Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 130
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Neurotoxicology ; 67: 112-120, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29778792

RESUMO

Exposure to elevated levels of manganese (Mn) causes manganism, a neurological disorder with similar characteristics to those of Parkinson's disease (PD). Valproic acid (VPA), an antiepileptic, is known to inhibit histone deacetylases and exert neuroprotective effects in many experimental models of neurological disorders. In the present study, we investigated if VPA attenuated Mn-induced dopaminergic neurotoxicity and the possible mechanisms involved in VPA's neuroprotection, focusing on modulation of astrocytic glutamate transporters (glutamate aspartate transporter, GLAST and glutamate transporter 1, GLT-1) and histone acetylation in H4 astrocyte culture and mouse models. The results showed that VPA increased promoter activity, mRNA/protein levels of GLAST/GLT-1 and glutamate uptake, and reversed Mn-reduced GLAST/GLT-1 in in vitro astrocyte cultures. VPA also attenuated Mn-induced reduction of GLAST and GLT-1 mRNA/protein levels in midbrain and striatal regions of the mouse brain when VPA (200 mg/kg, i.p., daily, 21 d) was administered 30 min prior to Mn exposure (30 mg/kg, intranasal instillation, daily, 21 d). Importantly, VPA attenuated Mn-induced dopaminergic neuronal damage by reversing Mn-induced decrease of tyrosine hydroxylase (TH) mRNA/protein levels in the nigrostriatal regions. VPA also reversed Mn-induced reduction of histone acetylation in astrocytes as well as mouse brain tissue. Taken together, VPA exerts attenuation against Mn-induced decrease of astrocytic glutamate transporters parallel with reversing Mn-induced dopaminergic neurotoxicity and Mn-reduced histone acetylation. Our findings suggest that VPA could serve as a potential neuroprotectant against Mn neurotoxicity as well as other neurodegenerative diseases associated with excitotoxicity and impaired astrocytic glutamate transporters.


Assuntos
Encéfalo/metabolismo , Dopamina/metabolismo , Transportador 1 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/biossíntese , Manganês/toxicidade , Ácido Valproico/farmacologia , Animais , Anticonvulsivantes/farmacologia , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Encéfalo/efeitos dos fármacos , Células Cultivadas , Transportador 1 de Aminoácido Excitatório/antagonistas & inibidores , Transportador 1 de Aminoácido Excitatório/genética , Transportador 2 de Aminoácido Excitatório/antagonistas & inibidores , Transportador 2 de Aminoácido Excitatório/genética , Expressão Gênica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos
2.
Neuropharmacology ; 135: 297-307, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29567092

RESUMO

Cocaine use disorder is a chronically relapsing disease without FDA-approved treatments. Using a rodent model of cocaine relapse, we and others have previously demonstrated that the beta-lactam antibiotic ceftriaxone attenuates cue- and cocaine-primed reinstatement of cocaine-seeking. Ceftriaxone restores cocaine-induced deficits in both system xc- and GLT-1 expression and function in the nucleus accumbens core (NAc). We recently demonstrated that restoration of GLT-1 expression in the NAc is necessary for ceftriaxone to attenuate reinstatement of cocaine-seeking. Here we used an adeno-associated virus (AAV) to overexpress GLT-1a in the NAc to investigate whether such restoration is sufficient to attenuate cue- and cocaine-primed reinstatement. Rats self-administered cocaine for two weeks and received injections of either AAV-GFAP-GLT-1a or AAV-GFAP-eGFP in the NAc following the last day of self-administration. Rats then underwent three weeks of extinction training (during which time transduction and expression occurred) before undergoing a cue- or cocaine-primed reinstatement test. Microdialysis for the quantification of glutamate efflux in the NAc was conducted during the cocaine-primed test. Rats that received AAV-GFAP-GLT-1a reinstated cue-primed cocaine-seeking in a similar manner as rats that received the control AAV-GFAP-eGFP. Upregulation of GLT-1a attenuated glutamate efflux during a cocaine-primed reinstatement test, but was not sufficient to attenuate reinstatement. We confirmed that GLT-1a upregulation resulted in functional upregulation of glutamate transport and expression, without affecting sodium-independent glutamate uptake, indicating system xc-was not altered. These results indicate that upregulation of NAc GLT-1 transporters alone is not sufficient to prevent the reinstatement of cocaine-seeking and implicate additional mechanisms in regulating glutamate efflux.


Assuntos
Cocaína/farmacologia , Transportador 2 de Aminoácido Excitatório/biossíntese , Ácido Glutâmico/metabolismo , Núcleo Accumbens/metabolismo , Animais , Dependovirus , Extinção Psicológica/efeitos dos fármacos , Vetores Genéticos , Masculino , Ratos , Recidiva , Autoadministração , Regulação para Cima
3.
J Neuroinflammation ; 15(1): 5, 2018 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-29304807

RESUMO

BACKGROUND: Laquinimod is an immunomodulatory drug under clinical investigation for the treatment of the progressive form of multiple sclerosis (MS) with both anti-inflammatory and neuroprotective effects. Excitotoxicity, a prominent pathophysiological feature of MS and of its animal model, experimental autoimmune encephalomyelitis (EAE), involves glutamate transporter (GluT) dysfunction in glial cells. The aim of this study was to assess whether laquinimod might exert direct neuroprotective effects by interfering with the mechanisms of excitotoxicity linked to GluT function impairments in EAE. METHODS: Osmotic minipumps allowing continuous intracerebroventricular (icv) infusion of laquinimod for 4 weeks were implanted into C57BL/6 mice before EAE induction. EAE cerebella were taken to perform western blot and qPCR experiments. For ex vivo experiments, EAE cerebellar slices were incubated with laquinimod before performing electrophysiology, western blot, and qPCR. RESULTS: In vivo treatment with laquinimod attenuated EAE clinical score at the peak of the disease, without remarkable effects on inflammatory markers. In vitro application of laquinimod to EAE cerebellar slices prevented EAE-linked glutamatergic alterations without mitigating astrogliosis and inflammation. Moreover, such treatment induced an increase of Slcla3 mRNA coding for the glial glutamate-aspartate transporter (GLAST) without affecting the protein content. Concomitantly, laquinimod significantly increased the levels of the glial glutamate transporter 1 (GLT-1) protein and pharmacological blockade of GLT-1 function fully abolished laquinimod anti-excitotoxic effect. CONCLUSIONS: Overall, our results suggest that laquinimod protects against glutamate excitotoxicity of the cerebellum of EAE mice by bursting the expression of glial glutamate transporters, independently of its anti-inflammatory effects.


Assuntos
Anti-Inflamatórios/administração & dosagem , Encefalomielite Autoimune Experimental/tratamento farmacológico , Transportador 2 de Aminoácido Excitatório/biossíntese , Ácido Glutâmico/metabolismo , Quinolonas/administração & dosagem , Animais , Cerebelo/efeitos dos fármacos , Cerebelo/metabolismo , Cerebelo/patologia , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/patologia , Transportador 1 de Aminoácido Excitatório/biossíntese , Feminino , Infusões Intraventriculares , Camundongos , Camundongos Endogâmicos C57BL , Técnicas de Cultura de Órgãos , Distribuição Aleatória
4.
Pharmacol Biochem Behav ; 159: 18-23, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28687200

RESUMO

Alcohol addiction is a chronic disease characterized by an inability to regulate drinking. A critical brain region involved in alcohol consumption is the nucleus accumbens (NA). Glutamate transmission in this region regulates alcohol consumption and relapse to alcohol-seeking. Across multiple alcohol-administration rodent models, basal extracellular glutamate levels are increased in the NA during early withdrawal. Glutamate transporter 1 (GLT-1) and system xC-, containing the subunit xCT, regulate NA glutamate levels. Ceftriaxone (Cef) increases expression and function of both transporters following extinction from cocaine self-administration and here we sought to determine if Cef would similarly decrease alcohol consumption while increasing xCT and GLT-1 in the NA core. We used the intermittent access to alcohol (IAA) paradigm to induce drinking in outbred Sprague-Dawley rats; this paradigm permits rats access to alcohol (20%v/v) for 24-h without water deprivation, followed by 24-h of abstinence. Following 17 24-h drinking sessions, Cef treatment (200mg/kg IP) was initiated and continued for 5days while a control group received vehicle (0.9% saline IP). Alcohol consumption was assessed for two 24-h periods during Cef and two 24-h periods after cessation of Cef treatment. In a separate cohort of rats, Cef's ability to alter blood alcohol levels (BALs) after a non-contingent alcohol injection (1g/kg) was assessed. We found that Cef decreased alcohol consumption during the period of Cef treatment and on the two days following injections, and this was accompanied by an increase in NA core xCT expression. Furthermore, a history of alcohol consumption did not alter xCT and GLT-1 expression relative to alcohol-naïve controls. Cef did not alter BALs, indicating that the reduction in alcohol consumption was not caused by altered alcohol clearance. These results indicate that while Cef reduces alcohol consumption in outbred rats, its ability to do so is not associated with an increase in GLT-1 expression.


Assuntos
Consumo de Bebidas Alcoólicas/tratamento farmacológico , Alcoolismo/tratamento farmacológico , Sistemas de Transporte de Aminoácidos Acídicos/biossíntese , Antibacterianos/uso terapêutico , Ceftriaxona/uso terapêutico , Transportador 2 de Aminoácido Excitatório/biossíntese , Núcleo Accumbens/efeitos dos fármacos , Núcleo Accumbens/metabolismo , Consumo de Bebidas Alcoólicas/psicologia , Alcoolismo/psicologia , Animais , Peso Corporal/efeitos dos fármacos , Depressores do Sistema Nervoso Central/sangue , Ingestão de Líquidos/efeitos dos fármacos , Etanol/sangue , Masculino , Ratos , Ratos Sprague-Dawley , Autoadministração , Regulação para Cima
5.
Artigo em Inglês | MEDLINE | ID: mdl-28347687

RESUMO

Alteration in glutamate neurotransmission has been found to mediate the development of drug dependence, including nicotine. We and others, through using western blotting, have reported that exposure to drugs of abuse reduced the expression of glutamate transporter-1 (GLT-1) as well as cystine/glutamate antiporter (xCT), which consequently increased extracellular glutamate concentrations in the mesocorticolimbic area. However, our previous studies did not reveal any changes in glutamate/aspartate transporter (GLAST) following exposure to drugs of abuse. In the present study, for the first time, we investigated the effect of chronic exposure to electronic (e)-cigarette vapor containing nicotine, for one hour daily for six months, on GLT-1, xCT, and GLAST expression in frontal cortex (FC), striatum (STR), and hippocampus (HIP) in outbred female CD1 mice. In this study, we also investigated the expression of alpha-7 nicotinic acetylcholine receptor (α-7 nAChR), a major pre-synaptic nicotinic receptor in the glutamatergic neurons, which regulates glutamate release. We found that inhalation of e-cigarette vapor for six months increased α-7 nAChR expression in both FC and STR, but not in the HIP. In addition, chronic e-cigarette exposure reduced GLT-1 expression only in STR. Moreover, e-cigarette vapor inhalation induced downregulation of xCT in both the STR and HIP. We did not find any significant changes in GLAST expression in any brain region. Finally, using liquid chromatography-tandem mass spectrometry (LC-MS/MS) techniques, we detected high concentrations of nicotine and cotinine, a major metabolite of nicotine, in the FC tissues of e-cigarette exposed mice. These data provide novel evidence about the effects of chronic nicotine inhalation on the expression of key glial glutamate transporters as well as α-7 nAChR. Our work may suggest that nicotine exposure via chronic inhalation of e-cigarette vapor may be mediated in part by alterations in the glutamatergic system.


Assuntos
Sistema X-AG de Transporte de Aminoácidos/biossíntese , Sistema y+ de Transporte de Aminoácidos/biossíntese , Sistemas Eletrônicos de Liberação de Nicotina , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Nicotina/administração & dosagem , Nicotina/farmacologia , Receptor Nicotínico de Acetilcolina alfa7/biossíntese , Administração por Inalação , Animais , Corpo Estriado/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Feminino , Lobo Frontal/metabolismo , Hipocampo/metabolismo , Camundongos , Neurônios/metabolismo , Nicotina/metabolismo
6.
Pharmacology ; 99(3-4): 153-159, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28049198

RESUMO

Glutamate transporter-1 (GLT-1), a major glutamate transporter expressed in astrocytes, takes up excess glutamate from the micro-environment in order to prevent excitotoxicity. Drugs that increase GLT-1 expression may have therapeutic effects in disorders associated with neuronal excitotoxicity. 2,3,4',5-tetrahydroxystilbene 2-O-ß-D-glucoside (TSG), a monomer of stilbene from polygonummultiflorum, exerts neuroprotection in a range of experimental models such as Alzheimer's disease and brain ischemia. In this study, we evaluated the effect of TSG on GLT-1 protein expression in mouse primary-cultured astrocytes. Results showed that TSG markedly increased the GLT-1 protein expression level in mouse primary-cultured astrocytes in a dose- and time-dependent manner, and this increase was mediated by the activation of protein kinase B (Akt) but not by the activation of extracellular signal-regulated protein kinase 1/2. Furthermore, inhibition of cAMP response element-binding protein, but not nuclear factor kappa B, abolished the TSG-mediated increase in GLT-1 protein expression in cultured astrocytes. Collectively, these findings may provide novel insights into the mechanism for TSG in neuroprotection, and would help search new agents targeting neurodegenerative disorders associated with impaired astrocytic glutamate transporters.


Assuntos
Astrócitos/efeitos dos fármacos , Transportador 2 de Aminoácido Excitatório/biossíntese , Glucosídeos/farmacologia , Estilbenos/farmacologia , Regulação para Cima/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Astrócitos/metabolismo , Células Cultivadas , Regulação da Expressão Gênica , Glucosídeos/química , Camundongos , Camundongos Endogâmicos C57BL , Estilbenos/química , Regulação para Cima/fisiologia
7.
J Ethnopharmacol ; 202: 138-146, 2017 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-28065779

RESUMO

Cerebral ischemia damages central neurons, and abnormal microenvironment in ischemic condition is the key factor to the damages. The increase of local concentration of glutamic acid, the overload of Ca2+, and the mitochondrial stress caused by release of cytochrome C are important factors of abnormal microenvironment in cerebral ischemia. In this study ginsenoside Rb1, a compound from Panax Notoginseng, was used to intervene abnormal environment of neurons in the hippocampal CA1 region in two animal models (microperfusion model and photothrombosis model). RESULTS: Compared with the vehicle in the sham group, ginsenoside had following effects. a) ginsenoside Rb1 increased the regional cerebral blood flow (rCBF) and the stability of neuronal ultrastructure in in the hippocampal CA1 region and improved the adaptability of neurons in two models. b) ginsenoside Rb1 improved the expression level of glial glutamate transporter1 (GLT-1) and reversed the uptake of glutamate (Glu) after ischemia, and as a result thereby decreased the excitability of Glu and the expression level of GLT-1 was proportional to the dose of ginsenoside Rb1 and similar to that of Nimodipine. c) ginsenoside Rb1 inhibited the expression level of NMDAR and the overload of Ca2+, thereby reducing neuronal damages. Meanwhile, the expression level of NMDAR was inversely proportional to the dose of ginsenoside Rb1, which was similar to that of Nimodipine. d) ginsenoside Rb1 decreased the release of cytochrome C (Cyt-C) and reduced the damages caused by neuronal mitochondrial stress. Meanwhile, the release of Cyt-C was inversely proportional to the dose of ginsenoside Rb1, which was similar to that of Nimodipine. Ginsenoside Rb1 may be as an effective drug for neuroprotection and improve cerebral blood flow after acute ischemia and prevent the secondary brain damage induced by stroke.


Assuntos
Microambiente Celular/efeitos dos fármacos , Ginsenosídeos/farmacologia , Hipocampo/efeitos dos fármacos , Animais , Região CA1 Hipocampal/citologia , Região CA1 Hipocampal/efeitos dos fármacos , Cálcio/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Circulação Cerebrovascular/efeitos dos fármacos , Citocromos c/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/genética , Ácido Glutâmico/metabolismo , Masculino , Neurônios/efeitos dos fármacos , Neurônios/ultraestrutura , Nimodipina/farmacologia , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores
8.
J Physiol Pharmacol ; 68(5): 709-714, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29375045

RESUMO

Though positive effects of exercise on mood and well being are well recognised, the central regulatory mechanisms are still not fully understood. The present study was aimed to testing the hypothesis that voluntary wheel running activates the gene expression of glutamate transporters in the brain cortex of rats. The animals were assigned to the control and voluntary wheel running groups. Voluntary wheel running rats had free access to a stainless steel activity wheel for 3 weeks. The daily running distance gradually increased to 6.21 ± 1.05 km by day 21. Vesicular glutamate transporter 3 (VGLUT3) mRNA levels in the frontal cortex were significantly elevated in the group of running animals compared to the values in sedentary controls, while the expression of other vesicular transporters were unchanged. The concentrations of mRNA coding for glial glutamate transporter 1 (GLT-1), but not glutamate aspartate transporter (GLAST) were increased by running. Voluntary wheel running resulted in an elevation of plasma corticosterone and increased expression of brain derived neurotrophic factor (BDNF) in the frontal cortex. In conclusion, chronic voluntary wheel running results in increased gene expression of VGLUT3 and GLT-1 in the brain cortex without changes in other glutamate transporter subtypes.


Assuntos
Transportador 2 de Aminoácido Excitatório/biossíntese , Lobo Frontal/metabolismo , Condicionamento Físico Animal/fisiologia , Proteínas Vesiculares de Transporte de Glutamato/biossíntese , Sistema X-AG de Transporte de Aminoácidos/biossíntese , Sistema X-AG de Transporte de Aminoácidos/genética , Animais , Fator Neurotrófico Derivado do Encéfalo/biossíntese , Fator Neurotrófico Derivado do Encéfalo/genética , Transportador 2 de Aminoácido Excitatório/genética , Expressão Gênica , Masculino , Condicionamento Físico Animal/métodos , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Proteína Vesicular 1 de Transporte de Glutamato/biossíntese , Proteína Vesicular 1 de Transporte de Glutamato/genética , Proteínas Vesiculares de Transporte de Glutamato/genética
9.
Mol Neurobiol ; 54(1): 58-71, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-26732590

RESUMO

Our previous study has proved that the up-regulation of glial glutamate transporter 1 (GLT-1) played an important role in the acquisition of brain ischemic tolerance after cerebral ischemic preconditioning (CIP) in rats. However, little is known about the mechanism involved in the up-regulation of GLT-1 in the process. The present study investigates whether p38 MAPK, ERK1/2, and/or JNK participates in the up-regulation of GLT-1 during the induction of brain ischemic tolerance by CIP. It was found that CIP significantly enhanced the expression of p-p38 MAPK without altering p-ERK1/2 and p-JNK expression in the CA1 hippocampus. Inhibition of p38 MAPK function by its selective inhibitor SB203580 or knockdown p38 MAPK expression by its antisense oligodeoxynucleotides (AS-ODNs) suppressed the induction of brain ischemic tolerance. Furthermore, p38 MAPK was activated earlier than the up-regulation of GLT-1 in the CA1 hippocampus after CIP. Meanwhile, the expression of p-p38 MAPK by astrocytes was increased, and p38 MAPK AS-ODNs dose-dependently inhibited the up-regulation of GLT-1 after CIP. Taken together, it could be concluded that p38 MAPK participates in the mediation of GLT-1 up-regulation during the induction of brain ischemic tolerance after CIP.


Assuntos
Isquemia Encefálica/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Hipocampo/metabolismo , Precondicionamento Isquêmico/métodos , Regulação para Cima/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Isquemia Encefálica/patologia , Hipocampo/patologia , Masculino , Ratos , Ratos Wistar
10.
J Neurotrauma ; 34(1): 220-234, 2017 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-27312729

RESUMO

We hypothesize that the primary mechanism for removal of glutamate from the extracellular space is altered after traumatic brain injury (TBI). To evaluate this hypothesis, we initiated TBI in adult male rats using a 2.0 atm lateral fluid percussion injury (LFPI) model. In the ipsilateral cortex and hippocampus, we found no differences in expression of the primary glutamate transporter in the brain (GLT-1) 24 h after TBI. In contrast, we found a decrease in glutamate uptake in the cortex, but not the hippocampus, 24 h after injury. Because glutamate uptake is potently regulated by protein kinases, we assessed global serine-threonine protein kinase activity using a kinome array platform. Twenty-five kinome array peptide substrates were differentially phoshorylated between LFPI and controls in the cortex, whereas 19 peptide substrates were differentially phosphorylated in the hippocampus (fold change ≥ ± 1.15). We identified several kinases as likely to be involved in acute TBI, including protein kinase B (Akt) and protein kinase C (PKC), which are well-characterized modulators of GLT-1. Exploratory studies using an inhibitor of Akt suggest selective activation of kinases in LFPI versus controls. Ingenuity pathway analyses of implicated kinases from our network model found apoptosis and cell death pathways as top functions in acute LFPI. Taken together, our data suggest diminished activity of glutamate transporters in the prefrontal cortex, with no changes in protein expression of the primary glutamate transporter GLT-1, and global alterations in signaling networks that include serine-threonine kinases that are known modulators of glutamate transport activity.


Assuntos
Lesões Encefálicas Traumáticas/metabolismo , Córtex Cerebral/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Ácido Glutâmico/metabolismo , Animais , Lesões Encefálicas Traumáticas/genética , Transportador 2 de Aminoácido Excitatório/genética , Expressão Gênica , Masculino , Ratos , Ratos Sprague-Dawley
11.
Mol Cell Biochem ; 425(1-2): 103-112, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27837432

RESUMO

Orexin-A, which is an endogenous neuropeptide, is reported to have a protective role in ischemic stroke. High-concentration glutamic acid (Glu) induced by hypoxia injury in ischemic stroke can be inhibited by glial glutamate transporter GLT-1 which is only expressed in astroglia cells. A previous study reported that Orexin-A may regulate GLT-1 expression. However, the role of orexin-A in the regulation of GLT-1 in ischemic stroke still remains unclear. In this study, we aimed to investigate the effect and the underlying mechanism of orexin-A on Glu uptake in astrocytes in vitro and this effect on protecting the neurons from anoxia/hypoglycemic injury. The expression of GLT-1 significantly increased in the astrocytes with orexin-A treatment under anoxia/hypoglycemic conditions, promoting the uptake of Glu and inhibiting the apoptosis of co-cultured cells of astrocytes and neurons. However, these effects were significantly weakened by treatment with orexin-A receptor 1 (OX1R) antagonist. Orexin-A significantly up-regulated the expressions of PKCα and ERK1/2 under anoxia/hypoglycemic conditions in astrocytes, whereas the OX1R antagonist markedly reversed the effect. Furthermore, PKCα or ERK1/2 inhibitor significantly constrained the GLT-1 expression in astrocytes and facilitated the apoptosis of co-cultured cells, and GLT-1 overexpression could reverse those effects of PKCα or ERK1/2 inhibitor. Taken together, orexin-A promoted the GLT-1 expression via OX1R/PKCα/ERK1/2 pathway in astrocytes and protected co-cultured cells against anoxia/hypoglycemic injury.


Assuntos
Apoptose/efeitos dos fármacos , Astrócitos/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Ácido Glutâmico/metabolismo , Hipoglicemia/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteína Quinase 3 Ativada por Mitógeno/sangue , Neurônios/metabolismo , Receptores de Orexina/metabolismo , Orexinas/farmacologia , Proteína Quinase C-alfa/biossíntese , Animais , Hipóxia Celular/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Regulação da Expressão Gênica/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley
12.
J Neurosci ; 36(41): 10529-10544, 2016 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-27733606

RESUMO

Ischemic stroke is the leading cause of disability, but effective therapies are currently widely lacking. Recovery from stroke is very much dependent on the possibility to develop treatments able to both halt the neurodegenerative process as well as to foster adaptive tissue plasticity. Here we show that ischemic mice treated with neural precursor cell (NPC) transplantation had on neurophysiological analysis, early after treatment, reduced presynaptic release of glutamate within the ipsilesional corticospinal tract (CST), and an enhanced NMDA-mediated excitatory transmission in the contralesional CST. Concurrently, NPC-treated mice displayed a reduced CST degeneration, increased axonal rewiring, and augmented dendritic arborization, resulting in long-term functional amelioration persisting up to 60 d after ischemia. The enhanced functional and structural plasticity relied on the capacity of transplanted NPCs to localize in the peri-ischemic and ischemic area, to promote the upregulation of the glial glutamate transporter 1 (GLT-1) on astrocytes and to reduce peri-ischemic extracellular glutamate. The upregulation of GLT-1 induced by transplanted NPCs was found to rely on the secretion of VEGF by NPCs. Blocking VEGF during the first week after stroke reduced GLT-1 upregulation as well as long-term behavioral recovery in NPC-treated mice. Our results show that NPC transplantation, by modulating the excitatory-inhibitory balance and stroke microenvironment, is a promising therapy to ameliorate disability, to promote tissue recovery and plasticity processes after stroke. SIGNIFICANCE STATEMENT: Tissue damage and loss of function occurring after stroke can be constrained by fostering plasticity processes of the brain. Over the past years, stem cell transplantation for repair of the CNS has received increasing interest, although underlying mechanism remain elusive. We here show that neural stem/precursor cell transplantation after ischemic stroke is able to foster axonal rewiring and dendritic plasticity and to induce long-term functional recovery. The observed therapeutic effect of neural precursor cells seems to underlie their capacity to upregulate the glial glutamate transporter on astrocytes through the vascular endothelial growth factor inducing favorable changes in the electrical and molecular stroke microenvironment. Cell-based approaches able to influence plasticity seem particularly suited to favor poststroke recovery.


Assuntos
Astrócitos/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Células-Tronco Neurais/transplante , Transplante de Células-Tronco/métodos , Acidente Vascular Cerebral/terapia , Animais , Comportamento Animal , Isquemia Encefálica/metabolismo , Infarto Cerebral/patologia , Transportador 2 de Aminoácido Excitatório/genética , Ácido Glutâmico/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Plasticidade Neuronal , Técnicas de Patch-Clamp , Recuperação de Função Fisiológica , Acidente Vascular Cerebral/patologia , Acidente Vascular Cerebral/psicologia , Regulação para Cima , Fator A de Crescimento do Endotélio Vascular/metabolismo
13.
Hum Mol Genet ; 25(14): 2923-2933, 2016 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-27179791

RESUMO

Glutamatergic dysfunction has been implicated in the pathogenesis of depressive disorders and Huntington's disease (HD), in which depression is the most common psychiatric symptom. Synaptic glutamate homeostasis is regulated by cystine-dependent glutamate transporters, including GLT-1 and system xc- In HD, the enzyme regulating cysteine (and subsequently cystine) production, cystathionine-γ-lygase, has recently been shown to be lowered. The aim of the present study was to establish whether cysteine supplementation, using N-acetylcysteine (NAC) could ameliorate glutamate pathology through the cystine-dependent transporters, system xc- and GLT-1. We demonstrate that the R6/1 transgenic mouse model of HD has lower basal levels of cystine, and showed depressive-like behaviors in the forced-swim test. Administration of NAC reversed these behaviors. This effect was blocked by co-administration of the system xc- and GLT-1 inhibitors CPG and DHK, showing that glutamate transporter activity was required for the antidepressant effects of NAC. NAC was also able to specifically increase glutamate in HD mice, in a glutamate transporter-dependent manner. These in vivo changes reflect changes in glutamate transporter protein in HD mice and human HD post-mortem tissue. Furthermore, NAC was able to rescue changes in key glutamate receptor proteins related to excitotoxicity in HD, including NMDAR2B. Thus, we have shown that baseline reductions in cysteine underlie glutamatergic dysfunction and depressive-like behavior in HD and these changes can be rescued by treatment with NAC. These findings have implications for the development of new therapeutic approaches for depressive disorders.


Assuntos
Acetilcisteína/administração & dosagem , Depressão/tratamento farmacológico , Transportador 2 de Aminoácido Excitatório/genética , Doença de Huntington/tratamento farmacológico , Receptores de N-Metil-D-Aspartato/genética , Animais , Autopsia , Comportamento Animal/efeitos dos fármacos , Pareamento Cromossômico/efeitos dos fármacos , Pareamento Cromossômico/genética , Cistationina gama-Liase/biossíntese , Cistationina gama-Liase/genética , Cistina/biossíntese , Depressão/genética , Depressão/fisiopatologia , Modelos Animais de Doenças , Transportador 2 de Aminoácido Excitatório/biossíntese , Ácido Glutâmico/genética , Ácido Glutâmico/metabolismo , Humanos , Doença de Huntington/genética , Doença de Huntington/fisiopatologia , Camundongos , Camundongos Transgênicos
14.
J Appl Toxicol ; 36(11): 1409-17, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-26988466

RESUMO

The widespread environmental contaminant, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), is considered one of the most toxic dioxin-like compounds. Although epidemiological studies have shown that TCDD exposure is linked to some neurological and neurophysiological disorders, the underlying mechanism of TCDD-mediated neurotoxicity has remained unclear. Astrocytes are the most abundant cells in the nervous systems, and are recognized as the important mediators of normal brain functions as well as neurological, neurodevelopmental and neurodegenerative brain diseases. In this study, we investigated the role of TCDD in regulating the expression of glutamate transporter GLT-1 in astrocytes. TCDD, at concentrations of 0.1-100 nm, had no significantly harmful effect on the viability of C6 glioma cells. However, the expression of GLT-1 in C6 glioma cells was downregulated in a dose- and time-dependent manner. TCDD also caused activation of protein kinase C (PKC), as TCDD induced translocation of the PKC from the cytoplasm or perinuclear to the membrane. The translocation of PKC was inhibited by one Ca(2+) blocker, nifedipine, suggesting that the effects are triggered by the initial elevated intracellular concentration of free Ca(2+) . Finally, we showed that inhibition of the PKC activity reverses the TCDD-triggered reduction of GLT-1. In summary, our results suggested that TCDD exposure could downregulate the expression of GLT-1 in C6 via Ca(2+) /PKC pathway. The downregulation of GLT-1 might participate in TCDD-mediated neurotoxicity. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Astrócitos/efeitos dos fármacos , Cálcio/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Dibenzodioxinas Policloradas/toxicidade , Proteína Quinase C/metabolismo , Animais , Astrócitos/metabolismo , Sinalização do Cálcio , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Regulação para Baixo , Ratos , Transdução de Sinais , Fatores de Tempo
15.
Development ; 143(7): 1170-81, 2016 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-26893340

RESUMO

Astrocytes are crucial in the formation, fine-tuning, function and plasticity of neural circuits in the central nervous system. However, important questions remain about the mechanisms instructing astrocyte cell fate. We have studied astrogenesis in the ventral nerve cord of Drosophila larvae, where astrocytes exhibit remarkable morphological and molecular similarities to those in mammals. We reveal the births of larval astrocytes from a multipotent glial lineage, their allocation to reproducible positions, and their deployment of ramified arbors to cover specific neuropil territories to form a stereotyped astroglial map. Finally, we unraveled a molecular pathway for astrocyte differentiation in which the Ets protein Pointed and the Notch signaling pathway are required for astrogenesis; however, only Notch is sufficient to direct non-astrocytic progenitors toward astrocytic fate. We found that Prospero is a key effector of Notch in this process. Our data identify an instructive astrogenic program that acts as a binary switch to distinguish astrocytes from other glial cells.


Assuntos
Astrócitos/citologia , Proteínas de Ligação a DNA/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/embriologia , Proteínas do Tecido Nervoso/genética , Neurópilo/citologia , Proteínas Nucleares/genética , Proteínas Proto-Oncogênicas/genética , Receptores Notch/genética , Fatores de Transcrição/genética , Animais , Astrócitos/metabolismo , Linhagem da Célula/fisiologia , Sistema Nervoso Central/embriologia , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/biossíntese , Proteínas de Drosophila/metabolismo , Transportador 1 de Aminoácido Excitatório/antagonistas & inibidores , Transportador 1 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas do Tecido Nervoso/metabolismo , Neurogênese/fisiologia , Neuroglia/citologia , Proteínas Proto-Oncogênicas/metabolismo , Interferência de RNA , RNA Interferente Pequeno/genética , Receptores Notch/metabolismo , Fatores de Transcrição/metabolismo
16.
J Neurosci ; 36(6): 2027-43, 2016 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-26865625

RESUMO

UNLABELLED: Growth-associated protein 43 (GAP43), a protein kinase C (PKC)-activated phosphoprotein, is often implicated in axonal plasticity and regeneration. In this study, we found that GAP43 can be induced by the endotoxin lipopolysaccharide (LPS) in rat brain astrocytes both in vivo and in vitro. The LPS-induced astrocytic GAP43 expression was mediated by Toll-like receptor 4 and nuclear factor-κB (NF-κB)- and interleukin-6/signal transducer and activator of transcription 3 (STAT3)-dependent transcriptional activation. The overexpression of the PKC phosphorylation-mimicking GAP43(S41D) (constitutive active GAP43) in astrocytes mimicked LPS-induced process arborization and elongation, while application of a NF-κB inhibitory peptide TAT-NBD or GAP43(S41A) (dominant-negative GAP43) or knockdown of GAP43 all inhibited astrogliosis responses. Moreover, GAP43 knockdown aggravated astrogliosis-induced microglial activation and expression of proinflammatory cytokines. We also show that astrogliosis-conditioned medium from GAP43 knock-down astrocytes inhibited GAP43 phosphorylation and axonal growth, and increased neuronal damage in cultured rat cortical neurons. These proneurotoxic effects of astrocytic GAP43 knockdown were accompanied by attenuated glutamate uptake and expression of the glutamate transporter excitatory amino acid transporter 2 (EAAT2) in LPS-treated astrocytes. The regulation of EAAT2 expression involves actin polymerization-dependent activation of the transcriptional coactivator megakaryoblastic leukemia 1 (MKL1), which targets the serum response elements in the promoter of rat Slc1a2 gene encoding EAAT2. In sum, the present study suggests that astrocytic GAP43 mediates glial plasticity during astrogliosis, and provides beneficial effects for neuronal plasticity and survival and attenuation of microglial activation. SIGNIFICANCE STATEMENT: Astrogliosis is a complex state in which injury-stimulated astrocytes exert both protective and harmful effects on neuronal survival and plasticity. In this study, we demonstrated for the first time that growth-associated protein 43 (GAP43), a well known growth cone protein that promotes axonal regeneration, can be induced in rat brain astrocytes by the proinflammatory endotoxin lipopolysaccharide via both nuclear factor-κB and signal transducer and activator of transcription 3-mediated transcriptional activation. Importantly, LPS-induced GAP43 mediates plastic changes of astrocytes while attenuating astrogliosis-induced microglial activation and neurotoxicity. Hence, astrocytic GAP43 upregulation may serve to indicate beneficial astrogliosis after CNS injury.


Assuntos
Astrócitos/efeitos dos fármacos , Proteína GAP-43/biossíntese , Proteína GAP-43/genética , Gliose/genética , Microglia/efeitos dos fármacos , NF-kappa B/genética , Síndromes Neurotóxicas/genética , Síndromes Neurotóxicas/patologia , Fator de Transcrição STAT3/genética , Receptor 4 Toll-Like/genética , Animais , Citocinas/biossíntese , Transportador 2 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/genética , Ativação de Macrófagos/efeitos dos fármacos , Neurônios , Fosforilação , Ratos , Ratos Sprague-Dawley , Transativadores/biossíntese , Transativadores/genética
17.
Psychopharmacology (Berl) ; 233(3): 405-15, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26514555

RESUMO

RATIONALE: Growing evidence suggests that downregulated clearance of glutamate and signaling pathways involving brain-derived neurotrophic factor (BDNF) and its receptor TrkB play a role in morphological changes in the hippocampus of depressed patients. The N-methyl-D-aspartate (NMDA) receptor antagonist ketamine is the most attractive antidepressant, although precise mechanisms are unknown. OBJECTIVE: In this study, we examined whether hippocampal BDNF-TrkB signaling underlies the antidepressant effects of ketamine via upregulating glutamate transporter 1 (GLT-1) in rats, subjected to the chronic unpredictable stress (CUS) for 42 days. The rats received a single injection of ketamine (10 mg/kg, i.p.) and/or a TrkB inhibitor, K252a (1 µl, 2 mM, intracerebroventicular (i.c.v.)) on day 43. Behavioral tests and brain sample collection were evaluated 24 h later. RESULTS: The CUS-exposed rats exhibited depression- and anxiety-like behaviors; decreased number of glial fibrillary acidic protein (GFAP)-positive (but not NeuN-positive) cells in the dentate gyrus (DG), CA1, and CA3 areas; increased number of cleaved caspase-3-positive astrocytes; reduced spine density; lower ratio of Bcl2 to Bax; and decreased levels of BDNF, phosphorylated cAMP response element binging protein (CREB), GLT-1, and postsynaptic density 95 (PSD95) proteins in the hippocampus. Ketamine alleviated the CUS-induced abnormalities. The effects of ketamine were antagonized by pretreatment with K252a. CONCLUSIONS: Our findings suggest that regulation of GLT-1 on astrocytes, responsible for 90 % of glutamate reuptake from the synapse, through BDNF-TrkB signaling is involved in mediation of the therapeutic effects of ketamine on behavioral abnormalities and morphological changes in the hippocampus of the CUS-exposed rats.


Assuntos
Antidepressivos/farmacologia , Apoptose/efeitos dos fármacos , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Transtorno Depressivo/tratamento farmacológico , Antagonistas de Aminoácidos Excitatórios/farmacologia , Transportador 2 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/genética , Ketamina/farmacologia , Receptor trkB/metabolismo , Estresse Psicológico/metabolismo , Estresse Psicológico/psicologia , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Comportamento Animal/efeitos dos fármacos , Carbazóis/farmacologia , Doença Crônica , Transtorno Depressivo/psicologia , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Alcaloides Indólicos/farmacologia , Injeções Intraventriculares , Masculino , Ratos , Ratos Sprague-Dawley , Receptor trkB/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos
18.
PLoS One ; 10(8): e0136111, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26301411

RESUMO

Glutamate is the major excitatory neurotransmitter, but prolonged exposure even at micromolar concentrations causes neuronal death. Extracellular glutamate is maintained at nanomolar level by glutamate transporters, which, however, may reverse transport and release glutamate. If and when the reverse occurs depends on glutamate transport stoichiometry (GTS). Previously we found that in the presence of chloride, the coupled GLT-1 glutamate transporter current and its relationship to radiolabeled glutamate flux significantly decreased when extracellular glutamate concentration increased above 0.2 mM, which implies a change in GTS. Such high concentrations are feasible near GLT-1 expressed close to synaptic release site during excitatory neurotransmission. The aim of this study was to determine GLT-1 GTS at both low (19-75 µM) and high (300-1200 µM) glutamate concentration ranges. GTS experiments were conducted in the absence of chloride to avoid contributions by the GLT-1 uncoupled chloride conductance. Mathematical analysis of the transporter thermodynamic equilibrium allowed us to derive equations revealing the number of a particular type of ion transported per elementary charge based on the measurements of the transporter reversal potential. We found that GLT-1a expressed in COS-7 cells co-transports 1.5 Na+, 0.5 Glu-, 0.5 H+ and counter-transports 0.6 K+ per elementary charge in both glutamate concentration ranges, and at both 37°C and 26°C temperatures. The thermodynamic parameter Q10 = 2.4 for GLT-1 turnover rate of 19 s-1 (37°C, -50 mV) remained constant in the 10 µM-10 mM glutamate concentration range. Importantly, the previously reported decrease in the current/flux ratio at high glutamate concentration was not seen in the absence of chloride in both COS-7 cells and cultured rat neurons. Therefore, only in the absence of chloride, GLT-1 GTS remains constant at all glutamate concentrations. Possible explanations for why apparent GTS might vary in the presence of chloride are discussed.


Assuntos
Transportador 2 de Aminoácido Excitatório/genética , Ácido Glutâmico/metabolismo , Neurônios/metabolismo , Transmissão Sináptica/genética , Animais , Células COS , Cloretos/metabolismo , Chlorocebus aethiops , Transportador 2 de Aminoácido Excitatório/biossíntese , Gluconatos/metabolismo , Ácido Glutâmico/química , Neurônios/patologia , Técnicas de Patch-Clamp , Ratos , Sódio/metabolismo
19.
Psychopharmacology (Berl) ; 232(13): 2333-42, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25619881

RESUMO

RATIONALE: Several studies have demonstrated a correlation between extracellular glutamate concentration in the mesolimbic reward pathway and alcohol craving. Extracellular glutamate concentration is regulated by several glutamate transporters. Glial glutamate transporter 1 (GLT1) is one of them that regulates the majority of extracellular glutamate concentration. In addition, cystine/glutamate antiporter (xCT) is another transporter that regulates extracellular glutamate. OBJECTIVES: We focus in this study to determine the effects of ceftriaxone, ß-lactam antibiotic, on glial proteins such as GLT1 isoforms, xCT, glutamate aspartate transporter (GLAST), and several associated signaling pathways as well as ethanol intake in P rats. Additionally, to examine the onset of signaling pathways associated with GLT1 upregulation following ceftriaxone treatment, we tested 2- versus 5-day daily dosing of ceftriaxone. RESULTS: Ceftriaxone treatment (100 mg/kg), 2 and 5 days, resulted in about five fold reduction in ethanol intake by P rats. The reduction in ethanol intake was associated with significantly enhanced expression of GLT1, GLT1a, GLT1b, and xCT in the nucleus accumbens (NAc) and prefrontal cortex (PFC) of 5-day ceftriaxone-treated P rats. Two-day-treated P rats showed marked changes in expression of these glutamate transporters in the PFC but not in the NAc. Importantly, ceftriaxone-treated P rats (2 and 5 days) demonstrated enhanced phosphorylation of Akt and nuclear translocation of nuclear factor kappaB (NFκB) in the NAc and PFC compared to control animals. CONCLUSIONS: These findings demonstrate that ceftriaxone treatment induced upregulation of GLT1, GLT1 isoforms, and xCT in association with activation of the Akt-NFκB signaling pathway.


Assuntos
Consumo de Bebidas Alcoólicas/tratamento farmacológico , Sistemas de Transporte de Aminoácidos Acídicos/biossíntese , Ceftriaxona/uso terapêutico , Etanol/administração & dosagem , Transportador 2 de Aminoácido Excitatório/biossíntese , Transdução de Sinais/efeitos dos fármacos , Consumo de Bebidas Alcoólicas/metabolismo , Animais , Ceftriaxona/farmacologia , Masculino , Núcleo Accumbens/efeitos dos fármacos , Núcleo Accumbens/metabolismo , Córtex Pré-Frontal/efeitos dos fármacos , Isoformas de Proteínas/biossíntese , Ratos , Transdução de Sinais/fisiologia , Resultado do Tratamento
20.
Neurochem Int ; 79: 33-43, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25454285

RESUMO

The glutamate transporters GLAST and GLT-1 are mainly expressed in glial cells and regulate glutamate levels in the synapses. GLAST and GLT-1 are the targets of several signaling pathways. In this study we explore the possible functional interaction between these transporters and GSK3ß. This kinase is involved in multiple cellular processes including neuronal development and synaptic plasticity. To evaluate whether GLT-1 and GLAST were regulated by GSK3ß, we coexpressed these proteins in heterologous expression systems. In both COS-7 cells and Xenopus laevis oocytes, GSK3ß stimulated the activity of GLT-1 and reduced that of GLAST. These effects were associated with corresponding changes in the amounts of GLT-1 or GLAST in the plasma membrane. These effects were suppressed by inhibitors of GSK3ß or a catalytically inactive form of the kinase. GSK3ß also decreases the incorporation of (32)Pi into GLT-1 and increases GLAST phosphorylation. Pharmacological inhibition of endogenous GSK3ß in primary cultures of rat brain cortex also leads to a differential modulation of GLT-1 and GLAST. Our results suggest that constitutively active GSK3ß is important in controlling the expression of functional glutamate transporters on the plasma membrane. This regulation might be relevant in physiological and pathological conditions in which glutamate transporters and GSK3ß signaling are involved.


Assuntos
Transportador 1 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/biossíntese , Quinase 3 da Glicogênio Sintase/metabolismo , Animais , Biotinilação , Células COS , Chlorocebus aethiops , Transportador 1 de Aminoácido Excitatório/genética , Transportador 2 de Aminoácido Excitatório/genética , Regulação da Expressão Gênica/fisiologia , Ácido Glutâmico/metabolismo , Glicogênio Sintase Quinase 3 beta , Humanos , Masculino , Oócitos/metabolismo , Técnicas de Patch-Clamp , Cultura Primária de Células , Ratos , Xenopus
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA