinal epithelial cells, can be extracted from intraluminal contents across the brush-border membrane and from arterial blood via the basolateral membrane. The purpose of the present study was to characterize glutamine transport by the basolateral membrane of rabbit epithelial cells. Transport of glutamine by isolated basolateral-membrane vesicles was mediated by both Na(+)-dependent and Na(+)-independent carriers. Tests were performed to distinguish glutamine uptake by likely transport agencies, including Systems A, ASC, N, IMINO, NBB, L and asc. The Na(+)-dependent glutamine uptake was strongly inhibited by an excess of 2-(methylamino)isobutyric acid (MeAIB), and glutamine was equally effective in inhibiting MeAIB transport. The reciprocal inhibition analysis, as well as a sensitivity to increased H+ concentration, indicates that Na(+)-dependent glutamine transport across the basolateral membrane is mediated by System A. The saturable Na(+)-independent glutamine transport was markedly inhibited by 2-aminobicyclo-[2,2,1]-heptane-2-carboxylic acid ('BCH') and insensitive to changes in assay pH, suggesting uptake via System L rather than System asc. The presence of a Na(+)-dependent carrier to mediate active transport of glutamine across the basolateral membrane is probably essential to ensure a continuous supply of this vital substrate to the enterocyte in the post-absorptive state.

Animals↗