The inflammatory microenvironment and neurotoxicity can hinder neuronal regeneration and functional recovery after spinal cord injury.Ruxolitinib,a JAK-STAT inhibitor,exhibits effectiveness in autoimmune diseases,arth...The inflammatory microenvironment and neurotoxicity can hinder neuronal regeneration and functional recovery after spinal cord injury.Ruxolitinib,a JAK-STAT inhibitor,exhibits effectiveness in autoimmune diseases,arthritis,and managing inflammatory cytokine storms.Although studies have shown the neuroprotective potential of ruxolitinib in neurological trauma,the exact mechanism by which it enhances functional recovery after spinal cord injury,particularly its effect on astrocytes,remains unclear.To address this gap,we established a mouse model of T10 spinal cord contusion and found that ruxolitinib effectively improved hindlimb motor function and reduced the area of spinal cord injury.Transcriptome sequencing analysis showed that ruxolitinib alleviated inflammation and immune response after spinal cord injury,restored EAAT2 expression,reduced glutamate levels,and alleviated excitatory toxicity.Furthermore,ruxolitinib inhibited the phosphorylation of JAK2 and STAT3 in the injured spinal cord and decreased the phosphorylation level of nuclear factor kappa-B and the expression of inflammatory factors interleukin-1β,interleukin-6,and tumor necrosis factor-α.Additionally,in glutamate-induced excitotoxicity astrocytes,ruxolitinib restored EAAT2 expression and increased glutamate uptake by inhibiting the activation of STAT3,thereby reducing glutamate-induced neurotoxicity,calcium influx,oxidative stress,and cell apoptosis,and increasing the complexity of dendritic branching.Collectively,these results indicate that ruxolitinib restores glutamate homeostasis by rescuing the expression of EAAT2 in astrocytes,reduces neurotoxicity,and effectively alleviates inflammatory and immune responses after spinal cord injury,thereby promoting functional recovery after spinal cord injury.展开更多
Glutamate excitotoxicity has been shown to play an important role in glaucoma, and glutamate can induce ferroptosis. The p38 mitogenactivated protein kinase(MAPK) pathway inhibitor SB202190 has a potential ability to ...Glutamate excitotoxicity has been shown to play an important role in glaucoma, and glutamate can induce ferroptosis. The p38 mitogenactivated protein kinase(MAPK) pathway inhibitor SB202190 has a potential ability to suppress ferroptosis, and its downstream targets, such as p53, have been shown to be associated with ferroptosis. However, whether ferroptosis also occurs in retinal ganglion cells in response to glutamate excitotoxicity and whether inhibition of ferroptosis reduces the loss of retinal ganglion cells induced by glutamate excitotoxicity remain unclear. This study investigated ferroptosis in a glutamate-induced glaucoma rat model and explored the effects and molecular mechanisms of SB202190 on retinal ganglion cells. A glutamate-induced excitotoxicity model in R28 cells and an N-methyl-D-aspartate-induced glaucoma model in rats were used. In vitro experiments showed that glutamate induced the accumulation of iron and lipid peroxide and morphological changes of mitochondria in R28 cells, and SB202190 inhibited these changes. Glutamate induced the levels of p-p38 MAPK/p38 MAPK and SAT1 and decreased the expression levels of ferritin light chain, SLC7A11, and GPX4. SB202190 inhibited the expression of iron death-related proteins induced by glutamate. In vivo experiments showed that SB202190 attenuated N-methyl-D-aspartate-induced damage to rat retinal ganglion cells and improved visual function. These results suggest that SB202190 can inhibit ferroptosis and protect retinal ganglion cells by regulating ferritin light chain, SAT1, and SLC7A11/Gpx4 pathways and may represent a potential retina protectant.展开更多
Epilepsy is a neurological disorder characterized by high morbidity,high recurrence,and drug resistance.Enhanced signaling through the excitatory neurotransmitter glutamate is intricately associated with epilepsy.Meta...Epilepsy is a neurological disorder characterized by high morbidity,high recurrence,and drug resistance.Enhanced signaling through the excitatory neurotransmitter glutamate is intricately associated with epilepsy.Metabotropic glutamate receptors(mGluRs)are G protein-coupled receptors activated by glutamate and are key regulators of neuronal and synaptic plasticity.Dysregulated mGluR signaling has been associated with various neurological disorders,and numerous studies have shown a close relationship between mGluRs expression/activity and the development of epilepsy.In this review,we first introduce the three groups of mGluRs and their associated signaling pathways.Then,we detail how these receptors influence epilepsy by describing the signaling cascades triggered by their activation and their neuroprotective or detrimental roles in epileptogenesis.In addition,strategies for pharmacological manipulation of these receptors during the treatment of epilepsy in experimental studies is also summarized.We hope that this review will provide a foundation for future studies on the development of mGluR-targeted antiepileptic drugs.展开更多
Poplar is one of the fastest-growing temperate trees in the world and is widely used in ornamental horticulture for shade.The root is essential for tree growth and development and its utilization potential is huge.Cal...Poplar is one of the fastest-growing temperate trees in the world and is widely used in ornamental horticulture for shade.The root is essential for tree growth and development and its utilization potential is huge.Calcium(Ca),as a signaling molecule,is involved in the regulation of plant root development.However,the detailed underlying regulatory mechanism is elusive.In this study,we analyzed the morphological and transcriptomic variations of 84K poplar(Populus alba×P.glandulosa)in response to different calcium concentrations and found that low Ca^(2+)(1 mmol·L^(-1))promoted lateral root development,while deficiency(0.1 mmol·L^(-1)Ca^(2+))inhibited lateral root development.Co-expression analysis showed that Ca^(2+)channel glutamate receptors(GLRs)were present in various modules with significance for root development.Two GLR paralogous genes,PagGLR3.3a and Pag GLR3.3b,were mainly expressed in roots and up-regulated under Ca^(2+)deficiency.The CRISPR/Cas9-mediated signal gene(crispr-PagGLR3.3a,PagGLR3.3b)and double gene(crispr-PagGLR3.3ab)mutants presented more and longer lateral roots.Anatomical analysis showed that crispr-PagGLR3.3ab plants had more xylem cells and promoted the development of secondary vascular tissues.Further transcriptomic analysis suggested that knockout of PagGLR3.3a and PagGLR3.3b led to the up-regulation of several genes related to protein phosphorylation,auxin efflux,lignin and hemicellulose biosynthesis as well as transcriptional regulation,which might contribute to lateral root growth.This study not only provides novel insight into how the Ca^(2+)channels mediated root growth and development in trees,but also provides a directive breeding of new poplar species for biofuel and bioenergy production.展开更多
Maintaining glutamate homeostasis after hypoxic ischemia is important for synaptic function and neural cell activity,and regulation of glutamate transport between astrocyte and neuron is one of the important modalitie...Maintaining glutamate homeostasis after hypoxic ischemia is important for synaptic function and neural cell activity,and regulation of glutamate transport between astrocyte and neuron is one of the important modalities for reducing glutamate accumulation.However,further research is needed to investigate the dynamic changes in and molecular mechanisms of glutamate transport and the effects of glutamate transport on synapses.The aim of this study was to investigate the regulatory mechanisms underlying Notch pathway mediation of glutamate transport and synaptic plasticity.In this study,Yorkshire neonatal pigs(male,age 3 days,weight 1.0–1.5 kg,n=48)were randomly divided into control(sham surgery group)and five hypoxic ischemia subgroups,according to different recovery time,which were then further subdivided into subgroups treated with dimethyl sulfoxide or a Notch pathway inhibitor(N-[N-(3,5-difluorophenacetyl-l-alanyl)]-S-phenylglycine t-butyl ester).Once the model was established,immunohistochemistry,immunofluorescence staining,and western blot analyses of Notch pathway-related proteins,synaptophysin,and glutamate transporter were performed.Moreover,synapse microstructure was observed by transmission electron microscopy.At the early stage(6–12 hours after hypoxic ischemia)of hypoxic ischemic injury,expression of glutamate transporter excitatory amino acid transporter-2 and synaptophysin was downregulated,the number of synaptic vesicles was reduced,and synaptic swelling was observed;at 12–24 hours after hypoxic ischemia,the Notch pathway was activated,excitatory amino acid transporter-2 and synaptophysin expression was increased,and the number of synaptic vesicles was slightly increased.Excitatory amino acid transporter-2 and synaptophysin expression decreased after treatment with the Notch pathway inhibitor.This suggests that glutamate transport in astrocytes-neurons after hypoxic ischemic injury is regulated by the Notch pathway and affects vesicle release and synaptic plasticity through the expression of synaptophysin.展开更多
Adipose mesenchymal stem cells(ADSCs)have protective effects against glutamate-induced excitotoxicity,but ADSCs are limited in use for treatment of optic nerve injury.Studies have shown that the extracellular vesicles...Adipose mesenchymal stem cells(ADSCs)have protective effects against glutamate-induced excitotoxicity,but ADSCs are limited in use for treatment of optic nerve injury.Studies have shown that the extracellular vesicles(EVs)secreted by ADSCs(ADSC-EVs)not only have the function of ADSCs,but also have unique advantages including non-immunogenicity,low probability of abnormal growth,and easy access to target cells.In the present study,we showed that intravitreal injection of ADSC-EVs substantially reduced glutamate-induced damage to retinal morphology and electroretinography.In addition,R28 cell pretreatment with ADSC-EVs before injury inhibited glutamate-induced overload of intracellular calcium,downregulation ofα-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid receptor(AMPAR)subunit GluA2,and phosphorylation of GluA2 and protein kinase C alpha in vitro.A protein kinase C alpha agonist,12-O-tetradecanoylphorbol 13-acetate,inhibited the neuroprotective effects of ADSC-EVs on glutamate-induced R28 cells.These findings suggest that ADSCEVs ameliorate glutamate-induced excitotoxicity in the retina through inhibiting protein kinase C alpha activation.展开更多
Glutamine and glutamate are major bioenergy substrates for normal and cancer cell growth.Cancer cells need more biofuel than normal tissues for energy supply,anti-oxidation activity and biomass production.Genes relate...Glutamine and glutamate are major bioenergy substrates for normal and cancer cell growth.Cancer cells need more biofuel than normal tissues for energy supply,anti-oxidation activity and biomass production.Genes related to metabolic chains in many cancers are somehow mutated,which makes cancer cells more glutamate dependent.Meanwhile,glutamate is an excitatory neurotransmitter for conducting signals through binding with different types of receptors in central neuron system.Interestingly,increasing evidences have shown involvement of glutamate signaling,guided through their receptors,in human malignancy.Dysregulation of glutamate transporters,such as excitatory amino acid transporter and cystine/glutamate antiporter system,also generates excessive extracellular glutamate,which in turn,activates glutamate receptors on cancer cells and results in malignant growth.These features make glutamate an attractive target for anti-cancer drug development with some glutamate targeted but blood brain barrier impermeable anti-psychosis drugs under consideration.We discussed the relevant progressions and drawbacks in this field herein.展开更多
The present study established a rat model of global cerebral ischemia induced by chest compression for six minutes to dynamically observe expressional changes of three glutamate transporters in the cerebral cortex and...The present study established a rat model of global cerebral ischemia induced by chest compression for six minutes to dynamically observe expressional changes of three glutamate transporters in the cerebral cortex and hippocampus. After 24 hours of ischemia, expression of glutamate transporter-1 significantly decreased in the cerebral cortex and hippocampus, which was accompanied by neuronal necrosis. At 7 days post-ischemia, expression of excitatory amino acid carrier 1 decreased in the hippocampal CA1 region and cortex, and was accompanied by apoptosis Expression of glutamate-aspartate transporter remained unchanged at 6 hours 7 days after ischemia. These results suggested that glutamate transporter levels were altered at different periods of cerebral ischemia.展开更多
Objective To investigate whether environmental cues associated with different properties of morphine could regulate the extracellular levels of glutamate and y-aminobutyric acid （GABA） in the hippocampal ventral sub...Objective To investigate whether environmental cues associated with different properties of morphine could regulate the extracellular levels of glutamate and y-aminobutyric acid （GABA） in the hippocampal ventral subiculum, which play a critical role in the reinstatement of drug-seeking behavior induced by environmental cues. Methods Conditioning place preference （CPP） and conditioning place aversion （CPA） models were used to establish environment associated with rewarding and aversive properties of morphine respectively. Microdialysis and high performance liquid chromatography were used to measure the extracelluar level of glutamate and GABA in the ventral subiculum under these environmental cues. Results Exposure to the environmental cues associated with rewarding properties of morphine resulted in a decrease （approximately 11%） of extracellular level of GABA in ventral subiculum, and exposure to the environmental cues associated with aversive properties of morphine resulted in an increase （approximately 230%） of extracellular level of glutamate in ventral subiculum. Conclusion Environmental cues associated with different properties of morphine modulate the release of distinct neurotransmitters in the hippocampal ventral subiculum possibly through different neural circuit.展开更多
A full_length cDNA has been cloned encoding nicotinamide adenine dinucleotide phosphate_specific glutamate dehydrogenase (NADP_GDH) from Chlorella sorokiniana with the RT_PCR method. The complete nucleotide sequence o...A full_length cDNA has been cloned encoding nicotinamide adenine dinucleotide phosphate_specific glutamate dehydrogenase (NADP_GDH) from Chlorella sorokiniana with the RT_PCR method. The complete nucleotide sequence of NADP_GDH gene had 94% homology to the previously reported one . The NADP_GDH gene was constructed into a vector highly expressed in plants. The specific activity of NADP_GDH in transformants was detected, but not in the control plants. All transformed shoots on MS medium containing lower concentration of nitrogen and the transformed seedlings grown in lower concentration of nitrogen vermiculite had higher growth rate and more leaves than the control plants. Transformed leaf discs cultured on MS medium containing different nitrogen concentrations had more chlorophyll contents compared to the controls. These results suggested that exogenous NADP_GDH may enhance the absorption and utilization to ammonium in plants. The increased weight of transformed leaf discs cultured on medium supplemented with different concentrations of phosphinothricin (PPT) was more than that of control discs. 0.5 μg/mL PPT could be used as a selecting drug instead of kanamycin to develop the transformants. These results suggested that the NADP_GDH gene might be used as a new selecting gene in the future research of plant gene engineering.展开更多
BACKGROUND： Injection of glutamate （Glu） in normal animals can cause neuronal c-Fos expression; however, whether Glu can induce spinal neuronal c-Fos expression in pain models is unclear. OBJECTIVE： To examine the...BACKGROUND： Injection of glutamate （Glu） in normal animals can cause neuronal c-Fos expression; however, whether Glu can induce spinal neuronal c-Fos expression in pain models is unclear. OBJECTIVE： To examine the effects of intraplantar and intrathecal injection of Glu on c-Fos expression in the L5 spinal cord dorsal horn Ⅰ/Ⅱ and Ⅲ/Ⅳ layers after spinal nerve ligation, and to study the effects of the N-methyI-D-aspartic acid （NMDA） receptor antagonist, D-2-amino-5-phosphonopentanoate （D-AP5）, and a selective group I mGluR antagonist, 7-hydroyiminocyclo propan[a]chromen-lacarboxylic acid ethyl ester （cpccoEt）. DESIGN, TIME AND SETTING： A randomized, controlled animal study was performed at the Department of Pharmacology, Oral Anatomy, and Neurobiology, Osaka University Graduate School of Dentistry, from December 2005 to December 2006. MATERIALS： Glu （5 μmol）, D-AP5 （50 nmot） and cpccoEt （250 nmol） were provided by Wako Pure Chemical Industries, Osaka, Japan, and diluted in saline （50 μL）. The pH of all solutions was adjusted to 7.4. METHODS： Twelve rats were randomly divided into sham operation （n = 6） and spinal nerve ligation （SNL; n = 6） groups for behavioral assessments of neuropathic pain after ligation surgery of the left L5-6 nerve segment. Another 60 rats were randomly divided into sham operation, SNL, saline-intraplantar, saline-intrathecal, Glu-intraplantar, Glu-intrathecal, D-AP5-intrathecal, Glu-D-AP5-intrathecal, cpccoEt-intrathecal, and Glu-cpccoEt-intrathecal groups, with 6 rats in each group. All groups except sham operation group received a similar SNL. On day 14, rats received a 50-μL injection of saline, Glu, D-AP5, and/or cpccoEt into the left intraplantar or intrathecal L5-4 segments. MAIN OUTCOME MEASURES： The number of c-Fos positive neurons in both Ⅰ/Ⅱ and Ⅲ/Ⅳ spinal layers at L6 was observed using immunohistochemistry 2 hours after administration. RESULTS： （1） SNL increased the level of c-Fos expression in two sides of the spinal cord, particularly on Ⅲ/Ⅳ spinal layers of the ligated side. （2） Intraplantar or intrathecal administration of saline significantly increased the c-Fos labeled neurons in Ⅰ/Ⅱ spinal layers of the ligated side, compared with SNL alone （P 〈 0.01）. （3） Intraplantar Glu （5 μmol） increased the number of c-Fos positive neurons in Ⅰ/Ⅱ spinal layers compared with intraplantar saline （P〈 0.01）. （4） The number of c-Fos neurons in Ⅰ/Ⅱ spinal layers on both the ipsilateral and contralateral side after intraplantar Glu was lower than intrathecal Glu （P〈 0.01）, with a 3-fold higher induction by intrathecal Glu. （5） Co-administration of D-AP5 or cpccoEt reduced the effects of intrathecal Glu （P 〈 0.01）. CONCLUSION： Intrathecal Glu increases c-Fos expression more than intraplantar Glu. Antagonists of NMDA and group I mGluRs block this effect.展开更多
Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system (CNS). Packaging and storage of glutamate into glutamatergic neuronal vesicles require ATP-dependent vesicular glutamate uptak...Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system (CNS). Packaging and storage of glutamate into glutamatergic neuronal vesicles require ATP-dependent vesicular glutamate uptake systems, which utilize the electrochemical proton gradient as a driving force. Three vesicular glutamate transporters (VGLUT1-3) have been recently identified from neuronal tissue where they play a key role to maintain the vesicular glutamate level. Recently, it has been demonstrated that glutamate signaling is also functional in peripheral neuronal and non-neuronal tissues, and occurs in sites of pituitary, adrenal, pineal glands, bone, GI tract, pancreas,skin, and testis. The glutamate receptors and VGLUTs in digestivesystem have been found in both neuronal and endocrinal cells. The glutamate signaling in the digestive system may have significant relevance to diabetes and GI tract motility disorders. This review will focus on the most recent update of molecular physiology of digestive VGLUTs.展开更多
Patients with liver disease may present hepatic enceph- alopathy (HE), a complex neuropsychiatric syndrome covering a wide range of neurological alterations, including cognitive and motor disturbances. HE reduces the ...Patients with liver disease may present hepatic enceph- alopathy (HE), a complex neuropsychiatric syndrome covering a wide range of neurological alterations, including cognitive and motor disturbances. HE reduces the quality of life of the patients and is associated with poor prognosis. In the worse cases HE may lead to coma or death. The mechanisms leading to HE which are not well known are being studied using animal models. The neurological alterations in HE are a consequence of impaired cerebral function mainly due to alterations in neurotransmission. We review here some studies indicating that alterations in neurotransmission associated to different types of glutamate receptors are responsible for some of the cognitive and motor alterations present in HE. These studies show that the function of the signal transduction pathway glutamate-nitric oxide-cGMP associated to the NMDA type of glutamate receptors is impaired in brain in vivo in HE animal models as well as in brain of patients died of HE. Activation of NMDA receptors in brain activates this pathway and increases cGMP. In animal models of HE this increase in cGMP induced by activation of NMDA receptors is reduced, which is responsible for the impairment in learning ability in these animal models. Increasing cGMP by pharmacological means restores learning ability in rats with HE and may be a new therapeutic approach to improve cognitive function in patients with HE. However, it is necessary to previously assess the possible secondary effects.Patients with HE may present psychomotor slowing, hypokinesia and bradykinesia. Animal models of HE also show hypolocomotion. It has been shown in rats with HE that hypolocomotion is due to excessive activation of metabotropic glutamate receptors (mGluRs) in substantia nigra pars reticulata. Blocking mGluR1 in this brain area normalizes motor activity in the rats, suggesting that a similar treatment for patients with HE could be useful to treat psychomotor slowing and hypokinesia. However, the possible secondary effects of mGluR1 antagonists should be previously evaluated. These studies are setting the basis for designing therapeutic procedures to specifically treat the individual neurological alterations in patients with HE.展开更多
BACKGROUND：Studies have reported that potassium channel openers exhibit a protective effect on cerebral ischemia-reperfusion injury and inhibit glutamate excitotoxicity in rats.However,the effects of the glutamate re...BACKGROUND：Studies have reported that potassium channel openers exhibit a protective effect on cerebral ischemia-reperfusion injury and inhibit glutamate excitotoxicity in rats.However,the effects of the glutamate receptor 1α and glutamate transporter 1 remain poorly understood.OBJECTIVE：To investigate the prophylactic use of the adenosine triphosphate-sensitive potassium channel opener cromakalim on neurological function and cerebral infarct size,as well as glutamate receptor 1α and glutamate transporter 1 expression,in rats with cerebral ischemia-reperfusion injury,and to explore action mechanisms underlying reduced glutamate excitotoxicity and neuroprotection in rats.DESIGN,TIME AND SETTING：Randomized,controlled,animal experiment was performed at the Brain Institute,Qingdao University Medical College,Between July 2008 and April 2009.MATERIALS：Cromakalim was purchased from Sigma,USA; rabbit anti-glutamate receptor 1α polyclonal antibody was offered by Wuhan Boster,China; rabbit anti-glutamate transporter 1 polyclonal antibody was offered by Santa Cruz Biotechnology,USA.METHODS：Sixty male,Wistar rats,aged 6 months,were randomly assigned to three groups （n =20）：sham-surgery,model,and cromakalim.Intraluminal thread methods were used to establish middle cerebral artery occlusion in rats from the model and cromakalim groups.Rats from the sham-surgery group were subjected to exposed common carotid artery,external carotid artery,and internal carotid artery,without occlusion.Cromakalim （10 mg/kg） was administered 30 minutes prior to middle cerebral artery occlusion,but there was no intervention in the model and sham-surgery groups.MAIN OUTCOME MEASURES：At 24 hours post-surgery,neurological behavioral functions were evaluated using Bederson＇s test,cerebral infarction volume was determined following tetrazolium chloride staining,and glutamate receptor 1a and glutamate transporter 1 expressions were detected using immunohistochemistry.RESULTS：Following cerebral ischemia-reperfusion injury,neurological behavioral malfunctions were obvious in all mice.Focal cerebral infarction was detected in ischemic hemispheres,glutamate receptor 1α expression increased,and glutamate transporter 1 expression decreased in the ischemic hemisphere （P〈 0.05）.Compared with the model group,neurological behavioral functions significantly improved,cerebral infarction volume was significantly reduced （P〈 0.05）,glutamate receptor 1α expression was significantly decreased,and glutamate transporter 1 expression was increased in the cromakalim group （P 〈 0.05）.CONCLUSION：Improved neurological function and reduced cerebral infarction volume in rats through the preventive use of cromakalim could be related to decreased glutamate receptor 1α expression and enhanced glutamate transporter 1 expression.展开更多
In recent years, more attention has been paid to the role of the glutamate transporter 1 （GLT-1, EAAT2） in major depressive disorder （MDD）. However, experimental data on brain GLT-1 levels are, to some extent, inc...In recent years, more attention has been paid to the role of the glutamate transporter 1 （GLT-1, EAAT2） in major depressive disorder （MDD）. However, experimental data on brain GLT-1 levels are, to some extent, inconsistent in human postmortem and animal studies, These discrepancies imply that the role of GLT-1 in the pathophysiology of MDD and the action of antidepressants remain obscure. This work was designed to study the impact of chronic unpredictable stress （CUS） for 2 ses- sions per day for 35 days and four weeks of fluoxetine （FLX） on depressive-like behaviors in rats, as well as the concomitant expression of the GLT-1 protein in the hippocampus. Behavioral changes were assessed by the sucrose preference and open field tests. GLT-1 levels were detected by immunohisto- chemistry and Western blot analysis. Our study demonstrated that the animals exposed to CUS showed depressive-like behaviors and exhibited a significant decrease in GLT-1 expression in the hippocampus. Chronic FLX treatment reversed the behavioral deficits and the CUS-induced decrease in GLT-1 levels. Taken together, our results support the reduction of GLT-1 in human postmortem studies in MDD and suggest that GLT-1 may be involved in the antidepressant activity of FLX. Our studies further support the notion that GLT-1 is an attractive candidate molecule associated with the fundamental processes of MDD and may be a potential, and novel pharmacological target for the treatment of MDD.展开更多
Glutamate is a regulated molecule in the mammalian testis. Extracellular regulation of glutamate in the body is determined largely by the expression of plasmalemmal glutamate transporters. We have examined by PCR, wes...Glutamate is a regulated molecule in the mammalian testis. Extracellular regulation of glutamate in the body is determined largely by the expression of plasmalemmal glutamate transporters. We have examined by PCR, western blotting and immunocytochemistry the expression of a panel of sodium-dependent plasmalemmal glutamate transporters in the rat testis. Proteins examined included： glutamate aspartate transporter （GLAST）, glutamate transporter 1 （GLT1）, excitatory amino acid carrier 1 （EAAC1）, excitatory amino acid transporter 4 （EAAT4） and EAAT5. We demonstrate that many of the glutamate transporters in the testis are alternately spliced. GLAST is present as exon-3- and exon-9-skipping forms. GLT1 was similarly present as the alternately spliced forms GLT1 b and GLTlc, whereas the abundant brain form （GLTla） was detectable only at the mRNA level. EAAT5 was also strongly expressed, whereas EAAC1 and EAAT4 were absent. These patterns of expression were compared with the patterns of endogenous glutamate localization and with patterns of D-aspartate accumulation, as assessed by immunocytochemistry. The presence of multiple glutamate transporters in the testis, including unusually spliced forms, suggests that glutamate homeostasis may be critical in this organ. The apparent presence of many of these transporters in the testis and sperm may indicate a need for glutamate transport by such cells.展开更多
Root biomass of rice seedlings was increased at lower concentration of exogenous NH 4 + , but it was decreased at higher concentration of exogenous NH 4 + . The level of free NH 4 + in the roots was accumulated gradua...Root biomass of rice seedlings was increased at lower concentration of exogenous NH 4 + , but it was decreased at higher concentration of exogenous NH 4 + . The level of free NH 4 + in the roots was accumulated gradually with the increase of NH 4 + concentration in the nutrient solution. The content of the soluble proteins was essentially constant at higher NH 4 + . The activities of glutamine synthetase (GS), NADH-dependent glutamate synthase (NADH-GOGAT), and NADH-dependent glutamate dehydrogenase (NADH-GDH) were risen with exogenous NH 4 + concentration at the lower NH 4 + concentration range. But the activities of GS and NADH-GOGAT were declined, and the level of NADH-GDH activity was kept constant under higher NH 4 + concentration. The GS/GDH ratio suggested that NH 4 + was assimilated by GS-GOGAT cycle under lower NH 4 + concentration, but NADH-GDH was more important for NH 4 + assimilation and detoxifying NH 4 + to the tissue cells at the higher NH 4 + level. According to the growth and the activity changes of these ammonium-assimilating enzymes of rice seedling roots, 10. 0 μg/mL NH 4 + -N in nutrient solution was more suitable to the rice growth.展开更多
Objective To study the effect of glutamate on the expression of vascular endothelial growth factor (VEGF) mRNA and protein in cultured rat astrocytes. Methods Cultured rat astrocytes were randomly divided into 6 group...Objective To study the effect of glutamate on the expression of vascular endothelial growth factor (VEGF) mRNA and protein in cultured rat astrocytes. Methods Cultured rat astrocytes were randomly divided into 6 groups:control group (C),glutamate group (G),QA group (Q),DCG-IV group (D),L-AP4 group (L) and glutamate+MCPG group (G+M). Cells were cultured under nomoxic condition (95% air,5% CO2). RT-PCR and ELISA methods were used to detect the expression of VEGF mRNA and protein in cultured astrocytes,respectively. G+M group was preincubated with 1mM MCPG for 30 min prior to the stimulation with glutamate. There were 7 time points at 0,4,8,12,16,24 and 48 h in each group except G+M group. Results The expression of VEGF mRNA and protein did not differ significantly among D group,L group and C group. Different from that in C group,the expression of VEGF mRNA and protein could be enhanced both in a dose-dependent and time-dependent manner in G group and Q group. Meanwhile,the enhanced expression of VEGF mRNA and protein in G group was completely suppressed by MCPG after 24 h. Conclusion Glutamate can increase the expression of VEGF mRNA and protein in cultured astrocytes,which may be due to the activation of group I metabotropic glutamate receptors in astrocytes.展开更多
<p style="margin-left:10.0pt;"> <br /> </p> <p style="margin-left:10.0pt;"> <span>The coenzyme B<sub>12</sub> dependent glutamate mutase is composed of two...<p style="margin-left:10.0pt;"> <br /> </p> <p style="margin-left:10.0pt;"> <span>The coenzyme B<sub>12</sub> dependent glutamate mutase is composed of two apoenzyme proteins subunits;S and E<sub>2</sub>, which while either fused or separate assemble with coenzyme B<sub>12</sub> to form an active holoenzyme (E<sub>2</sub>S<sub>2</sub>-B<sub>12</sub>) for catalyzing the reversible isomerization between (<i>S</i>)-glutamate and (2<i>S</i>, 3<i>S</i>)-3-methylas</span><span>- </span><span>partate. In order to assay the activity of glutamate mutase by UV spectrophotometry, this reaction is often coupled with methylaspartase which deaminates (2<i>S</i>, 3<i>S</i>)-3-methylaspartate to form mesaconate (<i>λ</i><sub>max</sub> = 240 nm, </span><span>Ɛ</span><sub><span>240</span></sub><span> = 3.8 mM<sup>-1</sup>·cm<sup>-1</sup>). The activities of different reconstitutions of glutamate mu<span>tase from separate apoenzyme components S and E in varied amount</span></span><span>s</span><span> of </span><span>coenzyme B<sub>12</sub> and adenosylpeptide B<sub>12</sub> as cofactors were measured by this assay and used to reveal the binding properties of the cofactor by the Michaelis</span><span>- </span><span>Menten Method. The values of <i>K<sub>m</sub></i> for coenzyme B<sub>12</sub> in due to reconstitutions of holoenzyme in 2, 7 and 14 S: E were determined as;1.12 ± 0.04 μM, 0.7 ± 0.05 μM and 0.52 ± 0.06 μM, respectively, so as those of adenosylpeptide B<sub>12</sub>;1.07 ± 0.04 μM and 0.35 ± 0.05 μM as obtained from respective 2 and 14 S: E compositions of holoenzyme. Analysis of these kinetics results curiously as<span>sociate</span></span><span>s</span><span> the increasing affinity of cofactors to apoenzyme with</span><span> </span><span>increased amount of component S used in reconstituting holoenzyme from separate</span><span> apoenzyme components and cofactor.</span><span> Moreover, in these studies a new method for assaying the activity of glutamate mutase was developed, whereby glutamate mutase activity is measured via depletion of NADH (<i>λ</i><sub>max</sub> = 340 nm, </span><span>Ɛ</span><sub><span>340</span></sub><span> = 6.3 mM<sup>-1</sup>·cm<sup>-1</sup>) as determined by UV spectrophotometry after addition of (2<i>S</i>,<span> 3<i>S</i>)-3-methylaspartate and pyruvate to a mixture of E<sub>2</sub>S<sub>2</sub>-B<sub>12</sub> and two auxiliary </span><span>holoenzymes system;pyridoxal-5-phosphate dependent glutamate-pyruvate </span><span>aminotransferase and N</span>ADH dependent (<i>R</i>)-2-hydroxyglutarate dehydrogenas<span>e. The activity of glutamate-pyruvate aminotransferase was relatively complete recovered upon the addition of (<i>S</i>)-glutamate and pyruvate to the mixtures of hologlutamate-pyruvate aminotransferase and (<i>R</i>)-2-hydroxylglutarate</span> dehydrogenase which were incubated with each putative inhibitor of glutamate mutase. Additionally, the new assay was used to determine the kinetic constants of (2<i>S</i>, 3<i>S</i>)-3-methylaspartate in the reaction of glutamate mutase as <i>K</i><sub>m</sub>= 7 ± 0.07 mM and <i>k</i><sub>cat</sub>= 0.54 ± 0.6 s<sup>-1</sup>. Application of Briggs-Haldane formula allowed the calculation of an equilibrium constant of the reversible isomerization, <i>K</i><sub>eq</sub> = [(<i>S</i>)-glutamate] × [(2<i>S</i>, 3<i>S</i>)-3-methylaspartate]<sup>-1</sup> = 16, where the kinetic constants of (<i>S</i>)-glutamate were determined by the standard methylaspartase coupled assay.<span></span></span> </p> <p> <br /> </p>展开更多
基金supported by the National Natural Science Foundation of China,No.82272484(to XC).
文摘The inflammatory microenvironment and neurotoxicity can hinder neuronal regeneration and functional recovery after spinal cord injury.Ruxolitinib,a JAK-STAT inhibitor,exhibits effectiveness in autoimmune diseases,arthritis,and managing inflammatory cytokine storms.Although studies have shown the neuroprotective potential of ruxolitinib in neurological trauma,the exact mechanism by which it enhances functional recovery after spinal cord injury,particularly its effect on astrocytes,remains unclear.To address this gap,we established a mouse model of T10 spinal cord contusion and found that ruxolitinib effectively improved hindlimb motor function and reduced the area of spinal cord injury.Transcriptome sequencing analysis showed that ruxolitinib alleviated inflammation and immune response after spinal cord injury,restored EAAT2 expression,reduced glutamate levels,and alleviated excitatory toxicity.Furthermore,ruxolitinib inhibited the phosphorylation of JAK2 and STAT3 in the injured spinal cord and decreased the phosphorylation level of nuclear factor kappa-B and the expression of inflammatory factors interleukin-1β,interleukin-6,and tumor necrosis factor-α.Additionally,in glutamate-induced excitotoxicity astrocytes,ruxolitinib restored EAAT2 expression and increased glutamate uptake by inhibiting the activation of STAT3,thereby reducing glutamate-induced neurotoxicity,calcium influx,oxidative stress,and cell apoptosis,and increasing the complexity of dendritic branching.Collectively,these results indicate that ruxolitinib restores glutamate homeostasis by rescuing the expression of EAAT2 in astrocytes,reduces neurotoxicity,and effectively alleviates inflammatory and immune responses after spinal cord injury,thereby promoting functional recovery after spinal cord injury.
基金supported by the National Natural Science Foundation of China,Nos.81974132,81770927Hunan Provincial Health Commission,No.20220702839+1 种基金the Natural Science Foundation of Hunan Province of China,No.2022JJ30076National Key R&D Program of China,No.2021YFA1101202(all to WS)。
文摘Glutamate excitotoxicity has been shown to play an important role in glaucoma, and glutamate can induce ferroptosis. The p38 mitogenactivated protein kinase(MAPK) pathway inhibitor SB202190 has a potential ability to suppress ferroptosis, and its downstream targets, such as p53, have been shown to be associated with ferroptosis. However, whether ferroptosis also occurs in retinal ganglion cells in response to glutamate excitotoxicity and whether inhibition of ferroptosis reduces the loss of retinal ganglion cells induced by glutamate excitotoxicity remain unclear. This study investigated ferroptosis in a glutamate-induced glaucoma rat model and explored the effects and molecular mechanisms of SB202190 on retinal ganglion cells. A glutamate-induced excitotoxicity model in R28 cells and an N-methyl-D-aspartate-induced glaucoma model in rats were used. In vitro experiments showed that glutamate induced the accumulation of iron and lipid peroxide and morphological changes of mitochondria in R28 cells, and SB202190 inhibited these changes. Glutamate induced the levels of p-p38 MAPK/p38 MAPK and SAT1 and decreased the expression levels of ferritin light chain, SLC7A11, and GPX4. SB202190 inhibited the expression of iron death-related proteins induced by glutamate. In vivo experiments showed that SB202190 attenuated N-methyl-D-aspartate-induced damage to rat retinal ganglion cells and improved visual function. These results suggest that SB202190 can inhibit ferroptosis and protect retinal ganglion cells by regulating ferritin light chain, SAT1, and SLC7A11/Gpx4 pathways and may represent a potential retina protectant.
基金supported by the Natural Science Foundation of Hunan Province,No.2021JJ30389(to JG)the Key Research and Development Program of Hunan Province of China,Nos.2022SK2042(to LL)and 2020SK2122(to ET)。
文摘Epilepsy is a neurological disorder characterized by high morbidity,high recurrence,and drug resistance.Enhanced signaling through the excitatory neurotransmitter glutamate is intricately associated with epilepsy.Metabotropic glutamate receptors(mGluRs)are G protein-coupled receptors activated by glutamate and are key regulators of neuronal and synaptic plasticity.Dysregulated mGluR signaling has been associated with various neurological disorders,and numerous studies have shown a close relationship between mGluRs expression/activity and the development of epilepsy.In this review,we first introduce the three groups of mGluRs and their associated signaling pathways.Then,we detail how these receptors influence epilepsy by describing the signaling cascades triggered by their activation and their neuroprotective or detrimental roles in epileptogenesis.In addition,strategies for pharmacological manipulation of these receptors during the treatment of epilepsy in experimental studies is also summarized.We hope that this review will provide a foundation for future studies on the development of mGluR-targeted antiepileptic drugs.
基金supported by the National Natural Science Foundation of China(Grant Nos.32371902,31901327)National Key Research and Development Program of China(Grant Nos.2019YFE0119100,2021YFD2200205)+1 种基金Overseas Expertise Introduction Project for Discipline Innovation(111 Project D18008)The researches foundation of Zhejiang A&F University(Grant No.2018FR013)。
文摘Poplar is one of the fastest-growing temperate trees in the world and is widely used in ornamental horticulture for shade.The root is essential for tree growth and development and its utilization potential is huge.Calcium(Ca),as a signaling molecule,is involved in the regulation of plant root development.However,the detailed underlying regulatory mechanism is elusive.In this study,we analyzed the morphological and transcriptomic variations of 84K poplar(Populus alba×P.glandulosa)in response to different calcium concentrations and found that low Ca^(2+)(1 mmol·L^(-1))promoted lateral root development,while deficiency(0.1 mmol·L^(-1)Ca^(2+))inhibited lateral root development.Co-expression analysis showed that Ca^(2+)channel glutamate receptors(GLRs)were present in various modules with significance for root development.Two GLR paralogous genes,PagGLR3.3a and Pag GLR3.3b,were mainly expressed in roots and up-regulated under Ca^(2+)deficiency.The CRISPR/Cas9-mediated signal gene(crispr-PagGLR3.3a,PagGLR3.3b)and double gene(crispr-PagGLR3.3ab)mutants presented more and longer lateral roots.Anatomical analysis showed that crispr-PagGLR3.3ab plants had more xylem cells and promoted the development of secondary vascular tissues.Further transcriptomic analysis suggested that knockout of PagGLR3.3a and PagGLR3.3b led to the up-regulation of several genes related to protein phosphorylation,auxin efflux,lignin and hemicellulose biosynthesis as well as transcriptional regulation,which might contribute to lateral root growth.This study not only provides novel insight into how the Ca^(2+)channels mediated root growth and development in trees,but also provides a directive breeding of new poplar species for biofuel and bioenergy production.
基金supported by the National Natural Science Foundation of China,Nos.81871408 and 81271631(to XMW)National Science Foundation for Young Scientists of China,No.81801658(to YZ)+1 种基金Outstanding Scientific Fund of Shengjing Hospital,No.201402(to XMW)345 Talent Support Project of Shengjing Hospital,No.30B(to YZ)。
文摘Maintaining glutamate homeostasis after hypoxic ischemia is important for synaptic function and neural cell activity,and regulation of glutamate transport between astrocyte and neuron is one of the important modalities for reducing glutamate accumulation.However,further research is needed to investigate the dynamic changes in and molecular mechanisms of glutamate transport and the effects of glutamate transport on synapses.The aim of this study was to investigate the regulatory mechanisms underlying Notch pathway mediation of glutamate transport and synaptic plasticity.In this study,Yorkshire neonatal pigs(male,age 3 days,weight 1.0–1.5 kg,n=48)were randomly divided into control(sham surgery group)and five hypoxic ischemia subgroups,according to different recovery time,which were then further subdivided into subgroups treated with dimethyl sulfoxide or a Notch pathway inhibitor(N-[N-(3,5-difluorophenacetyl-l-alanyl)]-S-phenylglycine t-butyl ester).Once the model was established,immunohistochemistry,immunofluorescence staining,and western blot analyses of Notch pathway-related proteins,synaptophysin,and glutamate transporter were performed.Moreover,synapse microstructure was observed by transmission electron microscopy.At the early stage(6–12 hours after hypoxic ischemia)of hypoxic ischemic injury,expression of glutamate transporter excitatory amino acid transporter-2 and synaptophysin was downregulated,the number of synaptic vesicles was reduced,and synaptic swelling was observed;at 12–24 hours after hypoxic ischemia,the Notch pathway was activated,excitatory amino acid transporter-2 and synaptophysin expression was increased,and the number of synaptic vesicles was slightly increased.Excitatory amino acid transporter-2 and synaptophysin expression decreased after treatment with the Notch pathway inhibitor.This suggests that glutamate transport in astrocytes-neurons after hypoxic ischemic injury is regulated by the Notch pathway and affects vesicle release and synaptic plasticity through the expression of synaptophysin.
基金supported by the National Key R&D Program of China,No.2016YFC1201800(to JFH)the Key Research and Development Program of Hunan Province,Nos.2018SK2090(to JFH),2022SK2079(to JFH)+2 种基金the Natural Science Foundation of Hu nan Province,No.2021JJ30891(to DC)the Human Resource Bank Program of Hunan Province,No.2020TP3003(to JFH)the School-Enterprise Joint Program of Central South University,No.2021XQLH092(to TQD)。
文摘Adipose mesenchymal stem cells(ADSCs)have protective effects against glutamate-induced excitotoxicity,but ADSCs are limited in use for treatment of optic nerve injury.Studies have shown that the extracellular vesicles(EVs)secreted by ADSCs(ADSC-EVs)not only have the function of ADSCs,but also have unique advantages including non-immunogenicity,low probability of abnormal growth,and easy access to target cells.In the present study,we showed that intravitreal injection of ADSC-EVs substantially reduced glutamate-induced damage to retinal morphology and electroretinography.In addition,R28 cell pretreatment with ADSC-EVs before injury inhibited glutamate-induced overload of intracellular calcium,downregulation ofα-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid receptor(AMPAR)subunit GluA2,and phosphorylation of GluA2 and protein kinase C alpha in vitro.A protein kinase C alpha agonist,12-O-tetradecanoylphorbol 13-acetate,inhibited the neuroprotective effects of ADSC-EVs on glutamate-induced R28 cells.These findings suggest that ADSCEVs ameliorate glutamate-induced excitotoxicity in the retina through inhibiting protein kinase C alpha activation.
基金supported by NIH/NCI ROICA140988-01 to JWpartially supported by Chinese Scholar Council to HY。
文摘Glutamine and glutamate are major bioenergy substrates for normal and cancer cell growth.Cancer cells need more biofuel than normal tissues for energy supply,anti-oxidation activity and biomass production.Genes related to metabolic chains in many cancers are somehow mutated,which makes cancer cells more glutamate dependent.Meanwhile,glutamate is an excitatory neurotransmitter for conducting signals through binding with different types of receptors in central neuron system.Interestingly,increasing evidences have shown involvement of glutamate signaling,guided through their receptors,in human malignancy.Dysregulation of glutamate transporters,such as excitatory amino acid transporter and cystine/glutamate antiporter system,also generates excessive extracellular glutamate,which in turn,activates glutamate receptors on cancer cells and results in malignant growth.These features make glutamate an attractive target for anti-cancer drug development with some glutamate targeted but blood brain barrier impermeable anti-psychosis drugs under consideration.We discussed the relevant progressions and drawbacks in this field herein.
基金supported by the National Natural Science Foundation of China, No. 81171168Shanghai Science and Technology Committee, No. 10140903200
文摘The present study established a rat model of global cerebral ischemia induced by chest compression for six minutes to dynamically observe expressional changes of three glutamate transporters in the cerebral cortex and hippocampus. After 24 hours of ischemia, expression of glutamate transporter-1 significantly decreased in the cerebral cortex and hippocampus, which was accompanied by neuronal necrosis. At 7 days post-ischemia, expression of excitatory amino acid carrier 1 decreased in the hippocampal CA1 region and cortex, and was accompanied by apoptosis Expression of glutamate-aspartate transporter remained unchanged at 6 hours 7 days after ischemia. These results suggested that glutamate transporter levels were altered at different periods of cerebral ischemia.
基金supported by the National Natural Science Foundation of China(No.30230130 and No.30400129)the Ministry of Science and Technology of China(No.2003CB515405,No.2005CB522406)+1 种基金the Program for Changjiang Scholars and Innovative Research Team of Ministry of Education of ChinaShanghai Municipal Commission for Science and Technology(No.06JC14008).
文摘Objective To investigate whether environmental cues associated with different properties of morphine could regulate the extracellular levels of glutamate and y-aminobutyric acid （GABA） in the hippocampal ventral subiculum, which play a critical role in the reinstatement of drug-seeking behavior induced by environmental cues. Methods Conditioning place preference （CPP） and conditioning place aversion （CPA） models were used to establish environment associated with rewarding and aversive properties of morphine respectively. Microdialysis and high performance liquid chromatography were used to measure the extracelluar level of glutamate and GABA in the ventral subiculum under these environmental cues. Results Exposure to the environmental cues associated with rewarding properties of morphine resulted in a decrease （approximately 11%） of extracellular level of GABA in ventral subiculum, and exposure to the environmental cues associated with aversive properties of morphine resulted in an increase （approximately 230%） of extracellular level of glutamate in ventral subiculum. Conclusion Environmental cues associated with different properties of morphine modulate the release of distinct neurotransmitters in the hippocampal ventral subiculum possibly through different neural circuit.
文摘A full_length cDNA has been cloned encoding nicotinamide adenine dinucleotide phosphate_specific glutamate dehydrogenase (NADP_GDH) from Chlorella sorokiniana with the RT_PCR method. The complete nucleotide sequence of NADP_GDH gene had 94% homology to the previously reported one . The NADP_GDH gene was constructed into a vector highly expressed in plants. The specific activity of NADP_GDH in transformants was detected, but not in the control plants. All transformed shoots on MS medium containing lower concentration of nitrogen and the transformed seedlings grown in lower concentration of nitrogen vermiculite had higher growth rate and more leaves than the control plants. Transformed leaf discs cultured on MS medium containing different nitrogen concentrations had more chlorophyll contents compared to the controls. These results suggested that exogenous NADP_GDH may enhance the absorption and utilization to ammonium in plants. The increased weight of transformed leaf discs cultured on medium supplemented with different concentrations of phosphinothricin (PPT) was more than that of control discs. 0.5 μg/mL PPT could be used as a selecting drug instead of kanamycin to develop the transformants. These results suggested that the NADP_GDH gene might be used as a new selecting gene in the future research of plant gene engineering.
基金the Scientific and Technological Research Project of Jiangxi Provincial Public Health Bureau,No.20071090
文摘BACKGROUND： Injection of glutamate （Glu） in normal animals can cause neuronal c-Fos expression; however, whether Glu can induce spinal neuronal c-Fos expression in pain models is unclear. OBJECTIVE： To examine the effects of intraplantar and intrathecal injection of Glu on c-Fos expression in the L5 spinal cord dorsal horn Ⅰ/Ⅱ and Ⅲ/Ⅳ layers after spinal nerve ligation, and to study the effects of the N-methyI-D-aspartic acid （NMDA） receptor antagonist, D-2-amino-5-phosphonopentanoate （D-AP5）, and a selective group I mGluR antagonist, 7-hydroyiminocyclo propan[a]chromen-lacarboxylic acid ethyl ester （cpccoEt）. DESIGN, TIME AND SETTING： A randomized, controlled animal study was performed at the Department of Pharmacology, Oral Anatomy, and Neurobiology, Osaka University Graduate School of Dentistry, from December 2005 to December 2006. MATERIALS： Glu （5 μmol）, D-AP5 （50 nmot） and cpccoEt （250 nmol） were provided by Wako Pure Chemical Industries, Osaka, Japan, and diluted in saline （50 μL）. The pH of all solutions was adjusted to 7.4. METHODS： Twelve rats were randomly divided into sham operation （n = 6） and spinal nerve ligation （SNL; n = 6） groups for behavioral assessments of neuropathic pain after ligation surgery of the left L5-6 nerve segment. Another 60 rats were randomly divided into sham operation, SNL, saline-intraplantar, saline-intrathecal, Glu-intraplantar, Glu-intrathecal, D-AP5-intrathecal, Glu-D-AP5-intrathecal, cpccoEt-intrathecal, and Glu-cpccoEt-intrathecal groups, with 6 rats in each group. All groups except sham operation group received a similar SNL. On day 14, rats received a 50-μL injection of saline, Glu, D-AP5, and/or cpccoEt into the left intraplantar or intrathecal L5-4 segments. MAIN OUTCOME MEASURES： The number of c-Fos positive neurons in both Ⅰ/Ⅱ and Ⅲ/Ⅳ spinal layers at L6 was observed using immunohistochemistry 2 hours after administration. RESULTS： （1） SNL increased the level of c-Fos expression in two sides of the spinal cord, particularly on Ⅲ/Ⅳ spinal layers of the ligated side. （2） Intraplantar or intrathecal administration of saline significantly increased the c-Fos labeled neurons in Ⅰ/Ⅱ spinal layers of the ligated side, compared with SNL alone （P 〈 0.01）. （3） Intraplantar Glu （5 μmol） increased the number of c-Fos positive neurons in Ⅰ/Ⅱ spinal layers compared with intraplantar saline （P〈 0.01）. （4） The number of c-Fos neurons in Ⅰ/Ⅱ spinal layers on both the ipsilateral and contralateral side after intraplantar Glu was lower than intrathecal Glu （P〈 0.01）, with a 3-fold higher induction by intrathecal Glu. （5） Co-administration of D-AP5 or cpccoEt reduced the effects of intrathecal Glu （P 〈 0.01）. CONCLUSION： Intrathecal Glu increases c-Fos expression more than intraplantar Glu. Antagonists of NMDA and group I mGluRs block this effect.
基金Supported by the National Institute of Diabetes and Digestive Kidney Diseases Grant R01-DK063142 and R01-DK33209
文摘Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system (CNS). Packaging and storage of glutamate into glutamatergic neuronal vesicles require ATP-dependent vesicular glutamate uptake systems, which utilize the electrochemical proton gradient as a driving force. Three vesicular glutamate transporters (VGLUT1-3) have been recently identified from neuronal tissue where they play a key role to maintain the vesicular glutamate level. Recently, it has been demonstrated that glutamate signaling is also functional in peripheral neuronal and non-neuronal tissues, and occurs in sites of pituitary, adrenal, pineal glands, bone, GI tract, pancreas,skin, and testis. The glutamate receptors and VGLUTs in digestivesystem have been found in both neuronal and endocrinal cells. The glutamate signaling in the digestive system may have significant relevance to diabetes and GI tract motility disorders. This review will focus on the most recent update of molecular physiology of digestive VGLUTs.
基金Supported by grants from the Ministerio de Ciencia y Tecnología, No. SAF2002-00851 and SAF2005-06089 and from Ministerio de Sanidad, No. Red G03-155 and PI050253 of Spain and by grants from Consellería de Empresa, Universidad y Ciencia, and de Sanidad, Generalitat Valenciana, No. Grupos03/001, GV04B-055, GV04B-012, GVS05/082 and ACOMP06/005 and AP-005/06
文摘Patients with liver disease may present hepatic enceph- alopathy (HE), a complex neuropsychiatric syndrome covering a wide range of neurological alterations, including cognitive and motor disturbances. HE reduces the quality of life of the patients and is associated with poor prognosis. In the worse cases HE may lead to coma or death. The mechanisms leading to HE which are not well known are being studied using animal models. The neurological alterations in HE are a consequence of impaired cerebral function mainly due to alterations in neurotransmission. We review here some studies indicating that alterations in neurotransmission associated to different types of glutamate receptors are responsible for some of the cognitive and motor alterations present in HE. These studies show that the function of the signal transduction pathway glutamate-nitric oxide-cGMP associated to the NMDA type of glutamate receptors is impaired in brain in vivo in HE animal models as well as in brain of patients died of HE. Activation of NMDA receptors in brain activates this pathway and increases cGMP. In animal models of HE this increase in cGMP induced by activation of NMDA receptors is reduced, which is responsible for the impairment in learning ability in these animal models. Increasing cGMP by pharmacological means restores learning ability in rats with HE and may be a new therapeutic approach to improve cognitive function in patients with HE. However, it is necessary to previously assess the possible secondary effects.Patients with HE may present psychomotor slowing, hypokinesia and bradykinesia. Animal models of HE also show hypolocomotion. It has been shown in rats with HE that hypolocomotion is due to excessive activation of metabotropic glutamate receptors (mGluRs) in substantia nigra pars reticulata. Blocking mGluR1 in this brain area normalizes motor activity in the rats, suggesting that a similar treatment for patients with HE could be useful to treat psychomotor slowing and hypokinesia. However, the possible secondary effects of mGluR1 antagonists should be previously evaluated. These studies are setting the basis for designing therapeutic procedures to specifically treat the individual neurological alterations in patients with HE.
基金Shandong Provincial Science and Technology Plan Foundation
文摘BACKGROUND：Studies have reported that potassium channel openers exhibit a protective effect on cerebral ischemia-reperfusion injury and inhibit glutamate excitotoxicity in rats.However,the effects of the glutamate receptor 1α and glutamate transporter 1 remain poorly understood.OBJECTIVE：To investigate the prophylactic use of the adenosine triphosphate-sensitive potassium channel opener cromakalim on neurological function and cerebral infarct size,as well as glutamate receptor 1α and glutamate transporter 1 expression,in rats with cerebral ischemia-reperfusion injury,and to explore action mechanisms underlying reduced glutamate excitotoxicity and neuroprotection in rats.DESIGN,TIME AND SETTING：Randomized,controlled,animal experiment was performed at the Brain Institute,Qingdao University Medical College,Between July 2008 and April 2009.MATERIALS：Cromakalim was purchased from Sigma,USA; rabbit anti-glutamate receptor 1α polyclonal antibody was offered by Wuhan Boster,China; rabbit anti-glutamate transporter 1 polyclonal antibody was offered by Santa Cruz Biotechnology,USA.METHODS：Sixty male,Wistar rats,aged 6 months,were randomly assigned to three groups （n =20）：sham-surgery,model,and cromakalim.Intraluminal thread methods were used to establish middle cerebral artery occlusion in rats from the model and cromakalim groups.Rats from the sham-surgery group were subjected to exposed common carotid artery,external carotid artery,and internal carotid artery,without occlusion.Cromakalim （10 mg/kg） was administered 30 minutes prior to middle cerebral artery occlusion,but there was no intervention in the model and sham-surgery groups.MAIN OUTCOME MEASURES：At 24 hours post-surgery,neurological behavioral functions were evaluated using Bederson＇s test,cerebral infarction volume was determined following tetrazolium chloride staining,and glutamate receptor 1a and glutamate transporter 1 expressions were detected using immunohistochemistry.RESULTS：Following cerebral ischemia-reperfusion injury,neurological behavioral malfunctions were obvious in all mice.Focal cerebral infarction was detected in ischemic hemispheres,glutamate receptor 1α expression increased,and glutamate transporter 1 expression decreased in the ischemic hemisphere （P〈 0.05）.Compared with the model group,neurological behavioral functions significantly improved,cerebral infarction volume was significantly reduced （P〈 0.05）,glutamate receptor 1α expression was significantly decreased,and glutamate transporter 1 expression was increased in the cromakalim group （P 〈 0.05）.CONCLUSION：Improved neurological function and reduced cerebral infarction volume in rats through the preventive use of cromakalim could be related to decreased glutamate receptor 1α expression and enhanced glutamate transporter 1 expression.
基金supported by the Key Technology Research of Major Mental Illness Prevention and Treatment for the Barriers to the Recognition and Prevention of Depression and Anxiety in the General Hospital,China(No.2012BAI01B05)
文摘In recent years, more attention has been paid to the role of the glutamate transporter 1 （GLT-1, EAAT2） in major depressive disorder （MDD）. However, experimental data on brain GLT-1 levels are, to some extent, inconsistent in human postmortem and animal studies, These discrepancies imply that the role of GLT-1 in the pathophysiology of MDD and the action of antidepressants remain obscure. This work was designed to study the impact of chronic unpredictable stress （CUS） for 2 ses- sions per day for 35 days and four weeks of fluoxetine （FLX） on depressive-like behaviors in rats, as well as the concomitant expression of the GLT-1 protein in the hippocampus. Behavioral changes were assessed by the sucrose preference and open field tests. GLT-1 levels were detected by immunohisto- chemistry and Western blot analysis. Our study demonstrated that the animals exposed to CUS showed depressive-like behaviors and exhibited a significant decrease in GLT-1 expression in the hippocampus. Chronic FLX treatment reversed the behavioral deficits and the CUS-induced decrease in GLT-1 levels. Taken together, our results support the reduction of GLT-1 in human postmortem studies in MDD and suggest that GLT-1 may be involved in the antidepressant activity of FLX. Our studies further support the notion that GLT-1 is an attractive candidate molecule associated with the fundamental processes of MDD and may be a potential, and novel pharmacological target for the treatment of MDD.
文摘Glutamate is a regulated molecule in the mammalian testis. Extracellular regulation of glutamate in the body is determined largely by the expression of plasmalemmal glutamate transporters. We have examined by PCR, western blotting and immunocytochemistry the expression of a panel of sodium-dependent plasmalemmal glutamate transporters in the rat testis. Proteins examined included： glutamate aspartate transporter （GLAST）, glutamate transporter 1 （GLT1）, excitatory amino acid carrier 1 （EAAC1）, excitatory amino acid transporter 4 （EAAT4） and EAAT5. We demonstrate that many of the glutamate transporters in the testis are alternately spliced. GLAST is present as exon-3- and exon-9-skipping forms. GLT1 was similarly present as the alternately spliced forms GLT1 b and GLTlc, whereas the abundant brain form （GLTla） was detectable only at the mRNA level. EAAT5 was also strongly expressed, whereas EAAC1 and EAAT4 were absent. These patterns of expression were compared with the patterns of endogenous glutamate localization and with patterns of D-aspartate accumulation, as assessed by immunocytochemistry. The presence of multiple glutamate transporters in the testis, including unusually spliced forms, suggests that glutamate homeostasis may be critical in this organ. The apparent presence of many of these transporters in the testis and sperm may indicate a need for glutamate transport by such cells.
文摘Root biomass of rice seedlings was increased at lower concentration of exogenous NH 4 + , but it was decreased at higher concentration of exogenous NH 4 + . The level of free NH 4 + in the roots was accumulated gradually with the increase of NH 4 + concentration in the nutrient solution. The content of the soluble proteins was essentially constant at higher NH 4 + . The activities of glutamine synthetase (GS), NADH-dependent glutamate synthase (NADH-GOGAT), and NADH-dependent glutamate dehydrogenase (NADH-GDH) were risen with exogenous NH 4 + concentration at the lower NH 4 + concentration range. But the activities of GS and NADH-GOGAT were declined, and the level of NADH-GDH activity was kept constant under higher NH 4 + concentration. The GS/GDH ratio suggested that NH 4 + was assimilated by GS-GOGAT cycle under lower NH 4 + concentration, but NADH-GDH was more important for NH 4 + assimilation and detoxifying NH 4 + to the tissue cells at the higher NH 4 + level. According to the growth and the activity changes of these ammonium-assimilating enzymes of rice seedling roots, 10. 0 μg/mL NH 4 + -N in nutrient solution was more suitable to the rice growth.
基金supported by the National Natural Science Foundation of China (N0.30770673)
文摘Objective To study the effect of glutamate on the expression of vascular endothelial growth factor (VEGF) mRNA and protein in cultured rat astrocytes. Methods Cultured rat astrocytes were randomly divided into 6 groups:control group (C),glutamate group (G),QA group (Q),DCG-IV group (D),L-AP4 group (L) and glutamate+MCPG group (G+M). Cells were cultured under nomoxic condition (95% air,5% CO2). RT-PCR and ELISA methods were used to detect the expression of VEGF mRNA and protein in cultured astrocytes,respectively. G+M group was preincubated with 1mM MCPG for 30 min prior to the stimulation with glutamate. There were 7 time points at 0,4,8,12,16,24 and 48 h in each group except G+M group. Results The expression of VEGF mRNA and protein did not differ significantly among D group,L group and C group. Different from that in C group,the expression of VEGF mRNA and protein could be enhanced both in a dose-dependent and time-dependent manner in G group and Q group. Meanwhile,the enhanced expression of VEGF mRNA and protein in G group was completely suppressed by MCPG after 24 h. Conclusion Glutamate can increase the expression of VEGF mRNA and protein in cultured astrocytes,which may be due to the activation of group I metabotropic glutamate receptors in astrocytes.
文摘<p style="margin-left:10.0pt;"> <br /> </p> <p style="margin-left:10.0pt;"> <span>The coenzyme B<sub>12</sub> dependent glutamate mutase is composed of two apoenzyme proteins subunits;S and E<sub>2</sub>, which while either fused or separate assemble with coenzyme B<sub>12</sub> to form an active holoenzyme (E<sub>2</sub>S<sub>2</sub>-B<sub>12</sub>) for catalyzing the reversible isomerization between (<i>S</i>)-glutamate and (2<i>S</i>, 3<i>S</i>)-3-methylas</span><span>- </span><span>partate. In order to assay the activity of glutamate mutase by UV spectrophotometry, this reaction is often coupled with methylaspartase which deaminates (2<i>S</i>, 3<i>S</i>)-3-methylaspartate to form mesaconate (<i>λ</i><sub>max</sub> = 240 nm, </span><span>Ɛ</span><sub><span>240</span></sub><span> = 3.8 mM<sup>-1</sup>·cm<sup>-1</sup>). The activities of different reconstitutions of glutamate mu<span>tase from separate apoenzyme components S and E in varied amount</span></span><span>s</span><span> of </span><span>coenzyme B<sub>12</sub> and adenosylpeptide B<sub>12</sub> as cofactors were measured by this assay and used to reveal the binding properties of the cofactor by the Michaelis</span><span>- </span><span>Menten Method. The values of <i>K<sub>m</sub></i> for coenzyme B<sub>12</sub> in due to reconstitutions of holoenzyme in 2, 7 and 14 S: E were determined as;1.12 ± 0.04 μM, 0.7 ± 0.05 μM and 0.52 ± 0.06 μM, respectively, so as those of adenosylpeptide B<sub>12</sub>;1.07 ± 0.04 μM and 0.35 ± 0.05 μM as obtained from respective 2 and 14 S: E compositions of holoenzyme. Analysis of these kinetics results curiously as<span>sociate</span></span><span>s</span><span> the increasing affinity of cofactors to apoenzyme with</span><span> </span><span>increased amount of component S used in reconstituting holoenzyme from separate</span><span> apoenzyme components and cofactor.</span><span> Moreover, in these studies a new method for assaying the activity of glutamate mutase was developed, whereby glutamate mutase activity is measured via depletion of NADH (<i>λ</i><sub>max</sub> = 340 nm, </span><span>Ɛ</span><sub><span>340</span></sub><span> = 6.3 mM<sup>-1</sup>·cm<sup>-1</sup>) as determined by UV spectrophotometry after addition of (2<i>S</i>,<span> 3<i>S</i>)-3-methylaspartate and pyruvate to a mixture of E<sub>2</sub>S<sub>2</sub>-B<sub>12</sub> and two auxiliary </span><span>holoenzymes system;pyridoxal-5-phosphate dependent glutamate-pyruvate </span><span>aminotransferase and N</span>ADH dependent (<i>R</i>)-2-hydroxyglutarate dehydrogenas<span>e. The activity of glutamate-pyruvate aminotransferase was relatively complete recovered upon the addition of (<i>S</i>)-glutamate and pyruvate to the mixtures of hologlutamate-pyruvate aminotransferase and (<i>R</i>)-2-hydroxylglutarate</span> dehydrogenase which were incubated with each putative inhibitor of glutamate mutase. Additionally, the new assay was used to determine the kinetic constants of (2<i>S</i>, 3<i>S</i>)-3-methylaspartate in the reaction of glutamate mutase as <i>K</i><sub>m</sub>= 7 ± 0.07 mM and <i>k</i><sub>cat</sub>= 0.54 ± 0.6 s<sup>-1</sup>. Application of Briggs-Haldane formula allowed the calculation of an equilibrium constant of the reversible isomerization, <i>K</i><sub>eq</sub> = [(<i>S</i>)-glutamate] × [(2<i>S</i>, 3<i>S</i>)-3-methylaspartate]<sup>-1</sup> = 16, where the kinetic constants of (<i>S</i>)-glutamate were determined by the standard methylaspartase coupled assay.<span></span></span> </p> <p> <br /> </p>
