The influence of cortical lesions on penicillin induced convulsive activity in the awake rat

1. Experiments were performed to investigate the effects of cortical lesions on convulsive behaviour. Rats were lesioned in the left motor or sensory cortex by aspirating cortical tissue 2 to 3 months prior to the elicitation of convulsions. Convulsions were induced in the awake rats by the GABA antagonist Na-penicillin (Na-PCN) which was applied into the superficial layer of the foreleg field of their right motor cortex. Convulsive activity was recorded by means of the EEG. 2. The time courses of convulsive cortical activity were similar in the animals without or with a cortical lesion. Generalized seizures belonged to the tonic-clonic type in both intact and lesioned rats. 3. The early period of convulsive activity was described by the time to the onset (latency) of the first convulsive potential, jerk and seizure, and by the mean repetition rate of jerks during the first ten minutes, and the duration of the first generalized seizure. None of these parameters was significantly affected by a cortical lesion. 4. The median duration of the convulsive activity in intact animals was 162 min. In rats with a lesion in the sensory cortex it raised to more than 540 min while a lesion of the motor cortex increased the median duration to more than 273 min. The differences between intact and lesioned rats were significant (p less than 0.01 and p = 0.05, respectively). 5. The median time to the onset of the last generalized seizure in intact rats corresponded to 92 min with respect to the time of Na-PCN application. In rats with a lesion of the sensory cortex the last seizure was generated 433 min and in animals with a lesion of the motor cortex 167 min after Na-PCN treatment of the motor cortex of one side. This increase of latency of the last seizure was significant for the rats with a lesioned sensory area (p less than 0.02) or motor area (p = 0.05) compared to that of the intact rats. Additionally, the number of generalized seizures was significantly (p less than 0.01) increased by both groups of rats with a lesion of the motor or sensory cortex. 6. It is suggested that a substantial lesion of the cortex decreases predominantly the intrinsic cortical inhibition thus destabilizing brain function. This destabilizing effect becomes pronounced under the condition of superimposed suppression of the GABAergic cortical component. It is concluded that the intrinsic cortical inhibitory mechanism which in the intact brain acts against hyperexcitation and prevents the development of neuronal synchronization, i.e. the formation of seizures, becomes less effective in performing this task once an abnormal brain activation has developed.     

Effect of aspartate and glutamate on experimental myocardial infarction in rats

Cardiac necrosis was produced in rats by administering isoproterenol sulphate (85 mg/kg, sc for 4 days). The myocardial damage was proved by observing the elevated levels of serum aspartate amino-transferase, lactate dehydrogenase and creatine phosphokinase and the changes were confirmed by histopathology of the tissue. Both aspartate and glutamate (100 mg/kg, ip) significantly reduced the elevated levels of these enzymes. The average degree of cardiac necrosis produced in these rats when observed macroscopically and histologically was also found to be significantly reduced on pretreatment with aspartate and glutamate.     

An examination of aspartate decarboxylase and glutamate decarboxylase activity in mosquitoes

A major pathway of beta-alanine synthesis in insects is through the alpha-decarboxylation of aspartate, but the enzyme involved in the decarboxylation of aspartate has not been clearly defined in mosquitoes and characterized in any insect species. In this study, we expressed two putative mosquito glutamate decarboxylase-like enzymes of mosquitoes and critically analyzed their substrate specificity and biochemical properties. Our results provide clear biochemical evidence establishing that one of them is an aspartate decarboxylase and the other is a glutamate decarboxylase. The mosquito aspartate decarboxylase functions exclusively on the production of beta-alanine with no activity with glutamate. Likewise the mosquito glutamate decarboxylase is highly specific to glutamate with essentially no activity with aspartate. Although insect aspartate decarboxylase shares high sequence identity with glutamate decarboxylase, we are able to closely predict aspartate decarboxylase from glutamate decarboxylase based on the difference of their active site residues.     

Effect of ionotropic glutamate receptors antagonists on the modifications in extracellular glutamate and aspartate levels during picrotoxin seizures: a microdialysis study in freely moving rats

Our previous studies have shown a local decrease in glutamate and aspartate levels during seizures, induced by picrotoxin microdialysis in the hippocampus of chronic freely moving rats. In this paper, we study the effect of continuous hippocampal microperfusion of the NMDA, AMPA and kainate glutamate receptor inhibitors 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine (MK-801); 6,7-dinitroquinoxaline-2,3-dione (DNQX), and 1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine hydrochloride (GYKI 52466). We also examine the action of L(-)-threo-3-hydroxyaspartic acid (THA), a glutamate and aspartate reuptake blocker, on the modification of extracellular glutamate and aspartate levels induced by picrotoxin, using the microdialysis method in freely moving rats. We found that changes in extracellular hippocampal concentrations in both amino acids are prevented by NMDA, AMPA and kainate receptor inhibitors. Seizures elicited under DNQX also induce a transient increase in aspartate extracellular levels coincident with seizure time. L(-)-threo-3-hydroxyaspartic acid increased the basal extracellular concentrations of both amino acids, but did not prevent the seizure-related decrease. Our results suggest that glutamate, the major neurotransmitter at the synaptic level, may also play an important role in non-synaptic transmission during seizures.     

Regional brain glutamate transport in rats at normal and raised concentrations of circulating glutamate

the permeability of the blood-brain barrier to glutamate was measured by quantitative autoradiography in brains of control rats (average plasma glutamate concentration of 95 ) and rats infused with glutamate (average plasma glutamate concentration of 837 m). Measurements of glutamate permeability were initiated by the injection of [¹⁴C]glutamate and stopped at 1 min to avoid the accumulation of [¹⁴C]glutamate metabolites. Glutamate entered the brain at a slow rate, with an average permeability-surface area product of 7 l.min^...g^-1, except in those areas known to have fenestrated capillaries. Glutamate accumulated in the choroid plexus of ventricles, but did not seem to enter the cerebrospinal fluid in detectable amounts regardless of the circulating concentration. Glutamate accumulated in circumventricular organs, such as the median eminence, where the radioactivity was localized without detectable spread. Infusion of glutamate to create high plasma concentrations did not result in greater spread of [¹⁴C]glutamate beyond the immediate vicinity of the circum ventricular organs.     

Electrophysiological properties of mesencephalic ventral tegmental area neurons in awake rats

A statistical analysis was made of spike activity and the shape and duration of individual action potentials in cells from the mesencephalic ventral tegmental area and adjacent regions during experiments on awake rats. Four groups of cells were identified in this test population whith electrophysiological features which may relate to their distinctive neurochemical profile and to their specific afferent connections.Institute of Pharmacology, Academy of Medical Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 18, No. 6, pp. 729–737, November–December, 1986.     

Stimulation induced recurrent epileptiform discharges block cortical and subcortical spreading depression in rats

Blockade of the spread depression [SD] in chemically or electrically stimulated areas of the cerebral cortex was analyzed in a series of experiments performed in 40 curarized, locally anaesthetised rats. Longlasting stimulation of the cerebral cortex (0.5 to 1.7 mA, 6 Hz, 0.1 ms) elicited recurrent episodes of enhanced evoked potentials propagating to remote cortical areas [Co], the caudate nucleus [Cd], the hippocampus [Hi] and the thalamus [Th] and was accompanied by marked slow potential shifts (4-6 mV amplitude, 2-3 min duration, at 2-5 min intervals]. The projected discharge interfered with SD initiation and propagation in all the examined structures. The SD blockade was most pronounced during the episodes and almost absent in the intervals between them. The block was manifested by reduced amplitude and duration of the slow potential of SD. Pentobarbital [20 mg/kg] suppressed the recurrent discharges and eliminated the corresponding SD blockade. Recurrent excitability changes induced by Cd and Th stimulation elicited similar effects but the threshold was higher in Cd and Th than in Co. SD was less effectively blocked by the projected discharge than by stimulation of the same structure, particularly in the vicinity of the recording electrodes, where the blockade could be observed even under pentobarbital. The SD blockade outlasted stimulation by a considerably longer period of time in Th (about 10 min) than in the Cd (about 3 min). The onset of stimulation and the projected episodes sometimes elicited SD waves but SD blockade prevailed with continued stimulation. The present findings support the hypothesis that excessive neural activity increases the potassium clearance and thus prevents the autoregenerative accumulation of potassium ions, mediating SD propagation.     

Penetration of the mitochondrial membrane by glutamate and aspartate

Evidence has been presented for the existence of systems located in mitochondrial membranes for the transport of phosphate (Chappell &; Crofts, 1966), dicarboxylic acids (Chappell &; Haarhoff, 1967; Robinson &; Chappell, 1967), citrate, isocitrate and cis-aconitate (Chappell, 1964, Chappell, 1966; Chappell &; Haarhoff, 1967; Chappell, Henderson, McGivan &; Robinson, 1967) and for oxoglutarate (Meijer &; Tager, 1966; Chappell et al., 1967). In this paper results are presented which reveal the probable existence of two more carrier-systems, one for L-glutamate and one for L-aspartate. The latter carrier requires the presence of L-glutamate, or certain analogues of this amino acid, before it is able to act. The evidence for the glutamate carrier is, firstly, that the rate of reduction of intramitochondrial NAD(P) by glutamate has an optimum below pH 6, whereas with the free dehydrogenase the pH optimum is greater than 8. Secondly, certain analogues of glutamate, namely 3-hydroxyglutamate, 2-aminoadipate and threo-hydroxyaspartate inhibit competitively the reduction of intramitochondrial NAD(P) by glutamate, but have no or negligible effect on the activity of enzyme in mitochondrial extracts. The evidence for the aspartate carrier rests on the fact that aspartate is unable to react rapidly with mitochondrial aspartate aminotransferase until glutamate, or of the compounds tested hydroxyglutamate, aminoadipate or threo-hydroxyaspartate, are present.     

Synthesis of glutamate and aspartate in rat brain synaptosomes

ATP and glutamine are the sources of endogenous ammonia in rat brain synaptosomes. The amount of endogenous ammonia formed from exogenous ATP is not sufficient to assure the maximum rate of aspartate and glutamate accumulation in the synaptosomes utilizing pyruvate + malate. Addition of exogenous NH4+ or depolarization of synaptosome plasma membranes with high K+ concentration led to a twofold increase in the rate of accumulation of these amino acids. This indicates that both exogenous and endogenous NH4+ is involved in the synthesis of aspartate and glutamate in nerve terminals. Accumulation of glutamate was stimulated by aminooxyacetate and inhibited by haloperidol which indicates that NH4+ is bound in the reaction catalysed by glutamate dehydrogenase. Endogenous oxaloacetate derived from pyruvate metabolism was the substrate for synthesis of aspartate. Additive inhibition of aspartate accumulation by fluorocitrate and (-) hydroxyacetate shows that, in addition to the tricarboxylic acid cycle, the reaction catalysed by ATP-citrate lyase serves in the synaptosomes as another source of oxaloacetate.     

The effect of dietary carbohydrate on the rise in plasma glutamate concentrations following oral glutamate ingestion in rats

Plasma glutamate concentrations were examined in male rats following oral intubation of monosodium L-glutamic acid (MSG, 250 mg/kg) soon after ingesting one of several meals differing in carbohydrate content. Intubation of MSG alone produced a 4-fold rise in plasma glutamate that peaked at 15 min, and returned to baseline by 60 min. Red blood cell glutamate concentrations were unchanged. The ingestion of a meal lacking carbohydrate produced a modest attenuation of the post-MSG intubation rise in plasma glutamate concentrations. This attenuating effect increased progressively with the carbohydrate content of the meal (and as the protein content declined, to maintain isocaloric meals), though as little as 5% carbohydrate marked attenuated the plasma glutamate rise. This effect diminished as the time interval between the meal and MSG intubation increased from 1 to 4 hrs. Similar, but not identical effects were noted when meals substituted fat (instead of protein) for carbohydrate. The intubation of MSG alone produced a slight increase in plasma alanine concentrations over the 60-min post-intubation period examined. The ingestion of any of the meals just prior to intubation did not influence this effect. Overall, the results indicate that although the ingestion of carbohydrate can markedly attenuate the rise in plasma glutamate that follows MSG consumption in rats, this effect is also influenced by the other macronutrients present. The absence of notable, meal related changes in plasma alanine suggests that this parameter does not provide a useful indication of gut glutamate transamination.     

Identification and cataloging of genes induced by long-lasting long-term potentiation in awake rats

Maintenance of long-term potentiation (LTP) requires de novo gene expression. Here we report the direct isolation, using PCR-differential display, of genes whose expression level was altered after induction of long-lasting LTP in the hippocampus of freely moving awake rats. Differential display using 480 primer combinations revealed 17 cDNA bands that showed a reproducible change in expression level. These cDNAs represented at least 10 different genes (termed RM1-10), all of which showed up-regulation at 75 min after LTP induction and a return to basal expression levels within 24 h. Three of these genes were known only from expressed sequence tags (RM1-3), two were known genes whose up-regulation by LTP has not been described (GADD153/CHOP and ler5), and five were known genes whose up-regulation by LTP has already been reported (MAPK phosphatase, NGFI-A/zif268, vesl-1S/homer-1a, Ag2, and krox-20). We characterized the expression profiles of genes in the two former categories with respect to NMDA receptor dependency, tissue specificity, and developmental regulation using northern blotting and semiquantitative RT-PCR. The up-regulation of all five of these genes was NMDA receptor-dependent and correlated with the persistence of LTP, suggesting that these genes may play functional roles in prolonged LTP maintenance.     

Changes in the spectrum of LDH isoenzymes in foci of epileptic activity induced by penicillin in the cerebral cortex of rats

The correlation between electrophysiological changes and isozymes of LDH of the rat brain cortex was studied in seizure foci induced by application of sodium penicillin. It was discovered that activity of LDH1 was suppressed, and that of LDH5 fraction was elevated in the determinant focus, which indicates the enhanced glucose anaerobic transformation. The spectrum of LDH isozymes did not practically differ from the indicators in control animals in a homotopic region of the contralateral hemisphere prior to creation of the mirror focus. The anaerobic processes were found to be increased in the mirror focus and in the determinant one as well. Similar pattern of changes in electrophysiological and neurochemical characteristics in the determinant and dependent mirror foci attests to the formation of a pathological system out of the two epileptic foci.     

Re-activation by glutamate or aspartate of amino-oxyacetate-inhibited aspartate aminotransferase in vitro and in isolated hepatocytes

Experiments with isolated rat hepatocytes and with cell extracts indicate, in contrast with previous reports, that pyruvate does not block or reverse the inhibition of aspartate aminotransferase (EC 2.6.1.1) by amino-oxyacetate. That inhibition, however, is partially overcome by glutamate or aspartate either in cell extracts or in whole cells incubated with substrate combinations that cause accumulation of those amino acids.     

Extracellular levels of brain aspartate, glutamate and GABA during an inhibitory avoidance response in the rat

The extracellular levels of aspartate, glutamate and GABA were measured by microdialysis, coupled with an HPLC method, in rat prefrontal cortex (mPFC) and ventral hippocampus (VH) before and during the performance of a step-down inhibitory task. The basal levels of glutamate were about 50% higher than those of aspartate, and GABA levels were about 20-folds smaller than those of the excitatory amino acids. There were no significant differences in the basal levels of any of the three amino acids between the two brain regions. The extracellular levels of aspartate increased during acquisition and recall trials in both VH and mPFC, whereas those of glutamate increased in the VH during acquisition only. A significant increase in GABA levels was also detected during acquisition but only in the mPFC. The neuronal origin of the increased extracellular levels of aspartate, glutamate and GABA was demonstrated by administering tetrodotoxin directly into the mPFC or VH by reverse dialysis. These findings, together with previous evidence from our and other laboratories, indicate a differential release of aspartate and glutamate from excitatory neurons during the performance of behavioral responses, and therefore, distinct roles for the two excitatory amino acids should be envisaged.     

Effect of excitant amino acid antagonists on glutamate receptors in the locust and on convulsions induced by glutamate, aspartate, kynurenine and quinolinic acid in mice

All excitatory amino acid antagonists studied: diethyl esters of aspartic (DEEA) and glutamic (DEEG) acids, 2-amino-3-phosphono-propionic acid (APPA) and 2-amino-4-phosphono-butanoic acid (APBA), diminished the amplitude of excitatory postsynaptic potentials (EPP) of the locust (Locusta migratoria migratorioides) muscle fibers and arbitrary blocked glutamate (GLU) and aspartate (ASP) responses. Kynurenine (KYN) and quinolinic (QUI) acid had no effect on EPP even at a concentration of 2 X 10(-2) M. The antagonists were not strictly selective against intracerebroventricularly administered endogenous convulsants: GLU, ASP, KYN and QUI and in simulation of experimental seizures in mice. The antagonists structurally similar to ASP prevented ASP- and KYN-induced seizures in lower doses than GLU derivatives. Anti-KYN, but not anti-QUI DEEA, DEEG, APPA and APBA efficacy suggests that KYN and QUI act on different structures or binding sites.