STRUCTURAL ALTERATIONS OF AMINO ACIDS AT THE LEVEL OF AMINOACYL-tRNAS: IDENTIFICATION OF THE TRANSFORMATION PRODUCTS OF DICARBOXYLIC AMINO ACIDS

Abstract— Glutamyl-, glutaminyl-, aspartyl- and asparaginyl-tRNAs were separated into different isoacceptor species by reverse phase column chromatography. RNase hydrolysates of any of the isoacceptor [¹⁴C]aminoacyl-tRNAs for a given amino acid gave radioactivity profiles, on paper electrophoresis, very similar to unfractionated tRNA. This suggested a lack of tRNA specificity for the transformation reaction involving the aminoacyl moieties of asparaginyl and glutaminyl-tRNAs. GnP₂ and AnP₂ detected in the products of deaminoacylation of glutaminyl and asparaginyl-tRNA showed a number of properties in common with GnE₃ and AnE₃ present in the RNase hydrolysates of the same tRNAs. Thus, GnP₂ and GnE₃ chromatographed in the same position in the phenol: water solvent and both yielded glutamate on acid hydrolysis and a mixture of glutamine and isoglutamine on alkaline hydrolysis. Similarly, AnP₂ and AnE₃ had the same R_F value in phenol :water chromatography and gave aspartate or a mixture of asparagine and isoasparagine when hydrolyzed with acid or alkali. On the basis of these results and other evidence, GnP₂ and GnE₃ were assigned the structure, α-aminoglutarimide; AnP₂ and AnE₃ were identified as α-aminosuccinimide. These cyclic compounds are presumed to be formed by nucleophilic attack of the amide nitrogen of asparagine or glutamine on the carbon of the aminoacyl ester carbonyl group. The cyclization-deesterification appeared to be facilitated by RNase hydrolysis of aminoacyl-tRNA indicating that the aminoacyl-tRNA is probably more resistant to this reaction than aminoacyladenosine. Neither the imides nor the amino acid isoamides were detected in the reaction mixture in which aminoacylation of tRNA was performed, suggesting that a mechanism may exist for inhibition of the cyclization reaction under conditions of active aminoacylation.     

REACTIONS OF FREE AND tRNA BOUND GLUTAMATE AND GLUTAMINE

—[¹⁴C]-Glutamate and [¹⁴C]-glutamine were incorporated into calf brain tRNA in the presence of homologous aminoacyl-tRNA synthetases. When the tRNAs were then deaminoacylated and chromatographed, a number of radioactive products were found in addition to the original amino acids. One of the products of glutamate transformation was identified to be glutamine. Formation of the radioactive products of glutamate in the presence and absence of tRNA indicated that glutamine was produced from glutamate at the level of the free amino acid followed by the incorporation of both substances into tRNA. Examination of the products of deaminoacylation of glutaminyl-tRNA showed that glutamine underwent structural alterations at the level of the aminoacyl-tRNAs to give rise to a cyclic derivative of glutarimide. This reaction was specific for glutamine, and constituted approximately 15 per cent of the total radioactivity in the deaminoacylation products of glutaminyl-tRNA.     

METABOLISM OF GLUCOSE AND FREE AMINO ACIDS IN BRAIN, STUDIED WITH 14C-LABELLED GLUCOSE AND BUTYRATE IN RATS INTOXICATED WITH CARBON DISULPHIDE

Abstract— The effect of 15 h continuous exposure to CS₂ on the metaboliam of glucose and free amino acids in the brain of rats was studied. CS₂ caused a moderate hypoglycaemia. There were also changes in the amounts of some amino acids in the brain. Glutamate and γ-aminobutyrate were lower whereas glutamine was markedly increased. Comparative studies in vivo of the metabolism of [2-¹⁴C]glucose and [1-¹⁴C]butyrate indicated that CS₂ did not affect glycolysis or the incorporation of ¹⁴C from glucose into amino acids except into γ-aminobutyrate which was reduced. Contrary to the findings with [¹⁴C]glucose, CS₂ provoked distinct changes in the labelling of amino acids when [¹⁴C]butyrate was the precursor. The most notable change was a markedly increased incorporation of ¹⁴C into glutamine. Based on the two-compartment model of brain glutamate the experimental findings indicated that CS₂ affected metabolism associated with the 'small' pool of glutamate but had a minimal effect on metabolism associated with the 'large' glutamate pool. The possibility is suggested that the changes observed involved an increased rate of ammonia removal. The low incorporation of ¹⁴C into γ-aminobutyrate from either precursor is consistent with other evidence showing that CS₂ interferes with pyridoxal phosphate-dependent enzymes.     

THE INCORPORATION OF DOUBLE-LABELLED ACETATE INTO GLUTAMATE AND RELATED AMINO ACIDS FROM ADULT MOUSE BRAIN: COMPARTMENTATION OF AMINO ACID METABOLISM IN BRAIN

Abstract– We have determined the incorporation of [³H]-, [1-¹⁴C]- and [2-¹⁴C]acetate into glutamate, glutamine and aspartate of the adult mouse brain. All these three acetates were incorporated more extensively into glutamine than into glutamate. This has been reported by several authors for each of these labelled acetates in separate experiments. It was shown that [³H, 2-¹⁴C]acetate can be used to obtain an acetate labelling ratio analogous to the previously used [2-¹⁴C]acetate/[1-¹⁴C]acetate labelling ratio. From these acetate labelling ratios of glutamine and glutamate conclusions can be deduced about the dynamic relationship of these amino acids with each other and with the tricarboxylic acid cycle.
A fairly large isotope effect between acetate and glutamate was observed. As this isotope effect is very likely caused by the citrate synthase reaction, it can be argued that citrate synthase involved in the conversion of labelled acetate into glutamate is far out of equilibrium in vivo. Comparing our data with literature data, the possibility can be suggested that citrate synthase in the acetate metabolizing compartment is in situ kinetically distinct from citrate synthase in other compartments of the brain.     

METABOLISM OF γ-HYDROXY-[1-14C] BUTYRATE BY RAT BRAIN: RELATIONSHIP TO THE KREBS CYCLE AND METABOLIC COMPARTMENTATION OF AMINO ACIDS

Abstract— Ninhydrin decarboxylation experiments were carried out on the labelled amino acids produced following intraventricular injection of either γ-hydroxy-[1-¹⁴C]butyric acid (GHB) or [1-¹⁴C] succinate. The loss of isotope (as ¹⁴CO₂) was similar for both substances. The [1-¹⁴C]GHB metabolites lost 75% of the label and the [1-¹⁴C] succinate metabolites lost 68%. This observation gives support to the hypothesis that the rat brain has the enzymatic capacity to metabolize [1-¹⁴C]GHB to succinate and to amino acids that have the isotope in the carboxylic acid group adjacent to the a-amino group. These results also indicate that the label from [1-¹⁴C]GHB does not enter the Krebs cycle as acetate. The specific activity ratio of radiolabelled glutamine to glutamic acid was determined in order to evaluate which of the two major metabolic compartments preferentially metabolize GHB. It was found that for [1-¹⁴C]GHB this ratio was 4.20 ± 0.18 (S.E. for n = 7) and for [l-¹⁴C]succinate this ratio was 7.71 (average of two trials, 7.74 and 7.69). These results suggest that the compartment thought to be associated with glial cells and synaptosomal structures is largely responsible for the metabolism of GHB. Metabolism as it might relate to the neuropharmacological action of GHB is discussed.     

14C-LABELLED AMINO ACIDS AND GLUCOSE IN RAT BRAIN AND LIVER AFTER INJECTION OF [2-14C]PROPIONATE

Abstract— [2-¹⁴C]Propionate injected into rats was metabolized into [¹⁴C]glucose and ¹⁴C-labelled aspartate, glutamate, glutamine and alanine. The results are consistent with the conversion of propionate into succinate and the oxidation of succinate into oxaloacetate, the precursor of labelled amino acids and the substrate for gluconeogenesis.
The ratio of the specific radioactivity of glutamine to glutamate was greater than 1 during the 30 min period in the brain, indicating that propionate taken up by the brain was metabolized mainly in the 'small glutamate compartment' in the brain. The results, therefore, support the previous conclusion (G aitonde , 1975) that the labelling of amino acids by [¹⁴C]propionate formed from [U-¹⁴C>]-threonine in thiamin-deficient rats was metabolized in the 'large glutamate compartment' of the brain.
The specific radioactivity ratio of glutamine to glutamate in the liver was less than 1 during the 10 min period but greater than 1 at 30min. These findings which gave evidence against metabolic compartments of glutamate in the liver, were interpreted as indicative of the entry of blood-borne [¹⁴C]glutamine synthesized in other tissues, e.g. brain. The labelling of amino acids when compared to that after injection of [U-¹⁴C]glucose showed that [2-¹⁴C]propionate was quantitatively a better source of amino acids in the liver. The concentration of some amino acids in the brain and liver was less in the adult than in the young rats, except for alanine and glutathione, where the liver content was more than double that in the adult.     

Influence of nitrate and ammonium on the growth and 2,4-diaminobutyric acid composition of flatpea (Lathyrus sylvestris L.)

Abstract. Presence of 2.4-diaminobutyric acid (A₂bu), a neurotoxin, in tissues of flatpea ( Lathyrus sylvestris L.) necessitates a thorough understanding of the regulation of this nonprotein amino acid before the species can be recommended to livestock producers for forage applications. To determine how different concentrations and ratios of NO₃ and NH⁺₄ in growth media influence the levels of A₂bu and other free amino acids in the 'Lathco'flatpea cultivar, plants were grown hydroponically in controlled environments. The concentration of A₂bu was highest in tissues when the NO₃ to NH⁺₄ ratio in the nutrient solution was low. Responses of amides and other nonprotein amino acids, especially in the roots, followed a similar trend. Free protein amino acids in leaves and stems were generally unaffected by changes in NO₃ to NH⁺₄ ratios. In roots, protein amino acids increased as the NO₃ to NH⁺₄ ratio in the growth medium increased. Ammonium inhibited shoot and root growth; NO₃ alleviated the toxic effects of NH⁺₄. Soluble protein concentrations were higher in the shoots of NO₃-fed plants and in the roots of plants supplied with NH⁺₄. These results suggest that accumulation of A₂bu and other nonprotein amino acids, as well as asparagine and glutamine, plays a role in detoxification of NH⁺₄ and storage of N.     

COMPARTMENTATION AND LABELLING OF AMINO ACIDS IN THE DEVELOPING RAT BRAIN AFTER INJECTION OF [U-14C]RIBOSE

Abstract— [U-¹⁴C]Ribose was given by subcutaneous injection to young rats aged 2–56 days. During the first week after birth ¹⁴C in the brain was found mainly combined in glucose, fructose and sedoheptulose which contained 46–57 per cent of the ¹⁴C in the acid soluble metabolites in the rat brain. In contrast, during the critical period (10–15 days after birth) the ¹⁴C in the free sugars decreased from 24 to 3 per cent, while the ¹⁴C content of amino acids in the brain increased from 11 to 44 per cent of the total perchloric acid-soluble ¹⁴C. The increase in labelling of amino acids during the critical period was attributed to increased glycolysis and increased oxidation of pyruvate. The relative specific radioactivity of y -aminobutyrate and aspartate in the rat brain at 28 days after birth was equal to or greater than the relative specific radioactivity of glutamate. Assuming that the increase in amino acid content following the cessation of cell proliferation in the brain is located mainly in cell processes (cytoplasm of axons, dendrites, glial processes and nerve terminals), tentative values were estimated for the pool sizes of glutamate, glutamine, aspartate and y -amino butyrate.     

EFFECTS OF POTASSIUM AND GLUTAMATE ON BRAIN CORTEX SLICES: UPTAKE AND RELEASE OF GLUTAMIC AND OTHER AMINO ACIDS

Abstract— Effects of an increased concentration of K⁺ (55 m m ) in the medium on fluxes of glutamate and other amino acids in the presence and absence of 10 m m -glutamate were studied. The following observations were made:
(1) The efflux of glutamate is slightly increased by excess K⁺. The glutamate efflux is smaller than the potassium fluxes.
(2) The K⁺-induced increase of glutamate efflux is enhanced under anoxia or in glutamate-containing media.
(3) The influx of glutamate is unaffected or slightly increased by excess K⁺.
(4) The efflux of GABA is increased by excess K⁺, both in the absence and in the presence of glutamate.
(5) Efflux of glutamine, leucine and lysine is increased by excess K⁺, but only provided that glutamate is also present in the medium.
(6) Efflux of glutamate and of GABA is increased by addition of 10 m m -glutamate.     

EFFLUX OF PHENYLALANINE FROM RAT CEREBRAL CORTEX SLICES AS INFLUENCED BY EXTRA- AND INTRACELLULAR AMINO ACIDS

Abstract— Effects of other amino acids on the efflux of l -[³H]phenylalanine from rat cerebral cortex slices were studied in a superfusion system. Extracellular large neutral amino acids caused a strong trans-stimulation of [³H]phenylalanine efflux. Some small neutral amino acids were less effective, whereas acidic and basic amino acids and the amino acids without an amino group in the α-position were ineffective. Any trans -inhibition was not detected. The stimulatory trans -effects of phenylalanine and tryptophan were additive, reversible and concentration-dependent. They were apparently mediated by the same mechanisms. The efflux of [³H]phenylalanine was much slower at 273 K than at 310 K, but the effects of unlabelled phenylalanine and tryptophan on it were qualitatively similar at both temperatures. Amino acids accumulated intracellularly at moderately high concentrations did not inhibit [³H]phenylalanine efflux, but phenylalanine, leucine, isoleucine and norleucine caused an enhancement. Spontaneous efflux of [³H]phenylalanine showed some similarities to physical diffusion, but its selective and specific modification by other amino acids strongly suggests the involvement of mediated processes.     

THE UPTAKE OF AMINO ACIDS BY THE INTACT OLFACTORY BULB OF THE MOUSE: A COMPARISON WITH TISSUE SLICE PREPARATIONS

Abstract— Intact olfactory bulbs from 8- to 15-day-old mice were compared to slices of olfactory bulb and cerebral hemisphere with respect to uptake of amino acids, respiratory rate, levels of ATP, retention of sodium and potassium, and extracellular space. The uptake of amino acids was lower in intact bulbs than in slice preparations, both in regard to initial rates of uptake and to final steady state levels, at external amino acid concentrations from 0·2 to 2·0mM. Uptake was lower in bulbs attached to brain than in those separated from it and somewhat higher in the half of the bulb closer to the cut surface. In all preparations the uptake of glutamic acid and glycine was highest, uptake of histidine and valine was intermediate, and uptake of lysine was lowest. These differences between intact bulbs and slices could not be correlated with differences in respiratory rate, levels of ATP, or changes in levels of Na⁺ or K⁺ ions. Increases in dextran and inulin spaces, however, were greatest in preparations having the highest rates of amino acid uptake. Although for several amino acids the maximal velocity of uptake (V_max) was 4-fold higher in slices of bulb than in intact bulbs, the affinity of amino acids to their carrier systems ( K _m) was similar, an indication that the same transport process was operative in both cases. On the basis of these results we propose that intact olfactory bulbs incubated in vitro possess a regulatory mechanism for the limitation of amino acid uptake that is absent or diminished in slices.     

Rapid assimilation of recently fixed N2 in root nodules of Myrica gale

In Myrica gale L. plants the assimilation of ammonia released by symbiotic Frankia was observed by ¹⁵N₂ labelling and subsequent analysis of the isotopic enrichment of nodule amino acids over time by single ion monitoring gas chromatography-mass spectrometry. In detached nodules of Myrica , glutamine was the first amino acid labelled at 30 s and subsequently the amino acids glutamate, aspartate, alanine and γ-amino butyric acid (GABA) became labelled. This pattern of labelling is consistent with the incorporation of ammonium via glutamine synthetase [GS; EC 6.3.1.2]. No evidence for the ammonium assimilation via glutamate dehydrogenase [GDH; EC 1.4.1.2] was observed as glutamate became labelled only after glutamine. Using attached nodules and pulse-chase labelling, we observed synthesis of glutamine, glutamate, aspartate, alanine, GABA and asparagine, and followed the transport of fixed nitrogen in the xylem largely as glutamine and asparagine. Estimation of the cost of nitrogen fixation and asparagine synthesis in Myrica nodules suggests a minimum of one sucrose required per asparagine produced. Rapid translocation of recently fixed nitrogen was observed in Myrica gale nodules as 80% of the nitrogen fixed during a 1-h period was translocated out of the nodules within 9 h. The large pool of asparagine that is present in nodules may buffer the transport of nitrogen and thus act to regulate nitrogen fixation via a feedback mechanism.     

ON THE SITE OF ORIGIN OF TRANSMITTER AMINO ACIDS RELEASED BY DEPOLARIZATION OF NERVE TERMINALS IN VITRO

Abstract— —The site of origin of transmitter amino acids released by depolarizing agents from nerve endings was studied. The model used was the incubated and depolarized synaptosome preparation from which the component soluble, synaptic vesicle, membrane and mitochondrial sub-fractions were obtained. Synaptosomal amino acids were radioactively labelled from D-[U-¹⁴C]glucose in vivo by intraventricular injection and in vitro during subsequent incubation. The specific radioactivities of amino acids released in response to K⁺ (56 mM) or veratrine (75 μM) were found to closely resemble those of the soluble cytoplasmic fraction, in most cases differing significantly from those of the other fractions. The specific radioactivity of the GABA and aspartate released by K⁺ stimulation and the GABA and glutamate released by veratrine were significantly different from that of the vesicles in each case. The specific radioactivities of glutamate released by both agents, and also GABA with K⁺ stimulation, were approximately double that of the amino acid released in control conditions. Depletion of the soluble cytoplasmic pools of glutamate, GABA and aspartate occurred following stimulation, corresponding to the induced-release of these compounds. Turnover of the amino acids in the other subfractions was too low to account for their participation in the release process in addition to the soluble cytoplasmic pool. A cytoplasmic origin of release of neurotransmitter amino acids from nerve endings is proposed.     

N-System Amino Acid Transport at the Blood-CSF Barrier

Abstract: Despite l -glutamine being the most abundant amino acid in CSF, the mechanisms of its transport at the choroid plexus have not been fully elucidated. This study examines the role of L-, A-, ASC-, and N-system amino acid transporters in l -[¹⁴C]glutamine uptake into isolated rat choroid plexus. In the absence of competing amino acids, approximately half the glutamine uptake was via a Na⁺-dependent mechanism. The Na⁺-independent uptake was inhibited by 2-amino-2-norbornane carboxylic acid, indicating that it is probably via an L-system transporter. Na⁺-dependent uptake was inhibited neither by the A-system substrate α-(methylamino)isobutyric acid nor by the ASC-system substrate cysteine. It was inhibited by histidine, asparagine, and l -glutamate γ-hydroxamate, three N-system substrates. Replacement of Na⁺ with Li⁺ had little effect on uptake, another feature of N-system amino acid transport. These data therefore indicate that N-system amino acid transport is present at the choroid plexus. The V _max and K _max for glutamine transport by this system were 8.1 ± 0.3 nmol/mg/min and 3.3 ± 0.4 m M , respectively. This system may play an important role in the control of CSF glutamine, particularly when the CSF glutamine level is elevated as in hepatic encephalopathy.     

TRANSAMINATION OF AMINO ACIDS IN HOMOGENATES OF RAT BRAIN

Abstract— The aminotransferase activity of homogenates of brains from adult and neonatal rats has been investigated. Aminotransferase activity was demonstrated wtih 15 of 22 amino acids incubated with seven keto acids. The basic amino acids exhibited little or no activity.

• 1 The greatest activity was obtained when glutamate or aspartate was incubated with α-ketoglutarate or oxaloacetate. Significant activity was also observed when the neutral aliphatic and aromatic amino acids were incubated with these two keto acids.
• 2 Activity with pyruvate was obtained principally upon incubation with glutamate and alanine. Most of the other amino acids that underwent transamination with α-ketoglutarate also did so with pyruvate, although at a lower rate.
• 3 When phenylpyruvate was added to the medium, glutamate, phenylalanine and tyrosine transaminated most actively.
• 4 Incubations with 11 amino acids and glyoxylic acid demonstrated aminotransferase activity, with glutamate and ornithine being the most active substrates.
• 5 α-Ketoisocaproate and α-ketoisovalerate accepted amino groups primarily from the branched-chain amino acids. Except for glutamate, activity with other amino acids was low or not detectable.
• 6 A comparison of aminotransferase activity in the newborn brain with that in the adult brain showed that the greatest change in activity occurred for glutamate with pyruvate or for alanine with α-ketoglutarate, these activities increasing about 10-fold from birth to adulthood; during this time activities with most other amino acids increased two- to threefold. Amino transfers from the branched-chain amino acids showed no increase with maturation, and some reactions, such as that with methionine and a number of keto acids, decreased from birth to adulthood.
• 7 Our results correspond in general to previous studies of aminotransferase activity in brain and in liver. However, our study also indicates a possible second aminotransferase acting on the branched-chain amino acids, the presence of aminotransferase activity for methionine and asparagine, and relatively high aminotransferase activity for glutamine or ornithine when incubated with glyoxylic acid rather than other keto acids. Moreover, phenylpyruvate and glyoxylate are active in amino transfers and may serve as substrates for a number of aminotransferases.

     

Translocation of amino acids from stem nodules of Sesbania rostrata demonstrated by GC-MS in planta 15N isotope dilution

We report a novel use of the ¹⁵N dilution technique to detail the translocation of amino compounds in the legume Sesbania rostrata . The conventional ¹⁵N dilution technique follows the dilution of ¹⁵N within a labelled plant, as ¹⁴N₂ is fixed by symbiotic bacteria. In our experiments, stem-nodulated Sesbania rostrata were enriched by feeding with ¹⁵N ammonium nitrate for 2 weeks, followed by a 1 week period where the only N available to the plants was via nitrogen fixation of atmospheric N₂. We measured the composition, concentration and ¹⁵N enrichment of amino compounds in various plant tissues, both above and below the stem nodules, using GC-MS and isotopic abundance mass spectrometry techniques. Approximately 28% of the total N in the stem nodules was derived from internal plant sources. The ureides allantoic acid and allantoin were not abundant in xylem, leaf or nodule tissues. The amides asparagine and glutamine were the major export products from stem nodules although a wide range of other amino compounds are also synthesized. Amino acids within the nodules had a low level of enrichment, demonstrating that a small fraction (≈ 11%) was derived from outside the nodules, and significant cycling of N (28% of xylem N) through the root system was revealed by measurements of ¹⁵N distribution and amino acid concentrations.     

STIMULATED INCORPORATION OF AMINO ACIDS INTO PROTEINS OF SYNAPTOSOMAL FRACTIONS INDUCED BY DEPOLARIZING TREATMENTS

Abstract— Rat cortical synaptosome preparations incorporated l -amino acids into protein at a linear rate over 30–60 min. Synaptosomes showed large increases in incorporation after treatment with electrical pulses, veratrine or K⁺. This was inhibited (controls, 73%; electrically stimulated 58%) by cycloheximide and by chloramphenicol (28 and 44% respectively). Omission of Ca²⁺ from the medium had no effect, but the absence of Na⁺ greatly diminished the stimulus-induced increase in incorporation due to pulses. Electrical pulses, veratrine and K⁺ (in order of effectiveness), all showed their greatest proportional effects on amino acid incorporation into the Triton-X-100 insoluble portions of the synaptosomal membrane: the 'junctional complex', soluble in SDS, and the residue. Tetrodotoxin (1 μ m ), although ineffective when potassium was the stimulus, prevented or reduced the respiratory response, K⁺ loss and differential amino acid release, as well as the amino acid incorporation into protein observed with pulses and veratrine.
C-6 glioma cells showed no stimulus-dependent increase in incorporation. Preliminary data, of gel electrophoretic analysis, on the proteins labelled under our conditions is presented.     

INTERACTION OF CATECHOLAMINE AND AMINO ACID METABOLISM IN BRAIN: EFFECT OF PARGYLINE AND l-DOPA

Abstract— The combination of l -DOPA and pargyline caused a decrease in level of aspartate and an increase in that of glutamine in vivo in cerebral cortex, cerebellum, brain stem, hypothalamus, neostriatum and cervical cord of rat. There was also a decreased incorporation of radioactivity from [1-¹⁴C]acetate into amino acids in vivo , most notably in cerebellum and brain stem. The labelling of glutamine was especially affected. In addition, cortical slices were prepared from guinea pigs which had been pretreated with pargyline. These slices were incubated with and without 1 m m l -DOPA in media containing [1-¹⁴C]acetate. Pargyline alone caused a stimulation of the labelling of glutamate and aspartate but not glutamine and GABA; the levels of aspartate and GABA were greater than in control slices. The addition of l -DOPA to slices from pargylinized animals caused a severe decrease in glutamine labelling but not in that of glutamate or aspartate; the level of glutamine was increased while that of glutamate was decreased. The results are discussed in terms of the known biochemical and morphological compartmentation of amino acids in brain. It is suggested that catecholamines, in the process of functioning as transmitters, may also function as metabolic regulators of other transmitters, e.g. amino acids, as well as of the energy required for balanced neuronal function.     

Major nitrogen compounds transported in xylem vessels from roots to top in Citrus trees

Four-year-old citrus trees ( Citrus unshiu Marcovitch) were fed via the roots with (¹⁵NH₄)2sO₄ or K¹⁵NO₃ as a nitrogen source. Nitrogenous compounds and their isotopic abundances in fine roots and xylem sap from trunks were assayed in order to obtain information on the species of nitrogen released by the root system into the ascending xyiem stream.
Arginine, asparagine, nitrate and proline in xylem sap accounted for 48, 21, 13 and 10%, respectively, of the total nitrogenous constituents tested in the sap. However, in the trees fed with labelled ammonium the main nitrogenous compound labelled with ¹⁵N in the xylem sap was asparagine and glutamine, which accounted for 79% and 18%, respectively, of total labelled nitrogen. In the xylem sap of trees fed with labelled nitrate, nitrate accounted for 94% of total labelled nitrogen. Nitrate and asparagine followed by glutamine showed the highest ratios of isotopic abundance in xylem sap as compared to fine roots. Proline and arginine had much lower ratios. These results indicate that nitrate, asparagine and glutamine are the main nitrogenous compounds released by the roots to the xylem stream, whereas arginine and proline are released into the xylern vessels by the trunk tissues. Furthermore, nitrate and asparagine are probably in steady movement upward in the trunk xylem, whereas glutamine is more easily taken up by the trunk tissues than nitrate and asparagine.     

METABOLISM OF GLUTAMATE AND RELATED AMINO ACIDS IN THE 10-DAY-OLD MOUSE BRAIN: EXPERIMENTS WITH LABELLED ACETATE AND β-HYDROXYBUTYRATE

Abstract– The pattern of incorporation of [³H, 1-¹⁴C]- and [³H. 2-¹⁴C]acetate into glutamate and related amino acids was studied in the brain of 10-day-old mice. A comparison of these patterns with those obtained for the adult brain led to the suggestion that the glutamate pool labelled directly by acetate is a much larger fraction of the total glutamate pool in the 10-day-old brain than it is in the adult brain.
Some data on the pattern of labelling of brain amino acids by 3-hydroxybutyrate. glucose and acetate support the hypothesis that direct carboxylation of pyruvate is somewhat more active in the immature than in the mature brain.
Differences in the labelling patterns of free and protein-bound brain amino acids by acetate, do indicate that the free amino acid pool labelled by acetate is not the precursor pool for protein synthesis.