Amino Acids In Cerebrospinal Fluid Research Articles

Huppke-Brendel syndrome: Novel cases and a therapeutic trial with ketogenic diet and N-acetylcysteine.

Huppke-Brendel syndrome (HBS) is an autosomal recessive disorder caused by SLC33A1 mutations, a gene coding for the acetyl-CoA transporter-1 (AT-1). So far it has been described in nine pediatric and one adult patient. Therapeutic trials with copper histidinate failed to achieve any clinical improvement. Here, we describe the clinical characteristics of two novel patients, one of them diagnosed by gene analysis and his sib postmortally based on clinical characteristics. We demonstrate a therapeutic trial with acetylation therapy, consisting of N-acetylcysteine and ketogenic diet, in one of them. We provide biochemical data on N-acetylated amino acids in cerebrospinal fluid (CSF) and plasma before and after starting this treatment regimen. Our results indicate that ketogenic diet and N-acetylcysteine do not seem to normalize the concentrations of N-acetylated amino acids in CSF or plasma. The overall metabolic pattern shows a trend toward lowered levels of N-acetylated amino acids in CSF and to a lesser extent in plasma. Although there are some assumptions, the function of AT-1 is still not clear and further studies are needed to better understand mechanisms underlying this complex disorder.

Metabolomics of Cerebrospinal Fluid Amino and Fatty Acids in Early Stages of Multiple Sclerosis.

Multiple sclerosis (MS) is a demyelinating and neurodegenerative autoimmune disease of the central nervous system (CNS) damaging myelin and axons. Diagnosis is based on the combination of clinical findings, magnetic resonance imaging (MRI) and analysis of cerebrospinal fluid (CSF). Metabolomics is a systematic study that allows us to track amounts of different metabolites in a chosen medium. The aim of this study was to establish metabolomic differences between the cerebrospinal fluid of patients in the early stages of multiple sclerosis and healthy controls, which could potentially serve as markers for predicting disease activity. We collected CSF from 40 patients after the first attack of clinical symptoms who fulfilled revised McDonald criteria of MS, and the CSF of 33 controls. Analyses of CSF samples were performed by using the high-performance liquid chromatography system coupled with a mass spectrometer with a high-resolution detector. Significant changes in concentrations of arginine, histidine, spermidine, glutamate, choline, tyrosine, serine, oleic acid, stearic acid and linoleic acid were observed. More prominently, Expanded Disability Status Scale values significantly correlated with lower concentrations of histidine. We conclude that these metabolites could potentially play a role as a biomarker of disease activity and predict presumable inflammatory changes.

Abnormal concentrations of acetylated amino acids in cerebrospinal fluid in acetyl-CoA transporter deficiency.

Acetyl-CoA transporter 1 (AT-1) is a transmembrane protein which regulates influx of acetyl-CoA from the cytosol to the lumen of the endoplasmic reticulum and is therefore important for the posttranslational modification of numerous proteins. Pathological variants in the SLC33A1 gene coding for AT-1 have been linked to a disorder called Huppke-Brendel syndrome, which is characterized by congenital cataracts, hearing loss, severe developmental delay and early death. It has been described in eight patients so far, who all had the abovementioned symptoms together with low serum copper and ceruloplasmin concentrations. The link between AT-1 and low ceruloplasmin concentrations is not clear, nor is the complex pathogenesis of the disease. Here we describe a further case of Huppke-Brendel syndrome with a novel and truncating homozygous gene variant and provide novel biochemical data on N-acetylated amino acids in cerebrospinal fluid (CSF) and plasma. Our results indicate that decreased levels of many N-acetylated amino acids in CSF are a typical metabolic fingerprint for AT-1 deficiency and are potential biomarkers for the defect. As acetyl-CoA is an important substrate for protein acetylation, we performed N-terminal proteomics, but found only minor effects on this particular protein modification. The acetyl-CoA content in patient's fibroblasts was insignificantly decreased. Our data may help to better understand the mechanisms underlying the metabolic disturbances, the pathophysiology and the clinical phenotype of the disease.

Is the Brain Undernourished in Alzheimer's Disease?

Cerebrospinal fluid (CSF) amino acid (AA) levels and CSF/plasma AA ratios in Alzheimer Disease (AD) in relation to nutritional state are not known. Methods: In 30 fasting patients with AD (46% males, 74.4 ± 8.2 years; 3.4 ± 3.2 years from diagnosis) and nine control (CTRL) matched subjects, CSF and venous blood samples were drawn for AA measurements. Patients were stratified according to nutritional state (Mini Nutritional Assessment, MNA, scores). Results: Total CSF/plasma AA ratios were lower in the AD subpopulations than in NON-AD (p < 0.003 to 0.017. In combined malnourished (16.7%; MNA < 17) and at risk for malnutrition (36.6%, MNA 17–24) groups (CG), compared to CTRL, all essential amino acids (EAAs) and 30% of non-EAAs were lower (p < 0.018 to 0.0001), whereas in normo-nourished ADs (46.7%, MNA > 24) the CSF levels of 10% of EAAs and 25% of NON-EAAs were decreased (p < 0.05 to 0.00021). CG compared to normo-nourished ADs, had lower CSF aspartic acid, glutamic acid and Branched-Chain AA levels (all, p < 0.05 to 0.003). CSF/plasma AA ratios were <1 in NON-AD but even lower in the AD population. Conclusions: Compared to CTRL, ADs had decreased CSF AA Levels and CSF/plasma AA ratios, the degree of which depended on nutritional state.

Cerebrospinal Fluid Amino Acid Metabolite Signatures of Diabetic Cognitive Dysfunction Based on Targeted Mass Spectrometry.

Diabetic cognitive dysfunction (DCD) is one of severe diabetic complications and might develop to irreversible dementia. Early diagnosis and detection of DCD is significant for prevention and treatment. The main objective of this study was to investigate the amino acid profiles of rat with DCD in the cerebrospinal fluid (CSF) to distinguish the early specific biomarkers. In total, rats were assigned into control and model groups. Model was induced by intraperitoneal injection of streptozotocin. The Morris water maze (MWM) method was used to evaluate learning and memory in rats on the 13th week after the model established. CSF samples were collected via cisterna magna puncture at the 0th, 5th, 9th, and 13th week, and amino acids profiling of CSF samples were performed via ultra performance liquid chromatography multiple reaction monitoring mass spectrometry (UPLC-MRM-MS). The amino acid profile was processed through multivariate analysis to identify potential biomarkers, and the related metabolic pathways were analyzed by MetaboAnalyst 5.0. Compared to the control group, the escape latency of the MWM was significantly prolonged in model group rats (p < 0.05). Different amino acid profiles were obtained between two groups. L-Alanine, L-Glutamine, L-Lysine, L-Serine, and L-Threonine were identified as potential biomarkers for DCD. These biomarkers are principally involved in glycine, serine, and threonine metabolism, aminoacyl-tRNA biosynthesis, alanine, aspartate, and glutamate metabolism, and glyoxylate and dicarboxylate metabolism. There are amino acid biomarkers in the CSF of rat with DCD. The mechanism of DCD is related to those pathways, which provide help for the early diagnosis and treatment and mechanism research.

Cerebrospinal fluid amino acids glycine, serine, and threonine in nonketotic hyperglycinemia.

Nonketotic hyperglycinemia (NKH) is caused by deficient glycine cleavage enzyme activity and characterized by elevated brain glycine. Metabolism of glycine is connected enzymatically to serine through serine hydroxymethyltransferase and shares transporters with serine and threonine. We aimed to evaluate changes in serine and threonine in NKH patients, and relate this to clinical outcome severity. Age‐related reference values were developed for cerebrospinal fluid (CSF) serine and threonine from 274 controls, and in a cross‐sectional study compared to 61 genetically proven NKH patients, categorized according to outcome. CSF d‐serine and l‐serine levels were stereoselectively determined in seven NKH patients and compared to 29 age‐matched controls. In addition to elevated CSF glycine, NKH patients had significantly decreased levels of CSF serine and increased levels of CSF threonine, even after age‐adjustment. The CSF serine/threonine ratio discriminated between NKH patients and controls. The CSF glycine/serine aided in discrimination between severe and attenuated neonates with NKH. Over all ages, the CSF glycine, serine and threonine had moderate to fair correlation with outcome classes. After age‐adjustment, only the CSF glycine level provided good discrimination between outcome classes. In untreated patients, d‐serine was more reduced than l‐serine, with a decreased d/l‐serine ratio, indicating a specific impact on d‐serine metabolism. We conclude that in NKH the elevation of glycine is accompanied by changes in l‐serine, d‐serine and threonine, likely reflecting a perturbation of the serine shuttle and metabolism, and of one‐carbon metabolism. This provides additional guidance on diagnosis and prognosis, and opens new therapeutic avenues to be explored.

Changes in the Cerebrospinal Fluid Amino Acid Profile of Patients with Anti-N2 Methyl-D-Aspartate Receptor Encephalitis

Changes in the Cerebrospinal Fluid Amino Acid Profile of Patients with Anti-N2 Methyl-D-Aspartate Receptor Encephalitis

Diet Effects on Cerebrospinal Fluid Amino Acids Levels in Adults with Normal Cognition and Mild Cognitive Impairment.

Background:Exploration of cerebrospinal fluid (CSF) amino acids and the impact of dietary intake on central levels may provide a comprehensive understanding of the metabolic component of Alzheimer’s disease.Objective:The objective of this exploratory study was to investigate the effects of two diets with varied nutrient compositions on change in CSF amino acids levels in adults with mild cognitive impairment (MCI) and normal cognition (NC). Secondary objectives were to assess the correlations between the change in CSF amino acids and change in Alzheimer’s disease biomarkers.Methods:In a randomized, parallel, controlled feeding trial, adults (NC, n = 20; MCI, n = 29) consumed a high saturated fat (SFA)/glycemic index (GI) diet [HIGH] or a low SFA/GI diet [LOW] for 4 weeks. Lumbar punctures were performed at baseline and 4 weeks.Results:CSF valine increased and arginine decreased after the HIGH compared to the LOW diet in MCI (ps = 0.03 and 0.04). This pattern was more prominent in MCI versus NC (diet by diagnosis interaction ps = 0.05 and 0.09), as was an increase in isoleucine after the HIGH diet (p = 0.05). Changes in CSF amino acids were correlated with changes in Alzheimer’s disease CSF biomarkers Aβ42, total tau, and p-Tau 181, with distinct patterns in the relationships by diet intervention and cognitive status.Conclusion:Dietary intake affects CSF amino acid levels and the response to diet is differentially affected by cognitive status.

LC–MS/MS analysis of twelve neurotransmitters and amino acids in mouse cerebrospinal fluid

BackgroundSo far, analytical investigation of neuroactive molecules in cerebrospinal fluid (CSF) of rodent models has been limited to rats, given the intrinsic anatomic difficulties related to mice sampling and the corresponding tiny amounts of CSF obtained. This poses a challenge for the research in neuroscience, where many, if not most, animal models for neuronal disorders rely on mice. New methodWe introduce a new, sensitive and robust LC–MS/MS method to analyze a panel of twelve neuroactive molecules (NM) from mouse CSF (aspartic acid, serine, glycine, glutamate, γ-aminobutyric acid, norepinephrine, epinephrine, acetylcholine, dopamine, serotonin, histamine and its metabolite 1-metylhistamine). The paper describes the sampling procedure that allows the collection of 1−2 microliters of pure CSF from individual mouse specimens. ResultsTo test its applicability, we challenged our method on the field, by sampling 37 individual animals, thus demonstrating its strength and reliability. Comparison with existing method(s)Compared to other methods, our procedure does not involve any extraction nor derivatization steps: samples are simply diluted and analyzed as such by LC–MS/MS, using a dedicated ion pairing agent in the chromatographic setup. The panel of neuroactive molecules that is analyzed in a single run is also significantly higher compared to other methods. ConclusionsGiven the number of mouse models used in the neuroscience research, we believe that our work will pave new ways to more advanced research in this field.

Diagnostic contribution of metabolic workup for neonatal inherited metabolic disorders in the absence of expanded newborn screening

Inherited metabolic disorders (IMDs) in neonates are a diagnostic and therapeutic challenge for the neonatologist, with the priority being to rapidly flag the treatable diseases. The objective of this study was to evaluate the contribution of targeted metabolic testing for diagnosing suspected IMDs on the basis of suggestive clinical setting or family history in neonates. We conducted an observational study over five years, from January 1st, 2010 to December 31, 2014 in the neonatal intensive care unit (NICU) at Robert Debré University Hospital, Paris, France. We assessed the number of neonates for whom a metabolic testing was performed, the indication for each metabolic test and the diagnostic yield of this selected metabolic workup for diagnosing an IMD. Metabolic testing comprised at least one of the following testings: plasma, urine or cerebrospinal fluid amino acids, urine organic acids, plasma acylcarnitine profile, and urine mucopolysaccharides and oligosaccharides. 11,301 neonates were admitted at the neonatal ICU during the study period. One hundred and ninety six neonates underwent metabolic testing. Eleven cases of IMDs were diagnosed. This diagnostic approach allowed the diagnosis, treatment and survival of 4 neonates (maple syrup urine disease, propionic acidemia, carnitine-acylcarnitine translocase deficiency and type 1 tyrosinemia). In total, metabolic testing was performed for 1.7% of the total number of neonates admitted in the NICU over the study period. These included 23% finally unaffected neonates with transient abnormalities, 5.6% neonates suffering from an identified IMD, 45.4% neonates suffering from a non-metabolic identified disease and 26% neonates with chronic abnormalities but for whom no final causal diagnosis could be made. In conclusion, as expected, such a metabolic targeted workup allowed the diagnosis of classical neonatal onset IMDs in symptomatic newborns. However, this workup remained normal or unspecific for 94.4% of the tested patients. It allowed excluding an IMD in 68.4% of the tested neonates. In spite of the high rate of normal results, such a strategy seems acceptable due to the severity of the symptoms and the need for immediate treatment when available in neonatal IMDs. However, its cost-effectiveness remains low especially in a clinically targeted population in a country where newborn screening is still unavailable for IMDs except for phenylketonuria in 2019.

Type-2-Diabetes Alters CSF but Not Plasma Metabolomic and AD Risk Profiles in Vervet Monkeys.

Epidemiological studies suggest that individuals with type 2 diabetes (T2D) have a twofold to fourfold increased risk for developing Alzheimer’s disease (AD), however, the exact mechanisms linking the two diseases are unknown. In both conditions, the majority of pathophysiological changes, including glucose and insulin dysregulation, insulin resistance, and AD-related changes in Aβ and tau, occur decades before the onset of clinical symptoms and diagnosis. In this study, we investigated the relationship between metabolic biomarkers associated with T2D and amyloid pathology including Aβ levels, from cerebrospinal fluid (CSF) and fasting plasma of healthy, pre-diabetic (PreD), and T2D vervet monkeys (Chlorocebus aethiops sabaeus). Consistent with the human disease, T2D monkeys have increased plasma and CSF glucose levels as they transition from normoglycemia to PreD and diabetic states. Although plasma levels of acylcarnitines and amino acids remained largely unchanged, peripheral hyperglycemia correlated with decreased CSF acylcarnitines and CSF amino acids, including branched chain amino acid (BCAA) concentrations, suggesting profound changes in cerebral metabolism coincident with systemic glucose dysregulation. Moreover, CSF Aβ40 and CSF Aβ42 levels decreased in T2D monkeys, a phenomenon observed in the human course of AD which coincides with increased amyloid deposition within the brain. In agreement with previous studies in mice, CSF Aβ40 and CSF Aβ42 were highly correlated with CSF glucose levels, suggesting that glucose levels in the brain are associated with changes in Aβ metabolism. Interestingly, CSF Aβ40 and CSF Aβ42 levels were also highly correlated with plasma but not CSF lactate levels, suggesting that plasma lactate might serve as a potential biomarker of disease progression in AD. Moreover, CSF glucose and plasma lactate levels were correlated with CSF amino acid and acylcarnitine levels, demonstrating alterations in cerebral metabolism occurring with the onset of T2D. Together, these data suggest that peripheral metabolic changes associated with the development of T2D produce alterations in brain metabolism that lead to early changes in the amyloid cascade, similar to those observed in pre-symptomatic AD.

Effects of L-theanine on Anxiety-like Behavior, Cerebrospinal Fluid Amino Acid Profile, and Hippocampal Activity in Wistar Kyoto Rats (P14-022-19)

ObjectivesL-theanine (N-ethyl-L-glutamine), an amino acid is well known as a relaxing agent. In the present study, we examined the effects of repeated L-theanine administration on behavior, hippocampal activity in Wistar Kyoto (WKY) rats, levels of amino acids in the cerebrospinal fluid (CSF), and studied an animal model of anxiety and depressive disorders. MethodsBehavioral tests were performed after 7–10 days of L-theanine (0.4 mg kg−1 day−1) or saline administration, followed by CSF sampling for high-performance liquid chromatography (HPLC) analysis. An independent set of animals was subjected to [18F] fluorodeoxyglucose positron emission tomography (PET) scanning after an identical dose of L-theanine or saline administration for 7 days. ResultsIn an elevated plus maze test, the time spent in open arms was significantly longer in the L-theanine group compared to saline group (P = 0.035). Also, significantly lower CSF glutamate (P = 0.039) and higher methionine (P = 0.024) concentrations were observed in the L-theanine group. Further, a significant increase in the standard uptake value ratio was observed in the hippocampus/cerebellum of the L-theanine group (P < 0.001). ConclusionsThese results suggest that L-theanine enhances hippocampal activity and exerts anxiolytic effects, which may be mediated by changes in glutamate and methionine levels in the brain. However, further study is required to effectively elucidate the mechanisms underlying the effects of L-theanine. Funding SourcesThis research study was funded by an unrestricted research grant provided by Taiyo Life Insurance, Himawari Foundation, Tokyo, Japan.

Abnormal Amino Acid Profiles of Blood and Cerebrospinal Fluid from Cystathionine β-Synthase-Deficient Mice, an Animal Model of Homocystinuria.

Mental retardation is the most common feature among inborn errors of amino acid metabolism. Patients with homocystinuria/homocysteinemia caused by cystathionine β-synthase (CBS) deficiency suffer from thromboembolism and mental retardation from early ages; therefore, detection by newborn screening is performed. Furthermore, elevated levels of serum homocysteine during pregnancy are associated with the occurrence of neural tube defects (NTDs) in newborns. However, the causes of such central nervous system (CNS) defects are unknown. We found previously impaired learning abilities in Cbs-deficient (Cbs-/-) mice (but not NTD births). Here, we investigated the amino acid profiles of serum and cerebrospinal fluid (CSF) from Cbs-/- mice. Mice deficient in cystathionine γ-lyase (Cth), a downstream enzyme of CBS in transsulfuration, as well as wild-type mice, were analyzed as controls. Cbs-/- and Cth-/- mice were smaller than wild-type mice, and CSF yields in Cbs-/- mice were lower than the others. CSF amino acid levels were generally lower than those in serum, and compared with the dramatic amino acid level alterations in Cbs-/- mouse serum, alterations in CSF were less apparent. However, marked upregulation (versus wild-type) of aspartic acid/asparagine (Asp/Asn), glutamine (Gln), serine (Ser), threonine (Thr), phenylalanine (Phe), tyrosine (Tyr), methionine (Met), total homocysteine, and citrulline, and downregulation of lysine (Lys) were found in Cbs-/- mouse CSF. Because similar regulation of total homocysteine/citrulline/Lys was observed in the CSF of Cth-/- mice, which are free of CNS dysfunction, the reduced CSF volumes and the level changes of other amino acids could be relevant to Cbs-/--specific CNS defects.

A Metabonomics Investigation into the Therapeutic Effects of BuChang NaoXinTong Capsules on Reversing the Amino Acid-Protein Interaction Network of Cerebral Ischemia.

Background Amino acids (AAs) in cerebrospinal fluid (CSF) play a pivotal role in cerebral ischemia (CI). BuChang NaoXinTong Capsules (BNC) are widely prescribed in Chinese medicine for the treatment of cerebrovascular and cardiovascular diseases. Methods In order to investigate the therapeutic effects and pharmacological mechanisms of BNC on reversing CI from a system level, an amino acid-protein interaction imbalanced network of CI containing metabolites of AAs, key regulatory enzymes, and proteins was constructed for the first time. Furthermore, a novel method for detecting the ten AAs in CSF was developed by UPLC-QQQ-MS in an effort to validate the imbalanced networks and the therapeutic effects of BNC via analysis of metabolites. Results Based on a middle cerebral artery occlusion (MCAO) rat model, the dynamic levels of amino acids in CSF 3, 6, 12, and 24 h after MCAO were analyzed. Up to 24 h, the accumulated nine AA biomarkers were found to significantly change in the MCAO group compared to the sham group and exhibited an obvious tendency for returning to baseline values after BNC treatment. In addition, based on the imbalanced network of CI, four key enzymes that regulate the generation of BNC-mediated AA biomarkers were selected and validated using an enzyme-linked immunosorbent assay and western blotting. Finally, aromatic-L-amino-acid decarboxylase (AADC) was found to be one of the putative targets for BNC-mediated protection against CI. Conclusion This study provides new strategies to explore the mechanism of cerebral ischemia and help discover the potential mechanism of BNC.

Aneurysmal Subarachnoid Hemorrhage Onset Alters Pyruvate Metabolism in Poor-Grade Patients and Clinical Outcome Depends on More: A Cerebrospinal Fluid Metabolomic Study.

Cerebral metabolism alterations influence cerebrospinal fluid (CSF) composition and are sensitive to brain injury. In subarachnoid hemorrhage (SAH) patients, Fisher scale, Hunt-Hess scale, and World Federation of Neurological Societies (WFNS) grading scale evaluating SAH severity are inadequate to predict long-term outcome; therefore, in an effort to determine metabolite pattern disparity and discover corresponding biomarkers, we designed an untargeted CSF metabolomic study covering a broad range of metabolites of SAH patients with different severity and outcome. The present study demonstrated the SAH altered the cerebrospinal fluid metabolome involving carbohydrate, lipid, and amino acid metabolism. Pyruvate metabolism was enhanced in SAH patients with Hunt-Hess scale above III, and the CSF pyruvate level was significantly associated with WFNS grading scale above III. There is no significant variation among CSF metabolome in SAH patients with merely different amounts and distribution of bleeding. SAH patients with unfavorable outcome present upregulated CSF amino acids level and enhanced lipid biosynthesis. The present study provides a novel possibility of early identification of patients who might possess unfavorable outcome and further clarification of the underlying pathophysiology.

Child with Susceptibility to Infection-Induced Acute Encephalopathy 3

AbstractA 10-month-old Bahraini boy born full-term to first-degree cousins, initially had normal developmental milestones, but presented with recurrent encephalopathy and seizures associated with upper respiratory tract infection. With each attack, the patient developmentally regressed further. Brain magnetic resonance imaging showed multiple old and new infarcts. Cerebrospinal fluid biochemistry, microbiology, and amino acids were unremarkable. Tandems mass spectrometry of urine organic acids was unremarkable as well. Electroencephalogram showed asymmetry with cortical irritability. Whole exome sequencing revealed a homozygous mutation of RAN-binding protein 2 gene, suggesting a diagnosis of susceptibility to infection-induced acute encephalopathy 3, which is an autosomal dominant condition. This is the first case to be reported in Bahrain.

P017 Impact of liver transplant on plasma and cerebrospinal fluid amino acids in patients with argininosuccinic aciduria

P017 Impact of liver transplant on plasma and cerebrospinal fluid amino acids in patients with argininosuccinic aciduria

Capillary electrophoresis-mass spectrometry of derivatized amino acids for targeted neurometabolomics – pH mediated reversal of diastereomer migration order

A targeted CE-MS approach was developed for the chiral analysis of biologically relevant amino acids in artificial cerebrospinal fluid (aCSF). In order to achieve chiral resolution, the five amino acids (Ser, Asn, Asp, Gln and Glu) were derivatized with (+)-1-(9-fluorenyl)ethyl chloroformate ((+)-FLEC). The diastereoselectivity was found to be highly dependent on pH for all analytes and the optimized background electrolyte (BGE) consisted of 150 mM acetic acid, adjusted to pH 3.7 with NH4OH. Furthermore, a reversal of the migration order of Asp derivatives was observed. This phenomenon seems to be caused by intra-molecular interactions affecting the pKa of the second ionizable group (the side chain carboxyl).The applicability of this method was evaluated using aCSF. A solid phase extraction (SPE) protocol was developed for the selective extraction of the FLEC derivatives. A full evaluation of the matrix effect and extraction yield was performed concluding that the matrix effect is marginal and the recoveries are between 46 and 92%. The method offers adequate sensitivity (limits of detection below 1 μM).

Protective effects of Ginkgo Terpene Lactones Meglumine Injection on focal cerebral ischemia in rats

To investigate the protective effects of ginkgo diterpene lactone meglumine injection (GDLMI) on cerebral focal ischemia reperfusion injury induced by middle cerebral artery occlusion (MCAO) in rats, and explore its possible mechanism. One hundred and forty male SD rats were randomly divided into sham operation group, model group, ginkgo biloba extract injection (Ginaton, 1.0 mL•kg⁻¹) group, nimodipine (0.4 mg•kg⁻¹) group, and GDLMI (5.2, 2.6, 1.3 mg•kg⁻¹) groups; All of rats received corresponding drugs by tail vein injection 4 days before operation (normal saline in model group and sham operation group). Except the sham operation group, the cerebral ischemic stroke model was established by MCAO method in right brain of the other rats. After 3 h of ischemia, all the animals received intravenous administration again. The neurobehavioral scores of rats after ischemia-reperfusion were evaluated and the infarct rate of brain tissue was observed by TTC staining. The super oxide dismutase (SOD), glutathione (GSH), malondialdehyde (MDA) and lactic acid (LA) contents in brain tissue homogenate and the concentration of Ca2+, glutamate (Glu) and aspartate (Asp), creatine phosphate kinase (CK-BB) and lactate dehydrogenase (LDH) content changes in cerebrospinal fluid were measured. As compared with the sham operation group, the cerebral infarction rate was increased significantly in the model group; the content of MDA and LA in the homogenate of brain tissue was increased, and the content of GSH and SOD was decreased; in cerebrospinal fluid, Ca2+ concentration was decreased, and the content of Glu and Asp, CK-BB and LDH increased significantly. As compared with the model group, the high and medium dose GDLMI groups can significantly reduce the cerebral infarction rate and improve the symptoms of neurological impairment; increase SOD and GSH activity, reduce MDA and LA content in serum; increase Ca2+ concentration in cerebrospinal fluid and decrease the content of neurotransmitter Glu and Asp as well as CK-BB and LDH. GDLMI could obviously improve neurologic impairment in model rats, and the mechanism may be related to recovering the blood brain barrier, scavenging free radicals, decreasing free Ca2+ inflow into the cells and the content of excitatory amino acid in cerebrospinal fluid to improve its protective effect on cerebral ischemia.

Effects of L-theanine on anxiety-like behavior, cerebrospinal fluid amino acid profile, and hippocampal activity in Wistar Kyoto rats.

The amino acid L-theanine (N-ethyl-L-glutamine) has historically been considered a relaxing agent. In the present study, we examined the effects of repeated L-theanine administration on behavior, levels of amino acids in the cerebrospinal fluid (CSF), and hippocampal activity in Wistar Kyoto (WKY) rats, an animal model of anxiety and depressive disorders. Behavioral tests were performed after 7-10days of L-theanine (0.4mgkg-1day-1) or saline administration, followed by CSF sampling for high-performance liquid chromatography (HPLC) analysis. An independent set of animals was subjected to [18F]fluorodeoxyglucose positron emission tomography (PET) scanning after the same dose of L-theanine or saline administration for 7days. In the elevated plus maze test, the time spent in the open arms was significantly longer in the L-theanine group than in the saline group (P=0.035). In addition, significantly lower CSF glutamate (P=0.039) and higher methionine (P=0.024) concentrations were observed in the L-theanine group than in the saline group. A significant increase in the standard uptake value ratio was observed in the hippocampus/cerebellum of the L-theanine group (P<0.001). These results suggest that L-theanine enhances hippocampal activity and exerts anxiolytic effects, which may be mediated by changes in glutamate and methionine levels in the brain. Further study is required to more fully elucidate the mechanisms underlying the effects of L-theanine.