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Social animals with a keen sense of smell, such as rats, can teach each other about the safety of new foods by a practice called social transmission of food preference (STFP). In this olfactory learning paradigm, a rat that has just eaten can familiarize another rat with the taste odor of a new food by allowing the naive rat to sniff its breath. The naive rat apparently infers that the novel food is safe to eat because another rat has eaten it, and exhibits greatly diminished fear of the new food at first exposure. On page 924 of this issue, Lesburgueres et al. ( 1 ) describe their discovery of two fascinating aspects of this form of long-term memory: Specific neurons or synapses in the cerebral cortex are specifically “tagged” and thereby allocated to participate in a memory for a particular food odor, and this process involves epigenetic molecular mechanisms.

Hippocampal cultures offer unique advantages for the study of neuronal development and synaptogenesis. Studies performed on this model enabled dissection of the temporal sequence of events which lead to the differentiation of pre- and postsynaptic compartments.

The time course of metabolism of (U-$\sp?$C) glutamate and (1-$\sp?$C) GABA by young transpiring wild type and transgenic tobacco (Nicotiana tabacum L. cv. Samsun. NN) shoots overexpressing glutamate decarboxylase was examined in the presence of the glycine decarboxylase inhibitor aminoacetonitrile. When the recycling of photorespiratory ammonium and the metabolism of (U-$\sp?$C) glutamate via GS/GOGAT was prevented by aminoacetonitrile, the size of the glycine and 2-oxoglutarate pools, respectively, increased to 300% and 1000-2800% of the controls after two hours, whereas the pool size and $\sp?$C-content of glutamine decreased to 60% and 30% of the controls, respectively. The proportion of $\sp?$C in Krebs-cycle organic acids increased from 10-25% to 30-40% of the controls. Succinate was labeled to a greater extent than citrate/2-oxoglutarate at the earliest sampling time of 15 min. The data were interpreted as support for an increased flux of carbon through the GABA shunt when glutamine metabolism was inhibited. I propose that under nitrogen limitation, glutamate metabolism via the GABA shunt is a mechanism to maintain the carbon:nitrogen balance in the plant.

Deficient activity of the alpha-ketoglutarate dehydrogenase complex (KGDHC) has been documented in Alzheimer disease (AD) brain and fibroblasts. We examined glutamate metabolism in intact AD fibroblasts previously shown to have reduced KGDHC activity to determine whether this enzyme deficiency leads to a metabolic deficit in extraneural tissues. After exposure to [13N]ammonia for 5 min, the ratio of [13N]glutamate to [13N]aspartate and the ratio of unlabeled glutamate to aspartate were comparable in AD and control fibroblasts. The ratio of [13N]glutamate/[13N]glutamine was 3.7 +/- 1.8 in AD and 6.8 +/- 2.8 in control cells, but the difference was not statistically significant. Unlabeled glutamate/glutamine ratios were similar in the AD and control cells. The rate of conversion of alpha-keto[1-14C]glutarate to 14CO2 was slightly but not statistically significantly less in the AD than in the control cells. The similarity in levels of glutamate and other amino acids in intact AD and control cells may be due in part to the high glutamate and glutamine content of the culture medium used. The similar incorporation of label derived from [13N]ammonia into glutamate/aspartate and the similar levels of alpha-ketoglutarate in AD and control cells may be due in part to the fact that KGDHC is not the rate-limiting enzyme for the tricarboxylic acid cycle in fibroblasts, although it does appear to be rate-limiting in brain. Cultured cells are established tools for identifying genetic and molecular defects in the host, but the studies above show the limitations in using them as a model of brain cell metabolism.

17. Ji, Z., Aas, 1.-E., Laake, J., Walberg, F. and Ottersen, 0. P. (1991) J. Comp. Neurol. 307,296-310 18. Bramham, C. R., Torp, R., Zhang, N., StormMathisen, J. and Ottersen, 0. P. (1990) Neuroscience 39,405-417 19. Torp, R., Haug, F. M., T~nder , N., Zimmer, J. and Ottersen, 0. P. (1992) Neuroscience 46,807-823 20. Burger, P. M., Mehl, E., Cameron, P. L., Maycox, P. R., Baumert, M., Lottspeich, F., De Camilli, P. and Jahn, R. (1989) Neuron 3,715-720 21. Shupliakov, O., Brodin, L., Ottersen, 0. P., StormMathisen, J. and Cullheim, s. (1992) J. Neurosci. 12, 3789-3803 22. Ottersen, 0. P., Laake, J. H. and Storm-Mathisen, J. (1990) Arch. Ital. Biol. 128, 11 1-125 23. Ottersen, 0. P., Storm-Mathisen, J., Bramham, C., Torp, R., Laake, J. and Gundersen, V. (1 990) Prog. Brain Res. 83,991 14 24. Nicholls, D. G. (1989) J. Neurochem. 52,331-341 25. Erecinska, M. and Silver, I. A. (1990) Prog. Neuro

The concentration of glutamate in the brain extracellular fluid must be kept low (approximately 3 microM) in order to maximize the signal-to-noise ratio upon the release of glutamate from neurons. In addition, the nerve endings require a supply of glutamate precursors that will not cause depolarization. The major precursor to neuronal glutamate is glutamine, which is synthesized in astrocytes and converted to glutamate in neurons. However, glutamine is not the sole source. Alanine also might serve as a precursor to glutamate via transamination, although this reaction is relatively inactive in synaptosomes. Finally, the branched-chain amino acids, and in particular leucine, appear to be very important precursors to glutamate and glutamine in astrocytes. By providing alpha-NH2 groups for the synthesis of glutamine, leucine also abets the uptake into brain of neutral amino acids, which are transported in exchange for brain glutamine. In addition, the branched-chain ketoacids are readily reaminated to the cognate amino acids, in the process consuming glutamate. Intraneuronal consumption of glutamate via ketoacid reamination might serve to buffer internal [glutamate] and to modulate the releasable pool.

The effects of maternal hypothyroxinemia during pregnancy on brain amino acid metabolism in adult rat progeny were investigated. Normal and partially thyroidectomised dams were mated and the progeny allowed to grow to adulthood. [U-14C]Glutamate, [U-14C]glucose and [4,5-3H]leucine metabolism were determined in cerebral cortical slices, and enzymes associated with glutamate metabolism were assayed in cerebral cortical homogenates.

The major excitatory neurotransmitters in the mammalian CNS--glutamate (Glu) and acetylcholine (ACh)--have vitally important beneficial functions but also harbor treacherous neurotoxic potential which, as described in this review, can be expressed in several different ways.

The biochemistry of synaptic transmission, especially the neurobiology of dopamine signaling, is discussed.

An earlier observation from this laboratory (J. Gen. Microbiol. 64, 423–427) that NAD-dependent glutamate dehydrogenase activity is modulated by rapid inactivation has been extended to show that this mechanism is completely reversible. Changes in properties of the enzyme accompany inactivation and two different forms, active (a) and inactive (b) of the enzyme with distinctive properties have been isolated. Incubation of the inactive enzyme with magnesium in vitro produced a rapid increase of activity; this was accompanied by a change in the properties of the enzyme to those of the a form. This control mechanism of enzyme interconversion appears widespread among yeasts. Its probable role in modulating glutamete synthesis and degradation is discussed.

Simultaneous measurements were made of net and unidirectional glutamate metabolism by the portal-drained viscera, liver, kidneys, and hindquarters of fed, acidotic, and fasted sheep. The contribution of glutamate to glutamine and glucose production also was estimated. Of the glutamate present in whole blood, 45% was in plasma and 55% in the cellular fraction. Acidosis and fasting reduced blood glutamate concentrations, but did not change the plasma:cellular ratio. [14C]glutamate exchanged only little between plasma and blood cells. Clearly, this demonstrates a lack of the alpha-amino acid transport system in blood cells. Net rates of plasma glutamate flux by the portal-drained viscera and kidneys were less than 0.5 mmol/h, but, in fed sheep, the liver released 2--3 mmol/h into the plasma and the hindquarters removed an average of 0.9 mmol/h. Both were reduced by acidosis and fasting. Unidirectional rates were highly significant and greater than net rates. Acidosis and fasting primarily seemed to affect production by the liver, but only utilization by the hindquarters. Plasma glutamate turnover averaged 6--9 mmol/h, but interconversions with glutamine were low; only 12--25% was converted to glutamine and most of this occurred in extrarenal tissues. A similar rate of 20--26% of the glutamate was converted to glucose, which accounted for about 4% of the total glucose produced. The kidneys seemed to play an important gluconeogenic role; whereas the liver possibly could account for 100% of the glucose produced from glutamate in fed sheep, the kidneys accounted for 40--45% during acidosis and fasting.

In structurally preserved cerebellar slices, the sites of high affinity uptake of acidic amino acids were analyzed using the non-metabolizable analogue, d -[3H]aspartate. Electron microscopic autoradiography showed the greatest accumulation of grains to be over glial structures. The labelling of the perikarya, dendrites and axonsof the putatively glutamatergic granule cells was very low. However ‘hypothetical grain’ analysis indicated that the terminals of these cells are probable sources of radioactivity even though they contained less than 9% of the total grains in the molecular layer. The resolution of the autoradiographic technique did not permit definitive conclusion, as the parallel fibre terminals are too small and are ensheathed by thin glial processes. Nevertheless, further supporting evidence for some d -[3H]aspartate uptake into parallel fibre terminals was obtained using mechanically chopped cerebellar slices in which compared with glia presynaptic structures are selectively preserved. It is concluded that, in line with hypotheses relating to the compartmentation of glutamate metabolism, the principal sites of uptake of acidic amino acids in the cerebellum are the glial cells. The results have clear implications regarding the use of high affinity uptake as a marker for glutamatergic nerve terminals.

Four hours following oral NH4Cl, renal glutamate concentrations in rats decreased from 31.1 µ moles/g dry wgt to 23.5 µ m/g dry weight (p

Dasinger , B. L. (University of Wisconsin, Madison) and J. B. Wilson . Glutamate metabolism in Brucella abortus strains of low and high virulence. J. Bacteriol. 84: 911–915. 1962.— Brucella abortus strains of low virulence oxidize glutamate at a high rate, whereas strains of high virulence oxidize glutamate at a relatively low rate. Results indicated that this observation was not related to differences in pathway of glutamate oxidation or to differences in total enzyme activity. Permeability studies showed that the maximal rates of glutamate accumulation were 2.8 μmoles per 2 min per g (wet wt) for a strain of high virulence and 6.2 μmoles per 2 min per g for a strain of low virulence, but equal intracellular steady-state concentrations were attained by both types of strains. Evidence is presented which suggests that the site of glutamate oxidation is separate from the pool of glutamate being measured. Unequal rates of permeability at these sites could be the reason for the differences in rate of glutamate oxidation.

The pathway from glutamic acid to α-hydroxyglutaric acid in Peptococcus aerogenes proceeds via α-ketoglutaric acid and is mediated by two NAD-dependent enzymes. One enzyme, an NAD-dependent glutamic acid dehydrogenase, oxidatively deaminates glutamic acid to α-ketoglutaric acid. The other enzyme, α-ketoglutaric acid reductase, reduces α-ketoglutaric acid to α-hydroxyglutaric acid in the presence of NADH. The demonstration of a very low level of α-ketoglutaric acid dehydrogenase activity in crude cell-free extracts indicates that the primary metabolic pathway for glutamic acid carbons proceeds via α-hydroxyglutaric acid and not via the TCA cycle. Potential energy-yielding mechanisms are discussed relative to the known specific distribution of glutamic acid carbon atoms in fermentation products.

Washed bovine spermatozoa were incubated with glutamate-1- and -5- 14 C in the presence of unlabeled pyruvate or α -ketoglutarate. In air, both pyruvate and α -ketoglutarate depressed 14 CO 2 production from glutamate-1- 14 C. Under anoxia, pyruvate tended to stimulate 14 CO 2 production while α -ketoglutarate depressed it. In other studies with uniformly labeled glutamate, 14 CO 2 production decreased while oxygen uptake increased as the α -ketoglutarate concentration was increased to 10mm; 20mm α -ketoglutarate inhibited both oxygen uptake and 14 CO 2 production. As unlabeled oxalacetate concentration was increased to 10mm, the 14 CO 2 production from glutamate-U- 14 C was only slightly depressed while oxygen uptake increased. Thus, glutamate degradation proceeded mainly via α -ketoglutarate and the citric acid cycle, but keto acid concentration influenced the pathway utilized.

A description is given of an inhibitor, propavane, which: 1. 1) is specific for glutamate oxidation in low concentrations, 2. 2) gives half maximal state 3 (i.e. ADP supplemented) respiration for about 12 uM, i.e. for 25 mu moles/mg protein, 3. 3) inhibits state 4 (i.e. ADP limited) respiration only very slightly, 4. 4) is not influenced by the presence of inorganic phosphate, 5. 5) cannot be reversed by uncouplers (2,4-dinitrophenol), 6. 6) maintains a completely oxidized steady state of the pyridine nucleotides in the presence of glutamate even when preincubated with amytal (2.7 mM).

Arterio-venous differences across forearm muscle in man in both prolonged starvation and in the postabsorptive state, show an uptake of glutamate and a relatively greater production of glutamine. Splanchnic arteriovenous differences in the postabsorptive state show a net uptake of glutamine and lesser rate of glutamate production. These data suggest that muscle is a major site of glutamine synthesis in man, and that the splanchnic bed is a site of its removal. The relative roles of liver and other tissues in the splanchnic circuit were not directly assessed, only the net balance. These data in man are in conflict with most previous studies in other species attributing the major proportion of glutamine production to the liver and, pari passu, to the splanchnic bed.

The basic observation which lead to the postulate of the metabolic compartmentation of glutamate metabolism in brain, as well as of the related Krebs cycles, has been the subject of two previous symposia in this series (Ralazs and Cremer, 1973; Berl et al., 1975) and is well known to most of the members of this Advanced Study Institute. In taking an overview of the field we can summarize by saying that labeled substrates fall into two classes, those yielding brain glutamine with a specific activity greater than glutamate and those giving the opposite behavior. Hence the two cycle models have been considered as the simplest possible ones which can lead to a reasonable fit to both sets of experimental data. By following other indices, e.g. the specific activity of aspartate relative to glutamate or the distribution of label between C-1 of glutamine and the rest of the molecule further sub-classifications can be made and these point to the need for further subdivision of the two cycle models. On the other hand, models with three to five cycles and associated pools of amino acids related to these Krebs cycles are extremely underdetermined with the experimental data currently available (van den Berg et al.,1975). Furthermore, it seems to be getting more difficult to generate the masses of data needed to refine such models. Accordingly we have been setting our sights lower as far as developing multicycle models and rather have been trying to examine pieces of existing models in more detail with a view to refining these sub-models further before tackling the bigger problems.

The possible involvement of the glutamine synthetase/glutamate synthase pathway in ammonia assimilation is reported in a monokaryon of Coprinus cinereus.

13 C-NMR has been used te follow the metabolism of 13 C-enriched substrates in isolated perfused rat liver. The fate of 90% enriched [2- 13 C]acetate has been studied in the perfused liver in order to investigate mitochondrial metabolism and the interrelations between cytosolic and mitochondrial pools of metabolites. Some compounds of the intermediary metabolism where found to be extensively labelled, e.g. glutamate, glutamine, acetoacetate and β-hydroxybutyrate. Under our experimental conditions, labelling of glutamate reached a steady-state within 30 min after the onset of perfusion of 20 mM acetate. In addition, the observed incorporation of 13 C into glutamine can be linked to the operation of the glutamate-glutamine antiport and to the high activity of cytosolic glutamate synthetase. The finding of both active glutaminase and glutamine synthetase activity in the same liver cells is evidence of the existence of an active glutamine-glutamate futile cycle.

The effect of kindling rats (with an electrical stimulus applied daily to the frontal cortex) on the concentrations of taurine, aspartate, glutamate, glutamine, and gamma-aminobutyric acid has been investigated. When compared with control groups, cortical glutamine concentrations were significantly decreased in kindled rats by approximately 20%. This decrease in glutamine directly correlated with the after-discharge duration (r = 0.84, p = 0.005). The significance of this in relation to glutamate metabolism and kindling is discussed here.

Glutamate and related amino acids in brain have long attracted the attention of many scientific investigators. The suggested beneficial effects of glutamate on behavior was one of the factors stimulating research on the metabolic aspects of glutamate in brain, a research area already actively pursued in the thirties by Krebs, Weil-Malherbe, and others (Weil-Malherbe, 1950). There is no doubt that Waelsch, through his involvement in the reported beneficial effects of glutamate on epileptic patients, was stimulated to concentrate on the metabolism of glutamate in the brain.

A single assay system has been developed for six enzymes of glutamate metabolism: glutamate dehydrogenase, glutaminase, asparate aminotransferase, γ-aminobutyrate aminotransferase, alanine aminotransferase, and glutamate decarboxylase. The first five are assayed by coupling them to Escherichia coli glutamate decarboxylase and measuring the release of 14 CO 2 from radioactive substrates. Glutamate decarboxylase is assayed directly. The assays are simple, use but one technique, and require very little working time. At a reasonable cost per assay, they are considerably more sensitive than other commonly used assays for the same enzymes. The sensitivity of the assay at a fixed price increases as the substrate concentration decreases.

1. (14)C from [1-(14)C]glucose injected intraperitoneally into mice is incorporated into glutamate, aspartate and glutamine in the brain to a much greater extent than (14)C from [2-(14)C]glucose. This difference for [1-(14)C]glucose and [2-(14)C]glucose increases with time. The amount of (14)C in C-1 of glutamate increases steadily with time with both precursors. It is suggested that a large part of the glutamate and aspartate pools in brain are in close contact with intermediates of a fast-turning tricarboxylic acid cycle. 2. (14)C from [1-(14)C]acetate and [2-(14)C]acetate is incorporated to a much larger extent into glutamine than into glutamate. An examination of the time-course of (14)C incorporated into glutamine and glutamate reveals that glutamine is not formed from the glutamate pool, labelled extensively by glucose, but from a small glutamate pool. This small glutamate pool is not derived from an intermediate of a fast-turning tricarboxylic acid cycle. 3. It is proposed that two different tricarboxylic acid cycles exist in brain.

Sirtuins are enzymes that respond to cellular or organismal stress by activation or inhibition of the activities of proteins via deacetylating or ADP-ribosylating them after translation. Previously, it was shown that sirtuins affect the age-related pathways in the cell via interfering with the metabolism. SIRT4 was also shown to affect the glutamate metabolism both in pancreatic cells and also in brain. The aim of this study is to identify the interacting proteins of mitochondrial SIRT4 in mouse brain. We identified a number of proteins that interact with SIRT4 in mouse brain. We also showed that the ATP levels are lower in SIRT4 KO mouse brain compared to controls. Revealing SIRT4 interacting proteins will help to understand its function better in brain and lead the path for developing therapies for agerelated brain diseases.

The glutamate-mediated synaptic responses of neocortical pyramidal cell to fast-spiking interneuron (pyramidal-FS) connections were studied by performing paired recordings at 30-33 degrees C in acute slices of 14- to 35-day-old rats (n = 39). Postsynaptic fast-spiking (FS) cells were recorded in whole cell configuration with a patch pipette, and presynaptic pyramidal cells were impaled with sharp intracellular electrodes. At a holding potential of -72 mV (near the resting membrane potential), unitary excitatory postsynaptic potentials (EPSPs) had a mean amplitude of 2.1 +/- 1.3 mV and a mean width at half-amplitude of 10.5 +/- 3.7 ms (n = 18). Bath application of the N-methyl-D-aspartate (NMDA) receptor antagonist D(-)2-amino-5-phosphonovaleric acid (D-AP5) had minor effects on both the amplitude and the duration of unitary EPSPs, whereas the alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA)/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) almost completely blocked the synaptic responses. In voltage-clamp mode, the selective antagonist of AMPA receptors 1-(4-aminophenyl)-3-methylcarbamyl-4-methyl-7,8-methylenedioxy-3, 4-dihydro-5H-2,3-benzodiazepine (GYKI 53655; 40-66 microM) blocked 96 +/- 1.9% of D-AP5-insensitive unitary excitatory postsynaptic currents (EPSCs), confirming the predominance of AMPA receptors, as opposed to kainate receptors, at pyramidal-FS connections (n = 3). Unitary EPSCs mediated by AMPA receptors had fast rise times (0.29 +/- 0.04 ms) and amplitude-weighted decay time constants (2 +/- 0.8 ms; n = 16). In the presence of intracellular spermine, these currents showed the characteristic rectifying current-voltage (I-V) curve of calcium-permeable AMPA receptors. A slower component mediated by NMDA receptors was observed when unitary synaptic currents were recorded at a membrane potential more positive than -50 mV. In response to short trains of moderately high-frequency (67 Hz) presynaptic action potentials, we observed only a limited temporal summation of unitary EPSPs, probably because of the rapid kinetics of AMPA receptors and the absence of NMDA component in these subthreshold synaptic responses. By combining paired recordings with extracellular stimulations (n = 11), we demonstrated that EPSPs elicited by two different inputs were summed linearly by FS interneurons at membrane potentials below the action potential threshold. We estimated that, in our in vitro recording conditions, 8 +/- 5 pyramidal cells (n = 18) should be activated simultaneously to make FS interneurons fire an action potential from -72 mV. The low level of temporal summation and the linear summation of excitatory inputs in FS cells favor the role of coincidence detectors of these interneurons in neocortical circuits.

The latest annual volume of a series entitled Current Topics in Eye Research appeared recently. Jose A. Zadunaisky and Hugh Davson are the editors of the series, and each volume contains four to six chapters about selected topics written by experts in the field. The chapters are reviews that primarily, but not exclusively, concentrate on the author's work and are aimed at the basic scientist in the same or a related field. This volume contains chapters about glutamate metabolism in the retina (Liane Reif-Lehrer, PhD), aqueous humor outflow (Paul Kaufman, MD), noninvasive corneal redox fluorophotometry (Barry Masters, PhD), and fluid and electrolyte transports across the corneal endothelium (Jorge Fischbarg, PhD, and Jong Jim Lim, PhD). The chapters are consistently informative, detailed, and well written. The chapter on glutamate metabolism in the retina is encyclopedic, with over 500 references. Dr Reif-Lehrer examines the molecule through many facets, including its metabolism and

International audience; The NMR structures of Corynebacterium glutamicum OdhI described by Barthe et al. in this issue of Structure reveal a major conformational rearrangement upon phosphorylation, suggesting an autoinhibition mechanism that accounts for the functional properties of the protein as a regulatory switch in glutamate metabolism.