L-glutamine Uptake Research Articles

A role for gamma‐glutamyl transpeptidase and the amino acid transport system xc‐ in cystine transport by a human pancreatic duct cell line.

1. The roles of the gamma-glutamyl cycle and the anionic amino acid transport system xc- in mediating L-cystine uptake were investigated in cultured human pancreatic duct PaTu 8902 cells. This cell line exhibits morphological features of normal pancreatic duct cells and expresses gamma-glutamyl transpeptidase (gamma-GT, EC 2.3.2.2), an enzyme involved in the metabolism and regulation of intracellular glutathione (GSH). 2. Uptake of L-cystine (10 microM) was linear for up to 10 min, temperature dependent, Na+ independent, saturable (Michaelis-Menten constant (Km), 86 +/- 25 microM; maximal velocity (Vmax), 109 +/- 33 nmol (mg protein)-1 h-1) and reduced by 80-90% by a 50-fold excess concentration of L-glutamate and L-homocysteic acid, but not L-aspartate. These transport properties resemble those described for system xc-, which exchanges cystine for intracellular glutamate. 3. Acivicin, a known inhibitor of gamma-GT, decreased gamma-GT activity from 2.58 +/- 0.96 to 0.97 +/- 0.11 mU (mg protein)-1 and decreased the initial rates of L-cystine and L-glutamine uptake by 41-55%. Anthglutin (1-gamma-L-glutamyl-2-(2-carboxyphenylhyl)hydrazine), a structurally different inhibitor of gamma-GT, also caused a concentration-dependent (0.01-1 mM) decrease in gamma-GT activity and L-cystine uptake. 4. Neither acivicin nor anthglutin inhibited the uptake of L-glutamate, a poor substrate for gamma-GT. 5. In the presence of a 500-fold excess concentration of glutamate, which should abolish entry of cystine via system xc-, the remaining fraction of cystine transport was inhibited by 50% by acivicin, suggesting that transport is, in part, dependent on the activity of gamma-GT. 6. Cystine transport was also 60-80% inhibited by a series of gamma-glutamyl amino acids (5 mM) including gamma-glutamyl-glutamate, gamma-glutamyl-glutamine and gamma-glutamyl-glycine. alpha-Dipeptides inhibited cystine transport by only 6-22%. 7. These findings demonstrate that in human pancreatic duct PaTu 8902 cells, cystine uptake is mediated by system xc- (50-60%) and the gamma-glutamyl cycle. Our results provide the first evidence linking gamma-GT with cystine transport in human epithelial cells and are of relevance in view of the importance of cystine as a sulphur amino acid source for GSH synthesis in cells exposed to oxidative stress.

Comparative study of the uptake of L-cysteine and L-cystine in the renal proximal tubule

1. The transport mechanisms of L-cysteine and L-cystine by luminal membrane vesicles isolated from either the proximal convoluted part (pars convoluta) or the proximal straight part (pars recta) of rabbit proximal tubuli were examined. 2. The uptake of L-cysteine in pars convoluta is characterized by a single sodium-dependent transport system (Km = 0.58 mM), whereas the sodiumdependent influx of L-cysteine in pars recta proceeds via transport systems with different affinities (Km1 = 0.03 mM, Km2 = 5.84 mM). An H(+)-gradient enhanced the uptake of L-cysteine in pars recta and only in the presence of a Na(+)-gradient. 3. The presence of a Na(+) gradient stimulated the influx of L-cystine in both parts of the nephron, but to a lesser extent than for L-cysteine. In addition a considerable amount of binding of L-cystine to membrane vesicles was observed in both pars convoluta and pars recta. 4. Stoichiometric studies indicated a coupling ratio of 1 Na(+): 1 amino acid for the transport components involved in the uptake of L-cysteine and L-cystine along the proximal tubule. 5. Competition experiments demonstrated that neutralα-amino acids inhibited the influx of L-cysteine in the proximal tubule. Basic and acidic amino acids had no effect on uptake of L-cysteine in pars convoluta, whereas a slight inhibitory effect of L-lysine and L-arginine was noted in pars recta. 6. Both basic and neutral amino acids, but not L-glutamate inhibited the uptake of L-cystine in pars convoluta. This was in general also the case in pars recta. However, addition of L-proline did not influenced the uptake of L-cystine, and L-phenylalanine, L-asparagine, L-glutamine, L-leucine and L-methionine only inhibited at a high concentration (5 mM).

Cultivation of attenuated hepatitis A virus antigen in a titanium static mixer reactor

The titanium static mixer reactor, demonstrated for a variety of vaccine processes during the late 197s, was investigated for the production of attenuated hepatitis A virus antigen from anchorage-dependent MRC-5 cells. This reactor system used Charles River Biotechnological Services cabinets for monitoring and process control. Cell inoculation protocols, using 6000-10,000 cells/cm(2), resulted on over 95% attachment at both the laboratory and pilot scales. Indirect monitoring techniques using oxygen, glucose, L-serine, and L-glutamine uptake rates were indicative of cell growth prior to virus inoculation as well as environmental and/or nutrient limitations. Seven laboratory-scale (3900 cm(2)) runs and one pilotscale (265,000 cm(2)) run were conducted to investigate refeeding regiments, parallel versus perpendicular element orientation, increased element surface area per unit volume, and scale-up performance. In general, lysate antigen yields achieved were similar to those of parallel T-flasks cultivated under similar conditions.

Transepithelial phosphate transport in rabbit proximal tubular cells adapted to phosphate deprivation.

Both renal and nonrenal cells in culture adapt to deprivation of Pi by increasing Na-dependent Pi uptake. We studied whether this change in uptake is reflected in an increased renal transepithelial Pi transport. We grew primary cultures of rabbit renal cortical cells in plastic flasks and subcultured them onto Millicell-HA filters. This produced cell monolayers, which structurally and functionally resembled proximal tubule. These cells performed Na-dependent net transepithelial transport of 32Pi in the apical-to-basolateral direction that was inhibited by phosphonoformic acid in the apical fluid or by ouabain in the basolateral fluid or by preincubation with parathyroid hormone. Overnight incubation at low Pi concentrations led to a progressive increase in 5-min Na-dependent Pi uptake into cell monolayers. Na-dependent Pi uptake was threefold higher following overnight incubation at 25 microM Pi, compared with 3 mM Pi, and the increase was one-half maximal with incubation at an extracellular Pi concentration ([Pi]) of 300 microM. This was associated with a decrease in Na-dependent transepithelial Pi flux to the basolateral fluid by the same cells, which fell dramatically following incubation at < or = 300 microM Pi. There was no change in Na-dependent uptake or transepithelial transport of L-glutamine. This adaptation to Pi deprivation in vitro appears to serve to restore depleted cell stores of Pi rather than to regulate transepithelial Pi transport.

Characteristics of L-glutamine transport in equine jejunal brush border membrane vesicles

SUMMARY The sodium-dependent transporter system responsible for L-glutamine uptake by brush border membrane vesicles prepared from equine jejunum was characterized. Vesicle purity was ascertained by a 14- to 17-fold increase in activity of the brush border enzyme markers. Glutamine uptake was found to occur into an osmotically active space with negligible membrane binding. The sodium-dependent velocity represented approximately 80% of total uptake and demonstrated overshoots. Kinetic studies of sodium-dependent glutamine transport at concentrations between 5 μM and 5 mM revealed a single saturable high-affinity carrier with a Michaelis constant of 519 ± 90 μM/ and a maximal transport velocity of 3.08 ± 0.97 nmol/mg of protein/10 s. Glutamine uptake was not affected by changes in environmental pH. Lithium could not substitute for sodium as a contransporter ion. 2-Methylaminoisobutyric acid inhibited the sodium-dependent carrier only minimally, but marked inhibition (&gt; 90%) was observed in the presence of histidine, alanine, cysteine, and nonradioactive glutamine. Kinetic analysis of the sodium-independent transporter revealed it to have a Michaelis constant = 260 ± 47 μM and a maximal transport velocity of 0.32 ± 0.06 nmol/mg of protein/10 s. We conclude that glutamine transport in equine jejunal brush border membrane vesicles occurs primarily via the system B transporter and, to a lesser extent, by a sodium-independent carrier.

Regulation of rat erythrocyte l-glutamine, 1-glutamate and l-lysine uptake by short term starvation

1. 1. The kinetic parameters ( K m , V max and K d of l-glutamine, l-glutamate and l-lysine uptake by isolated red blood cells in fed and 24 hr starved rats have been determined. 2. 2. l-Lysine and l-glutamine uptake was best fitted by a two transport component: a saturable component and a diffusion one. 3. 3. Starvation brought about important decreases in the K m and V max for both l-lysine and l-glutamine uptake. 4. 4. The K d for l-glutamine showed a significant increase whereas that corresponding to l-lysine did not change by starvation. 5. 5. l-Glutamate uptake adjusted to diffusion kinetics, with a K d which did not change due to starvation. 6. 6. It is concluded that the amino acid uptake showed specific regulation by starvation. 7. 7. The mechanism involved is not dependent on protein synthesis—given the unnucleated nature of mammal red cells. 8. 8. The magnitude of the changes observed in the uptake kinetic parameters may account for the extent of the blood amino acid pool changes as those produced in vivo over physiological limits.

Kinetics of the sodium-dependent glutamine transporter in human intestinal cell confluent monolayers

The intestinal epithelium metabolism of glutamine plays a critical role in inter-organ nitrogen flow. Although it is known that glutamine is the primary oxidative energy source and nucleotide precursor in intestinal cells, the luminal uptake of glutamine by the apical surface of enterocytes is poorly understood. In this study we have uncovered the sodium-dependent transporter system responsible for L-glutamine uptake by the apical membrane of a human intestinal epithelial cell line. The sodium-dependent Michaelis constant (Km) = 247 ± 45 μM glutamine, and Jmax = 4.44 ± 0.65 × 10 −9 mole min −1(mg protein) −1 (37°C). Glutamine shares the transporter with alanine, as demonstrated by unlabeled glutamine inhibition of [ 3H]alanine uptake kinetics with a purely competitive-type inhibition pattern, and glutamine inhibition Ki = 20 ± 18 μM by Dixon analysis. The inhibition pattern for a series of amino acid analogs indicated that this intestinal apical membrane sodium-dependent transporter for glutamine is distinct from any other transport system found in membranes of non-intestinal cells.

Amino acid uptake by liver of genetically obese Zucker rats

Alanine and glutamine uptake by the liver of 50-52-day-old genetically obese Zucker rats and their lean littermates has been studied. The net uptake in vivo of L-alanine is 2-fold higher in the obese animals. No significant change in L-glutamine net balance was found. We also studied the Na(+)-dependent uptake of L-alanine and L-glutamine into plasma-membrane vesicles isolated from either obese- or lean-rat livers. Vmax. values of both L-alanine and L-glutamine transport were 2-fold higher in those preparations from obese rats. No change in Km was observed. As suggested by inhibition studies, this seemed to be mediated by an enhancement of the activities of systems A, ASC and N. We conclude that the liver of the obese Zucker rat is extremely efficient in taking up neutral amino acids from the afferent blood, which results in an enhanced net uptake of L-alanine in vivo. The changes in transport activities at the plasma-membrane level might contribute to increase amino acid disposal by liver, probably for lipogenic purposes, as recently reported by Terrettaz & Jeanrenaud [Biochem. J. (1990) 270, 803-807].

Kinetics and Na independence of amino acid uptake by blood side of perfused sheep choroid plexus

The kinetic constants and sodium independence of amino acid uptake by the basolateral face (blood side) of the isolated perfused choroid plexus of the sheep were investigated. Uptake of 3H-labeled L-alanine, glycine, L-glutamine, L-leucine, L-glutamate, and L-lysine was not significantly inhibited when the sodium level of the perfusate was lowered to less than 6 mM. The cerebrospinal fluid (CSF) secretion rate was, however, reduced by 65.6%. The Michaelis constants for L-serine, glycine, L-phenylalanine, L-glutamate, and L-arginine were measured and varied between 25.4 microM for L-arginine and 2.6 microM for L-glutamate. The Vmax for the amino acids varied by about the same range as Km, with L-serine having the highest Vmax of 28.8 nmol.min-1.g-1, and L-phenylalanine having the lowest at 4.6 nmol.min-1.g-1. At normal plasma concentrations of 30-155 microM for individual amino acids, the carriers for glycine, L-phenylalanine, and L-glutamate would be almost fully saturated (greater than 90%), while those for L-serine and L-arginine would be greater than 60% saturated. This restricted uptake of amino acids by the basolateral face of the choroid plexus, coupled to clearance of these substances out of the CSF (5), could account for the low levels in the CSF compared with that in the plasma.

L-glutamine and d-glucose uptake by developing endosperms of maize

Developing maize ( Zea mays) kernels were incubated with l-[G- 3H]glutamine and d-[U- 14C]glucose to characterize mechanisms of amino acid and sugar uptake by the endosperm tissue. The results indicate that mechanisms Of l-glutamine uptake are different from those of d-glucose. In contrast to d-glucose uptake, l-glutamine uptake showed substrate-saturable kinetics and was inhibited by metabolic inhibitors and protein reagents. The pH optimum for l-glutamine uptake(pH 5.0–5.5) was lower than the optimum of d-glucose uptake (pH 7). EDTA (10mM) stimulated l-glutamine uptake but inhibited d-glucose uptake. The uptake of d-glucose is not a simple diffusion process. l-Glucose, a stereoisomer of d-glucose, was taken up by endosperms at a much slower rate than was d-glucose, indicating a high degree of specificity of the sugar uptake system. The involvement of basal endosperm transfer cells in transport of assimilates into developing maize endosperms is discussed.

Amino Acid Metabolism and Water Transport Across the Seawater Eel Intestine

ABSTRACT To elucidate how intracellular L-alanine enhances water transport across the seawater eel intestine, effects of various metabolic inhibitors were examined. The L-alanine-induced water flux was inhibited by amino-oxyacetate, an inhibitor of aminotransferase. After blocking the synthesis of pyruvate from L-alanine with this drug, water transport was stimulated with pyruvate, whose effects were inhibited by oxythiamine, an inhibitor of pyruvate dehydrogenase. 2,4-Dinitro-phenol (DNP) also inhibited the effects of L-alanine. Furthermore, L-alanine enhanced ouabain-sensitive O2 consumption in this tissue, and the enhancement in O2 consumption preceded that in the transepithelial potential difference (PD) and the net water flux. These results indicate that L-alanine is metabolized through the citric acid cycle to produce ATP, and that a metabolic product stimulates ion and water transport. L-Glutamine also seems to be metabolized just like L-alanine because: L-glutamine acted from inside the enterocyte; DNP inhibited the effects of L-glutamine; neither of the effects of L-glutamine and L-alanine were additive but they were mutually complementary; L-glutamine also enhanced ouabain-sensitive O2 consumption; and the increment in O2 consumption preceded that in the PD and the net water flux. The effects of L-glutamine on the PD and the net water flux depended on glutamine concentration and the concentration-response curve was of the Michaelis-Menten type, indicating that the rate of L-glutamine uptake into the enterocyte limits the overall rate of L-glutamine metabolism. A regulatory role of amino acids for ion and water transport is discussed.

Characteristics of L-glutamine transport in perfused rat skeletal muscle.

1. We have investigated glutamine transport in the perfused rat hindlimb using the paired-tracer isotope dilution technique. 2. Uptake of L-glutamine was stereospecific, saturable, sodium dependent, insulin sensitive and pH insensitive in the physiological range. The maximum capacity of transport (Vmax) under normal perfusate conditions at 37 degrees C, 145 mM-Na+ and in the absence of insulin was 1156 +/- 193 nmol min-1 g-1 with transport being half-maximal at a perfusate glutamine concentration of 9.25 +/- 1.15 mM. 3. The kinetics of Na+ dependence strongly suggested co-transport of Na+ and glutamine with a stoichiometry of 1:1; furthermore, Na+ activated the carrier without any change in the concentration of glutamine at which transport was half-maximal, i.e. a 'Vmax effect' rather than a 'Km effect'. 4. The characteristics of glutamine transport, especially its substrate specificity and the pattern of competitive and non-competitive inhibition of glutamine transport by other amino acids, suggest that it is mediated by a carrier or carriers for which asparagine and histidine are also suitable substrates. 5. The characteristics of muscle glutamine transport are related but distinct from those of system N identified in hepatocytes; we suggest that they are sufficiently distinct to justify the identification of a new variant of mammalian amino acid transport systems which may be identified by the symbol Nm. 6. The kinetic characteristics of system Nm are such that glutamine is likely to be the most rapidly exchanging amino acid across the muscle membrane at physiological intra- and extracellular glutamine concentrations. Its hormone and ion sensitivities are likely to be important in the physiological modulation of whole-body glutamine metabolism and also during derangements observed in disease and after injury.

Effects of fasting on citrate transport by the brush-border membrane of rat kidney.

Fasting in rats decreases plasma citrate levels and reduces urinary citrate excretion by the kidney. After 72 h of fasting, the endogenous renal citrate clearance was decreased and the fractional citrate excretion was 0.026 +/- 0.008 compared with 0.218 +/- 0.030 in control fed rats. To determine whether these findings result from an adaptation in citrate transport across the plasma membrane of the renal tubular cell, Na+ gradient-dependent [14C]citrate uptake was examined in brush-border membrane vesicles (BBMW) prepared from kidneys of fed and 72-h fasted rats. The initial rate (10 s) of Na+ gradient-stimulated uptake of 100 microM citrate was significantly increased in BBMW from kidneys of fasted rats (380 +/- 24.9 pmol/mg prot) compared with fed rats (255 +/- 24.9 pmol/mg prot). Arterial acid-base parameters from conscious animals were similar between the two groups. There was no significant difference in Na+-independent citrate uptake or in L-glutamine uptake measured at 20 s in BBMV from the kidneys of fasted compared with fed rats. An adaptation occurs in the brush-border membrane of the renal tubular cell of fasting rats, unrelated to systemic acidosis, that may result in increased reabsorption of citrate.

Amino Acid Transport in Protoplasts Isolated from Soybean Leaves

We isolated large quantities of mesophyll protoplasts from source and sink leaves of soybean plants and examined them for amino acid uptake. Accumulation of amino acids in isolated protoplasts was linear for at least 40 minutes. Uptake kinetics revealed the presence of both saturable and linear components. Increasing external pH decreases the uptake. The uncoupler, carbonyl cyanide p-trifluoromethoxyphenylhydrazone at 15 micromolar inhibited and fusicoccin at 10 micromolar stimulated amino acid uptake. Our data are consistent with a proton-cotransport mechanism for the uptake of l-glutamine and alpha-amino isobutyric acid into soybean mesophyll cells.

The Transport of L-Glutamate by Rhizobium leguminosarum Involves a Common Amino Acid Carrier

Rhizobium leguminosarum MNF3841 grown on glucose/NH4Cl constitutively transported l-asparagine, l-aspartate, l-glutamate, l-glutamine, glycine, l-leucine, l-methionine and l-phenylalanine. Transport rates were increased 1.5-4-fold by growth on glucose/l-glutamate. Uptake of L-glutamate, l-glutamine, l-asparagine and l-leucine was inhibited to varying extents by a broad range of l-amino acids. Analogues of l-glutamate in which the amino group or α-hydrogen was methylated inhibited L-glutamate transport much less effectively. Also while 2-and 3-amino acids interfered with l-glutamate uptake, d-glutamate did not. Inhibition by 2,4-dinitrophenol, carbonyl cyanide m-chlorophenylhydrazone and cyanide indicated that amino acid transport was active. The ratio of the intracellular to extracellular concentration of l-leucine after 5 min accumulation was 768. Cells loaded with l-[14C]leucine exhibited exchange not only with external l-leucine but also with l-glutamate. The apparent K m for l-glutamate transport was 0·081 μm. Both l-aspartate and l-alanine were competitive inhibitors of l-glutamate uptake with apparent K i values of 0·164 μm and 2·3 μm, respectively. These results suggest that there is an extremely high affinity carrier for l-glutamate that is not only very sensitive to inhibition by l-aspartate but also capable of being inhibited by a broad range of amino acids at an order of magnitude higher concentration.

Glutamine transport in renal basolateral vesicles from dogs with metabolic acidosis.

To determine whether the increased ammonia production per nephron in chronic metabolic acidosis is accompanied by augmented L-glutamine transport across the basolateral membrane of the renal cortical cell and consequent increased availability of this ammoniagenic amino acid, we measured L-[3H]glutamine transport in basolateral membrane vesicles (BLMV) isolated from kidneys of normal and acidotic dogs. Na+ -dependent electrogenic transport of L-[3H]glutamine was demonstrated in BLMV from kidneys of normal dogs that exhibited saturability over the concentration range of 25 microM to 2 mM L-glutamine. The apparent Km was 416 +/- 114 microM and Vmax was 536 +/- 129 pmol X mg protein-1 X 15 s-1. The initial rate of Na+ -dependent L-[3H]glutamine transport was increased in BLMV from kidneys of acidotic dogs, as reflected by an increased apparent Vmax. We conclude that an adaptation resulting in greater uptake of L-glutamine across the basolateral membrane of the renal cortical cell may underlie, in part, the increased rate of ammonia production per nephron seen in chronic metabolic acidosis.

Neutral amino acid transport in isolated rat pancreatic islets.

The neutral amino acid transport systems of freshly isolated rat pancreatic islets have been studied by first examining the transport of L-alanine and the nonmetabolizable analogue 2-(methylamino)isobutyric acid (MeAIB). By comparing the uptake of MeAIB and L-alanine for their pH dependency profile, choline and Li+ substitution for Na+, tolerance to N-methylation, and competition with other amino acids, the existence in pancreatic islets of both A and ASC amino acid transport systems was established. The systems responsible for the inward transport of five natural amino acids was studied using competition analysis and Na+ dependency of uptake. These studies defined three neutral amino acid transport systems: A and ASC (Na+-dependent) and L (Na+-independent). L-Proline entered rat islet cells mainly by system A; L-leucine by the Na+-independent system L. The uptake of L-alanine, L-serine, and L-glutamine was shared by systems ASC and L, the participation of system A being negligible for these three amino acids. An especially broad substrate specificity for systems L and ASC is therefore suggested for the rat pancreatic islet cells. The regulation of amino acid transport was also investigated in two conditions differing as to glucose concentration and/or availability, i.e. islets from fasted rats and islets maintained in tissue culture at high or low glucose concentrations. Neither alanine nor MeAIB transport was altered by fasting of the islet-donor rats. On the other hand, pancreatic islets maintained for 2 days in tissue culture at high (16.7 mM) glucose transported MeAIB at twice the rate of islets maintained at low (2.8 mM) glucose. Amino acid starvation of pancreatic islets during 11 h of tissue culture resulted in a 2-fold increase in MeAIB transport.

Uptake of L-glutamine into synaptosomes. Is the γ-glutamyl cycle involved?

Transport of L-glutamine into rat cortical synaptosomes has been investigated by ( 14C)L-glutamine uptake experiments. This amino acid enters synaptosomes both by an active carrier mediated system, which may be the result of γ-glutamyl cycle activity and by a Na +-dependent transport system. This view is supported by the following observations : a) as demonstrated previously (10), glutamine inside synaptosomes reaches concentrations higher than those of the incubation medium, and initial rates of uptake approach saturation kinetics ; b) the uptake of glutamine is inhibited by uncouplers ; c) the uptake is inhibited by methionine sulfoximine, a suicide-inhibitor of an enzyme of the γ-glutamyl cycle ; d) the initial rate of uptake is lowered by decreasing the Na +-level of the incubation medium or by adding ouabain. The validity of this hypothesis is discussed.

Multiple drug resistance in the fission yeast Schizosaccharomyces pombe: Correlation between drug and amino acid uptake and membrane ATPase activities

Cyh3 and cyh4, multiple drug resistant strains of Schizosaccharomyces pombe, show a much reduced uptake of trichodermin, chloramphenicol, cycloheximide, L-lysine, glycine, L-threonine, L-glutamine, L-arginine and L-glutamic acid when compared to wild type. The plasma membrane and mitochondrial ATPase activities of these mutants are also greatly reduced. Since the uptake of such compounds is likely to be driven by a proton electrochemical gradient set up by the membrane ATPase it is suggested that the primary effect of these mutations is at the level of the membrane ATPase. Another drug resistant strain, cyh1, which is resistant only to high levels of cycloheximide, shows increased uptake of trichodermin, L-lysine, glycine, L-threonine, L-glutamine when compared to wild type. The plasma membrane and mitochondrial ATPases of cyh1 are considerably greater than those of wild type. It has been shown previously that cyh1 possesses an altered 60S ribosonal subunit protein when compared to wild type and this makes it resistant to cycloheximide. There is no obvious explanation as to how this change could lead to the alterations in drug and amino acid uptake and in ATPase activities observed.

Glutamine and glutamate transport in cultured neuronal and glial cells

The uptake of l-glutamine in neuronal and glial cultures derived from rat cerebral hemispheres was found to be mediated by a low affinity-high capacity mechanism which was concentrative in both cell types; the calculated K m and V max were twice as high in glial than in neuronal cultures. In contrast l-glutamate was taken up by a high affinity system which was particularly efficient and concentrative in the glial cells. Different transport mechanisms for l-glutamine appeared to operate in the two cell types: l-glutamine uptake in neurons was sodium-dependent, specifically inhibited by l-glutamine but not affected by high potassium concentrations in the external medium; on the other hand, glial glutamine transport was decreased when potassium concentration increased, was sodium-independent and significantly inhibited by 3 structurally related amino acids. No significant contribution of homoexchange could be detected in either cell type. After [ 14C]glutamine preincubation, the radioactivity released into the superfusion medium by neuronal cells was increased in the presence of a high potassium concentration; no such effect could be seen in the case of glial cultures. A regulatory mechanism is suggested where astrocyte depolarization and repolarization would channel a flux of glutamine toward the neurons, subsequent to a glutamate flux in the opposite direction.