Age-dependent telomere attrition as a potential indicator of racial differences in renal growth patterns.

BACKGROUND: Racial differences in the predilection to salt sensitivity may arise from different renal growth patterns. To test this idea, we monitored age-dependent telomere attrition rate, reflecting largely the replicative history of somatic cells, in the outer renal cortex and the inner renal medulla of African Americans and Caucasians. METHODS: Telomere length, determined by the mean length of the terminal restriction fragments (TRF), was measured in specimens from 58 African-American and 63 Caucasian males, ages 1 day to 71 years. RESULTS: In the outer renal cortex, TRF length attrition rate was significantly slower in African Americans (-0.021 +/- 0.0064 kb/year) than in Caucasians (-0.060 +/- 0.0094 kb/year) (p = 0.0007). In both ethnic groups the TRF length attrition rate was slower in the inner medulla than in the outer renal cortex, but without significant racial differences. CONCLUSIONS: The proximal tubule is the most abundant nephron structure in the outer renal cortex. Less proliferative growth of proximal tubular cells in kidneys from African Americans may be one factor explaining the slower age-dependent telomere attrition rate in the outer renal cortex of African Americans than in Caucasians.

Glutamate-pyruvate transaminase protects against glutamate toxicity in hippocampal slices.

Elimination of glutamate through enzymatic degradation is an alternative to glutamate receptor blockade in preventing excitotoxic neuronal injury. Glutamate pyruvate transaminase (GPT) is a highly active glutamate degrading enzyme that requires pyruvate as a co-substrate. This study examined the ability of GPT to protect neurons of the hippocampal slice preparation against glutamate toxicity. Two methods were used to elevate the concentration of glutamate in the peri-neuronal space. In an endogenous release paradigm, slices were incubated with 100-500 microM L-trans-pyrrolidine-2,4-dicarboxylate (PDC), an inhibitor of glutamate re-uptake. One hour of exposure to PDC in normal, pyruvate-free slice maintenance medium caused a dose dependent increase in neuronal death assessed 24 h later by propidium iodide uptake in dead cell nuclei. GPT (10 U/ml) decreased neuronal death caused by exposure to PDC at all PDC concentrations tested. Neuroprotection in this model was not dependent on added or non-physiologic levels of pyruvate. In a different paradigm, glutamate was added directly to the normal, pyruvate-free slice maintenance medium and not rinsed away, exposing the slices to a range of 1-5 mM glutamate for an extended period. Twenty-four hours later, neuronal death was again assessed by propidium iodide uptake. GPT was again neuroprotective, decreasing neuronal death in the range from 3 to 5 mM glutamate. In the setting of incubation with this large load of glutamate, neuroprotection by GPT was enhanced by adding pyruvate to the medium. GPT is an effective neuroprotectant against glutamate excitotoxicity. When exposure is limited to endogenously released glutamate, neuroprotection by GPT is not dependent on added pyruvate.

Kynurenate production by cultured human astrocytes.

In the rodent brain, astrocytes are known to be the primary source of kynurenate (KYNA), an endogenous antagonist of both the glycine(B) and the alpha7 nicotinic acetylcholine receptor. In the present study, primary human astrocytes were used to examine the characteristics and regulation of de novo KYNA synthesis in vitro. To this end, cells were exposed to KYNA's bioprecursor L-kynurenine, and newly formed KYNA was recovered from the extracellular milieu. The production of KYNA was stereospecific and rose with increasing L-kynurenine concentrations, reaching a plateau in the high microM range. In an analogous experiment, astrocytes also readily produced and liberated the potent, specific glycine(B) receptor antagonist 7-chlorokynurenate from L-4-chlorokynurenine. KYNA synthesis was dose-dependently reduced by L-leucine or L-phenylalanine, two amino acids that compete with L-kynurenine for cellular uptake, and by aminooxyacetate, a non-specific aminotransferase inhibitor. In contrast, KYNA formation was stimulated by 5 mM pyruvate or oxaloacetate, which act as co-substrates of the transamination reaction. Aglycemic or depolarizing (50 mM KCl or 100 microM veratridine) conditions had no effect on KYNA synthesis. Subsequent studies using tissue homogenate showed that both known cerebral kynurenine aminotransferases (KAT I and KAT II) are present in astrocytes, but that KAT II appears to be singularly responsible for KYNA formation under physiological conditions. Taken together with previous results, these data suggest that very similar mechanisms control KYNA synthesis in the rodent and in the human brain. These regulatory events are likely to influence the neuromodulatory effects of astrocyte-derived KYNA in the normal and diseased human brain.

Enzymatic degradation protects neurons from glutamate excitotoxicity.

Several enzymes with the capacity to degrade glutamate have been suggested as possible neuroprotectants. We initially evaluated the kinetic properties of glutamate pyruvate transaminase (GPT; also known as alanine aminotransferase), glutamine synthetase, and glutamate dehydrogenase under physiologic conditions to degrade neurotoxic concentrations of glutamate. Although all three enzymes initially degraded glutamate rapidly, only GPT was able to reduce toxic (500 microM) levels of glutamate into the physiologic (<20 microM) range. Primary cultures of fetal murine cortical neurons were subjected to paradigms of either exogenous or endogenous glutamate toxicity to evaluate the neuroprotective value of GPT. Neuronal survival after exposure to added glutamate ranging from 100 to 500 microM was improved significantly in the presence of GPT (> or =1 U/ml). Cultures were also exposed to the glutamate transporter inhibitor L-trans-pyrrolidine-2,4-dicarboxylate (PDC), which produces neuronal injury by elevating extracellular glutamate. GPT significantly reduced the toxicity of PDC. This reduction was associated with a reduction in the PDC-dependent rise in the medium concentration of glutamate. These results suggest that enzymatic degradation of glutamate by GPT can be an alternative to glutamate receptor blockade as a strategy to protect neurons from excitotoxic injury.

In vivo glutamine hydrolysis in the formation of extracellular glutamate in the injured rat brain.

Hydrolysis of extracellular glutamine as a potential source of increased extracellular glutamate in the quinolinic acid (QUIN)-injured brain of the unanesthetized, free-moving rat was examined by microdialysis and HPLC analysis. Injury was initiated by injection of 100 nmoles of QUIN into the hippocampus. Immediately postinjury or 24 hr postinjury, the injection site was perfused with artificial cerebrospinal fluid + (14)C-glutamine to measure its conversion to (14)C-glutamate. L-trans-pyrrolidine-2,4-dicarboxylate (L-PDC), a glutamate uptake inhibitor, was added to the perfusate to enhance the detection of extracellular (14)C-glutamate. QUIN injury was followed by an immediate increase in extracellular glutamate that persisted 24 hr later. When (14)C-glutamine was added to the perfusate, a significant amount of (14)C-glutamate was recovered, and it was greater following QUIN injury than in control animals (P < 0.001). Up to 32% of the extracellular (14)C-glutamine was converted to (14)C-glutamate following QUIN injury. Considering the high concentration of glutamine normally present in the extracellular fluid, glutamine hydrolysis is a potential and important source for the increase in extracellular glutamate after neuronal injury in vivo.

Affinity purification and partial characterization of the zonulin/zonula occludens toxin (Zot) receptor from human brain.

The intercellular tight junctions (TJs) of endothelial cells represent the limiting structure for the permeability of the blood-brain barrier (BBB). Although the BBB has been recognized as being the interface between the bloodstream and the brain, little is known about its regulation. Zonulin and its prokaryotic analogue, zonula occludens toxin (Zot) elaborated by Vibrio cholerae, both modulate intercellular TJs by binding to a specific surface receptor with subsequent activation of an intracellular signaling pathway involving phospholipase C and protein kinase C activation and actin polymerization. Affinity column purification revealed that human brain plasma membrane preparations contain two Zot binding proteins of approximately 55 and approximately 45 kDa. Structural and kinetic studies, including saturation and competitive assays, identified the 55-kDa protein as tubulin, whereas the 45-kDa protein represents the zonulin/Zot receptor. Biochemical characterization provided evidence that this receptor is a glycoprotein containing multiple sialic acid residues. Comparison of the N-terminal sequence of the zonulin/Zot receptor with other protein sequences by BLAST analysis revealed a striking similarity with MRP-8, a 14-kDa member of the S-100 family of calcium binding proteins. The discovery and characterization of this receptor from human brain may significantly contribute to our knowledge on the pathophysiological regulation of the BBB.

2-Oxoacids regulate kynurenic acid production in the rat brain: studies in vitro and in vivo.

This study was designed to examine the role of 2-oxoacids in the enzymatic transamination of L-kynurenine to the excitatory amino acid receptor antagonist, kynurenate, in the rat brain. In brain tissue slices incubated in Krebs-Ringer buffer with a physiological concentration of L-kynurenine, pyruvate, and several other straight- and branched-chain 2-oxoacids, substantially restored basal kynurenate production in a dose-dependent manner without increasing the intracellular concentration of L-kynurenine. All 2-oxoacids tested also reversed or attenuated the hypoglycemia-induced decrease in kynurenate synthesis, but only pyruvate and oxaloacetate also substantially restored intracellular L-kynurenine accumulation. Thus, 2-oxoacids increase kynurenate formation in the brain primarily by functioning as co-substrates of the transamination reaction. This was supported further by the fact that the nonspecific kynurenine aminotransferase inhibitors (aminooxy)acetic acid and dichlorovinylcysteine prevented the effect of pyruvate on kynurenate production in a dose-dependent manner. Moreover, all 2-oxoacids tested attenuated or prevented the effects of veratridine, quisqualate, or L-alpha-aminoadipate, which reduce the transamination of L-kynurenine to kynurenate. Finally, dose-dependent increases in extracellular kynurenate levels in response to an intracerebral perfusion with pyruvate or alpha-ketoisocaproate were demonstrated by in vivo microdialysis. Taken together, these data show that 2-oxoacids can directly augment the de novo production of kynurenate in several areas of the rat brain. 2-Oxoacids may therefore provide a novel pharmacological approach for the manipulation of excitatory amino acid receptor function and dysfunction.

Compartmentation of [14C]glutamate and [14C]glutamine oxidative metabolism in the rat hippocampus as determined by microdialysis.

Metabolic compartmentation of amino acid metabolism in brain is exemplified by the differential synthesis of glutamate and glutamine from the identical precursor and by the localization of the enzyme glutamine synthetase in glial cells. In the current study, we determined if the oxidative metabolism of glutamate and glutamine was also compartmentalized. The relative oxidation rates of glutamate and glutamine in the hippocampus of free-moving rats was determined by using microdialysis both to infuse the radioactive substrate and to collect 14CO2 generated during their oxidation. At the end of the oxidation experiment, the radioactive substrate was replaced by artificial CSF, 2 min-fractions were collected, and the specific activities of glutamate and glutamine were determined. Extrapolation of the specific activity back to the time that artificial CSF replaced 14C-amino acids in the microdialysis probe yielded an approximation of the interstitial specific activity during the oxidation. The extrapolated interstitial specific activities for [14C]glutamate and [14C]glutamine were 59 +/- 18 and 2.1 +/- 0.5 dpm/pmol, respectively. The initial infused specific activities for [U-14C]glutamate and [U-14C]glutamine were 408 +/- 8 and 387 +/- 1 dpm/pmol, respectively. The dilution of glutamine was greater than that of glutamate, consistent with the difference in concentrations of these amino acids in the interstitial space. Based on the extrapolated interstitial specific activities, the rate of glutamine oxidation exceeds that of glutamate oxidation by a factor of 5.3. These data indicate compartmentation of either uptake and/or oxidative metabolism of these two amino acids. The presence of [14C]glutamine in the interstitial space when [14C]glutamate was perfused into the brain provided further evidence for the glutamate/glutamine cycle in brain.

Alpha-ketoisocaproate alters the production of both lactate and aspartate from [U-13C]glutamate in astrocytes: a 13C NMR study.

The present study determined the metabolic fate of [U-13C]glutamate in primary cultures of cerebral cortical astrocytes from rat brain and also in cultures incubated in the presence of 1 or 5 mM alpha-ketoisocaproate (alpha-KIC). When astrocytes were incubated with 0.2 mM [U-13C]glutamate, 64.1% of the 13C metabolized was converted to glutamine, and the remainder was metabolized via the tricarboxylic acid (TCA) cycle. The formation of [1,2,3-(13)C3]glutamate demonstrated metabolism of the labeled glutamate via the TCA cycle. In control astrocytes, 8.0% of the [13C]glutamate metabolized was incorporated into intracellular aspartate, and 17.2% was incorporated into lactate that was released into the medium. In contrast, there was no detectable incorporation of [13C]glutamate into aspartate in astrocytes incubated in the presence of alpha-KIC. In addition, the intracellular aspartate concentration was decreased 50% in these cells. However, there was increased incorporation of [13C]glutamate into the 1,2,3-(13)C3-isotopomer of lactate in cells incubated in the presence of alpha-KIC versus controls, with formation of lactate accounting for 34.8% of the glutamate metabolized in astrocytes incubated in the presence of alpha-KIC. Altogether more of the [13C]glutamate was metabolized via the TCA cycle, and less was converted to glutamine in astrocytes incubated in the presence of alpha-KIC than in control cells. Overall, the results demonstrate that the presence of alpha-KIC profoundly influences the metabolic disposition of glutamate by astrocytes and leads to altered concentrations of other metabolites, including aspartate, lactate, and leucine. The decrease in formation of aspartate from glutamate and in total concentration of aspartate may impair the activity of the malate-aspartate shuttle and the ability of astrocytes to transfer reducing equivalents into the mitochondria and thus compromise overall energy metabolism in astrocytes.

Effect of alpha-ketoisocaproate and leucine on the in vivo oxidation of glutamate and glutamine in the rat brain.

Leucine and alpha-ketoisocaproate (alpha-KIC) were perfused at increasing concentrations into rat brain hippocampus by microdialysis to mimic the conditions of maple syrup urine disease. The effects of elevated leucine or alpha-KIC on the oxidation of L-[U-14C]glutamate and L-[U-14C]glutamine in the brain were determined in the non-anesthetized rat. 14CO2 generated by the metabolic oxidation of [14C]glutamate and [14C]glutamine in brain was measured following its diffusion into the eluant during the microdialysis. Leucine and alpha-KIC exhibited differential effects on 14CO2 generation from radioactive glutamate on glutamine. Infusion of 0.5 mM alpha-KIC increased [14C]glutamate oxidation approximately 2-fold; higher concentrations of alpha-KIC did not further stimulate [14C]glutamate oxidation. The enhanced oxidation of [14C]glutamate may be attributed to the function of alpha-KIC as a nitrogen acceptor from [14C]glutamate yielding [14C]alpha-ketoglutarate, an intermediate of the tricarboxylic acid cycle. [14C]glutamine oxidation was not stimulated as much as [14C]glutamate oxidation and only increased at 10 mM alpha-KIC reflecting the extra metabolic step required for its oxidative metabolism. In contrast, leucine had no effect on the oxidation of either [14C]glutamate or [14C]glutamine. In maple syrup urine disease elevated alpha-KIC may play a significant role in altered energy metabolism in brain while leucine may contribute to clinical manifestations of this disease in other ways.

Induction of astrocyte argininosuccinate synthetase and argininosuccinate lyase by dibutyryl cyclic AMP and dexamethasone.

Arginine is an intermediate in the elimination of excess nitrogen and is the substrate for nitric oxide synthesis. Arginine synthesis has been reported in brain tissue. We have studied the activity of the arginine biosynthetic enzymes argininosuccinate synthetase and argininosuccinate lyase in dexamethasone and/or dibutyryl cyclic AMP treated rat astrocyte cultures. Argininosuccinate lyase activity was stimulated by treatment with either effector and an additive effect was obtained when both agents were added simultaneously. Argininosuccinate synthetase was also increased in dexamethasone treated astrocytes. The effect of dibutyryl cyclic AMP on argininosuccinate synthetase was variable, suggesting a role for additional factors in its regulation as compared to argininosuccinate lyase. Regulation of arginine synthesis in astrocytes may be important to insure that arginine is not limiting for nitric oxide synthesis in neural tissue.

Increased expression of 72-kd type IV collagenase (MMP-2) in human aortic atherosclerotic lesions.

MMP-2, a secreted 72-kd metalloproteinase that specifically degrades type IV collagen as well as denatured collagens, has been implicated in smooth muscle cell migration. To evaluate the possible contribution of this enzyme to the formation and progression of the atherosclerotic lesion, the expression of MMP-2 was studied in human aortic tissue. MMP-2 was visualized in frozen sections of the aortic wall by an immunofluorescent technique with a polyclonal antibody. Expression of MMP-2 in the aortic extracts was also studied by zymography and Western blotting. Our results reveal that a greater amount of MMP-2 is present in fatty streaks and atherosclerotic plaques as compared with normal regions of the aorta. Immunoblotting analysis showed that MMP-2 was expressed in atherosclerotic plaque > fatty streak > normal aortic wall in a ratio of approximately 4:2:1. Zymograms show that both forms (activated and latent) of MMP-2 increased in the atherosclerotic plaques. The presence of macrophages, detected by an immunohistochemical technique in some areas of higher MMP-2 expression suggests that these cells are a possible source of MMP-2. We conclude that MMP-2 collagenase may have a role in the formation and progression of the atherosclerotic lesion and may be involved in clinical complications of atherosclerosis, such as fissure and rupture, leading to thrombosis.

Exogenous glutamate concentration regulates the metabolic fate of glutamate in astrocytes.

The metabolic fate of glutamate in astrocytes has been controversial since several studies reported > 80% of glutamate was metabolized to glutamine; however, other studies have shown that half of the glutamate was metabolized via the tricarboxylic acid (TCA) cycle and half converted to glutamine. Studies were initiated to determine the metabolic fate of increasing concentrations of [U-13C] glutamate in primary cultures of cerebral cortical astrocytes from rat brain. When astrocytes from rat brain were incubated with 0.1 mM [U-13C] glutamate 85% of the 13C metabolized was converted to glutamine. The formation of [1,2,3-13C3] glutamate demonstrated metabolism of the labeled glutamate via the TCA cycle. When astrocytes were incubated with 0.2-0.5 mM glutamate, 13C from glutamate was also incorporated into intracellular aspartate and into lactate that was released into the media. The amount of [13C] lactate was essentially unchanged within the range of 0.2-0.5 mM glutamate, whereas the amount of [13C] aspartate continued to increase in parallel with the increase in glutamate concentration. The amount of glutamate metabolized via the TCA cycle progressively increased from 15.3 to 42.7% as the extracellular glutamate concentration increased from 0.1 to 0.5 mM, suggesting that the concentration of glutamate is a major factor determining the metabolic fate of glutamate in astrocytes. Previous studies using glutamate concentrations from 0.01 to 0.5 mM and astrocytes from both rat and mouse brain are consistent with these findings.

Use of intracellular versus extracellular specific activities in calculation of glutamine metabolism in astrocytes: effect of dibutyryl cyclic AMP.

The rate of glutaminase-dependent metabolism of glutamine in intact astrocytes was determined under conditions in which the extracellular concentration of glutamine was varied between 0.2 and 3.2 mM glutamine for control and dibutyryl cyclic AMP (dBcAMP)-treated cells. Glutamine metabolism by intact cells increased with increasing extracellular glutamine when calculations were based on the extracellular specific activity of glutamine. However, when the rate was based on the intracellular specific activity of glutamine, the rate of glutamine metabolism was independent of the media glutamine concentration. Similar results were obtained when cells were treated with dBcAMP, although the rates were approximately twice as high compared to untreated cells. The rate of formation of 14CO2 from [1-14C]glutamine and [1-14C]glutamate, based on the extracellular specific activities, were 93 +/- 5 and 40 +/- 4 nmol/mg protein/h, respectively. Oxidation rates based on the experimentally determined intracellular specific activity of glutamine and glutamate were 144 +/- 8 and 209 +/- 18 nmol/mg protein/h, respectively. In dBcAMP-treated astrocytes, the oxidation rates were higher than in untreated cells. These studies demonstrate that determination of the specific activity of compounds inside the cell aids in the interpretation of metabolic studies with intact cells and that both the initial steps of glutamine metabolism and the rate of 14CO2 formation from 14C-glutamine via the TCA cycle were increased in dBcAMP-treated astrocytes.

Elevation of amino acids in the interstitial space of the rat brain following infusion of large neutral amino and keto acids by microdialysis: alpha-ketoisocaproate infusion.

alpha-Ketoisocaproate was infused into the brain of free-moving, awake rats by microdialysis to create a microenvironment similar to that found in maple syrup urine disease. The eluate of the probe was analyzed for amino acids to determine if alpha-ketoisocaproate was transaminated to leucine and if the amino acid homeostasis was altered. The interstitial levels of leucine were increased up to 11-fold and other large neutral amino acids were increased 2- to 3-fold indicating an active branched chain keto acid transaminase activity and enhanced hetero-exchange across cell membranes. The elevation of large neutral amino acids in the interstitial space is discussed in terms of the synthesis of leucine and neurotransmitters in maple syrup urine disease.

Elevation of amino acids in the interstitial space of the rat brain following infusion of large neutral amino and keto acids by microdialysis: leucine infusion.

A microenvironment similar to that found in maple syrup urine disease was created in the brain of free-moving, awake rats by the infusion of leucine into the brain using microdialysis. To determine the effects on amino acid homeostasis the eluate of the probe was analyzed. Perfusion with leucine elevated the interstitial levels of large neutral amino acids suggesting hetero exchange of large neutral amino acids from neuronal cells into the interstitial space. The data also demonstrated the inter relationship of leucine and glutamine, both of which may be nitrogen sinks in the brain. Elevation of large neutral amino acids in the interstitial space suggests a decreased concentration in neurons which might have an effect on the synthesis of serotonin and catecholamines and suggests a mechanism by which elevated leucine may affect neuronal function in maple syrup urine disease.

Effect of dibutyryl cyclic AMP and dexamethasone on glutamine synthetase gene expression in rat astrocytes in culture.

Astrocytes are the primary site of glutamate conversion to glutamine in the brain. We examined the effects of treatment with either dibutyryl cyclic AMP and/or the synthetic glucocorticoid dexamethasone on glutamine synthetase enzyme activity and steady-state mRNA levels in cultured neonatal rat astrocytes. Treatment of cultures with dibutyryl cyclic AMP alone (0.25 mM-1.0 mM) increased glutamine synthetase activity and steady state mRNA levels in a dose-dependent manner. Similarly, treatment with dexamethasone alone (10(-7)-10(-5) M) increased glutamine synthetase mRNA levels and enzyme activity. When astrocytes were treated with both effectors, additive increases in glutamine synthetase activity and mRNA were obtained. However, the additive effects were observed only when the effect of dibutyryl cyclic AMP alone was not maximal. These findings suggest that the actions of these effectors are mediated at the level of mRNA accumulation. The induction of glutamine synthetase mRNA by dibutyryl cyclic AMP was dependent on protein synthesis while the dexamethasone effect was not. Glucocorticoids and cyclic AMP are known to exert their effects on gene expression by different molecular mechanisms. Possible crosstalk between these effector pathways may occur in regulation of astrocyte glutamine synthetase expression.

Valproate increases glutaminase and decreases glutamine synthetase activities in primary cultures of rat brain astrocytes.

It has been proposed that hyperammonemia may be associated with valproate therapy. As astrocytes are the primary site of ammonia detoxification in brain, the effects of valproate on glutamate and glutamine metabolism in astrocytes were studied. It is well established that, because of compartmentation of glutamine synthetase, astrocytes are the site of synthesis of glutamine from glutamate and ammonia. The reverse reaction is catalyzed by the ubiquitous enzyme glutaminase, which is present in both neurons and astrocytes. In astrocytes exposed to 1.2 mM valproate, glutaminase activity increased 80% by day 2 and remained elevated at day 4; glutamine synthetase activity was decreased 30%. Direct addition of valproate to assay tubes with enzyme extracts from untreated astrocytes had significant effects only at concentrations of 10 and 20 mM. When astrocytes were exposed for 4 days to 0.3, 0.6, or 1.2 mM valproate and subsequently incubated with L-[U-14C]glutamate, label incorporation into [14C]glutamine was decreased by 11, 25, and 48%, respectively, and is consistent with a reduction in glutamine synthetase activity. Label incorporation from L-[U-14C]glutamate into [14C]aspartate also decreased with increasing concentrations of valproate. Following a 4-day exposure to 0.6 mM valproate, the glutamine levels increased 40% and the glutamate levels 100%. These effects were not directly proportional to valproate concentration, because exposure to 1.2 mM valproate resulted in a 15% decrease in glutamine levels and a 25% increase in glutamate levels compared with control cultures. Intracellular aspartate was inversely proportional to all concentrations of extracellular valproate, decreasing 60% with exposure to 1.2 mM valproate.(ABSTRACT TRUNCATED AT 250 WORDS)

A glutamatergic mechanism for aluminum toxicity in astrocytes.

The effect of aluminum on the metabolism of glutamate and glutamine in astrocytes was studied to provide information about a possible biochemical mechanism for aluminum neurotoxicity and its potential contribution to neurodegenerative disease. Exposure of cultured rat brain astrocytes for 3-4 d to 5-7.5 mM aluminum lactate increased glutamine synthetase activity by 100-300% and diminished glutaminase activity by 50-85%. Increased glutamine synthetase enzyme activity was accompanied by an elevated level of glutamine synthetase mRNA. Alterations in glutaminase and glutamine synthetase following aluminum exposure caused increased intracellular glutamine levels, decreased intracellular glutamate levels, and increased conversion of glutamate to glutamine and the release of the latter into the extracellular space. The results of these changes may alter the availability of neurotransmitter glutamate in vivo and may be a mechanism for the aluminum neurotoxicity observed in individuals exposed to the metal during dialysis procedures and other situations.