Amino Acids In Brain Research Articles

Biochemical, Metabolic, and Behavioral Characteristics of Immature Chronic Hyperphenylalanemic Rats.

Phenylketonuria and hyperphenylalanemia are inborn errors in metabolism of phenylalanine arising from defects in steps to convert phenylalanine to tyrosine. Phe accumulation causes severe mental retardation that can be prevented by timely identification of affected individuals and their placement on a Phe-restricted diet. In spite of many studies in patients and animal models, the basis for acquisition of mental retardation during the critical period of brain development is not adequately understood. All animal models for human disease have advantages and limitations, and characteristics common to different models are most likely to correspond to the disorder. This study established similar levels of Phe exposure in developing rats between 3 and 16 days of age using three models to produce chronic hyperphenylalanemia, and identified changes in brain amino acid levels common to all models that persist for ~16 h of each day. In a representative model, local rates of glucose utilization (CMRglc) were determined at 25-27 days of age, and only selective changes that appeared to depend on Phe exposure were observed. CMRglc was reduced in frontal cortex and thalamus and increased in hippocampus and globus pallidus. Behavioral testing to evaluate neuromuscular competence revealed poor performance in chronically-hyperphenylalanemic rats that persisted for at least 3 weeks after cessation of Phe injections and did not occur with mild or acute hyperphenylalanemia. Thus, the abnormal amino acid environment, including hyperglycinemia, in developing rat brain is associated with selective regional changes in glucose utilization and behavioral abnormalities that are not readily reversed after they are acquired.

Changes in free amino acid and monoamine concentrations in the chick brain associated with feeding behavior.

Domesticated chicks are precocial and therefore have relatively well-developed feeding behavior. The role of hypothalamic neuropeptides in food-intake regulation in chicks has been reported for decades. However, we hypothesized that nutrients and their metabolites in the brain may be involved in food intake in chicks because these animals exhibit a very frequent feeding pattern. Therefore, the purpose of this study was to examine the feeding behavior of chicks as well as the associated changes in free amino acid and monoamine concentrations in the chick brain. The feeding behavior of chicks was recorded continuously for 6 h. The next day, brain and blood samples were collected when the chicks either attempted to have food (hungry group) or turned food down (satiated group), in order to analyze the concentrations of the free amino acids and monoamines. We confirmed that the feeding behavior of neonatal chicks was characterized by short resting periods between very brief times spent on food intake. Several free amino acids in the mesencephalon were significantly lower in the satiated group than in the hungry group, while l-histidine and l-glutamine were significantly higher. Notably, there was no change in the free amino acid concentrations in other brain regions or plasma. As for monoamines, serotonin and norepinephrine were significantly lower in the mesencephalon of the hungry group compared with the satiated group, but 5 hydroxyindolacetic acid (5-HIAA) was higher. In addition, serotonin and norepinephrine levels were significantly higher in the brain stem of the hungry chicks compared with the satiated group, but levels of 5-HIAA and homovanillic acid were lower. Levels of both dopamine and its metabolite, 3,4-dihydroxyphenylacetic acid, were significantly higher in the diencephalon and telencephalon of the chicks in the hungry group. In conclusion, the changes in the free amino acids and monoamines in the brain may have some role in the feeding behavior of neonatal chicks.

Voluntary Exercise Prevents Oxidative Stress in the Brain of Phenylketonuria Mice.

High phenylalanine levels in phenylketonuria (PKU) have been associated with brain oxidative stress and amino acid imbalance. Exercise has been shown to improve brain function in hyperphenylalaninemia and neurodegenerative diseases. This study aimed to verify the effects of exercise on coordination and balance, plasma and brain amino acid levels, and brain oxidative stress markers in PKU mice. Twenty wild-type (WT) and 20 PAH(enu2) (PKU) C57BL/6 mice were placed in cages with (exercise, Exe) or without (sedentary, Sed) running wheels during 53 days. At day 43, a balance beam test was performed. Plasma and brain were collected for analyses of amino acid levels and the oxidative stress parameters superoxide dismutase (SOD) activity, sulfhydryl and reduced glutathione (GSH) contents, total radical-trapping antioxidant potential (TRAP), and total antioxidant reactivity (TAR). SedPKU showed poor coordination (p < 0.001) and balance (p < 0.001), higher plasma and brain phenylalanine (p < 0.001), and increased brain oxidative stress (p < 0.05) in comparison to SedWT. ExePKU animals ran less than ExeWT (p = 0.018). Although no improvement was seen in motor coordination and balance, exercise in PKU restored SOD, sulfhydryl content, and TRAP levels to controls. TAR levels were increased in ExePKU in comparison to SedPKU (p = 0.012). Exercise decreased plasma and brain glucogenic amino acids in ExePKU, but did not change plasma and brain phenylalanine in both WT and PKU. Exercise prevents oxidative stress in the brain of PKU mice without modifying phenylalanine levels. Hence, exercise positively affects the brain, demonstrating its value as an intervention to improve brain quality in PKU.

Review Of The Main Welfare Risks Related To Electrical Stunning Of Small Ruminants (Ovine And Caprine Species)

EFSA commissioned a comprehensive review of the welfare aspects of electrical stunning methods for small ruminants with an emphasize on low ampere stunning to establish the state of the art in the field and to assess whether scientific studies would address criteria outlined in an EFSA guidance on the assessment criteria for studies evaluating the effectiveness of stunning interventions regarding animal protection at the time of killing (EFSA Journal 2013;11(12):3486). The review was not formulated as a systematic review with a focused question instead the review followed the approach to assessing the literature described by the EFSA guidance. The key databases searched were: Science Citation Index (1900-2014), CAB Abstracts (1910-2014) and Medline (1990-2014). Key conferences proceedings and the bibliographies of review articles were manually searched. The search yielded 1599 records. 706 duplicate records were removed and 894 records assessed for relevance. Relevant studies reported electronic stunning of small ruminants and outcomes associated with onset and duration of unconsciousness. Eighteen papers reported electrical approaches to stunning in sheep. No goats were studied. None of the papers reported all of the parameters detailed in the EFSA guidance (EFSA, 2013) and a risk of bias assessment was not conducted. No studies reported the appearance of the electrodes. When the frequency (Hz) applied to the animal was reported, it was not specified whether this represented a minimum or maximum frequency. Only one study explicitly reported an effect size for amperes. The study suggested that the odds of a poor stun were higher for amperes of 0.6 (odds ratio (OD) of 6.27 with 95% confidence interval (CI) of 1.98-20.7) and 0.8 (OR of 24.4 with 95% CI of 6.98-85.2) when compared to a poor stun at 1.25 ampere. © European Food Safety Authority, 2015

Alterations in brain amino acid metabolism and inhibitory effects on PKC are possibly correlated with anticonvulsant effects of the isomeric mixture of α- and β-amyrin from Protium heptaphyllum

Context: α- and β-Amyrin (AMY) from Protium heptaphyllum (Aubl) March (Burseraceae) is found in Brazil and used in diverse inflammation-related diseases. This species presents a central action, as previously described.Objective: The objectives were to evaluate the anticonvulsant effect of AMY in mice and to verify the mechanism of action.Material and methods: Seizures were induced by pentylenetetrazole followed by acute or subchronic treatments (5–25 mg/kg, p.o. and i.p.) and determination of brain amino acids (10 and 25 mg/kg, i.p., 7 d).Results: In the acute treatment, AMY (10, 25, and 50 mg/kg, p.o.) increased the latency to the first convulsion (FC) by 30, 44, and 40% and time to death (TD) by 36, 52, and 42%, respectively. When administered intraperitoneally, the same doses increased FC by 62, 75, and 73% and TD by 76, 82, and 119%, respectively. Combined with polymixin or staurosporine, AMY (25 mg/kg, i.p.) increased TD by 61 and 63%, respectively, as related to each drug alone. When subchronically administered (25 and 50 mg/kg, i.p.) increased FC by 75 and 101% and TD by 86 and 124%, respectively. AMY increased taurine (116 and 76%) and tyrosine concentrations (135 and 110%) in basal ganglia and hippocampus, respectively, and decreased by 68, 65, and 62% glutamate, aspartate, and GABA in basal ganglia.Conclusion: Thus, the AMY anticonvulsant activity is related to the GABAergic system and may be linked to the inhibition of the signaling cascade of PKC as well as to alterations in amino acids metabolism.

Effects of eslicarbazepine acetate on acute and chronic latrunculin A-induced seizures and extracellular amino acid levels in the mouse hippocampus.

BackgroundLatrunculin A microperfusion of the hippocampus induces acute epileptic seizures and long-term biochemical changes leading to spontaneous seizures. This study tested the effect of eslicarbazepine acetate (ESL), a novel antiepileptic drug, on latrunculin A-induced acute and chronic seizures, and changes in brain amino acid extracellular levels. Hippocampi of Swiss mice were continuously perfused with a latrunculin A solution (4 μM, 1 μl/min, 7 h/day) with continuous EEG and videotape recording for 3 consecutive days. Microdialysate samples were analyzed by HPLC and fluorescence detection of taurine, glycine, aspartate, glutamate and GABA. Thereafter, mice were continuously video monitored for two months to identify chronic spontaneous seizures or behavioral changes. Control EEG recordings (8 h) were performed in all animals at least once a week for a minimum of one month.ResultsOral administration of ESL (100 mg/kg), previous to latrunculin A microperfusion, completely prevented acute latrunculin A-induced seizures as well as chronic seizures and all EEG chronic signs of paroxysmal activity. Hippocampal extracellular levels of taurine, glycine and aspartate were significantly increased during latrunculin A microperfusion, while GABA and glutamate levels remained unchanged. ESL reversed the increases in extracellular taurine, glycine and aspartate concentrations to basal levels and significantly reduced glutamate levels. Plasma and brain bioanalysis showed that ESL was completely metabolized within 1 h after administration to mainly eslicarbazepine, its major active metabolite.ConclusionESL treatment prevented acute latrunculin A-induced seizures as well as chronic seizures and all EEG chronic signs of paroxysmal activity, supporting a possible anti-epileptogenic effect of ESL in mice.

P.1.g.077 The protective role of arylpiperazine in neuroinflammation

Objetive: The purpose of the present work was to study the effects of standardized extract of Erythrina velutina (SEEV) in the determination of amino acids in mice brain undergoing global cerebral ischemia (ISQ). Methods: For this purpose, animals (swiss mice, males, 30−35 g) were subjected to transient cerebral ischemia by occlusion of both carotid arteries during 30 minutes and treated for 5 days with SEEV 200 or 400mg/kg and Memantine (MEM) 10mg/kg, 2 or 24 hours (2 or 24 h) after ischemia. The same procedure was done in false-operated group (FO) + dimethylsulfoxide (DMSO) with the exception of clamping the carotid arteries. On day 6 after induction of ischemia, the animals were subjected to sacrificed and dissected brains on ice the prefrontal cortex (PFC), determination of glutamate (GLU), and gammaamino-butirico acid (GABA). Statistical analysis were performed using two-way ANOVA considering significant when p< 0.05. Results: The results show there was an increase in the amount of glutamate in the PFC of ischemic animals treated with DMSO (ISQ + DMSO: 25.6±4.6) compared to the control group (FO + DMSO: 3.28±0.9). The amount GLU was reduced by both doses of SEEV (ISQ + SEEV 200mg 24 h: 21.9±6.1) and (ISQ + SEEV 400mg 24 h: 9.6±2.1) with only 24 hours. Regarding the inhibitory amino acid in the PFC groups (ISQ + SEEV 200mg 24 h: 355.7±71.0) and (ISQ + SEEV 400mg 2 h: 606.8±45.1) showed a significant increase when compared to ischemic control. Groups (ISQ + SEEV 200mg 2 h: 263.5±37.2), (ISQ + SEEV 400mg 2 h: 235.1±52.8), (ISQ + MEM 10mg 24 h: 300.4±49.7) showed an increase in GABA concentrations compared with the control group (ISQ + DMSO: 115.6±27.4). An increase in the concentrations of this amino acid in relation to time of administration of the drug can also be seen when comparing the group (ISQ + MEM 10mg 2 h: 138.0±21.8) and (ISQ + MEM 10mg 24 h: 300.4±49.7). Conclusion: In conclusion, these results corroborate the SEEV at both doses developed a neuroprotective action, possibly by reducing the levels of excitatory amino acids and increase inhibitory amino acids after on cerebral ischemia in mice.

Toxicological study of Zaleplon (Anxiolytic drug) in experimental animals.

Anxiety disorders are among the most prevalent mental disorders in the general population. Nearly 30 million persons are affected in the United States, with women affected nearly twice as frequently as men. These disorders are frequently treated using benzodiazepines (Diazepam) and non-benzodiazepines (Zaleplon). Non-benzodiazepines have replaced the benzodiazepines for anxiety treatment due to low risk of dependence and CNS effects encouraging drug abuse. This work was designed to investigate the toxicological overdose effect of zaleplon and diazepam on behavior, blood pressure, heart rate, brain neurotransmitter content and liver & kidney functions in rats. Rats were divided into five groups: 1% Tween 80, p.o. (control), diazepam (1 & 4 mg/kg, p.o.) and zaleplon (1 & 4 mg/kg, p.o.). All groups were injected daily for four weeks to perform the following: behavioral study (assessment of motor co-ordination and anxiety performance), measurement of blood pressure & heart rate, determination of the free amino acid and monoamine contents in the whole brain, determination of serum aminotransferases activity, total protein, total and direct billirubin, creatinine & blood urea nitrogen of experimental rats. Zaleplon decreased motor co-ordination, blood pressure and heart rate with extent lesser than diazepam. However, it decreased anxiety performance and brain monoamine content with extent more than diazepam. While, diazepam increased brain amino acid content, serum aminotransferases activity, total protein, total & direct billirubin, creatinine and blood urea nitrogen by magnitude more than zaleplon.

The contribution of ketone bodies to basal and activity-dependent neuronal oxidation in vivo.

The capacity of ketone bodies to replace glucose in support of neuronal function is unresolved. Here, we determined the contributions of glucose and ketone bodies to neocortical oxidative metabolism over a large range of brain activity in rats fasted 36 hours and infused intravenously with [2,4-(13)C₂]-D-β-hydroxybutyrate (BHB). Three animal groups and conditions were studied: awake ex vivo, pentobarbital-induced isoelectricity ex vivo, and halothane-anesthetized in vivo, the latter data reanalyzed from a recent study. Rates of neuronal acetyl-CoA oxidation from ketone bodies (V(acCoA-kbN)) and pyruvate (V(pdhN)), and the glutamate-glutamine cycle (V(cyc)) were determined by metabolic modeling of (13)C label trapped in major brain amino acid pools. V(acCoA-kbN) increased gradually with increasing activity, as compared with the steeper change in tricarboxylic acid (TCA) cycle rate (V(tcaN)), supporting a decreasing percentage of neuronal ketone oxidation: ∼100% (isoelectricity), 56% (halothane anesthesia), 36% (awake) with the BHB plasma levels achieved in our experiments (6 to 13 mM). In awake animals ketone oxidation reached saturation for blood levels >17 mM, accounting for 62% of neuronal substrate oxidation, the remainder (38%) provided by glucose. We conclude that ketone bodies present at sufficient concentration to saturate metabolism provides full support of basal (housekeeping) energy needs and up to approximately half of the activity-dependent oxidative needs of neurons.

Prepuberal Stimulation of 5-HT7-R by LP-211 in a Rat Model of Hyper-Activity and Attention-Deficit: Permanent Effects on Attention, Brain Amino Acids and Synaptic Markers in the Fronto-Striatal Interface

The cross-talk at the prefronto-striatal interface involves excitatory amino acids, different receptors, transducers and modulators. We investigated long-term effects of a prepuberal, subchronic 5-HT7-R agonist (LP-211) on adult behaviour, amino acids and synaptic markers in a model for Attention-Deficit/Hyperactivity Disorder (ADHD). Naples High Excitability rats (NHE) and their Random Bred controls (NRB) were daily treated with LP-211 in the 5th and 6th postnatal week. One month after treatment, these rats were tested for indices of activity, non selective (NSA), selective spatial attention (SSA) and emotionality. The quantity of L-Glutamate (L-Glu), L-Aspartate (L-Asp) and L-Leucine (L-Leu), dopamine transporter (DAT), NMDAR1 subunit and CAMKIIα, were assessed in prefrontal cortex (PFC), dorsal (DS) and ventral striatum (VS), for their role in synaptic transmission, neural plasticity and information processing. Prepuberal LP-211 (at lower dose) reduced horizontal activity and (at higher dose) increased SSA, only for NHE but not in NRB rats. Prepuberal LP-211 increased, in NHE rats, L-Glu in the PFC and L-Asp in the VS (at 0.250 mg/kg dose), whereas (at 0.125 mg/kg dose) it decreased L-Glu and L-Asp in the DS. The L-Glu was decreased, at 0.125 mg/kg, only in the VS of NRB rats. The DAT levels were decreased with the 0.125 mg/kg dose (in the PFC), and increased with the 0.250 mg/kg dose (in the VS), significantly for NHE rats. The basal NMDAR1 level was higher in the PFC of NHE than NRB rats; LP-211 treatment (at 0.125 mg/kg dose) decreased NMDAR1 in the VS of NRB rats. This study represents a starting point about the impact of developmental 5-HT7-R activation on neuro-physiology of attentive processes, executive functions and their neural substrates.

Pharmacologic inhibition of L‐tyrosine degradation ameliorates cerebral dopamine deficiency in murine phenylketonuria (PKU)

Monoamine neurotransmitter deficiency has been implicated in the etiology of neuropsychiatric symptoms associated with chronic hyperphenylalaninemia in phenylketonuria (PKU). Two proposed explanations for neurotransmitter deficiency in PKU include first, that chronically elevated blood L-phenylalanine (Phe) inhibits the transport of L-tyrosine (Tyr) and L-tryptophan (Trp), the substrates for dopamine and serotonin synthesis respectively, into brain. In the second hypothesis, elevated Phe competitively inhibits brain tyrosine hydroxylase (TH) and tryptophan hydroxylase (TPH) activities, the rate limiting steps in dopamine and serotonin synthesis. Dietary supplementation with large neutral amino acids (LNAA) including Tyr and Trp has been recommended for individuals with chronically elevated blood Phe in an attempt to restore amino acid and monoamine homeostasis in brain. As a potential alternative treatment approach, we demonstrate that pharmacologic inhibition of Tyr degradation through oral administration of nitisinone (NTBC) yielded sustained increases in blood and brain Tyr, decreased blood and brain Phe, and consequently increased dopamine synthesis in a murine model of PKU. Our results suggest that Phe-mediated inhibition of TH activity is the likely mechanism of impaired dopamine synthesis in PKU. Pharmacologic inhibition of Tyr degradation may be a promising adjunct therapy for CNS monoamine neurotransmitter deficiency in hyperphenylalaninemic individuals with PKU.

Effect of Glutamine Synthetase Inhibition on Brain and Interorgan Ammonia Metabolism in Bile Duct Ligated Rats

Ammonia has a key role in the development of hepatic encephalopathy (HE). In the brain, glutamine synthetase (GS) rapidly converts blood-borne ammonia into glutamine which in high concentrations may cause mitochondrial dysfunction and osmolytic brain edema. In astrocyte-neuron cocultures and brains of healthy rats, inhibition of GS by methionine sulfoximine (MSO) reduced glutamine synthesis and increased alanine synthesis. Here, we investigate effects of MSO on brain and interorgan ammonia metabolism in sham and bile duct ligated (BDL) rats. Concentrations of glutamine, glutamate, alanine, and aspartate and incorporation of (15)NH(4)(+) into these amino acids in brain, liver, muscle, kidney, and plasma were similar in sham and BDL rats treated with saline. Methionine sulfoximine reduced glutamine concentrations in liver, kidney, and plasma but not in brain and muscle; MSO reduced incorporation of (15)NH(4)(+) into glutamine in all tissues. It did not affect alanine concentrations in any of the tissues but plasma alanine concentration increased; incorporation of (15)NH(4)(+) into alanine was increased in brain in sham and BDL rats and in kidney in sham rats. It inhibited GS in all tissues examined but only in brain was an increased incorporation of (15)N-ammonia into alanine observed. Liver and kidney were important for metabolizing blood-borne ammonia.

Inhibition of Mitochondrial Pyruvate Transport by Zaprinast Causes Massive Accumulation of Aspartate at the Expense of Glutamate in the Retina

Transport of pyruvate into mitochondria by the mitochondrial pyruvate carrier is crucial for complete oxidation of glucose and for biosynthesis of amino acids and lipids. Zaprinast is a well known phosphodiesterase inhibitor and lead compound for sildenafil. We found Zaprinast alters the metabolomic profile of mitochondrial intermediates and amino acids in retina and brain. This metabolic effect of Zaprinast does not depend on inhibition of phosphodiesterase activity. By providing (13)C-labeled glucose and glutamine as fuels, we found that the metabolic profile of the Zaprinast effect is nearly identical to that of inhibitors of the mitochondrial pyruvate carrier. Both stimulate oxidation of glutamate and massive accumulation of aspartate. Moreover, Zaprinast inhibits pyruvate-driven O2 consumption in brain mitochondria and blocks mitochondrial pyruvate carrier in liver mitochondria. Inactivation of the aspartate glutamate carrier in retina does not attenuate the metabolic effect of Zaprinast. Our results show that Zaprinast is a potent inhibitor of mitochondrial pyruvate carrier activity, and this action causes aspartate to accumulate at the expense of glutamate. Our findings show that Zaprinast is a specific mitochondrial pyruvate carrier (MPC) inhibitor and may help to elucidate the roles of MPC in amino acid metabolism and hypoglycemia.

Paracetamol impairs the profile of amino acids in the rat brain

In our experiment we investigated the effect of subcutaneous administration of paracetamol on the levels of amino acids in the brain structures. Male Wistar rats received for eight weeks paracetamol at two doses: 10mg/kg b.w. (group P10, n=9) and 50mg/kg b.w. per day s.c. (group P50, n=9).The regional brain concentrations of amino acids were determined in the prefrontal cortex, hippocampus, hypothalamus and striatum of control (Con, n=9) and paracetamol-treated groups using HPLC. Evaluation of the biochemical results indicated considerable decrease of the content of amino acids in the striatum (glutamine, glutamic acid, taurine, alanine, aspartic acid) and hypothalamus (glycine) between groups treated with paracetamol compared to the control. In the prefrontal cortex paracetamol increased the level of γ-aminobutyric acid (GABA). The present study demonstrated significant effect of the long term paracetamol treatment on the level of amino acids in the striatum, prefrontal cortex and hypothalamus of rats.

Involvement of the Neutral Amino Acid Transporter SLC6A15 and Leucine in Obesity-Related Phenotypes

Brain pathways, including those in hypothalamus and nucleus of the solitary tract, influence food intake, nutrient preferences, metabolism and development of obesity in ways that often differ between males and females. Branched chain amino acids, including leucine, can suppress food intake, alter metabolism and change vulnerability to obesity. The SLC6A15 (v7-3) gene encodes a sodium-dependent transporter of leucine and other branched chain amino acids that is expressed by neurons in hypothalamus and nucleus of the solitary tract. We now report that SLC6A15 knockout attenuates leucine's abilities to reduce both: a) intake of normal chow and b) weight gain produced by access to a high fat diet in gender-selective fashions. We identify SNPs in the human SLC6A15 that are associated with body mass index and insulin resistance in males. These observations in mice and humans support a novel, gender-selective role for brain amino acid compartmentalization mediated by SLC6A15 in diet and obesity-associated phenotypes.

Stimulation of the Po-shen and Shen-hun scalp-acupuncture bands modifies levels of inhibitory and excitatory amino acids in the immature rat brain

ObjectiveIn the present study, the effect of stimulation of the Po-shen and Shen-hun scalp-acupuncture bands on tissue amino acid concentrations in several brain regions in awake and pentobarbital-sedated immature rats was evaluated. Materials and methodsSprague–Dawley rats (aged 15days) were organized in four groups of at least eight animals: control groups received saline solution 0.9% or sodium pentobarbital at 30mg/kg dosage via intraperitoneal. Experimental groups received saline solution or sodium pentobarbital plus stimulation in Po-shen and Shen-hun scalp-acupuncture bands for one continuous hour during 10 sessions by using scalp-acupuncture. ResultsAs compared to rats receiving saline solution, scalp-acupuncture produced significant changes in amino acid concentrations, depending on the analyzed region, as follows: in inhibitory amino acids, a GABA increase was observed in amygdala and hippocampus (491 and 184%, respectively), but a decrease in the substantia nigra (80%); glycine showed decrease in all the analyzed regions, except for an increase in brainstem(78%); glutamine presented an increase in hippocampus and cortex (42 and 149%, respectively). In the case of excitatory amino acids, glutamate decreased in all the analyzed regions; whereas aspartate decreased in substantia nigra and brainstem (77.08 and 35%, correspondingly) but increased in hippocampus and cortex (32 and 54%, respectively). The combined treatment of scalp-acupuncture and a GABAergic depressant drug like pentobarbital resulted in almost all changes induced in amino acids for scalp-acupuncture alone being significantly reverted. ConclusionStimulation of the Po-shen and Shen-hun scalp-acupuncture bands by using scalp-acupuncture alone might produce depressant activity by changes in amino acids, but the combination with a GABAergic tranquilizer like sodium pentobarbital can interfere with this response.

Differential levels of brain amino acids in rat models presenting learned helplessness or non-learned helplessness

Glutamatergic and γ-aminobutyric acid (GABA)ergic abnormalities have recently been proposed to contribute to depression. The learned helplessness (LH) paradigm produces a reliable animal model of depression that expresses a deficit in escape behavior (LH model); an alternative phenotype that does not exhibit LH is a model of resilience to depression (non-LH model). We measured the contents of amino acids in the brain to investigate the mechanisms involved in the pathology of depression. LH and non-LH models were subjected to inescapable electric footshocks at random intervals following a conditioned avoidance test to determine acquirement of predicted escape deficits. Tissue amino acid contents in eight brain regions were measured via high-performance liquid chromatography. The non-LH model showed increased GABA levels in the dentate gyrus and nucleus accumbens and increased glutamine levels in the dentate gyrus and the orbitofrontal cortex. The LH model had reduced glutamine levels in the medial prefrontal cortex. Changes in the ratios of GABA, glutamine, and glutamate were detected in the non-LH model, but not in the LH model. Reductions in threonine levels occurred in the medial prefrontal cortex in both models, whereas elevated alanine levels were detected in the medial prefrontal cortex in non-LH animals. The present study demonstrates region-specific compensatory elevations in GABA levels in the dentate gyrus and nucleus accumbens of non-LH animals, supporting the implication of the GABAergic system in the recovery of depression.

Long term compulsivity on the 5-choice serial reaction time task after acute Chlorpyrifos exposure

Pesticide exposure has been associated with neuropsychological and psychiatric impairments and neurodegenerative disorders. Pesticide exposure commonly causes a deficit in inhibitory control behaviours. In the present study, we investigated whether acute exposure to organophosphate (OP) chlorpyrifos (CPF) is related to long-term lack of inhibitory control; we also examined the possible neurochemical basis of this association. Lister Hooded rats were exposed to an acute dose of CPF (250mg/kg). Seven months later, we tested inhibitory control with the 5-choice serial reaction time task (5-CSRTT). We manipulated the baseline conditions of this task and also systemically pre-administered d-amphetamine, quinpirole, dizocilpine (MK-801) or ketanserin. We also analysed the post-mortem baseline levels of monoamines and amino acids in different brain regions. On the 5-CSRT task, CPF-exposed rats showed elevated perseverative responses that persisted across manipulation of baseline conditions of the task and under most of the pharmacological challenges tested. Only d-amphetamine induced a dose-dependent amelioration of the increased perseverative responses in the CPF group. The CPF group also exhibited increased levels of dopamine metabolism in the hippocampus and decreased levels of gamma-aminobutyric acid (GABA) and glutamate in the striatum compared to the vehicle group. These findings suggest that CPF induced a long-term compulsivity that was apparent in the 5-CSRT task and associated with changes in monoaminergic and amino acid brain systems of inhibitory control function. Exposure to high doses of OP should be taken into account in studies of environmental causes for neurodegenerative, neuropsychological and neuropsychiatric disorders.

Hepatocyte Transplantation Improves Blood and Brain Phenylalanine and Neurotransmitter Imbalances in a Mouse Model of Phenylketonuria (PKU)

Introduction: PKU (OMIM 261600), one of the most common inborn errors of metabolism (IEM) occurring in approximately 1 in 16,000 births in the United States, is characterized by deficiency of the phenylalanine hydroxylase (PAH) enzyme, the rate limiting step in phenylalanine (Phe) catabolism. Resulting chronic elevation of Phe in the blood and tissues, most importantly the brain, is responsible for PKU pathophysiology (mental retardation, seizures, growth delay). With good compliance, a special Phe-restricted diet can improve Phe imbalance and prevent most severe phenotypes, however the diet is unpalatable, expensive, and some patients still endure frequent hospitalizations and persistent cognitive defects. Partial correction of another IEM mouse model by hepatic transplantation of allogenic hepatocytes (Mol. Ther., 2009, 17:1266-1273; Biochimica et Biophysica. Acta., 2009, 1792:1004-1010) despite low functional engraftment provided proof-of-principle for these studies. Methods: Neonatal PAH -/- mice were given three direct hepatic injections of 1×106 syngenic mouse hepatocytes during the first 10 days of life (DOL). In addition, animals were given a single splenic transplant (2×106 cells) after 21 DOL. Mice were sacrificed at 1 or 3 months for blood and tissue analysis. All animals were maintained on normal mouse diet throughout the experiment. Results: Phe in the blood and brains of PAH-/- mice was severely elevated above wildtype. Surprisingly, Phe sera levels in untreated PKU females was elevated >35% higher than PKU males. Phe was improved up to 25% in the sera of PAH-/- females following hepatocyte transplant (HTx) while males were not improved. Importantly, amino acid levels in the brain showed multiple improvements following HTx, and no sex difference was apparent. Phe levels in PKU-HTx mice were reduced ˜50% compared to PKU controls. Additional brain amino acids (alanine, serine, histidine, ethanolamine) as well as amino acids that can act as neurotransmitters (taurine, glycine, aspartate) were normalized in PAH-/- animals after HTx. Dopamine (DA) metabolites (DOPAC, 3-MT, HVA) and dopamine turnover (DOPAC+HVA/DA) was disrupted in PAH-/- brains, though DA and tyrosine were unaffected. HTx normalized 3-MT/DA, suggesting dopamine release was improved. DOPAC was improved ˜35% and DA turnover was improved ˜25%, some to a level similar to wildtype, though statistical significance was not achieved. Severe disruptions were found in PAH-/- brains along the serotonin pathway and in serotonin turnover which were not resolved through transplant. Conclusions: HTx was able to significantly reduce brain Phe in PAH-/- animals and in the sera of PAH-/- females. The discrepancy in serum Phe concentrations observed between male and female mice is likely unique to the model, as this has not been reported in patients. Importantly, in addition to reduction of brain Phe levels following transplant, multiple other amino acids and neurotransmitters were normalized. Metabolites along the dopamine pathway, dopamine turnover, and dopamine release were also improved. Therefore, hepatocyte transplantation may be a viable therapy for PKU.

Quantitative LC–MS/MS analysis of arachidonoyl amino acids in mouse brain with treatment of FAAH inhibitor

An additional class of endogenous lipid amides, N-arachidonoyl amino acids (Ara-AAs), is growing in significance in the field of endocannabinoids. The development, validation, and application of a sensitive and selective method to simultaneously monitor and quantify the level of Ara-AAs along with anandamide (AEA) and 2-arachidonoyl glycerol (2-AG) in mouse brain has been established. The linearity of the method over the concentration ranges of 0.2–120pg/μl for the standards of N-arachidonoyl amino acids, N-arachidonoyl alanine (NAAla), serine (NASer), γ-aminobutyric acid (NAGABA), and glycine (NAGly); 0.7–90pg/μl for AEA-d0/d8; and 7.5–950pg/μl for 2-AG was determined with R2 values of 0.99. Also the effects of the FAAH inhibitor URB 597 on the endogenous levels of these analytes were investigated. AEA and NASer brain levels exhibit a dose-dependent increase after systemic administration of URB 597, whereas NAGly and NAGABA were significantly decreased after treatment. NAAla and 2-AG were not altered after URB 597 treatment. The potential benefit of establishing this assay extends beyond the quantification of the Ara-AAs along with AEA and 2-AG in mouse brain, to reveal a variety of pharmacological effects and physiological roles of these analytes.