ALTERATION OF GABA SYSTEM AND PURKINJE CELLS IN RAT CEREBELLUM BY THE CONVULSANT 3-MERCAPTOPROPIONIC ACID

Abstract— The effect of the convulsant, 3-mercaptopropionic acid (MP) on the content of free amino acids and on the activity of some enzymes related to their metabolism was studied in the rat cerebellum. A decrease in the activity of glutamate decarboxylase (EC 4.1.1.15) and in the level of GABA was found; at the same time, the activity of GABA-aminotransferase (EC 2.6.1.19) was increased. These changes coincided with a profound alteration of the morphology of the Purkinje cells which was related to the dose of MP. These findings, plus some changes in the content of other free amino acids and the activities of related enzymes, suggest that 3-mercaptopropionic acid induces in the cerebellum an imbalance among the amino acids involved in the excitation-inhibition mechanisms.     

Hormonal control of metabolism of gamma-aminobutyric acid in the rat hypothalamus and hippocampus

It has been established that hydrocortisone administration increased the amount of total, free, bound and synaptosomal GABA in the hypothalamus, glutamate decarboxylase activity in the homogenate and synaptosomes and time of the mediator turnover. ACTH administration increased the GABA content and glutamate decarboxylase activity in synaptosomes. The total amino acid content and time of its turnover got higher only with single hormone administration. In the hippocamp hydrocortisone administration increased the total and free GABA contents, its turnover time, glutamate decarboxylase activity in the homogenate and decreased GABA-aminotransferase activity in the homogenate and synaptosomes. The GABA level in synaptosomes grew only with multiple hormone administration. Single administration of ACTH decreased the total GABA content, glutamate decarboxylase activity in the homogenate, while its multiple administration increased the GABA level in synaptosomes followed by a decrease of GABA-aminotransferase activity in the homogenate and synaptosomes. The GABA turnover time fell with single hormone administration and grew with the multiple one. Adrenalectomy induced no changes in the GABA content and activity of its metabolism enzymes in the hypothalamus, however the bound GABA level decreased, while the turnover time increased. In the hippocamp adrenalectomy decreased total, free and synaptosomal GABA contents, glutamate decarboxylase activity in a homogenate and turnover time. Subsequent hydrocortisone administration only partly normalized the revealed changes of the GABA metabolism in the brain structures under adrenalectomy.     

Developmental change of endogenous glutamate and gamma-glutamyl transferase in cultured cerebral cortical interneurons and cerebellar granule cells,and in mouse cerebral cortex and cerebellum in vivo

The developmental change of endogenous glutamate, as correlated to that of gamma-glutamyl transferase and other glutamate metabolizing enzymes such as phosphate activated glutaminase, glutamate dehydrogenase and aspartate, GABA and ornithine aminotransferases, has been investigated in cultured cerebral cortex interneurons and cerebellar granule cells. These cells are considered to be GABAergic and glutamatergic, respectively. Similar studies have also been performed in cerebral cortex and cerebellum in vivo. The developmental profiles of endogenous glutamate in cultured cerebral cortex interneurons and cerebellar granule cells corresponded rather closely with that of gamma-glutamyl transferase and not with other glutamate metabolizing enzymes. In cerebral cortex and cerebellum in vivo the developmental profiles of endogenous glutamate, gamma-glutamyl transferase and phosphate activated glutaminase corresponded with each other during the first 14 days in cerebellum, but this correspondence was less good in cerebral cortex. During the time period from 14 to 28 days post partum the endogenous glutamate concentration showed no close correspondence with any particular enzyme. It is suggested that gamma-glutamyltransferase regulates the endogenous glutamate concentration in culture neurons. The enzyme may also be important for regulation of endogenous glutamate in brain in vivo and particularly in cerebellum during the first 14 days post partum. Gamma-glutamyl transferase in cultured neurons and brain tissue in vivo appears to be devoid of maleate activated glutaminase.Abbreviations used Asp-T aspartate aminotransferase (EC 2.6.1.1) - GABA-T GABA aminotransferase (EC 2.6.1.19) - GAD glutamate decarboxylase (EC 4.1.1.15) - gamma-GT gamma-glutamyl transferase (gamma-glutamyl transpeptidase) (EC. 2.3.2.2) - Glu glutamate - GDH glutamate dehydrogenase (EC 1.4.1.3) - GS glutamine synthetase (EC 6.3.1.2) - MAG maleate activated glutaminase - Orn-T ornithine aminotransferase (EC 2.6.1.13) - PAG phosphate activated glutaminase (EC 3.5.1.1)     

Mercaptopropionic acid: a convulsant that inhibits glutamate decarboxylase

—3-Mercaptopropionic (MP) and 4-mercaptobutyric (MB) acids caused convulsions in rats after the intraperitoneal administration of 32 and 200 mg/kg body wt. respectively. These compounds competitively inhibited glutamate decarboxylase (L-glutamate 1-carboxy-lyase; EC 4.1.1.15) of rat brain and bacterial GABA transaminase (4-aminobutyrate: 2-oxoglutarate aminotransferase; EC 2.6.1.c). Glutamine synthetase (L-glutamate: ammonia ligase (ADP); EC 6.3.1.2) was not affected. Pyridoxal phosphate added in vitro did not reverse the inhibition. Action of these compounds is compared to methionine sulphoximine, a bacterial exotoxin (lactylaminopimelic acid) and to hydrazinopropionic acid.     

FORMATION OF GABA FROM PUTRESCINE IN THE BRAIN OF FISH (SALMO IRIDEUS GIBB.)

Abstract— It was demonstrated after intraperitoneal and intracerebral injections of [1,4-¹⁴C]-putrescine.2 HCl that GABA is formed in vivo in the trout brain via a pathway in which glutamic acid is not an intermediate. Intraperitoneal and intracerebral injections of both thiosemicarbazide and 3-mercaptopropionic acid had no measurable effects on GABA concentration, transformation of glutamic acid into GABA in vivo , or on glutamate de-carboxylase activity in the brain within the first 3 h after the application of the inhibitors. Only a small decrease in concentration of pyridoxal phosphate was noticed in the fish brain after thiosemicarbazide administration. The relatively high concentrations of pyridoxal phosphate in the trout brain may be one of the reasons for the ineffectiveness of thiosemicarbazide in inhibiting glutamate decarboxylase in vivo. After intracerebral injections of [1-¹⁴C]GABA, a half-life of 7 h was determined for GABA. The slow turnover rate of GABA in trout brain, which can be assumed from this observation, may give a further explanation of the ineffectiveness of the glutamate decarboxylase inhibitors in lowering the GABA content ot fish brain within a few hours.     

EFFECT OF UNILATERAL VISUAL DEPRIVATION AND VISUAL STIMULATION ON THE ACTIVITIES OF GLUTAMATE DECARBOXYLASE, GABA-α KETOGLUTARATE TRANSAMINASE, ASPARTATE AMINOTRANSFERASE AND HEXOKINASE OF THE OPTIC LOBE OF THE ADULT PIGEON

Abstract— A study was made of the effect of unilateral visual deprivation and stimulation upon the activities of glutamate decarboxylase (EC 4.1.1.15), GABA-α-ketoglutarate transaminase (EC 2.6.1.19). aspartate aminotransferase (EC 2.6.1.1) and hexokinase (EC 2.7.1.1) in the optic lobe of the adult pigeon ( Columba Livia ). Visual deprivation was achieved by eyelid suturing or by enucleation and maintained for 1–9 weeks. Unilateral visual stimulation was maintained for 75 min following 72 h in the dark. A small but significant increase was observed in the activities of glutamate decarboxylase and aspartate aminotransferase after unilateral visual stimulation and a decrease after unilateral enucleation. The activities of GABA-α-ketoglutarate transaminase and hexokinase decreased after unilateral visual stimulation and increased after enucleation. Unilateral eyelid suturing resulted in a significant reduction in the activity of glutamate decarboxylase and an increase in the activity of GABA-α-ketoglu-tarate transaminase. Hexokinase activity was, however, unchanged following unilateral eyelid suturing.     

Effect of pyridoxal phosphate on gamma-aminobutyric acid metabolism in different sections of the brain in irradiated animals]

A study was made of the effect of X-rays (4,5 Gy) and pyridoxal phosphate (3 mg/kg, v/v) on the activity of pyridoxal enzymes of GABA metabolism (e.g. glutamate decarboxylase, E.C. 4.1.1.15) and aminobutyrate aminotransferase (GABA-T, E.C. 2.6.1.19), as well as on GABA and glutamate content of the hemisphere cortex, brain stem and cerebellum of rabbits 6 and 10 days following irradiation and injection of a coenzyme. The height of the radiation sickness in rabbits was characterized by the manifest changes in glutamate decarboxylase and GABA-T activity, as well as in GABA and glutamate content of various brain parts differing in the structural and functional functions. The administration of pyridoxal phosphate produced pronounced activation of glutamate decarboxylase, particularly 6 days after irradiation and administration of the co-enzyme, and, to a lesser extent, influenced GABA-T function. Pyridoxal phosphate favored maintaining the GABA level above the control level in the hemisphere cortex and brain stem 6 and 10 days after exposure. The injection of pyridoxal phosphate did not normalize the glutamate content of the brain parts 6 days after exposure, but favored the normalization of GABA-T activity on day 10.     

Distribution of Glycine, γ-Aminobutyric Acid, Glutamate Decarboxylase, and γ-Aminobutyric Acid Transaminase in Rabbit and Mudpuppy Retinas

Abstract: The distributions of glycine, γ-aminobutyric acid (GABA), glutamate decarboxylase (EC 4.1.1.15), and GABA transaminase (EC 2.6.1.19) were determined in rabbit and mudpuppy retinas. In both species, peak levels of the amino acids and the enzymes occurred in the inner plexiform layer. Glutamate decarboxylase was almost entirely confined to the inner plexiform layer. Determinations were also made of the GABA content of 107 individual putative amacrine cell somas from mudpuppy retina. About 30% of those somas were found to have high endogenous GABA levels.     

THE EFFECTS OF SOME NEWER ANAESTHETICS ON THE IN VITRO ACTIVITY OF GLUTAMATE DECARBOXYLASE AND GABA TRANSAMINASE IN CRUDE BRAIN EXTRACTS AND ON THE LEVELS OF AMINO ACIDS IN VIVO

—The effects of several anaesthetic and hypnotic compounds with well-defined excitatory side-effects on glutamate decarboxylase and γ-aminobutyric acid transaminase activity have been examined. The dissociative anaesthetics ketamine and γ-hydroxybutyric acid produced competitive inhibition of glutamate decarboxylase with respect to glutamate at concentrations which had no effect on GABA transaminase activity. The inhibitor constant (K_i) values were, ketamine: 13.3 mm , γ-hydroxybutyric acid; 8.8 mm . The steroid anaesthetic alphaxalone was also a potent competitive inhibitor of glutamate decarboxylase K_i= 4.1 mm ). Pentobarbitone, thiopentone and methohexitone non-competitively inhibited both glutamate decarboxylase and GABA-transaminase but only at high concentration (> 20 mm ). None of the drugs tested produced any significant change in brain GABA or glutamate levels following the injection of an hypnotic or anaesthetic dose. It is proposed that an alteration in the rate of GABA synthesis as a result of the inhibition of glutamate decarboxylase could explain the convulsive properties of the dissociative anaesthetics when given at high doses.     

THE EFFECT OF dl-ALLYLGLYCINE ON POLYAMINE AND GABA METABOLISM IN MOUSE BRAIN

Abstract— dl -Allylglycine, a potent inhibitor of glutamate decarboxylase in vivo when given intraperitoneally, causes a marked decrease in brain GABA concentration and at the same time a dramatic increase in l -ornithine decarboxylase activity and a simultaneous decrease in S -adenosyl- l -methionine decarboxylase activity followed by putrescine accumulation. It does not, however, alter the degree of GABA formation from putrescine. The timing of the recovery of glutamate decarboxylase activity after the injection of dl -allylglycine is concomitant with that of the GABA concentration, indicating that it is probably glutamate decarboxylase that is solely responsible for making up the GABA deficit caused by dl -allylglycine, and that the changes in polyamine metabolism are associated in some indirect way with the recovery process.     

Developmental changes in the brain levels of neurotransmitters as influenced by maternal ethanol consumption in the rat.

Abstract— Effects of prolonged maternal ethanol consumption by pregnant and lactating rats were investigated on the cerebral contents of certain neurotransmitters and the activities of their enzymes in fetal and neonatal brains. Ethanol consumption by the pregnant rats resulted in a significant decrease in the fetal brain contents of acetylcholine and an increase in γ-aminobutyric acid and glutamate. The cerebral levels of 5-hydroxytryptamine (5-HT), norepinephrine of choline in the fetal brains from the ethanol group were not significantly different from those observed in the control group. With the increase in gestational age, an increase in the levels of 5-HT, norepinephrine, acetylcholine, γ-amino-butyric acid (GABA) and glutamate was observed in the fetal brains from both ethanol and control groups. The brains of neonates suckling on ethanol-fed mothers showed a significant decrease in acetylcholine levels and an increase in GABA and glutamate levels as compared to the corresponding controls. The activities of L-glutamate decarboxylase and GABA-aminotransferase in the fetal as well as neonatal brains were decreased in the ethanol-fed group as compared to the controls. A single injection of acetaldehyde resulted in a decrease in acetylcholine, GABA and glutamate levels in both neonatal and adult brains.     

THE TOXIC EFFECT OF SODIUM GLUTAMATE ON RAT RETINA: CHANGES IN PUTATIVE TRANSMITTERS AND THEIR CORRESPONDING ENZYMES

Abstract– Subcutaneous administration of high doses of sodium glutamate to rats during their first week after birth produced an almost total loss of choline acetyltransferase, a 90% reduction in glutamate decarboxylase and 70% reductions in acetylcholinesterase and DOPA decarboxylase activities in the adult retina. In addition there was a 70% decrease in GABA and 35-55% decrease in aspartate, glutamate, glycine, alanine and glutamine. No reduction in taurine was observed. The results support the view that the enzymes are mainly localized in the interneurons of retina and that taurine is present in the photoreceptor cells.
Glutamate treatment was also followed by a small reduction in choline acetyltransferase and glutamate decarboxylase of the superior colliculus and in choline acetyltransferase of hippocampus, whereas no changes could be detected in the lateral geniculate body of the adult rat. Unilateral enucleation performed on 1-day-old animals did not alter choline acetyltransferase, acetylcholinesterase, glutamate decarboxylase and DOPA decarboxylase activities in the superior colliculus and in the lateral geniculate body of the adult rat.     

Effect of Inhibitors of GABA Aminotransferase on the Metabolism of GABA in Brain Tissue and Synaptosomal Fractions

Abstract: Five inhibitors of the GABA degrading enzyme GABA-aminotransferase (GABA-T), viz., gabaculine, γ-acetylenic GABA, γ-vinyl GABA, ethanolamine O -sulphate, and aminooxyacetic acid, as well as GABA itself and the antiepileptic sodium vdproate were administered to mice in doses equieffective to raise the electroconvulsive threshold by 30 V. The animals were killed at the time of maximal anticonvulsant effect of the respective drugs and GABA, GABA-T and glutamate decarboxylase (GAD) were determined in whole brain and synaptosomes, respectively. The synaptosomal fraction was prepared from brain by conventional ultracentrifugation procedures. All drugs studied brought about significant increases in both whole brain and synaptosomal GABA concentrations, and, except GABA itself, inhibited the activity of GABA-T. Furthermore, all drugs, except GABA and γ-acetylenic GABA, activated GAD in the synaptosomal fraction. This was most pronounced with ethanolamine O -sulphate, which induced a twofold activation of this enzyme but exerted only a weak inhibitory effect on GABA-T. The results suggest that activation of GAD is an important factor in the mechanism by which several inhibitors of GABA-T and also valproate increase GABA concentrations in nerve terminals, at least in the relatively non-toxic doses as used in this study.     

An Ascochlorin Derivative,AS-6, Potentiates Insulin Action in Streptozotocin Diabetic Mice and Rats

γ-Aminobutyric acid (GABA), a hypotensive compound, and alanine accumulated in tea leaves under anaerobic conditions. Since the ¹⁵N in ¹⁵N-glutamic acid was well incorporated in GABA and alanine during anaerobic incubation, glutamic acid seemed to be a source of nitrogen for the increased GABA and alanine. GOT and GPT were the predominant amino acid transaminases in tea leaves. Although glutamate decarboxylase and GPT seemed to be important for GABA and alanine accumulation, the activities of these enzymes did not increase under anaerobic conditions. Glutamate decarboxylase, which formed GABA from glutamate, was purified 52.4-fold. This enzyme, with an optimum pH at 5.8, was activated by pyridoxal phosphate and used only l-glutamic acid as a substrate.     

Metabolic routes of GABA formation in chick embryo brain

Sobue and Nakajima (1978) reported that GABA formation from putrescine is significant in chick embryo brain between days 6 and 8 of incubation. They attributed an important functional role to the putrescine-derived GABA. We found that depletion of putrescine and spermidine in chick embryos by inhibition of ornithine decarboxylase activity did not decrease the in vivo rate of GABA formation, showing that putrescine is, from a quantitative point of view, a negligible source for GABA in chick embryo brain. The changes of brain GABA levels obtained after administration of glutamate decarboxylase inhibitors and in vitro determinations of glutamate decarboxylase activity were compatible with the assumption that GABA is mainly formed by decarboxylation of l-glutamate, even during early brain development. Participation of the NAD⁺-dependent, aerobic transformation of glutamate into GABA (Seiler and Wagner, 1976) in the overall GABA production of chick embryo brain could, however, not be excluded.     

Acute metabolic effects of taurine on the enzymes metabolizing glutamate and gaba

100 mg of taurine per kg body weight had been administered intraperitoneally and 30 min after the administration the animals were sacrificed. Glutamate dehydrogenase, aspartate aminotransferase, alanine aminotransferase, glutaminase, glutamine synthetase, glutamate decarboxylase and GABA aminotransferase along with the content of glutamate and GABA in cerebral cortex, cerebellum and brain stem were studied and compared with the same obtained in the rats treated with normal saline in place of taurine. The results indicated a significant decrease in the activity of glutamate dehydrogenase in cerebral cortex and cerebellum and a significant increase in brain stem. Glutaminase and glutamine synthetase were found to increase significantly both in cerebral cortex and cerebellum. The activities of glutamate decarboxylase was found to increase in all the three regions along with a significant decrease in GABA aminotransferase while the content of glutamate showed a decrease in all the three brain regions, the content of GABA was observed to increase significantly. The above effects of taurine on the metabolism of glutamate and GABA are discussed in relation to the functional role of GABA and glutamate. The results indicate that taurine administration would result in a state of inhibition in brain.     

Pentylenetetrazole inhibits glutamate dehydrogenase and aspartate aminotransferase, and stimulates GABA aminotransferase in homogenates from rat cerebral cortex

The mechanism by which pentylenetetrazole provokes convulsions in animals has been investigated by measuring its influence in vitro on the activities of several enzymes of glutamate metabolism in rat brain homogenates. Pentylenetetrazole does not affect the specific activities of glutamine synthetase, glutaminase, or glutamate decarboxylase; it inhibits those of glutamate dehydrogenase and aspartate aminotransferase, and stimulates that of gamma-aminobutyric acid (GABA) aminotransferase. The overall consequence of the action of pentylenetetrazole on the activities of these enzymes should be an increase in the concentration of glutamate and a decrease in that of GABA. This modulation of glutamate and GABA metabolism by pentylenetetrazole could contribute to the triggering of convulsions.     

GABA and its related enzymes in the lower auditory system of the guinea pig

The possible existence of GABA-transmitter neurons in the lower auditory system of the guinea pig has been investigated by means of three different experimental approaches: (1) the regional distribution of GABA and its related enzymes, (2) the subcellular distribution of glutamate decarboxylase, and (3) the effect of selected nerve lesions on glutamate decarboxylase concentrations in the auditory nuclei. Within the regions investigated considerable variations in glutamate decarboxylase activity and GABA concentration were found, with the highest values observed in the inferior colliculus. The dorsal cochlear nucleus also contained significant amounts of both glutamate decarboxylase and GABA, in addition to high concentrations of GABA transaminase. The subcellular distribution of glutamate decarboxylase was bimodal in both the cochlear nucleus and inferior colliculus with most enzyme activity recovered in the soluble and synaptosomal fractions. Neither end organ (cochlea) nor trapezoid body lesions induced a significant loss of glutamate decarboxylase activity in either the cochlear nucleus or inferior colliculus. The results suggest the presence of short axon GABAergic interneurons in the cochlear nucleus, most of which appear to terminate within the dorsal cochlear nucleus.     

THE TOXIC EFFECT OF SODIUM GLUTAMATE AND DL-α-AMINOADIPIC ACID ON RAT RETINA: CHANGES IN HIGH AFFINITY UPTAKE OF PUTATIVE TRANSMITTERS

Subcutaneous administration of high doses of sodium glutamate to new born rats was used to destroy retinal interneurons and ganglion cells. Such treatment was accompanied by 90% reduction in the high affinity uptake of choline, 60–70% reductions in the uptakes of GABA, diamino-n-butyric acid and glycine and 30–40% reductions in the uptakes of asparatate and glutamate measured on retinal homogenates from 30-day-old rats. The high affinity uptakes of β-alanine and taurine were unchanged. Preincubation of retinal homogenates with 1 mM β-alanine or 100 μM diamino-n-butyric acid severely reduced the high affinity GABA uptake in control and experimental animals. In intact retinae, however, the glutamate treatment increased the high affinity uptake of β-alanine by 70%, whereas that of diamino-n-butyric acid was reduced by 40% and the high affinity uptakes of GABA and glutamate were unchanged. Four hours after injection of the gliotoxic compound DL-α-aminoadipic acid into the vitreous body of 30-day-old rats, the Müller cells could no longer be identified. This lesion was accompanied by 55% reduction in the high affinity uptake of β-alanine and 25% reduction in the uptakes of GABA and glutamate on intact retinae. The high affinity uptakes of diamino-n-butyric acid, choline and the enzyme activities of choline acetyltransferase and glutamate decarboxylase were unchanged under these conditions. After 24 h, however, the Müller cells could be recognized again, and the β-alanine uptake had normalized.     

Metabolism and functions of gamma-aminobutyric acid

Gamma-aminobutyric acid (GABA), a four-carbon non-protein amino acid, is a significant component of the free amino acid pool in most prokaryotic and eukaryotic organisms. In plants, stress initiates a signal-transduction pathway, in which increased cytosolic Ca²⁺ activates Ca²⁺/calmodulin-dependent glutamate decarboxylase activity and GABA synthesis. Elevated H⁺ and substrate levels can also stimulate glutamate decarboxylase activity. GABA accumulation probably is mediated primarily by glutamate decarboxylase. However, more information is needed concerning the control of the catabolic mitochondrial enzymes (GABA transaminase and succinic semialdehyde dehydrogenase) and the intracellular and intercellular transport of GABA. Experimental evidence supports the involvement of GABA synthesis in pH regulation, nitrogen storage, plant development and defence, as well as a compatible osmolyte and an alternative pathway for glutamate utilization. There is a need to identify the genes of enzymes involved in GABA metabolism, and to generate mutants with which to elucidate the physiological function(s) of GABA in plants.