Ontogenetic Study of the Effects of Energetic Nutrients on Amino Acid Metabolism of Rat Cerebral Cortex

We studied the effect of various energetic nutrients on metabolism of l-[U-¹⁴C]leucine and [1–¹⁴C]glycine in cerebral cortex of rats at different ages. At gestational age, glucose and lactate stimulated protein synthesis from l-[U-¹⁴C]leucine and [1–¹⁴C]glycine and from l-[U-¹⁴C]leucine, respectively; glucose, -OH-butyrate and lactate stimulated lipid synthesis from l-[U-¹⁴C]leucine. At 10 days of age, glucose, mannose, and fructose stimulated protein synthesis, and glucose and mannose stimulated oxidation to CO₂ as well as lipid synthesis from l-[U-¹⁴C]leucine. In adult rats, glucose, mannose, and fructose stimulated protein synthesis from l-[U-¹⁴C]leucine and [1–¹⁴C]glycine; glutamine also markedly decreased the oxidation of l-[U-¹⁴C]leucine and [1–¹⁴C]glycine in 10–day-old and adult rats.     

Increased γ-Aminobutyric Acid Receptor Function in the Cerebral Cortex of Myoclonic Calves with an Hereditary Deficit in Glycine/Strychnine Receptors

Inherited congenital myoclonus (ICM) of Poll Hereford cattle is a neurological disease in which there are severe alterations in spinal cord glycine-mediated neurotransmission. There is a specific and marked decrease, or defect, in glycine receptors and a significant increase in neuronal (synaptosomal) glycine uptake. Here we have examined the characteristics of the cerebral gamma-aminobutyric acid (GABA) receptor complex, and demonstrate that the malfunction of the spinal cord inhibitory system is accompanied by a change in the major inhibitory system in the cerebral cortex. In synaptic membrane preparations from ICM calves, both high-and low-affinity binding sites for the GABA agonist [3H]muscimol were found (KD = 9.3 +/- 1.5 and 227 +/- 41 nM, respectively), whereas only the high-affinity site was detectable in controls (KD = 14.0 +/- 3.1 nM). The density and affinity of benzodiazepine agonist binding sites labelled by [3H]diazepam were unchanged, but there was an increase in GABA-stimulated benzodiazepine binding. The affinity for t-[3H]butylbicyclo-o-benzoate, a ligand that binds to the GABA-activated chloride channel, was significantly increased in ICM brain membranes (KD = 148 +/- 14 nM) compared with controls (KD = 245 +/- 33 nM). Muscimol-stimulated 36Cl- uptake was 12% greater in microsacs prepared from ICM calf cerebral cortex, and the uptake was more sensitive to block by the GABA antagonist picrotoxin. The results show that the characteristics of the GABA receptor complex in ICM calf cortex differ from those in cortex from unaffected calves, a difference that is particularly apparent for the low-affinity, physiologically relevant GABA receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ischemia-Induced Alterations in Lipid Metabolism of the Gerbil Cerebral Cortex: I. Changes in Free Fatty Acid Liberation

Abstract: Does the impaired lipid metabolism during nonlethal transient ischemia truly recover within a few hours after recirculation? In an attempt to answer this question, we first investigated the time course of the changes in the amount and composition of free fatty acids (FFAs) accumulated during 5-min ischemia and after various postischemic recirculation durations (3 min, 1 h, 24 h, 3 days, and 6 days) in the gerbil cerebral cortex. Then those of FFAs liberated in response to the second 5-min ischemia at various recirculation intervals (3 min, 1 h, 3 days, and 6 days) following the initial one were also measured to evaluate the changes in the cellular response. The former study disclosed that the FFA levels transiently returned to the control levels at 1-h recirculation, increased again a few days after the onset of recirculation, followed by the final return to the control levels after 6-day recirculation. The latter study disclosed that the cellular response to the second ischemia was quite different from that to the initial one even after 6-day recirculation, suggesting that membrane lipid metabolism had not yet been recovered even at such a late period. We discuss the significance of the alterations in lipid metabolism.     

Glutamate and γ-Aminobutyric Acid Neurotransmitter Systems in the Acute Phase of Maple Syrup Urine Disease and Citrullinemia Encephalopathies in Newborn Calves

Cerebral cortex tissue was obtained at autopsy from neonatal Poll Hereford calves with clinically confirmed maple syrup urine disease (MSUD), neonatal Holstein-Friesian calves with clinically confirmed citrullinemia, and matched controls. From this, synaptosomes were prepared for studies of neurotransmitter amino acid uptake and stimulus-induced release, and synaptic plasma membranes were obtained for studies of associated postsynaptic receptor binding sites. As well as having abnormal brain tissue concentrations of the pathognomic plasma amino acids (markedly increased levels of the branched-chain compounds valine, isoleucine, and leucine in MSUD; marked elevation of citrulline levels in citrullinemia), both groups of diseased animals showed reduced brain tissue concentrations of each of the transmitter amino acids glutamate, aspartate, and gamma-aminobutyric acid (GABA). Nontransmitter amino acids were generally unaffected in either disease. Citrullinemic calves showed a marked increase in brain glutamine concentration; in calves with MSUD, the glutamine concentration was raised, but to a much lesser extent. The Na(+)-dependent synaptosomal uptake of both glutamate and GABA was markedly reduced (to less than 50% of control values in both cases) in citrullinemic calves but was unaltered in calves with MSUD. Whereas synaptosomes from normal calves showed the expected stimulus-coupled release of transmitter amino acids, especially glutamate and aspartate, and no response to stimulus of nontransmitter amino acids, there was no increased release of transmitter amino acids in response to depolarization in synaptosomes from citrullinemic calves.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cerebral Cortex Ammonia and Glutamine Metabolism During Liver Insufficiency-Induced Hyperammonemia in the Rat

Hyperammonemia has been suggested to induce enhanced cerebral cortex ammonia uptake, subsequent glutamine synthesis and accumulation, and finally net glutamine release into the blood stream, but this has never been confirmed in liver insufficiency models. Therefore, cerebral cortex ammonia- and glutamine-related metabolism was studied during liver insufficiency-induced hyperammonemia by measuring plasma flow and venous-arterial concentration differences of ammonia and amino acids across the cerebral cortex (enabling estimation of net metabolite exchange), 1 day after portacaval shunting and 2, 4, and 6 h after hepatic artery ligation (or in controls). The intra-organ effects were investigated by measuring cerebral cortex tissue ammonia and amino acids 6 h after liver ischemia induction or in controls. Arterial ammonia and glutamine increased in portacaval-shunted rats versus controls, and further increased during liver ischemia. Cerebral cortex net ammonia uptake, observed in portacaval-shunted rats, increased progressively during liver ischemia, but net glutamine release was only observed after 6 h of liver ischemia. Cerebral cortex tissue glutamine, gamma-aminobutyric acid, most other amino acids, and ammonia levels were increased during liver ischemia. Glutamate was equally decreased in portacaval-shunted and liver-ischemia rats. The observed net cerebral cortex ammonia uptake, cerebral cortex tissue ammonia and glutamine accumulation, and finally glutamine release into the blood suggest that the rat cerebral cortex initially contributes to net ammonia removal from the blood during liver insufficiency-induced hyperammonemia by augmenting tissue glutamine and ammonia pools, and later by net glutamine release into the blood. The changes in cerebral cortex glutamate and gamma-aminobutyric acid could be related to altered ammonia metabolism.     

Amino Acid Release from Cerebral Cortex in Experimental Acute Liver Failure, Studied by In Vivo Cerebral Cortex Microdialysis

Both increased gamma-aminobutyric acid (GABA)-ergic and decreased glutamatergic neurotransmission have been suggested relative to the pathophysiology of hepatic encephalopathy. This proposed disturbance in neurotransmitter balance, however, is based mainly on brain tissue analysis. Because the approach of whole tissue analysis is of limited value with regard to in vivo neurotransmission, we have studied the extracellular concentrations in the cerebral cortex of several neuroactive amino acids by application of the in vivo microdialysis technique. During acute hepatic encephalopathy induced in rats by complete liver ischemia, increased extracellular concentrations of the neuroactive amino acids glutamate, taurine, and glycine were observed, whereas extracellular concentrations of aspartate and GABA were unaltered and glutamine decreased. It is therefore suggested that hepatic encephalopathy is associated with glycine potentiated glutamate neurotoxicity rather than with a shortage of the neurotransmitter glutamate. In addition, increased extracellular concentration of taurine might contribute to the disturbed neurotransmitter balance. The observation of decreasing glutamine concentrations, after an initial increase, points to a possible astrocytic dysfunction involved in the pathophysiology of hepatic encephalopathy.     

Increased Receptor for Advanced Glycation Endproducts Immunocontent in the Cerebral Cortex of Vitamin A-Treated Rats

Vitamin A, beyond its biological role, is an alternative choice in treating some life threatening pathologies, for instance leukemia and immunodeficiency. On the other hand, vitamin A therapy at moderate to high doses has caused concern among public health researchers due to the toxicological aspect resulting from such habit. It has been described hepatotoxicity, cognitive disturbances and increased mortality rates among subjects ingesting increased levels of vitamin A daily. Then, based on the previously reported data, we investigated here receptor for advanced glycation endproducts (RAGE) immunocontent and oxidative damage levels in cerebral cortex of vitamin A-treated rats at clinical doses (1,000–9,000 IU/kg day⁻¹). RAGE immunocontent, as well as oxidative damage levels, were observed increased in cerebral cortex of vitamin A-treated rats. Whether increased RAGE levels exert negative effects during vitamin A supplementation it remains to be investigated, but it is very likely that deleterious consequences may arise from such alteration.     

Noradrenalin-Inducible Cyclic-AMP Accumulation in Rat Cerebral Cortex: Changes During Complete Global Ischemia

Neurologic dysfunction after cerebral ischemic insults may be due not only to neuronal death, but also to a possibly reversible failure in synaptic transmission. Because noradrenaline (NA)-inducible cyclic-AMP (cAMP) accumulation in brain may reflect the integrity of synaptic transmission mechanisms and brain viability, we studied its changes in cerebral cortex after various durations of decapitation ischemia. Unanesthetized rats were decapitated and the brains were kept at 37 degrees C for times ranging from 0 to 60 min. Cerebral cortical slices were incubated in vitro and NA (11.2 microM)-induced cAMP accumulation was evaluated over 10 min. At 0 min of ischemia, NA-induced cAMP accumulation was 56 pmol/mg protein/10 min. Between 0 and 20 min of ischemia, a linear eightfold increase, to 435 +/- 49 pmol/mg protein/10 min, occurred in NA-induced cAMP accumulation, with no further increase after longer durations of ischemia. The mechanisms modulating the increase in cortical NA-inducible cAMP accumulation with a maximum response after 20 min of ischemia remain to be defined.     

On the Status of Lysolecithin in Rat Cerebral Cortex During Ischemia

Abstract: Lysolecithin (lysoglycerophosphocholine, LPC) was isolated from rat cerebral cortex and quantitatively analyzed at various times after postdecapitative ischemic treatment. In addition, different procedures for extraction and analysis of the LPC in brain were evaluated. Results indicated that LPC can be quantitatively extracted into the organic phase using the conventional extraction procedure with chloroform-methanol (2:1, vol/ vol). However, care should be taken to avoid using strong acids, which can hydrolyze the alkenylether side chain of the plasmalogens, resulting in the release of 2-acyl-phospholipids. Quantitative GLC analysis using myris-toyl-LPC as internal standard revealed a level of 1.8 nmol LPC/mg protein in brain with acyl groups comprised mainly of 16:0, 18:0, and 18:1. The acyl group profile reflects that the LPC are derived mainly from phospho-lipase A₂ action. An increase of 46% in the LPC level was observed at 1 min after ischemic treatment, but this was followed by a steady decline. Ischemia induced an increase in the LPC species that are enriched in 18:0 and 18:1 fatty acids. The transient appearance of LPC during ischemia further suggests that this phospholipid is undergoing active turnover, possibly hydrolysis by the lysophospholipase. This mechanism of action may account, at least in part, for the increase in both saturated and unsaturated fatty acids during the early phase of the ischemic treatment.     

The Deduced Amino Acid Sequences of Human Platelet and Frontal Cortex Monoamine Oxidase B Are Identical

Abstract: Monoamine oxidases (MAOs) A and B play important roles in the metabolism of neuroactive, vasoactive amines. Human platelets contain only MAO B, often used as an indicator of brain MAO B. The validity of this model remained to be evaluated. This report describes the molecular cloning of human MAO B from frontal cortex and platelets. Two overlapping PCR-amplified clones of human platelet MAO B and four PCR-amplified clones of human frontal cortex MAO B covering the entire coding region were sequenced using five internal oligomers and M13 reverse and forward primers. The nucleotide sequences of human MAO B cDNA from platelet and frontal cortex were identical to that of human liver MAO B except for three nucleotides that differed in frontal cortex: nucleotides 440 A → G, 794 C → T, and 825 C → T. Whether or not these differences are artifactual, all three represent silent mutations, which would not alter the amino acid of the encoded polypeptides. Thus, the deduced amino acid sequences of MAO B from frontal cortex, platelet, and liver are identical. These findings indicate the validity of using platelet MAO B mRNA as a marker for brain MAO B and provide a new approach to study the role of brain MAO B in humans.     

Changes in Regional Neurotransmitter Amino Acid Levels in Rat Brain During Seizures Induced by l-Allylglycine, Bicuculline, and Kainic Acid

Changes in amino acid concentrations were studied in the cortex, cerebellum, and hippocampus of the rat brain, after 20 min of seizure activity induced by kainic acid, 47 mumol/kg i.v.; L-allylglycine, 2.4 mmol/kg i.v.; or bicuculline, 3.27 mumol/kg i.v. in paralysed, mechanically ventilated animals. Metabolic changes associated with kainic acid seizures predominate in the hippocampus, where there are decreases in aspartate (-26%), glutamate (-45%), taurine (-20%), and glutamine (-32%) concentrations and an increase in gamma-aminobutyric acid (GABA) concentration (+ 26%). L-Allylglycine seizures are associated with generalized decreases in GABA concentrations (-32 to -54%), increases in glutamine concentrations (+10 to +53%), and a decrease in cortical aspartate concentration (-14%). Bicuculline seizures, in fasted rats, are associated with marked increases in the levels of hippocampal GABA (+106%) and taurine (+40%). In the cerebellum, there are increases in glutamine (+50%) and taurine concentrations (+36%). These changes can be explained partially in terms of known biochemical and neurophysiological mechanisms, but uncertainties remain, particularly concerning the cerebellar changes and the effects of kainic acid on dicarboxylic amino acid metabolism.     

Neurotensin Regulation of Endogenous Acetylcholine Release from Rat Cerebral Cortex: Effect of Quinolinic Acid Lesions of the Basal Forebrain

The effects of neurotensin (NT) on endogenous acetylcholine (ACh) release from basal forebrain, frontal cortex, and parietal cortex slices were tested. The results show that NT differentially regulates evoked ACh release from frontal and parietal cortex slices without altering either spontaneous or evoked ACh release from basal forebrain slices. In the frontal cortex, NT significantly inhibited evoked ACh release by a tetrodotoxin (TTX)-insensitive mechanism, suggesting an action directly on cholinergic terminals. In the parietal cortex, NT enhanced evoked ACh release by a TTX-sensitive mechanism, suggesting an action of NT on the cholinergic neuron or in close proximity to the cholinergic neuron. The effects of NT on ACh release were confined to evoked ACh release; that is, spontaneous ACh release was not affected. NT did not affect spontaneous or potassium-evoked ACh release from occipital cortex slices. The second set of experiments tested the effects of quinolinic acid (QUIN) lesions of the basal forebrain cell bodies on the NT-induced regulation of evoked ACh release in the cerebral cortex. QUIN lesions of basal forebrain cell bodies caused decreases in choline acetyltransferase activity (27 and 28%), spontaneous ACh release (14 and 21%), and evoked ACh release (38 and 44%) in frontal and parietal cortex, respectively. In addition, 11 days following QUIN lesions of basal forebrain cell bodies, the action of NT to regulate evoked ACh release in frontal cortex or parietal cortex was no longer observed. The results suggest that in the rat frontal and parietal cortex, NT differentially regulates the activity of cholinergic neurons by decreasing and increasing evoked ACh release, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Rate of Protein Glycosylation in Rat Cerebral Cortex

Quantitative aspects of the pathway leading to the formation of asparagine-linked oligosaccharides were investigated in rat cerebral cortex. Steady-state labeling conditions were achieved with [2-3H]mannose by developing a micromethod of incubation of cerebral cortex particles in the presence of physiological concentrations of glucose (1 g/L). The rate of [2-3H]mannose uptake and incorporation into protein was markedly affected when the concentration of glucose was lowered to 0.05 g/L. It was found that in the presence of glucose (1 g/L), a minor fraction of the utilized [2-3H]mannose is used in glycoprotein formation and the remaining labeled sugar enters the other major metabolic pathways, generating tritiated water which is rapidly exchanged with that of the medium. Under these conditions, the intracellular isotopic dilution of [2-3H]mannose-labeled precursors was calculated to be about 11.5-fold. These data allow determination of the rate of the net transfer of mannose into proteins. Comparison of the rate of glycosylation between 5- and 30-day-old cerebral cortex revealed a striking difference: 2.1 and 0.3 ng of mannose/mg protein/h, respectively.     

蔬菜腌制过程中的氨基酸组成变化

在腌制过程中,芥菜的精氨酸和组氨酸等碱性氨基酸含量和氨基酸总量逐渐减少,酸性氨基酸和疏水性氨基酸含量增加。在各种氨基酸中,苏氨酸含量最高,占４４．７４％,其次是谷氨酸和丙氨酸。在腌制后,芥菜的氨基酸组成发生了显著的变化,苏氨酸下降幅度最大,而蛋氨酸、苯丙氨酸、亮氨酸、异亮氨酸、缬氨酸和赖氨酸等必需氨基酸摩尔组分显著提高。理想氨基酸模式谱的相关系数腌制前为０．０６０１７,腌制２１６小时后增到０．４９９７８,表明腌制加工可显著提高蔬菜蛋白质的营养效价。     

A Global Multiregional Proteomic Map of the Human Cerebral Cortex

The Brodmann area(BA)-based map is one of the most widely used cortical maps for studies of human brain functions and in clinical practice; however, the molecular architecture of BAs remains unknown. The present study provided a global multiregional proteomic map of the human cerebral cortex by analyzing 29 BAs. These 29 BAs were grouped into 6 clusters based on similarities in proteomic patterns: the motor and sensory cluster, vision cluster, auditory and Broca’s area cluster, Wernicke’s area c...     

Intracerebroventricular Administration of Ouabain to Rats Changes the Expression of NMDA Receptor Subunits in Cerebral Cortex and Hippocampus

Ouabain exerts neurotoxic action and activates the population of NMDA receptors. Herein the effect of ouabain on the expression of NMDA subunits was evaluated. Adult Wistar rats were administered intracerebroventricularly with 0.1, 10 and 100 nmol ouabain or saline solution (control). Two days later, membranes of cerebral cortex and hippocampus were isolated. Western blots with antibodies for the NMDA receptor subunits: NR1; NR2A; NR2B; NR2C and NR2D were carried out. In cerebral cortex, NR2D subunit increased 30% with 10 nmol ouabain dose. With 100 nmol ouabain, NR1 and NR2D subunits enhanced 40 and 20%, respectively. In hippocampus, with the dose of 0.1 nmol ouabain, NR1 subunit enhanced roughly 50% whereas NR2B subunit decreased 30%. After administration of 10 nmol ouabain dose, NR2A, NR2B and NR2C subunits decreased 40, 50 and 30%, respectively. With the dose of 100 nmol of ouabain, NR1, NR2A and NR2B subunits diminished 10–20%. It is concluded that ouabain administration led to a differential regulation in the expression of NMDA subunits. These results may be correlated with the modulatory action of ouabain on NMDA receptor.     

新型氨基酸制剂对创伤大鼠血游离氨基酸的影响

观察了富含牛磺酸 (Tau)、谷氨酰胺 (Gln)以及高支链氨基酸 (HBCAA)的新型氨基酸制剂对创伤大鼠血中游离氨基酸浓度的影响。结果表明 ,创伤后三天起 ,血浆游离氨基酸总和均显著降低 ,对照组基本无改变 ;创伤后Tau、BCAA、精氨酸以及天冬氨酸等具有抗氧化和免疫调节作用的氨基酸含量明显降低 ,新处方使用一周后其浓度有效回升 ,且效果好于 17种氨基酸 ,从而有利于机体伤口的愈合。这些结果为进一步阐明复合氨基酸制剂促进创伤愈合的作用及其开发应用提供了理论依据     

富含BCAA 的PN 对肝硬化大鼠肝部分切除术后肝脏NK 的影响

目的:探讨富含支链氨基酸的肠外营养对肝硬化大鼠肝部分切除术后肝脏自然杀伤细胞(NK细胞)的影响。方法:20只肝硬化大鼠随机分为肝部分切除术后行8.5%Novamin的肠外营养5 d组10只,肝部分切除术后行10%Hepa的肠外营养5 d组10只。应用流式细胞仪测定大鼠肝脏NK细胞的百分率;应用4小时51Cr释放法测定肝脏NK细胞的杀伤活性。结果:与8.5%Novamin的肠外营养5 d组比较,10%Hepa的肠外营养5 d组肝脏NK细胞占全部淋巴细胞的百分比和肝脏NK细胞杀伤活性明显升高(P<0.05)。结论:富含支链氨基酸的肠外营养可以增加肝脏NK细胞的百分率和肝脏NK细胞的杀伤活性。     

Modulation of the Electrically Evoked Release of 5-[3H]Hydroxytryptamine from Rat Cerebral Cortex: Effects of Alpidem, CL 218872, and Diazepam

The effect of omega (benzodiazepine)-receptor agonists, antagonists, and inverse agonists on the electrically evoked release of 5-[3H]hydroxytryptamine ([3H]5-HT) was studied in superfused slices of the rat frontal cerebral cortex. The electrically evoked release of [3H]5-HT was enhanced by nanomolar concentrations of diazepam and the selective omega 1-receptor agonists alpidem and CL 218872. The omega 1/omega 2- and omega 1-receptor antagonists flumazenil and CGS 8216, respectively, did not modify the electrically evoked release of [3H]5-HT. The omega 3-receptor agonist Ro 5-4864 and the omega 1-receptor inverse agonist ethyl-beta-carboline-3-carboxylate on their own did not affect the electrically evoked release of [3H]5-HT. On the other hand, the inverse agonist 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylic acid methyl ester (DMCM), at micromolar concentrations, inhibited both the spontaneous and the evoked release of [3H]5-HT. The facilitation of the electrically evoked release of [3H]5-HT by diazepam, alpidem, or CL 218872 was potentiated by gamma-aminobutyric acid (GABA). Exposure to flumazenil and CGS 8216 antagonized the facilitation by diazepam, alpidem, or CL 218872 of [3H]5-HT release. The inhibition of the release of [3H]5-HT by DMCM was not modified by exposure to either flumazenil, CGS 8216, or GABA. The inhibitory effect of DMCM was not observed when monoamine oxidase activity was inhibited by pargyline.(ABSTRACT TRUNCATED AT 250 WORDS)