The absorption, tissue distribution and excretion of enteraly administered alpha-ketoglutarate in rats.

The absorption, tissue distribution and excretion of enteral alpha-ketoglutarate (AKG) was studied in four experiments. Six male Sprague Dawley rats were used to investigate the excretion of AKG in urine and faeces. Thirty rats, randomly assigned to five groups, were used to investigate the distribution of AKG in body tissues. They were gavaged with AKG enriched with 3 muCi/kg BW of (14)C uniformly marked AKG. Fourteen male Sprague Dawley rats were used to study the absorption of AKG (duodenum vs. ileum). Intestinal recovery of NaAKG vs. CaAKG was investigated in 36 rats. There was no significant excretion of non-metabolized AKG in the urine and faeces. There was no significant difference in the systemic levels of AKG when comparing the proximal to distal small intestine infusion. Up to 50%, 30% and 20% of gastrically delivered AKG was recovered in the stomach, 0.5, 1 and 2 h after gavage; the jejunal recovery achieved a maximum of 3%, 30 min after gavage, and was not detectable 2 h later. There was a relatively high distribution of (14)C-AKG in the tissues (e.g. liver, brain, bones, skin, muscles), 3 h after gavage, up to 70% of the administered dose. In conclusion, the high rate of retention of the carbon from AKG allows the postulation that there is a non-energetic mode of metabolism of intragastrically administered AKG. After conversion to final metabolites, AKG penetrates into all tissues and organs of rats, including the bone tissue. Intestinal absorption of AKG does not depend on the type of AKG salt administered.

Pharmacokinetics, safety, and effects on exercise performance of L-arginine alpha-ketoglutarate in trained adult men.

OBJECTIVE: We evaluated the pharmacokinetics, safety, and efficacy of l-arginine alpha-ketoglutarate (AAKG) in trained adult men. METHODS: Subjects participated in two studies that employed a randomized, double-blind, controlled design. In study 1, 10 healthy men (30-50 y old) fasted for 8 h and then ingested 4 g of time-released or non-timed-released AAKG. Blood samples were taken for 8 h after AAKG ingestion to assess the pharmacokinetic profile of L-arginine. After 1 wk the alternative supplement was ingested. In study 2, which was placebo controlled, 35 resistance-trained adult men (30-50 y old) were randomly assigned to ingest 4 g of AAKG (three times a day, i.e., 12 g daily, n = 20) or placebo (n = 15). Participants performed 4 d of periodized resistance training per week for 8 wk. At 0, 4, and 8 wk of supplementation the following tests were performed: clinical blood markers, one repetition maximum bench press, isokinetic quadriceps muscle endurance, anaerobic power, aerobic capacity, total body water, body composition, and psychometric parameters tests. Data were analyzed by repeated measures analysis of variance. RESULTS: In study 1, significant differences were observed in plasma arginine levels in subjects taking non-timed-release and timed-release AAKG. In study 2, significant differences were observed in the AAKG group (P < 0.05) for 1RM bench press, Wingate peak power, blood glucose, and plasma arginine. No significant differences were observed between groups in body composition, total body water, isokinetic quadriceps muscle endurance, or aerobic capacity. CONCLUSION: AAKG supplementation appeared to be safe and well tolerated, and positively influenced 1RM bench press and Wingate peak power performance. AAKG did not influence body composition or aerobic capacity.

Fate of enterally administered ornithine in healthy animals: interactions with alpha-ketoglutarate.

In order to improve our understanding of the metabolic interactions between alpha-ketoglutarate (alpha KG) and ornithine (Orn), which constitute the two parts of ornithine alpha-ketoglutarate (OKG) used as an adjuvant in enteral nutrition, we have investigated the plasma appearance and tissue distribution (qualitative and quantitative) of enterally administered 14C-Orn and 14C-alpha KG in healthy mice and rats. The influence of unlabelled alpha KG or Orn on 14C-Orn or 14C-alpha KG metabolism, respectively, was also studied. Unlabelled alpha KG was able to reduce strongly the rate of intestinal absorption of 14C-Orn, whereas the inverse was not true. This alpha KG-induced loss in plasma radioactivity after a load of Orn was associated with a decrease of radioactivity in tissue with no modification of the qualitative distribution in organs. In this study, a direct interaction between alpha KG and Orn was demonstrated at the intestinal level. The mechanisms involved in this phenomenon probably involve the regulation of metabolic conversions among alpha KG, Glu, pyrroline-5-carboxylate, and Orn. This is of importance in the therapeutic use of ornithine salts in clinical nutrition.

Toxicokinetic profiles of α-ketoglutarate cyanohydrin, a cyanide detoxification product, following exposure to potassium cyanide.

Poisoning by cyanide can be verified by analysis of the cyanide detoxification product, α-ketoglutarate cyanohydrin (α-KgCN), which is produced from the reaction of cyanide and endogenous α-ketoglutarate. Although α-KgCN can potentially be used to verify cyanide exposure, limited toxicokinetic data in cyanide-poisoned animals are available. We, therefore, studied the toxicokinetics of α-KgCN and compared its behavior to other cyanide metabolites, thiocyanate and 2-amino-2-thiazoline-4-carboxylic acid (ATCA), in the plasma of 31 Yorkshire pigs that received KCN (4mg/mL) intravenously (IV) (0.17 mg/kg/min). α-KgCN concentrations rose rapidly during KCN administration until the onset of apnea, and then decreased over time in all groups with a half-life of 15 min. The maximum concentrations of α-KgCN and cyanide were 2.35 and 30.18 µM, respectively, suggesting that only a small fraction of the administered cyanide is converted to α-KgCN. Although this is the case, the α-KgCN concentration increased >100-fold over endogenous concentrations compared to only a three-fold increase for cyanide and ATCA. The plasma profile of α-KgCN was similar to that of cyanide, ATCA, and thiocyanate. The results of this study suggest that the use of α-KgCN as a biomarker for cyanide exposure is best suited immediately following exposure for instances of acute, high-dose cyanide poisoning.

Development of nano alpha-ketoglutarate nebulization formulation and its pharmacokinetic and safety evaluation in healthy human volunteers for cyanide poisoning.

Development of nano alpha-ketoglutarate (A-KG) nebulization formulation for neutralization of inhaled cyanide ion toxicity. Objectives of the present study were to (a) develop a novel A-KG nebulization formulation against cyanide poisoning, particularly hydrogen cyanide gas (b) validate its respiratory fraction in vitro and in vivo, and (c) create its pharmacokinetic data in human volunteers. The formulation was optimized on the basis of particle size of aerosolized droplets after nebulization in 6 volunteers. Gamma scintigraphy was used to quantify total and regional lung deposition of nebulized A-KG after radiolabeling it with Technetium-99m. The formulation was optimized using 30% ethanol-saline with particle size in the range of 300-500 nm. In vitro and in vivo studies showed that drug nebulization resulted in a significant respirable fraction of 65 ± 0.6% with whole lung deposition of 13 ± 1%. Human pharmacokinetic data was derived in 6 healthy human volunteers with peak serum concentration (C(max)) of 39 ± 3 µg/ml, while the area under curve (AUC) after inhalation was 376 ± 23 µg × h/ml indicating that the drug was rapidly and completely absorbed when targeted directly to lungs. Significant lung deposition of A-KG was achieved with the developed formulation. The formulation appears to have several advantages, including the potential of neutralizing inhaled CN(-) ions in the lungs themselves. It is a safe and efficacious procedure, suitable for hospital or ambulance use in accidental cyanide poisoning cases, or as a preventive approach for fire-rescue teams.

Radiolabeling and dose fixation study of oral alpha-ketoglutarate as a cyanide antidote in healthy human volunteers.

CONTEXT: Radiolabeling and dose fixation study of alpha-ketoglutarate (A-KG). OBJECTIVE: A-KG is a potential oral antidote for cyanide poisoning. Its protective efficacy in animals was best exhibited at a dose of 2.0 g/kg body weight, which when extrapolated to human is very high. The objective of this study was to reduce the dose of A-KG in humans with concomitant increase in its bioavailability, employing pharmacoscintigraphic techniques to assess kinetics in man. MATERIALS AND METHODS: A-KG was radiolabeled with technetium-99m pertechnetate (Tc-99m) and its purity, labeling efficiency, and stability in vitro were determined by instant thin layer chromatography. Time-dependent bio-absorption of the drug in rats and rabbits was assessed by gamma scintigraphy after oral administration of a tracer dose of (99m)Tc-A-KG mixed with nonradioactive A-KG at a concentration of 0.1-2.0 g/kg in the presence or absence of aqueous dilution. Furthermore, scintigraphy and radiometry studies were performed in healthy human volunteers using 5-20 g of A-KG, given in single or split doses followed by different quantity of water. Drug bioavailability was estimated periodically. RESULTS: High radiolabeling (>97%) of A-KG with a stability of 24 h in vitro was obtained. Less than 1% absorption of the drug occurred within 20 min after A-KG was administered in animals at a concentration of 2.0 g/kg body weight. One-tenth reduction in dose increased the bioavailability to 15%. Significant improvement in gastric emptying of the drug was achieved when the drug was administered along with 1-5 mL of water. In humans, two doses of 10 g A-KG given at an interval of 10 min, followed by 300 mL of water, increased the drug bioavailability to 40% as compared to a single dose of 20 g. DISCUSSION: Significant reduction in A-KG dose was achieved in humans as compared to the recommended dose in animals. CONCLUSION: Aqueous dilution improves the bioavailability of A-KG in humans.

Triggering and amplification of insulin secretion by dimethyl alpha-ketoglutarate, a membrane permeable alpha-ketoglutarate analogue.

Cytosolic alpha-ketoglutarate is a potential signalling compound at late steps of stimulus-secretion-coupling in the course of insulin secretion induced by glucose and other fuels. This hypothesis is mainly based on the insulin-releasing effect of the membrane permeable ester dimethyl alpha-ketoglutarate which enters the beta-cell and is cleaved to produce cytosolic monomethyl alpha-ketoglutarate and eventually alpha-ketoglutarate. The present study tested this hypothesis. Insulin release, K(ATP) channel currents, membrane potential, ATP/ADP ratio and fluorescence of NAD(P)H (reduced pyridine nucleotides) were measured in mouse pancreatic islets and beta-cells. At a substimulatory glucose concentration (5 mM), dimethyl alpha-ketoglutarate (15 mM) produced a sustained insulin release, but no change of the islet ATP/ADP ratio and NAD(P)H fluorescence. In the absence of glucose, however, dimethyl alpha-ketoglutarate (15 mM) did not stimulate insulin release although it increased the ATP/ADP ratio and NAD(P)H fluorescence. Insulin secretion induced by a maximally effective concentration of the K(ATP) channel-blocking sulfonylurea glipizide was strongly amplified by dimethyl alpha-ketoglutarate in the presence of 5 mM glucose, but only moderately in the absence of glucose. Dimethyl alpha-ketoglutarate directly inhibited K(ATP) channels in inside-out membrane patches, depolarized the plasma membrane of intact beta-cells and generated action potentials. In conclusion, the stimulation of insulin secretion by extracellularly applied dimethyl alpha-ketoglutarate depends on inhibition of beta-cell K(ATP) channels by direct action of dimethyl alpha-ketoglutarate. The metabolism of alpha-ketoglutarate generated intracellularly by ester cleavage contributes to stimulation of insulin secretion both by indirect K(ATP) channel inhibition (via activation of ATP production) and by an amplifying effect.

First-pass metabolism limits the intestinal absorption of enteral alpha-ketoglutarate in young pigs.

Our results in a previous study indicated that the portal absorption of intragastrically fed alpha-ketoglutarate (AKG) was limited in young pigs. Our aim was to quantify the net portal absorption, first-pass metabolism, and whole-body flux of enterally infused AKG. In study 1, we quantified the net portal nutrient absorption in young pigs (n = 9) given an intraduodenal infusion of milk replacer [10 mL/(kg . h)] and either saline (control) or 930 micromol/(kg . h) AKG for 4 h. In study 2, we quantified the luminal disappearance of a duodenal AKG bolus in young pigs (n = 7). In study 3, we quantified the whole-body kinetics of (13)C-AKG metabolism when infused either enterally (n = 9) or intravenously (n = 9) in young pigs. In study 1, when compared with the control group, enteral AKG infusion increased (P < 0.01) the arterial (13.8 +/- 1.7 vs. 27.4 +/- 3.6 micromol/L) and portal (22.0 +/- 1.4 vs. 64.6 +/- 5.9 micromol/L) AKG concentrations and the net portal absorption of AKG [19.7 +/- 2.8 vs. 95.2 +/- 12.0 micromol/(kg . h)]. The mean fractional portal appearance of enterally infused AKG was 10.23 +/- 1.3%. In study 2, the luminal disappearance of AKG was 663 micromol/(kg . h), representing 63% of the intraduodenal dose. In study 3, the whole-body (13)C-AKG flux [4685 +/- 666 vs. 801 +/- 67 micromol/(kg . h)] was higher (P < 0.05) when given enterally than intravenously, but (13)CO(2) recovery was not different (37.3 +/- 1.0 vs. 36.2 +/- 0.7%dose). The first-pass splanchnic (13)C-AKG utilization was approximately 80%, of which 30% was oxidized to (13)CO(2). We conclude that the intestinal absorption of AKG is limited in young pigs largely due to substantial first-pass gastrointestinal metabolism.

alpha-Ketoglutarate (AKG) absorption from pig intestine and plasma pharmacokinetics.

To study the absorption, metabolism and kinetics, the AKG (in different concentrations) was administered intravenously, intra-portally, orally and directly into the ileum or duodenum of pigs, chronically fitted with portal and jugular catheters and T-shaped cannula at the duodenum and ileum. Additionally, this study was conducted to determine the influence of low pH, Fe(2+) or/and SO on AKG gut absorption and conversely FeSO(4) and FeSO(4)/AKG on Fe(2+) gut absorption. It is concluded that AKG was significantly better absorbed from the upper small intestine than from the distal sections. Furthermore, low pH, Fe(2+) and/or SO ions enhanced AKG absorption. The AKG administered to the portal vein was rapidly eliminated from the blood (half-life less than 5 min). The short lifetime for AKG is probably dependent on quick metabolism in the enteorcyetes and liver. However, the prolonged half-life can be related to its low AKG blood concentration. The Fe(2+) concentrations in blood increased after FeSO(4) and FeSO(4)/AKG duodenal infusion. The implication of above observations is important for practical application of the AKG in animal and human nutrition as well in medicine.

Synaptosomal uptake of alpha-ketoglutarate and glutamine in thioacetamide-induced hepatic encephalopathy in rats.

The kinetics of uptake of two astroglia-derived glutamate (GLU) precursors, alpha-ketoglutarate (alpha-KG) and glutamine (GLN) were determined in synaptosomes derived from rats with acute hepatic encephalopathy (HE) induced with a hepatotoxin, thioacetamide (TAA). TAA treatment increased by 33% Vmax for high affinity, low capacity alpha-KG uptake, without influencing its Km. The increase of the uptake capacity for alpha-KG may represent a response of the GLUergic nerve terminals to the decreased cerebral alpha-KG content, which during HE is associated with depressed activity of pyruvate carboxylase, an enzyme that replenishes alpha-KG in astrocytes. The result is thus consistent with the notion that HE affects the astroglial control of GLUergic neurotransmission. The Km and Vmax for the low affinity, high capacity GLN uptake was not affected by TAA treatment.

p-Aminohippurate transport in apical and basolateral membranes of the OK kidney epithelial cells.

p-Aminohippurate (PAH) transport in apical and basolateral membranes of OK cells was studied to characterize each membrane process and to determine which process is important for the vectorial transport of PAH across the OK cell monolayers. PAH uptake from the basal side into OK cells was much higher than from the apical side, and was inhibited almost completely by anion transport inhibitors (probenecid and furosemide) and low temperature. On the other hand, PAH efflux from OK cells to the apical side was higher than to the basal side. The efflux across the apical membrane also was inhibited by the transport inhibitors and low temperature. The uptake of PAH across the basolateral membrane was inhibited by dicarboxylates such as alpha-ketoglutarate, and kinetic analysis showed that the inhibition was a mixed-type. These findings suggest that PAH transport in apical and basolateral membranes of OK cells is a specifically mediated process, and both processes contribute to the vectorial transport of PAH across the cell monolayers. PAH/dicarboxylate exchange may be involved in PAH transport in the basolateral membrane of OK cells.

Net portal absorption of enterally fed alpha-ketoglutarate is limited in young pigs.

Our aim was to quantify the intestinal metabolic fate of dietary alpha-ketoglutarate (AKG). Female pigs (n = 6; 21 d old) were implanted with arterial, venous, portal and gastric catheters and an ultrasonic portal flow probe and fed a corn and soybean meal-based diet. On the day of the experiment, the pigs received a 4-h intragastric infusion of sodium AKG at a rate equivalent to 0, 2.5, 5 or 10% of dietary intake. The net portal AKG balance of the control and 2.5% treatments did not differ and were not different from zero. However, the net portal AKG balance of both the 5 [163 micro mol/(kg. h)] and 10% [159 micro mol/(kg. h)] treatments were greater (P < 0.05) than the control. Despite significant net AKG absorption at the 5 and 10% levels, the net portal appearance represented only 10.8 and 6.7%, respectively, of the enteral input. The net portal appearances of glutamate, glutamine, ammonia and the branched-chain amino acids were not affected by dietary AKG level. We conclude that the absorption of dietary AKG is limited in young pigs and does not change the net portal balance of amino acids or ammonia.

Absorption of alpha-ketoglutarate by the gastrointestinal tract of pigs.

Only a small percentage of alpha-ketoglutarate (AKG) administered lumenally to pigs appears in the portal circulation. This has been attributed to mucosal metabolism, and possibly by limited absorption. Although transporters for di- and tricarboxylic acids, which includes the sodium-dependent transporter NaDC-1, have been detected in the small intestine, correlations with functional assays are lacking. Therefore, intact tissues from three regions of the small intestine, stomach, and colon of weaned pigs were used to measure rates of AKG absorption. Western analysis was used to detect NaDC-1 in the three regions of small intestine. Rates of AKG absorption were highest in the small intestine, lowest in the colon, and intermediate in the stomach. Immunoreactive NaDC-1 was detected in the small intestine and this coincided with a component of AKG absorption that was inhibited by AKG and succinate. In contrast, absorption of AKG was inhibitable by unlabeled AKG, but not succinate, in the stomach, and by neither in the colon. Feeding studies indicated that the amounts of AKG that might be included in practical diets for pigs would not (1) upregulate rates of AKG absorption or (2) exceed estimated capacities of the small intestine to absorb AKG. The present findings indicate that the efficacy of AKG as an alternative metabolic fuel for enterocytes to spare dietary amino acids is not limited by absorption.

Glutamine and 2-oxoglutarate as metabolic precursors of the transmitter pools of glutamate and GABA: correlation of regional uptake by rat brain synaptosomes.

To more clearly define the roles of glutamine and 2-oxoglutarate as metabolic precursors of the transmitter pools of glutamate and GABA we have determined the relative rates at which these four substances, and adenosine and serotonin are accumulated by synaptosomes derived from twelve regions of the rat brain. Initial transport conditions and low substrate concentrations were used to maximize uptake by high-affinity systems, except the uptake of glutamine was determined at both low and high concentrations. Because the uptake of 2-oxoglutarate is markedly enhanced by glutamine, 2-oxoglutarate uptake was determined with and without glutamine (0.2 mM) added to the incubation medium. For each substrate, regional differences in uptake ranged from approximately two- to fourteen-fold. An anaylsis of uptake kinetics revealed that the regional differences were due primarily to differences in transport capacity rather than substrate affinities, at least for glutamate, GABA, and 2-oxoglutarate. Thirty-four correlation analyses of relative uptake values were performed. Strong correlations were found between 2-oxoglutarate and glutamate, and between glutamine and glutamate, whereas no strong correlations occurred between these substrates and GABA. Our results support the view that both glutamine and 2-oxoglutarate are major precursors of the transmitter pool of glutamate throughout the rat brain, but their relative contributions toward replenishing the transmitter pool of GABA are less certain.

Efflux of intracellular alpha-ketoglutarate via p-aminohippurate/dicarboxylate exchange in OK kidney epithelial cells.

The involvement of intracellular alpha-ketoglutarate (alpha-KG) in p-aminohippurate (PAH) transport was investigated in OK kidney epithelial cells. Efflux of intracellular alpha-KG from the OK cells to the basolateral side was increased by applying PAH to the basolateral side of the cells. In contrast, the intracellular alpha-KG concentration was not influenced by the addition of PAH. The alpha-KG efflux across the basolateral membrane induced by PAH was higher than that across the apical membrane. Probenecid inhibited the PAH-dependent alpha-KG efflux. The alpha-KG efflux to the basolateral side was saturable with increasing concentration of PAH in the basolateral medium. Antimycin A, a metabolic inhibitor, inhibited [14C]PAH uptake across the basolateral membrane of OK cells in a dose-dependent manner. In addition, both the alpha-KG efflux induced by PAH and the intracellular alpha-KG concentration were decreased by antimycin A dose-dependently. These results directly show that alpha-KG generated by intracellular metabolism is effluxed via PAH/dicarboxylate exchange in the basolateral membrane of OK cells.

Effect of lithium on renal transport and utilization of alpha-ketoglutarate in the rat.

Experiments were carried out in the intact functioning rat kidney to study the effect of lithium on both the renal transport of alpha-ketoglutarate (alpha-KG) along the nephron by micropuncture techniques and the renal uptake and peritubular transport of alpha-KG by measuring the renal blood flow, the urinary flow and the rate of renal alpha-KG delivery, filtration, reabsorption or secretion and excretion. At endogenous plasma alpha-KG concentration, 2.3 mM plasma lithium caused an increase in the fractional excretion of alpha-KG, whereas 4.6 mM plasma lithium led to a net secretion of alpha-KG. The micropuncture data indicate that this secretion occurred between the late proximal and the distal tubule, i.e., in the pars recta and/or in the loop of Henle. When plasma alpha-KG concentration was elevated, the two doses of lithium used inhibited the reabsorption of alpha-KG both in the proximal tubule and in the pars recta and/or the loop of Henle. Renal arteriovenous measurements reveal that, at low plasma alpha-KG concentrations, lithium caused a significant decrease in both the renal uptake of alpha-KG and the peritubular transport of this organic anion. These results suggest that the alpha-KG secreted in the pars recta and/or the loop of Henle was synthesized within the renal cells of the latter segments and not transported from the blood to the tubular lumen. At higher plasma alpha-KG concentrations, both the peritubular transport and the renal reabsorption were reduced by lithium.

Absorption and metabolism of alpha-ketoglutarate in growing pigs.

The portal appearance of enteral alpha-ketoglutarate (AKG) and the effect of enteral or parenteral AKG on portal net appearance of glucose, short-chain fatty acids, alanine, aspartate, glutamate, glutamine, proline and insulin were investigated in three growing pigs. During the experimental samplings the pigs were fed hourly with a standard feed mix with 5% glucose (control), 5% AKG (enteral) or no feed additive but continuously infused with AKG into the mesenteric vein in an amount equivalent to 5% of feed intake (parenteral). The arterial plasma concentration of AKG increased (p < 0.05) following both enteral (from 16+/-2 to 22+/-3 micromol/l) and parenteral (from 16+/-2 to 425+/-27 micromol/l) administration of AKG. With the enteral treatment 4+/-1% of the AKG could be accounted for in the portal vein, however, with the parenteral treatment 86+/-5% could be accounted for in the portal vein. The arterial plasma concentration of proline increased (p < 0.05) with the enteral treatment (365 +/- 3 to 443 +/- 39 micromol/l), but was not affected by the parenteral treatment (p > 0.10). The plasma concentration glutamine decreased (p < 0.05) with the parenteral treatment only. The portal net appearance of proline showed a numerical increase with the enteral treatment but no other affects on arterial concentrations or portal net appearance were found. A small accompanying study showed that only small amounts of enteral AKG was present in the small intestine. It was therefore concluded that enteral AKG has a low availability to peripheral tissues either because it is absorbed and metabolized in the stomach and duodenum or because it is metabolized by microbes in the stomach. The study showed that AKG is metabolized differently following enteral and parenteral application in growing pigs.

Uptake of 2-oxoglutarate in Synechococcus strains transformed with the Escherichia coli kgtP gene.

A number of cyanobacteria from different taxonomic groups exhibited very low levels of uptake of 2-[U-(14)C]oxoglutarate. Synechococcus sp. strain PCC 7942 was transformed with DNA constructs carrying the Escherichia coli kgtP gene encoding a 2-oxoglutarate permease and a kanamycin resistance gene cassette. The Synechococcus sp. strains bearing the kgtP gene incorporated 2-oxoglutarate into the cells through an active transport process. About 75% of the radioactivity from the 2-[U-(14)C]oxoglutarate taken up that was recovered in soluble metabolites was found as glutamate and glutamine. 2-Oxoglutarate was, however, detrimental to the growth of a Synechococcus sp. strain bearing the kgtP gene.

Kinetics of 2-oxoglutarate uptake by synaptosomes from bovine and rat retina and cerebral cortex and regulation by glutamate and glutamine.

2-Oxoglutarate (2-OG) is a metabolic precursor of glutamate and may be utilized to replenish the neurotransmitter pool. 2-OG is rapidly transported into neurons by a high-affinity carrier that is particularly prevalent in glutamatergic terminals. Here we report the kinetics of [U-14C]2-OG uptake by crude synaptosomal preparations from bovine and rat retina and brain, and the modulatory effects of glutamate and glutamine. In all four tissues, 2-OG uptake was mediated by a high-affinity system (Kt approximately 1 microM) that was subject to negative feedback control by glutamate and biphasic modulation by glutamine (another precursor of neurotransmitter glutamate).

Urate/alpha-ketoglutarate exchange in avian basolateral membrane vesicles.

Membrane transport pathways for transcellular secretion of urate across the proximal tubule were investigated in avian kidney. The presence of coupled urate/alpha-ketoglutarate exchange was investigated in basolateral membrane vesicles (BLMV) by [(14)C]urate and [alpha-(3)H]ketoglutarate flux measurements. An inward Na gradient induced accumulation of alpha-ketoglutarate of sufficient magnitude to suggest a Na-dicarboxylate cotransporter. An inward Na gradient also induced concentrative accumulation of urate in the presence of alpha-ketoglutarate but not in its absence, suggesting urate/alpha-ketoglutarate exchange. alpha-Ketoglutarate-dependent stimulation of urate uptake was not observed in brush-border membrane vesicles. An outward urate gradient induced concentrative accumulation of alpha-ketoglutarate. alpha-Ketoglutarate-coupled urate uptake was specific for alpha-ketoglutarate, Cl dependent, and insensitive to membrane potential. alpha-Ketoglutarate-coupled urate uptake was inhibited by increasing p-aminohippurate (PAH) concentrations, and alpha-ketoglutarate-coupled PAH uptake was observed. alpha-Ketoglutarate-coupled PAH uptake was inhibited by increasing urate concentrations, and an outward urate gradient induced concentrative accumulation of PAH. These results suggest a Cl-dependent, alpha-ketoglutarate-coupled anion exchange mechanism as a pathway for active urate uptake across the basolateral membrane of urate-secreting proximal tubule cells.