Significant Increase In Alanine Research Articles

FC091: Changes in the Gut Microbiome and Systemic Metabolome in an In Vivo Model of Peritoneal Dialysis Supplemented with Alanyl-Glutamine

Abstract BACKGROUND AND AIMS Long-term usage of peritoneal dialysis (PD) leads to morphological and functional changes—such as progressive thickening, vascularization and vasculopathy—of the peritoneum, thereby limiting PD efficiency. Vasculopathy induces changes in permeability of the endothelial barrier, which has been suggested to influence the gut microbiome (gMB). Microbial dysbiosis in turn can potentiate inflammation, a risk factor for cardiovascular disease (CVD). The complex interplay of molecules associated with CKD and PD with the gMB is hypothesized to be in causal relationship with occurrences of cardiovascular events and progression of CVD. Novel PD fluids that slow or prevent these processes are in need. The immunomodulatory and cytoprotective compound alanyl-glutamine (AG) has been shown in other model systems to improve the endothelial barrier function, supposedly protecting the gMB from the extraintestinal environment. Our aim is to identify changes of the gMB and the systemic metabolome caused by the addition of AG to PD under conditions of CKD. METHOD Mice (C57/Bl6 NCrl) underwent a 5/6 nephrectomy to induce uraemia. Via a subcutaneously implanted catheter the animals received five times per week a daily injection of 2 mL PD fluid (3.86% glucose with/without 8 mM AG) for six weeks to a total of 30 injections. During the experiment, mice received standard chow and tap water ad libitum. At the end of the experiment, fecal samples from three different intestinal sites and stool were collected and a serum sample was drawn from each mouse. Tissue samples from parietal and visceral peritoneum and relevant organs were preserved for paraffin and cryo sections. Peritoneal effluent cells were spun down and snap-frozen in liquid nitrogen and the supernatant was stored on −80°C. Bacterial ribosomal RNA (rRNA) was isolated from each fecal sample and the V4 region of the 16S rRNA was amplified. Assessment of the plasma metabolome was performed by targeted mass spectrometry (MxP Quant 500, Biocrates, Austria). The experiment was approved by the institutional animal ethics committee. RESULTS Elevated creatinine levels after subtotal nephrectomy confirmed the uremic status of the mice. Descriptive analysis of the gMB showed decreased Shannon diversity, indicating reduced bacterial diversity, in uremic and PD fluid receiving mice over time, whereas AG receiving and control mice retained a stable Shannon diversity. Furthermore, the gMB of mice exposed to PD fluid with AG showed five bacterial classes and 13 bacterial orders to be differently abundant versus mice exposed to PD fluid without AG. In the colon of uremic mice Clostridia was decreased versus control. Serum concentrations of 630 metabolites were assessed by our metabolomics approach. A significant increase in alanine and glutamine serum concentrations was seen in mice exposed to AG supplemented PD fluid. Further, 17 amino acids and four biogenic amines showed to be differently abundant in mice exposed to AG supplemented PD fluid versus without AG. Lastly, plasma of uremic mice showed a significant increase of the toxic amino acid symmetric dimethyl arginine (SDMA) and citrulline. CONCLUSION Exposure to PD fluid and having chronic kidney disease both affect the gut microbiome and systemic metabolome. Supplementation of AG to PD fluid showed changes at overall microbiome diversity and serum metabolites. Further analysis of these changes in the context of tissue alterations will clarify functional differences of the cytoprotective additive AG. Preservation of the mesothelial and endothelial barrier function may represent a mechanistic explanation for the observed improvement in protein loss and systemic inflammation in a recent phase II clinical trial in PD patients. Better understanding of the gut-peritoneal barrier may open up new therapeutic targets for improving clinical outcome of PD treatment and its long-term efficacy.

Metabolomic Variation Aligns with Two Geographically Distinct Subpopulations of Brachypodium Distachyon before and after Drought Stress.

Brachypodium distachyon (Brachypodium) is a non-domesticated model grass that has been used to assess population level genomic variation. We have previously established a collection of 55 Brachypodium accessions that were sampled to reflect five different climatic regions of Turkey; designated 1a, 1c, 2, 3 and 4. Genomic and methylomic variation differentiated the collection into two subpopulations designated as coastal and central (respectively from regions 1a, 1c and the other from 2, 3 and 4) which were linked to environmental variables such as relative precipitation. Here, we assessed how far genomic variation would be reflected in the metabolomes and if this could be linked to an adaptive trait. Metabolites were extracted from eight-week-old seedlings from each accession and assessed using flow infusion high-resolution mass spectrometry (FIE-HRMS). Principal Component Analysis (PCA) of the derived metabolomes differentiated between samples from coastal and central subpopulations. The major sources of variation between seedling from the coastal and central subpopulations were identified. The central subpopulation was typified by significant increases in alanine, aspartate and glutamate metabolism and the tricarboxylic acid (TCA) cycle. Coastal subpopulation exhibited elevated levels of the auxin, indolacetic acid and rhamnose. The metabolomes of the seedling were also determined following the imposition of drought stress for seven days. The central subpopulation exhibited a metabolomic shift in response to drought, but no significant changes were seen in the coastal one. The drought responses in the central subpopulation were typified by changes in amino acids, increasing the glutamine that could be functioning as a stress signal. There were also changes in sugars that were likely to be an osmotic counter to drought, and changes in bioenergetic metabolism. These data indicate that genomic variation in our Turkish Brachypodium collection is largely reflected as distinctive metabolomes (“metabolotypes”) through which drought tolerance might be mediated.

Antioxidant defenses and metabolic responses of blue mussels (Mytilus edulis) exposed to various concentrations of erythromycin

Erythromycin, one of the most widely used macrolide antibiotics, has been detected in various aquatic environments, so erythromycin ecotoxicity should deserve more attention. In this study, blue mussels (Mytilus edulis) were exposed to erythromycin to explore its potential physiological toxicity. After 2d acute and 7d sub-acute exposure to erythromycin, blue mussel glutathione S-transferase (GST) and catalase (CAT) activities were determined with microplate methods and metabolic responses were analyzed using 1H nuclear magnetic resonance (1H NMR). The results revealed that GST was approximately 1.6 times higher in exposed mussels at 200 mg/L and higher concentrations. CAT was about 1.9 times higher in exposed mussels at 200 mg/L, indicating that erythromycin exposure led that blue mussels enhanced antioxidant responses. Low doses of erythromycin exposure had a relatively small impact on the metabolism, while high doses of erythromycin exposure (200 and 400 mg/L) disturbed metabolic balance. With the increase of erythromycin concentrations, the individual metabolic differences within the same treatment groups also increased. The significant increase in alanine, glutamate, taurine, glycine and betaine were observed after acute and subacute exposure. Betaine played an important role in protecting antioxidant enzyme activities through adjusting osmotic pressure. The metabolomic results also showed the modes of erythromycin acted on the energy metabolism, osmoregulation, nerve activities and amino acid metabolism. This study highlighted how metabolomics can provide a comprehensive picture of metabolic responses, although significant antioxidant and metabolic responses were observed at high exposure concentrations.

Abstract 5258: 1 H MRS analysis of pancreas metabolites altered by cachexia

Abstract Cachexia is a multifactorial syndrome characterized by skeletal muscle weight loss and reduced physical activity. The extreme weight loss due to cachexia results in a particularly poor quality of life causing profound weakness, listlessness, and inability to function [1, 2]. Severe weight loss decreases the tolerance to treatments and anticancer therapies, and leads to the reduced survival of patients. In pancreatic cancer, the syndrome affects nearly 80% of patients [3]. Cancers can stimulate cachexia through the dysfunction of multiple organs [4]. Metabolic abnormalities may interfere with the regular functioning of these organs to increase the severity of the disease. Here we determined metabolic changes in the pancreas with cachexia-inducing and non-cachexia inducing tumor growth using high-resolution quantitative 1H magnetic resonance spectroscopy (MRS) of pancreas tissue obtained from normal, non-cachectic (Panc1) and cachectic (Pa04C) mice bearing pancreatic ductal adenocarcinoma (PDAC) xenografts. Cancer cells were inoculated in the right flank of six to eight week old male severe combined immunodeficient mice. The pancreas was removed from mice following euthanization once tumors were ~300 mm3, snap frozen and stored at -80°C prior to dual phase extraction. 1H MRS was performed on the water phase. All 1H MR spectra with water suppression were acquired on a 750 MHz MR spectrometer using a single pulse sequence. All data processing analyses and quantification were performed with TOPSPIN 3.5 software. All statistical analysis were performed with MetaboAnalyst software [5]. Multivariate analyses performed to analyze the differences in the metabolic profiles among the groups (Control n = 9, Pa04C = 10 and Panc1 = 8) revealed differences in the overall metabolic pattern in the pancreas from normal, cachectic and non-cachectic groups. A significant decrease of leucine, isoleucine, valine, BCAA, glutamate, choline, pyruvate, glycine, niacinamide, fumarate and increase of alanine, pyruvate, and NAD was detected in cachectic mouse pancreas compared to control pancreas. A significant increase in alanine, phenylalanine, pyruvate, phosphocholine and decrease in glycine, NAD, niacinamide was observed in cachectic mouse pancreas compared to non-cachectic mouse pancreas. Pathway impact analysis using MetaboAnalyst web software indicated alterations in branch chain amino acid pathways and glutamate, glutamine metabolism. These results provide new insights into changes in pancreas metabolism with cachexia, and support investigating metabolic targets and biomarkers to reduce cachexia-associated morbidity. Supported by NIH R01CA193365 and R35CA209960. Ref: 1. Fearon K et al, The Lancet Oncology. 2011; 2. Penet MF, Bhujwalla ZM. Cancer journal. 2015; 3. Winnard PT, Jr. et al, Cancer research. 2016; 4. Argiles JM et al, Nature reviews Cancer. 2014; 5. Xia J, Wishart DS. Nature protocols. 2011. Citation Format: Raj Kumar Sharma, Santosh K. Bharti, Paul T. Winnard Jr, Yelena Mironchik, Marie-France Penet, Zaver M. Bhujwalla. 1 H MRS analysis of pancreas metabolites altered by cachexia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5258.

THE EFFECT OF SILICON DIOXIDE NANOPARTICLES ON THE LEAD HEPATOTOXICITY

Nanoparticles are known to facilitate transport of chemicals and medications through membrane barriers into cells. This results in the raising of toxic effect when two contaminants act on the body at the same time. The aim of this study was to determine how silicon dioxide (SiO2) nanoparticles affect the hepatotoxic properties of lead. Alanine and aspartate transaminases, alkaline phosphatase activities, as well as total protein, total bilirubin, creatinine and urea contents were measured in rat blood plasma. It was shown that SiO2 nanoparticles did not significantly affect the above listed markers. Administration of lead acetate to the animals caused cytolysis of hepatocytes, as evidenced by significant increase of alanine and aspartate transaminases, alkaline phosphatase activities, along with significant decrease of total plasma protein and increase of total bilirubin levels. The increase of aminotransferases and alkaline phosphatase activities in animals that received both SiO2 nanoparticles and lead acetate was significant comparedto the animals that received only lead acetate. This suggests that SiO2 nanoparticles, which are normally present inthe environment and food products as contaminants, can enhance the negative hepatotoxic effects of lead.

Impact of microbial communities on floral nectar chemistry: Potential implications for biological control of pest insects

Nectar-producing plants are increasingly used in agroecosystems to provide biological control agents (BCAs) such as predatory insects and parasitoids with necessary sugars to fulfil their nutritional requirements. However, it has recently been shown that nectar is commonly infested by microorganisms which may change the chemistry of the nectar and therefore its attractiveness and nutritional suitability for insects. The aim of this study was to investigate the impact of nectar-inhabiting microorganisms on nectar chemistry of three co-flowering plant species that are commonly used to provide BCAs with sugar resources (Borago officinalis L., Centaurea cyanus L. and Symphytum officinale L.). First, we assessed the abundance and visitation rate of flower-visiting insects on these plant species. Further, we identified the culturable microbes inhabiting the floral nectar and assessed changes in pH, total sugar concentration, main nectar sugars and amino acids. Flowers of all three plant species were found to be visited by a wide range of insects, with B. officinalis being the most attractive plant species for BCAs. Nectar microbial community structure differed significantly between plant species, irrespective of experimental field site. Microbial contamination affected both sugar and amino acid concentration and composition. Especially, sucrose and the fructan sugars 1-kestose and neokestose were found at significantly lower concentrations in contaminated nectar. Further, microbial contamination resulted in a significant increase of alanine and glycine, and a decrease in threonine and valine. Further research is needed to investigate the precise impact of these changes on the efficacy of biological control agents to control pest insects.

D-Galactosamine Intoxication in Experimental Animals: Is it Only an Experimental Model of Acute Liver Failure?

BackgroundShort-term administration of Galactosamine to experimental animals causes liver damage and acute liver failure (ALF), as well as acute renal failure in some cases. The aim of our study was to describe kidney disorders that developed in the course of galactosamine-induced liver failure.Material/MethodsSprague-Dawley rats were randomly divided into 2 groups: a study group administered galactosamine intraperitoneally and a control group administered saline.ResultsAll the animals in the study group developed liver damage and failure within 48 h, with significant increase of alanine (p<0.001), aspartate aminotransferases (p<0.0001), bilirubin (p<0.004), and ammonia (p<0.005) and decrease of albumin (p<0.001) concentrations. Acute renal failure was observed in all test animals, with a significant increase in creatinine (p<0.001) and urea (p<0.001) concentrations and a decrease in creatinine clearance (p<0.0012). Moreover, osmotic clearance (p<0.001), daily natriuresis (p<0.003), and fractional sodium excretion (p<0.016) decreased significantly in this group of animals. The ratio of urine osmolality to serum osmolality did not change. Histopathology of the liver revealed massive necrosis of hepatocytes, whereas renal histopathology showed no changes.ConclusionsAcute renal failure that developed in the course of galactosamine-induced ALF was of a functional nature, with the kidneys retaining the ability to concentrate urine and retain sodium, and there were no renal changes in the histopathological examination. It seems that the experimental model of ALF induced by galactosamine can be viewed as a model of hepatorenal syndrome that occurs in the course of acute damage and liver failure.

Differential toxicological effects induced by mercury in gills from three pedigrees of Manila clam Ruditapes philippinarum by NMR-based metabolomics

Mercury is a hazardous pollutant in the Bohai marine environments due to its high toxicity to the marine organisms and subsequent ecological risk. Manila clam Ruditapes philippinarum is one of important sentinel organisms in 'Mussel Watch Program' launched in China and therefore used as a bioindicator in marine and coastal ecotoxicology. There are dominantly distributed three pedigrees of clam (White, Liangdao Red and Zebra) in Yantai population endowed with different tolerances to environmental stressors. In this study, gill tissues were collected from both untreated and mercury exposed White, Liangdao Red and Zebra clams, and the extracts were analyzed by NMR-based metabolomics to compare the original metabolomes and the toxicological effects induced by mercury exposure in three pedigrees. The major abundant metabolites in White clam sample were branched-chain amino acids, lactate, alanine, arginine, acetoacetate, glutamate, succinate, citrate, malonate and taurine, while the metabolite profile of Liangdao Red clam sample comprises relative high levels of alanine, arginine, glutamate, succinate and glycogen. For Zebra clam sample, the metabolite profile exhibited relatively high amount of aspartate, acetylcholine and homarine. After 48 h exposure of 20 μg l(-1) Hg(2+), the metabolic profiles from all the three pedigrees of clams commonly showed significant increases in alanine, arginine, glutamate, aspartate, α-ketoglutarate, glycine and ATP/ADP, and decreases in citrate, taurine and homarine. The unique metabolic differences between the metabolomes of gill tissues from Hg(2+)-exposed White, Liangdao Red and Zebra clams were found, including elevated acetylcholine and branched-chain amino acids in White clams, and the declined succinate in both White and Liangdao Red samples as well as the declined betaine in Zebra and White clams. Overall, our findings showed the differential toxicological responses to mercury exposure and that White clams could be a preferable bioindicator for the metal pollution monitoring based on the metabolic changes from gill compared with other two (Liangdao Red and Zebra) pedigrees of clams.

DETERMINATION OF ESTIVATION-INDUCED CHANGES IN THE AMINO ACID CONTENT OF BIOMPHALARIA GLABRATA SNAILS BY HIGH PERFORMANCE THIN-LAYER CHROMATOGRAPHY-DENSITOMETRY

High performance thin-layer chromatography-densitometry (HPTLC-Dens) was used to determine changes in the amino acid content of the digestive-gland gonad complex (DGG) of Biomphalaria glabrata snails as a function of estivation. Amino acids were extracted in ethanol-water (70:30) from the DGG of B. glabrata snails estivated for 7 days, and determined on silica gel or cellulose layers developed with either 2-butanol-pyridine-glacial acetic acid-deionized water (39:34:10:26) or 2-butanol-pyridine-25% ammonia-deionized water (39:34:10:26). Separated zones were detected by postchromatographic derivitization with ninhydrin and quantified by visible mode slit-scanning densitometry at 610 nm. Alanine, arginine, glycine, leucine/isoleucine, lysine, serine, and valine were unequivocally identified in chromatograms from both estivated and unestivated snails. HPTLC-Dens showed a significant increase in alanine in estivated snails compared to unestivated controls (Student's t-test, P < 0.05). The finding of increased alanine levels in the DGG during estivation is in agreement with the hypothesis that anaerobic pathways are an important part of overall metabolism in B. glabrata during estivation.

Ecophysiological and Metabolic Adaptations to Sulphide Exposure of the Oyster Crassostrea gigas

The purpose of this study is to examine the response of the oyster Crassostrea gigas to sulphide at different biological scales. A first experiment was designed to measure for 20 h the clearance (CR) and oxygen consumption (OC) rates of oysters exposed at different concentrations of sulphide (0, 3, 6, 12, 20, and 40 mM). The second experiment was carried out to evaluate the metabolic adaptations to chronic sulphide exposure for 10 days by measuring PK and PEPCK enzyme activities, succinate and alanine content, the adenylate energy charge (AEC), and the activity of the electron transport system (ETS) in whole oysters at above 20 mM. PK and PEPCK mRNAs and enzyme activities have been measured in adductor muscle. When exposed to sulphide above 20 mM, CR stopped, whereas oyster maintained their average OC rate between 1 and 2 mg O2 h -1 g -1 dw. In the second experiment, sulphide exposure of oysters resulted in higher mortality, reduced the glycolytic flux by inhibiting the PK activity and decreased the ETS activity leading to a lower AEC. At day 10, a kind of recovery took place: the ETS activity and AEC of oysters exposed to sulphide increased to the level of the control oysters. In the adductor muscle, sulphide did not affect the transcriptional level of PK; PK was only regulated at enzyme level. Conversely, PEPCK was regulated only at transcriptional level. Nevertheless, muscle exhibited a wholly anaerobic metabolism by the significant increase of alanine and succinate contents followed by the significantly decrease of the ATP content during the 10 days of the experiment. This experiment showed that sulphide involved partially the whole oyster and wholly the adductor muscle in an anaerobic pathway of energy production; but oxygen was needed for the detoxification of the organism.

Region-dependent alterations in glutamate and GABA measured by high-resolution magnetic resonance spectroscopy following acute binge inhalation of toluene in juvenile rats

Little is known about the neurochemical effects accompanying the high-concentration inhalant exposures characteristic of binge solvent abuse. In adult animals, prior studies with other patterns of exposure indicate that toluene, a commonly abused household and industrial solvent, has significant effects on the glutamatergic and GABAergic neurotransmitter systems and on other neurotransmitter systems as well. In the current investigation, high-resolution “magic angle” spinning proton magnetic resonance spectroscopy (HR-MAS 1H-MRS) was used to assess the effect of acute binge toluene inhalation on regional brain concentrations of various neurochemicals including glutamate (GLU), GABA, and glutamine (GLN) in juvenile male and female rats. Acute toluene (8000 ppm or 12,000 ppm) significantly reduced levels of hippocampal GABA (− 12%) and GLU (− 8%), and the GLU/GLN ratio, an index of glutamatergic tone, was significantly reduced (− 22%) in the dorsal anterior striatum, driven largely by a 28% increase in GLN. Significant increases in alanine and lactate in several brain regions after acute toluene may be indicative of altered oxygen-dependent metabolism associated with the inhalation of higher concentrations of toluene ( e.g., > 5000 ppm). Other components of the MR-visible neurochemical profile, such as N-acetylaspartate (NAA), myo-inositol, creatine, and various choline containing compounds, were unchanged by acute toluene. The results are consistent with the notion that binge toluene exposure affects juvenile neurochemistry in systems mediating the rewarding and emotional aspects of substance abuse. Moreover the results provide a framework to understand further 1H-MRS studies in clinical populations.

Cerebral amino acid changes in an animal model of intrauterine growth retardation

As part of a series of experiments to ascertain the effects of prenatal malnutrition on brain development, we measured brain amino acids in an animal model of intrauterine growth retardation (IUGR) obtained by restriction of blood supply to the fetus in utero during the last 5 days of gestation. In the present study, amino acids were measured during development by HPLC as their O-phthaldialdehyde derivatives in cerebral cortex, cerebellum and hippocampus. In rats with IUGR, significant increase of alanine (by 20% to 50%) and taurine (by 20% to 80%) were observed prior to weaning in the cerebellum and the cerebral cortex respectively. Alanine levels were also increased in hippocampus. In control animals, at birth, activities of the GABA nerve terminal marker enzyme glutamic acid decarboxylase (GAD) were found to be 32%, 17%, and 11% of adult values in cerebellum, hippocampus and cerebral cortex respectively. Two-day-old rats with IUGR had significantly lower GAD activities in all brain regions. Thus, impairment of nutrient supply to fetal brain results in selective regional abnormalities of amino acids particularly in the cerebral cortex.

A biochemical and NMR spectroscopic study of hydrazine in the isolated rat hepatocyte.

Using isolated rat hepatocytes the biochemical effects of hydrazine have been investigated using both conventional assay techniques and high resolution proton NMR. High resolution proton NMR revealed that hydrazine caused a significant increase in alanine and lactate levels in the incubation buffer, whereas levels of beta-hydroxybutyrate were decreased. NMR also detected metabolites of hydrazine notably acetylhydrazine and a cyclised hydrazone formed with alpha-ketoglutarate. Changes were detected in NADH and NADPH, ATP, succinate dehydrogenase (SDH) and total non-protein sulphydryl groups (TNPSH). However, the changes in pyridine nucleotides occurred at higher concentrations than those affecting succinate dehydrogenase and ATP. Similarly, the depletion of TNPSH occurred at a higher concentration and with a different time course to that seen with ATP depletion and inhibition of succinate dehydrogenase.

The relationship between the cardiac contractile function, adenine nucleotides and amino acids of cardiac tissue and mitochondria at acute respiratory hypoxia.

The effect of asphyxia and subsequent resumption of respiration on the content of adenine nucleotides and some amino acids in heart tissue and mitochondria, as well as respiration of heart mitochondria was studied in rats. The depression of cardiac contractile function during asphyxia showed a better correlation with losses in mitochondrial adenine nucleotides (ATP + ADP + AMP) than those in cardiac tissue. The decrease in the heart work index was accompanied by a decrease in state 3 respiration with glutamate and malate as well as uncoupled respiration with these substrates. This did not occur with succinate. Nonphosphorylating (state 4) respiratory rates and ADP/O ratios were slightly affected by asphyxia, when respiratory substrates of both types were used. The decreased level of glutamic acid in the tissue and mitochondria of asphyxic hearts was simultaneously observed with a significant increase of alanine in cardiac tissue and of aspartic acid in the mitochondria. The losses of intramitochondrial ATP and respiratory activity with NAD-dependent substrates during asphyxia were associated with a reduction of glutamic acid level in mitochondria. The recovery of cardiac function during resumption of respiration was related to the restoration of mitochondrial respiration supported by glutamate and malate, as well as to the restoration of mitochondrial adenine nucleotides and glutamic acid. The results suggest that the depression of cardiac function caused by acute respiratory hypoxia may be attributed to impairment of electron transport, particularly in complex I of the respiratory chain and changes in metabolism of glutamic acid.

Influence of nutritional status on exertion-induced forearm amino acid metabolism in normal man.

Normal volunteers were evaluated in the postabsorptive state, following 10 days of protein-calorie starvation, and during intravenous feeding ( ivf ) to determine the impact of nutritional status upon exertion-induced muscle amino acid metabolism. An isolated forearm model allowed an evaluation of recovery of metabolism following 1 min of submaximal isotonic exercise. Forearm blood flow returned to near basal levels within 15 min after exertion during postabsorptive and ivf conditions, but remained greater than or equal to 150% of basal at 1 hr after exercise during starvation. At 30 and 60 min after exercise, forearm plasma flux of total and essential amino acids were unchanged from basal in the postabsorptive state. However, the pattern of essential amino acid flux demonstrated a relative reduction in isoleucine and leucine efflux compared with basal, and this pattern persisted throughout 1 hr of recovery. During starvation, a significant (P less than 0.05) increase in total and essential amino acid efflux was observed throughout the recovery period. Starvation was also associated with significant increases in alanine and lysine efflux during recovery. Intravenous feeding was associated with a significant (P less than 0.05) uptake of essential amino acids with respect to basal levels at 30 min after exercise. At 60 min, there was a shift to total amino acid efflux but no change from basal flux for essential amino acids. During ivf , the pattern of essential amino acid uptake returned to basal within 1 hr after exertion.

Chronic metabolic effects of ammonia in mouse brain.

Chronic ammonia toxicity in experimental mice was induced by exposing them for 2 and 5 days to 5 % (v/v) ammonia solution. The enzymes concerned with glutamate metabolism (aspartate-, alanine- and tyrosine aminotransferases, glutamate dehydrogenase and glutamine synthetase) and (Na+ + K+)-ATPase were estimated in the three regions of brain (cerebellum, cerebral cortex and brain stem) and in liver. Glutamate, aspartate, alanine, glutamine and GABA, RNA and protein were also estimated in the three regions of brain and liver. A significant rise in the activity of (Na+ + K+)-ATPase in all the three regions of brain along with a fall in the activity of alanine aminotransferase was noticed. Changes in the activities of other enzymes were also observed. A significant increase in alanine and a decrease in glutamic acid was observed while no change was observed in the content of other amino acids belonging to the glutamate family. As a result of this, changes in the ratios of glutamate/glutamine and glutamate + aspartate/GABA was observed. The results indicated that the brain was in a state of more depression and less of excitation. Under these conditions the liver tissue was showing a profound rise in the activity of the enzymes of glutamate metabolism. The results are further discussed.

Cerebral metabolism in hypoxic hypoxia. II. Citric acid cycle intermediates and associated amino acids

In order to study the influence of hypoxia upon the metabolism of citric acid cycle intermediates and associated amino acids in the cerebral cortex, lightly anesthetized rats were subjected to a lowering of arterial Po 2 to about 25 mm Hg, and brains were frozen in situ for metabolite analyses either 1, 2, 5, 15 or 30 min following the induction of hypoxia. The size of the citric acid cycle pool as calculated from the concentrations of citrate, isocitrate, α-ketoglutarate, succinate, fumarate, malate and oxaloacetate increased gradually during the course of hypoxia. It is concluded that the main anaplerotic reactions were CO 2 fixation at the pyruvate carboxylase step and a shift in the alanine aminotransferase reaction. There was a decrease in α-ketoglutarate at 1 min, in all probability due to a primary change in malate-oxaloacetate ratio and a secondary shift in the aspartate aminotransferase reaction. Later, α-ketoglutarate increased in parallel with the generalized increase in pool size and with the rise in alanine concentration. The cytoplasmic NADH-NAD + ratio, as calculated from intracellular pH i and malate-oxaloacetate atio, increased by about 50% at 1 min and remained constant during the course of hypoxia. The results suggest that the lactate dehydrogenase reaction was off equilibrium when glycolytic flux was maximally increased. Aspartate decreased already at 1 min, a significant increase in alanine occurred at 15 min, and elevated levels of GABA and glutamine were observed at 30 min. The increase in GABA was unrelated to loss of α-ketoglutarate and may (like the rise in glutamine concentration) have resulted from enhanced amination of α-ketoglutarate, possibly combined with a relative loss of NAD +.

Cerebral amino acids and lipids in drug-induced status epilepticus.

Abstract—Dogs were given repeated doses of pentylenetetrazol to maintain a condition of status epilepticus for periods of 30–40 min. Analyses of cerebral tissue frozen in situ showed significant increases in alanine, arginine, γ‐aminobutyric acid, glycine, histidine, leucine, lysine, phenylalanine, serine, tyrosine, and valine. Decreases occurred in the glutamic and aspartic acid levels. Other measured amino compounds were unchanged. Ammonia was increased, but not more than occurs in seizures of brief duration. A large decrease was noted in the ganglioside fraction, and a decrease also in a fraction containing the lecithins and sphingomyelins. The cephalin, cerebroside–sulphatide, and cholesterol fractions were not affected.Similar repetitive seizures induced by bemegride and continuing for shorter periods (5–9 min) brought about less extensive changes. Glutamic acid was increased, in contrast to the pentylenetetrazol effect, and the apparent decrease in aspartic acid was not statistically significant. Increases were noted in alanine, γ‐aminobutyric acid, lysine, and ammonia. The ganglioside and lecithin‐sphingomyelin fractions were decreased.

Free Amino Acids and Related Compounds in Brain and Other Tissues: Effects of Convulsant Drugs

Publisher Summary This chapter summarizes a series of studies on free amino acids and related compounds in the brain. Cerebral tissue was analyzed by ion-exchange chromatography and other methods, and the technique of freezing the brain in situ with liquid air was used to avoid post-mortem increases that otherwise occur in alanine, ammonia, γ-aminobutyric acid and lactic acid. Dogs were used for most of the experiments, but supplemental studies on the effects of fluoro compounds in mice are included in the chapter. Anoxia induced by the administration of 4.5% oxygen in nitrogen for 12–13 min brought about significant increase in alanine, y-aminobutyric acid, glutamic acid, lactic acid, leucine, and tyrosine and decrease in aspartic acid and a fraction consisting of methionine and cystathionine. Ammonia was not significantly increased. Infusion of buffered ammonium chloride for a 10 min or 45 min period generally caused some degree of depression of cortical electrical activity, although in one animal a seizure developed. Amino-oxyacetic acid, which is known to antagonize the convulsant actions of thiosemicarbazide and methionine sulfoximine, was found to induce increase in brain γ-aminobutyrate, alanine, ammonia, glutamine, lactate, lysine, and tyrosine and a decrease in aspartate. Methionine sulfoximine was found to have extensive effects on the nitrogenous metabolism of the brain, causing increases in alanine, ammonia, lysine, phosphoethanolamine, and serine and decrease in aspartate, glutamate, glutamine, leucine, and the methionine and cystathionine fraction.