Reduction of large neutral amino acid levels in plasma and brain of hyperleucinemic rats

Neurological dysfunction is common in patients with maple syrup urine disease (MSUD). However, the mechanisms underlying the neuropathology of this disorder are poorly known. In the present study we investigated the effect of acute hyperleucinemia on plasma and brain concentrations of amino acids. Fifteen-day-old rats were injected subcutaneously with 6 micromol L-leucine per gram body weight. Controls received saline in the same volumes. The animals were sacrificed 30--120 min after injection, blood was collected and their brain rapidly removed and homogenized. The amino acid concentrations were determined by HPLC using orthophtaldialdehyde for derivatization and fluorescence for detection. The results showed significant reductions of the large neutral amino acids (LNAA) L-phenylalanine, L-tyrosine, L-isoleucine, L-valine and L-methionine, as well as L-alanine, L-serine and L-histidine in plasma and of L-phenylalanine, L-isoleucine, L-valine and L-methionine in brain, as compared to controls. In vitro experiments using brain slices to study the influence of leucine on amino acid transport and protein synthesis were also carried out. L-Leucine strongly inhibited [14C]-L-phenylalanine transport into brain, as well as the incorporation of the [14C]-amino acid mixture, [14C]-L-phenylalanine and [14C]-L-lysine into the brain proteins. Although additional studies are necessary to evaluate the importance of these effects for MSUD, considering previous findings of reduced levels of LNAA in plasma and CSF of MSUD patients during crises, it may be speculated that a decrease of essential amino acids in brain may lead to reduction of protein and neurotransmiter synthesis in this disorder.     

Large Neutral Amino Acid Supplementation Exerts Its Effect through Three Synergistic Mechanisms: Proof of Principle in Phenylketonuria Mice

Background

Phenylketonuria (PKU) was the first disorder in which severe neurocognitive dysfunction could be prevented by dietary treatment. However, despite this effect, neuropsychological outcome in PKU still remains suboptimal and the phenylalanine-restricted diet is very demanding. To improve neuropsychological outcome and relieve the dietary restrictions for PKU patients, supplementation of large neutral amino acids (LNAA) is suggested as alternative treatment strategy that might correct all brain biochemical disturbances caused by high blood phenylalanine, and thereby improve neurocognitive functioning.

Objective

As a proof-of-principle, this study aimed to investigate all hypothesized biochemical treatment objectives of LNAA supplementation (normalizing brain phenylalanine, non-phenylalanine LNAA, and monoaminergic neurotransmitter concentrations) in PKU mice.

Methods

C57Bl/6 Pah-enu2 (PKU) mice and wild-type mice received a LNAA supplemented diet, an isonitrogenic/isocaloric high-protein control diet, or normal chow. After six weeks of dietary treatment, blood and brain amino acid and monoaminergic neurotransmitter concentrations were assessed.

Results

In PKU mice, the investigated LNAA supplementation regimen significantly reduced blood and brain phenylalanine concentrations by 33% and 26%, respectively, compared to normal chow (p<0.01), while alleviating brain deficiencies of some but not all supplemented LNAA. Moreover, LNAA supplementation in PKU mice significantly increased brain serotonin and norepinephrine concentrations from 35% to 71% and from 57% to 86% of wild-type concentrations (p<0.01), respectively, but not brain dopamine concentrations (p = 0.307).

Conclusions

This study shows that LNAA supplementation without dietary phenylalanine restriction in PKU mice improves brain biochemistry through all three hypothesized biochemical mechanisms. Thereby, these data provide proof-of-concept for LNAA supplementation as a valuable alternative dietary treatment strategy in PKU. Based on these results, LNAA treatment should be further optimized for clinical application with regard to the composition and dose of the LNAA supplement, taking into account all three working mechanisms of LNAA treatment.     

Kinetic Constants for Blood—Brain Barrier Amino Acid Transport in Conscious Rats

The kinetic constants for large neutral amino acid (LNAA) transport across the blood-brain barrier (BBB) of conscious rats were determined in four brain regions: cortex, caudate-putamen, hippocampus, and thalamus-hypothalamus. Indwelling external carotid artery catheters allowed for single-bolus (200 microliters) injections directly into the arterial system of unanesthetized and lightly restrained animals. Our results showed lower brain uptake index values for conscious rats compared to previous reports for anesthetized animals which are consistent with higher rates of cerebral blood flow in the conscious animals. Km values were lower in the conscious animals and ranged from 29% to 87% of the Km values in pentobarbital-anesthetized animals whereas the KD values were about twofold higher in the conscious animals. No apparent regional differences were observed. Influx rates were determined which take into consideration flow rates and plasma amino acid concentrations. Our results showed an average amino acid influx value of 5.2 nmol/min/g, which is 53% higher than the average influx in pentobarbital-anesthetized animals. The present results in conscious animals regarding the low Km of LNAA transport across the BBB lend further support to the importance of fluctuations in plasma amino acid concentrations and LNAA transport competitive effects on brain amino acid availability.     

Relationship Between Plasma and Brain Large Neutral Amino Acids in Rats Fed Diets with Different Compositions at Different Times of the Day

Large neutral amino acids (LNAAs) compete with each other for carrier-mediated transport through the blood-brain barrier into the brain. The relative plasma concentration, expressed as the ratio of each LNAA to the sum of LNAAs, is considered the main regulator of brain LNAA concentrations. In order to investigate the consistency of this assumption throughout a 24-h period, we have compared the relationship of plasma LNAAs to brain LNAAs among groups of rats fed diets containing various amounts of protein (in order to obtain a wide range of plasma LNAA levels) at two different phases of the light/dark cycle (0900 and 2100 hours). The relationship between plasma and brain LNAAs was found to be dependent on both diet and the time of day. Similar plasma amino acid concentrations in the morning and in the evening contrasted with different brain concentrations. Furthermore, previous findings that brain LNAA concentrations are influenced by plasma amino acid concentrations were confirmed.     

Beta-adrenergic control of brain uptake of large neutral amino acids

The amino acids tyrosine and tryptophan are precursors of physiologically active amines in the central nervous system. To reach the brain they have to compete with other large neutral aminio acids (LNAA) for the normally saturated carrier by which these amino acids are transported into the brain. The beta-adrenergic agonist isoprenaline is demonstrated to cause an increase in the brain concentration of most LNAA without a concomitant decrease in any of them. This finding indicates that the transport of LNAA into the brain is regulated by a beta-adrenergic mechanism.     

Large neutral amino acids: dietary effects on brain neurochemistry and function

The ingestion of large neutral amino acids (LNAA), notably tryptophan, tyrosine and the branched-chain amino acids (BCAA), modifies tryptophan and tyrosine uptake into brain and their conversion to serotonin and catecholamines, respectively. The particular effect reflects the competitive nature of the transporter for LNAA at the blood–brain barrier. For example, raising blood tryptophan or tyrosine levels raises their uptake into brain, while raising blood BCAA levels lowers tryptophan and tyrosine uptake; serotonin and catecholamine synthesis in brain parallel the tryptophan and tyrosine changes. By changing blood LNAA levels, the ingestion of particular proteins causes surprisingly large variations in brain tryptophan uptake and serotonin synthesis, with minimal effects on tyrosine uptake and catecholamine synthesis. Such variations elicit predictable effects on mood, cognition and hormone secretion (prolactin, cortisol). The ingestion of mixtures of LNAA, particularly BCAA, lowers brain tryptophan uptake and serotonin synthesis. Though argued to improve physical performance by reducing serotonin function, such effects are generally considered modest at best. However, BCAA ingestion also lowers tyrosine uptake, and dopamine synthesis in brain. Increasing dopamine function in brain improves performance, suggesting that BCAA may fail to increase performance because dopamine is reduced. Conceivably, BCAA administered with tyrosine could prevent the decline in dopamine, while still eliciting a drop in serotonin. Such an LNAA mixture might thus prove an effective enhancer of physical performance. The thoughtful development and application of dietary proteins and LNAA mixtures may thus produce treatments with predictable and useful functional effects.     

EVIDENCE FOR SEPARATE SYSTEMS FOR THE TRANSPORT OF NEUTRAL AND BASIC AMINO ACIDS ACROSS THE BLOOD-BRAIN BARRIER

Abstract— The effects of high circulating concentrations of several amino acids on the free amino acids of rat brain were measured, to see whether or not the results followed any consistent pattern. High circulating concentrations of large, neutral amino acids (phenylalanine, valine or isoleucine) caused significantly decreased values only of other large, neutral amino acids in the brains. High circulating concentrations of the basic amino acids lysine or arginine caused significantly decreased values only of each other. The data suggest that there are separate systems for the transport of neutral and basic amino acids across the blood-brain barrier. The effects of valine and lysine on the uptake by brain and the con-vulsant action of allylglycine (a neutral amino acid) were consistent with the concept of separate systems for the transport of amino acids across the blood-brain barrier. Valine inhibited the uptake by brain and the convulsant action of allylglycine in mice, but lysine did not. The data suggest that allylglycine and valine are transported into the brain by a common mechanism that does not transport lysine.     

Diurnal rhythms in rat plasma amino acids

To obtain detailed data on the diurnal rhythm in rat plasma amino acids, groups of rats were killed every two hours during 24 hours and the amino acids in plasma were measured. By using such a short interval between the blood samples, it was possible to reveal differences in rhythmicity between the various amino acids, more detailed than those previously described. Furthermore, it was found that those large neutral amino acids (LNAA) which compete with each other for the carrier mediated transport from plasma into the brain demonstrated different rhythms, whereby also the relation between these competing amino acids varied during the day. This finding might have implications for the transport of the various LNAAs into the brain, and secondarily also for the synthesis of the monoaminergic neurotransmitters in the neurons, for which the LNAAs tyrosine and tryptophan serve as precursors.     

Na+-dependent transport of large neutral amino acids occurs at the abluminal membrane of the blood-brain barrier

Several Na+-dependent carriers of amino acids exist on the abluminal membrane of the blood-brain barrier (BBB). These Na+-dependent carriers are in a position to transfer amino acids from the extracellular fluid of brain to the endothelial cells and thence to the circulation. To date, carriers have been found that may remove nonessential, nitrogen-rich, or acidic (excitatory) amino acids, all of which may be detrimental to brain function. We describe here Na+-dependent transport of large neutral amino acids across the abluminal membrane of the BBB that cannot be ascribed to currently known systems. Fresh brains, from cows killed for food, were used. Microvessels were isolated, and contaminating fragments of basement membranes, astrocyte fragments, and pericytes were removed. Abluminal-enriched membrane fractions from these microvessels were prepared. Transport was Na+ dependent, voltage sensitive, and inhibited by 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid, a particular inhibitor of the facilitative large neutral amino acid transporter 1 (LAT1) system. The carrier has a high affinity for leucine (Km 21 +/- 7 microM) and is inhibited by other neutral amino acids, including glutamine, histidine, methionine, phenylalanine, serine, threonine, tryptophan, and tyrosine. Other established neutral amino acids may enter the brain by way of LAT1-type facilitative transport. The presence of a Na+-dependent carrier on the abluminal membrane capable of removing large neutral amino acids, most of which are essential, from brain indicates a more complex situation that has implications for the control of essential amino acid content of brain.     

Inhibition of Neutral Amino Acid Transport Across the Human Blood-Brain Barrier by Phenylalanine

Abstract: The delivery of large neutral amino acids (LNAAs) to brain across the blood-brain barrier (BBB) is mediated by the L-type neutral amino acid transporter present in the membranes of the brain capillary endothelial cell. In experimental animals, the L-system transporter is saturated under normal conditions, and therefore an elevation in the plasma concentration of one LNAA will reduce brain uptake of others. In this study, we used positron emission tomography (PET) to determine the effect of elevated plasma phenylalanine concentrations on the uptake of an artificial neutral amino acid, [¹¹C]-aminocyclohexanecarboxylate ([¹¹C]ACHC), in human brain. PET scans were performed on six normal male subjects after an overnight fast and again 60 min after oral administration of 100 mg/kg of phenylalanine. The plasma phenylalanine concentration increased by an average of 11-fold between the first and second scans. This increase produced a reduction in [¹¹C]ACHC uptake in all brain regions but not in scalp. The mean ± SD influx rate constant for whole brain decreased after phenylalanine ingestion from 0.036 ± 0.002 to 0.019 ± 0.004 ml/g/min. Kinetic analysis of the effect of plasma phenylalanine concentration on the rate of [¹¹C]ACHC uptake is compatible with a model of competitive inhibition so that large increases in the concentration of one LNAA in plasma will reduce the brain uptake of other LNAAs across the human BBB.     

Neutral amino acid transport at the human blood-brain barrier

The kinetics of human blood-brain barrier neutral amino acid transport sites are described using isolated human brain capillaries as an in vitro model of the human blood-brain barrier. Kinetic parameters of transport (Km, Vmax, and KD) were determined for eight large neutral amino acids. Km values ranged from 0.30 +/- 0.08 microM for phenylalanine to 8.8 +/- 4.6 microM for valine. The amino acid analogs N-methylaminoisobutyric acid and 2-aminobicyclo[2.2.1]heptane-2-carboxylic acid were used as model substrates of the alanine- and leucine-preferring transport systems, respectively. Phenylalanine is transported solely by the L-system (which is sensitive to 2-aminobicyclo[2.2.1]heptane-2-carboxylic acid), and leucine is transported equally by the L- and ASC-system (which is sodium-dependent and N-methylaminoisobutyric acid-independent). Dose-dependent inhibition of the high affinity transport system by p-chloromercuribenzenesulfonic acid is demonstrated for phenylalanine, similar to the known sensitivity of blood-brain barrier transport in vivo. The Km values for the human brain capillary in vitro correlate significantly (r = 0.83, p less than 0.01) with the Km values for the rat brain capillary in vivo. The results show that the affinity of human blood-brain barrier neutral amino acid transport is very high, i.e. very low Km compared to plasma amino acid concentrations. This provides a physical basis for the selective vulnerability of the human brain to derangements in amino acid availability caused by a selective hyperaminoacidemia, e.g. hyperphenylalaninemia.     

Isolated Brain Microvessels as In Vitro Equivalents of the Blood-Brain Barrier: Selective Removal by Collagenase of the A-System of Neutral Amino Acid Transport

On treatment with collagenase, brain microvessels, together with several protein components, lose some enzymatic activities such as alkaline phosphatase and gamma-glutamyltranspeptidase, whereas no change occurs in the activities of 5'-nucleotidase and glutamine synthetase. The energy-requiring "A-system" of polar neutral amino acid transport is also severely inactivated, whereas the L-system for the facilitated exchange of branched chain and aromatic amino acids is preserved. In the collagenase-digested microvessels, this leads to loss of the transtimulation effect of glutamine on the transport of large neutral amino acids, because such transtimulation is due to a cooperation between the A- and L-systems. By contrast, NH4+ maintains (and even enhances) its ability to stimulate the L-system of amino acid transport, presumably through glutamine synthesis within the endothelial cells.     

Ventriculo–cisternal perfusion of twelve amino acids in the rabbit

The clearances of twelve amino acids from the ventricles during ventriculocisternal perfusion in the rabbit have been measured; uptake by the brain was also measured and this permitted the separate computation of loss to brain and loss to blood during the perfusion. Clearance under carrier-free conditions was greater than when a concentration of 5mM unlabeled amino acid was present in the perfusion fluid. Brain uptake was also usually reduced by the presence of unlabeled amino acid due presumably to suppression of accumulation by brain cells. Reduction of transport across the blood-brain barrier would tend to increase brain uptake, and there was some evidence for a balance between the two opposing tendencies. Inhibition of clearance of a given labeled amino acid could be brought about by unlabeled amino acids of different molecular species. In general, the amino acids fell into three categories: neutral, acidic, and basic, and there was some overlap between them; of the neutral amino acids the A- and L-classification of Christensen was valid, although once again there was some overlap. If, during ventriculo-cisternal perfusion of a labeled amino acid, the activity of this labeled amino acid in the blood was raised well above that in the inflowing perfusion fluid, the labeled amino acid continued to be cleared from the perfusion fluid, suggesting uphill transport. On this basis it was suggested that the normally low concentrations of amino acids in the cerebrospinal fluid (CSF), by comparison with those in plasma, were due to an active transport from the CSF to the blood. Substrate-facilitated transport, whereby the penetration of labeled amino acid into the perfusion fluid from blood could be accelerated by adding unlabeled amino acid to the perfusion fluid, or vice versa, was demonstrated.     

Methionine Sulfoximine Prevents the Accumulation of Large Neutral Amino Acids in Brain of Portacaval-Shunted Rats

Portal-systemic shunting and hyperammonemia lead to an accumulation of the large neutral amino acids in brain and apparently alter transport of neutral amino acids across the blood-brain barrier. It has been proposed that portal-systemic shunting leads to a high brain concentration of glutamine, a product of cerebral ammonia detoxification, and thereby affects the transport of other neutral amino acids across the blood-brain barrier. To test this hypothesis, rats with a portacaval shunt were treated with L-methionine-dl-sulfoximine (MSO), an inhibitor of glutamine synthesis. Treatment with MSO resulted in lower concentrations of the neutral amino acids in brain of portacaval-shunted rats and a higher brain ammonia concentration, compared with untreated shunted rats. These results suggest that the accumulation of neutral amino acids in brain after portacaval shunt depends on the increased synthesis of glutamine in brain.     

Effects of hemorrhagic hypotension on tyrosine concentrations in rat spinal cord and plasma

Tyrosine is the precursor for catecholamine neurotransmitters. When catecholamine-containing neurons are physiologically active (as sympathoadrenal cells are in hypotension), tyrosine administration increases catecholamine synthesis and release. Since hypotension can alter plasma amino acid composition, we examined the effects of an acute hypotensive insult on tyrosine concentrations in plasma and spinal cord. Rats were cannulated and bled until the systolic blood pressure was 50 mmHg, or were kept normotensive for 1 h. Tyrosine and other large neutral amino acids (LNAA) known to compete with tyrosine for brain uptake were assayed in plasma and spinal cord. The rate at which intra-arterial [3H]tyrosine disappeared from the plasma was also estimated in hemorrhaged and control rats. In plasma of hemorrhaged animals, both the tyrosine concentration and the tyrosine/LNAA ratio was elevated; moreover, the disappearance of [3H]tyrosine was slowed. Tyrosine concentrations also increased in spinal cords of hemorrhaged-hypotensive rats when compared to normotensive controls. Changes in plasma amino acid patterns may thus influence spinal cord concentrations of amino acid precursors for neurotransmitters during the stress of hemorrhagic shock.     

Site-directed mutagenesis of rabbit LAT1 at amino acids 219 and 234

The availability of amino acids in the brain is regulated by the blood-brain barrier (BBB) large neutral amino acid transporter type 1 (LAT1) isoform, which is characterized by a high affinity (low Km) for substrate large neutral amino acids. The hypothesis that brain amino acid transport activity can be altered with single nucleotide polymorphisms was tested in the present studies with site-directed mutagenesis of the BBB LAT1. The rabbit has a high Km LAT1 large neutral amino acid transporter, as compared to the low Km neutral amino acid transporter at the human or rat BBB. The rabbit LAT1 was cloned from a rabbit brain capillary cDNA library. Alignment of the amino acid sequences of rabbit, human, and rat LAT1 revealed two radical amino acid residues that differ in the rabbit relative to the rat or human LAT1. The G219D mutation had a modest effect on the Km and Vmax of tryptophan transport via cloned rabbit LAT1 in frog oocytes, but the W234L variant reduced the Km by 64% and the Vmax by 96%. Conversely, LAT1 transport of either tryptophan or phenylalanine was nearly normalized when the double mutation W234L/G219D variant was produced. These studies show that marked changes in the affinity and capacity of the LAT1 are caused by single nucleotide polymorphisms and that phenotype can be restored with a double mutation.     

Identification of a membrane protein, LAT-2, that Co-expresses with 4F2 heavy chain, an L-type amino acid transport activity with broad specificity for small and large zwitterionic amino acids.

We have identified a new human cDNA, L-amino acid transporter-2 (LAT-2), that induces a system L transport activity with 4F2hc (the heavy chain of the surface antigen 4F2, also named CD98) in oocytes. Human LAT-2 is the fourth member of the family of amino acid transporters that are subunits of 4F2hc. The amino acid transport activity induced by the co-expression of 4F2hc and LAT-2 was sodium-independent and showed broad specificity for small and large zwitterionic amino acids, as well as bulky analogs (e.g. BCH (2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid)). This transport activity was highly trans-stimulated, suggesting an exchanger mechanism of transport. Expression of tagged N-myc-LAT-2 alone in oocytes did not induce amino acid transport, and the protein had an intracellular location. Co-expression of N-myc-LAT-2 and 4F2hc gave amino acid transport induction and expression of N-myc-LAT-2 at the plasma membrane of the oocytes. These data suggest that LAT-2 is an additional member of the family of 4F2 light chain subunits, which associates with 4F2hc to express a system L transport activity with broad specificity for zwitterionic amino acids. Human LAT-2 mRNA is expressed in kidney > placenta > brain, liver > spleen, skeletal muscle, heart, small intestine, and lung. Human LAT-2 gene localizes at chromosome 14q11.2-13 (13 cR or approximately 286 kb from marker D14S1349). The high expression of LAT-2 mRNA in epithelial cells of proximal tubules, the basolateral location of 4F2hc in these cells, and the amino acid transport activity of LAT-2 suggest that this transporter contributes to the renal reabsorption of neutral amino acids in the basolateral domain of epithelial proximal tubule cells.     

Saturable Retention of Vasopressin by Hippocampus Vessels In Vivo, Associated with Inhibition of Blood-Brain Transfer of Large Neutral Amino Acids

Vasopressin receptors have been reported in the endothelium of brain capillaries. The function of these receptors is not known. To test the prediction that vasopressin receptors in brain capillary endothelium affect amino acid transport across the blood-brain barrier and to assess the role of vasopressin transport across the cerebral vascular endothelium, we measured (a) the endothelial permeability to the large neutral amino acid leucine in the absence and presence of arginine vasopressin (AVP) and (b) the permeability of the blood-brain barrier to AVP relative to manitol. In brain regions protected by the blood-brain barrier, after circulation for 20 s, coinjection of leucine and AVP intravenously led to a decrease of leucine transport unrelated to changes of blood flow. The decrease was most pronounced in hippocampus (42%) and least pronounced in olfactory bulb and colliculi (17 and 19%, respectively). In the latter regions, the endothelial permeability to AVP did not significantly exceed that of mannitol. In hippocampus and in regions with no blood-brain barrier (pituitary and pineal glands), AVP retention in excess of mannitol retention was blocked by unlabeled AVP. The findings do not contradict the hypothesis of a role for AVP in the regulation of large neutral amino acid transfer into brain tissue.     

Transport of threonine and tryptophan by rat brain slices: relation to other amino acids at concentrations found in plasma

Threonine content of brain decreases in young rats fed a threonine-limiting, low protein diet containing a supplement of small neutral amino acids (serine, glycine and alanine), which are competitors of threonine transport in other systems (Tews et al., 1977). Threonine transport by brain slices was inhibited more by a complex amino acid mixture resembling plasma from rats fed the small neutral amino acid supplement than by mixtures resembling plasma from control rats or from rats fed a supplement of large neutral amino acids. Greater inhibition was seen with mixtures containing only the small neutral amino acids than with mixtures containing only large neutral amino acids. On an equimolar basis, serine and alanine were the most inhibitory; large neutrals were moderately so; and glycine and lysine were without effect. Threonine transport was also strongly inhibited by α-amino-n-butyric acid and homoserine, less so by α-aminoisobutyric acid, and not at all by GABA. The complex amino acid mixtures strongly inhibited α-aminoisobutyric acid transport by brain or liver slices but, in contrast to effects in brain, the extent of the inhibition in liver was not much affected by altering the composition of the mixture. Tryptophan accumulation by brain slices was effectively inhibited by other large neutral amino acids in physiologically occurring concentrations. Threonine, or a mixture of serine, glycine and alanine only slightly inhibited tryptophan uptake; basic amino acids were without effect and histidine stimulated tryptophan transport slightly. These results support the conclusion that a diet-induced decrease in the concentration in brain of a specific amino acid may be related to increased inhibition of its transport into brain by increases in the concentrations of transport-related, plasma amino acids.     

Uptake of Amino Acids by Brain Micro vessels Isolated from Rats with Experimental Chronic Renal Failure

The neurological disorders seen in patients with chronic renal failure and liver cirrhosis are analogous. Previous in vivo studies have shown that the impaired blood-brain amino acid transport seen in rats with chronic renal failure is similar to that of rats with portocaval anastomosis. To elucidate whether a comparable underlying pathogenic mechanism plays a role in both pathological conditions, blood and brain amino acid levels together with amino acid transport by isolated brain microvessels have been studied in rats with chronic renal failure and in sham-operated rats. Brain microvessels isolated from rats with experimental chronic renal failure showed that the uptake of labeled large neutral amino acid, i.e., leucine or phenylalanine, but not of lysine or alpha-methylaminoisobutyric acid, was significantly increased with respect to sham-operated rats; conversely, the uptake of glutamic acid in rats with chronic renal failure was significantly lower compared with values in controls. Kinetic analysis indicated that this was mainly due to increased exchange transport activity (Vmax) of the L-system, rather than to changes in the affinity (Km) of the carrier system for the relative substrate. These data, together with the significant rise of brain glutamine levels and an increased brain-to-plasma ratio of the sum of large neutral amino acids, are analogous to what was previously observed in rats with portocaval anastomosis.(ABSTRACT TRUNCATED AT 250 WORDS)