RESUMO
Alzheimer's disease (AD) is the leading cause of dementia. Non-competitive N-Methyl-D-aspartate (NMDA) receptor antagonist memantine improved cognition and molecular alterations after preclinical treatment. Nevertheless, clinical results are discouraging. In vivo efficacy of the RL-208, a new NMDA receptor blocker described recently, with favourable pharmacokinetic properties was evaluated in Senescence accelerated mice prone 8 (SAMP8), a mice model of late-onset AD (LOAD). Oral administration of RL-208 improved cognitive performance assessed by using the three chamber test (TCT), novel object recognition test (NORT), and object location test (OLT). Consistent with behavioural results, RL-208 treated-mice groups significantly changed NMDAR2B phosphorylation state levels but not NMDAR2A. Calpain-1 and Caspase-3 activity was reduced, whereas B-cell lymphoma-2 (BCL-2) levels increased, indicating reduced apoptosis in RL-208 treated SAMP8. Superoxide Dismutase 1 (SOD1) and Glutathione Peroxidase 1 (GPX1), as well as a reduction of hydrogen peroxide (H2O2), was also determined in RL-208 mice. RL-208 treatment induced an increase in mature brain-derived neurotrophic factor (mBDNF), prevented Tropomyosin-related kinase B full-length (TrkB-FL) cleavage, increased protein levels of Synaptophysin (SYN) and Postsynaptic density protein 95 (PSD95). In whole, these results point out to an improvement in synaptic plasticity. Remarkably, RL-208 also decreased the protein levels of Cyclin-Dependent Kinase 5 (CDK5), as well as p25/p35 ratio, indicating a reduction in kinase activity of CDK5/p25 complex. Consequently, lower levels of hyperphosphorylated Tau (p-Tau) were found. In sum, these results demonstrate the neuroprotectant role of RL-208 through NMDAR blockade.
RESUMO
Resveratrol is a polyphenol that is mainly found in grapes and red wine and has been reported to be a caloric restriction (CR) mimetic driven by Sirtuin 1 (SIRT1) activation. Resveratrol increases metabolic rate, insulin sensitivity, mitochondrial biogenesis and physical endurance, and reduces fat accumulation in mice. In addition, resveratrol may be a powerful agent to prevent age-associated neurodegeneration and to improve cognitive deficits in Alzheimer's disease (AD). Moreover, different findings support the view that longevity in mice could be promoted by CR. In this study, we examined the role of dietary resveratrol in SAMP8 mice, a model of age-related AD. We found that resveratrol supplements increased mean life expectancy and maximal life span in SAMP8 and in their control, the related strain SAMR1. In addition, we examined the resveratrol-mediated neuroprotective effects on several specific hallmarks of AD. We found that long-term dietary resveratrol activates AMPK pathways and pro-survival routes such as SIRT1 in vivo. It also reduces cognitive impairment and has a neuroprotective role, decreasing the amyloid burden and reducing tau hyperphosphorylation.
Assuntos
Envelhecimento , Doença de Alzheimer/dietoterapia , Biomarcadores/metabolismo , Restrição Calórica , Suplementos Nutricionais , Longevidade/efeitos dos fármacos , Estilbenos/administração & dosagem , Doença de Alzheimer/metabolismo , Doença de Alzheimer/fisiopatologia , Animais , Western Blotting , Cognição/efeitos dos fármacos , Modelos Animais de Doenças , Inibidores Enzimáticos/administração & dosagem , Imuno-Histoquímica , Camundongos , Resveratrol , Ribonucleotídeo Redutases/antagonistas & inibidoresRESUMO
OBJECTIVES: We investigated the neuroprotective effects of lithium in an experimental neurodegeneration model gated to kainate (KA) receptor activation. METHODS: The hippocampus from KA-treated mice and hippocampal cell cultures were used to evaluate the pathways regulated by chronic lithium pretreatment in both in vivo and in vitro models. RESULTS: Treatment with KA, as measured by fragmentation of alpha-spectrin and biochemically, induced the activation of calpain resulting in p35 cleavage to p25, indicating activation of cyclin-dependent kinase 5 (cdk5) and glycogen synthase kinase-3ss (GSK-3ss) and an increase in tau protein phosphorylation. Treatment with lithium reduced calpain activation and reduced the effects of cdk5 and GSK-3ss on tau. KA treatment of cultures resulted in neuronal demise. According to nuclear condensed cell counts, the addition of lithium to neuronal cell cultures (0.5-1 mM) a few days before KA treatment had neuroprotective and also antiapoptotic effects. The action of lithium on calpain/cdk5 and GSK-3ss pathways produced similar results in vivo. As calpain is activated by an increase in intracellular calcium, we showed that lithium reduced calcium concentrations in basal and KA-treated hippocampal cells, which was accompanied by an increase in NCX3, a Na+/Ca2+ exchanger pump. CONCLUSION: A robust neuroprotective effect of lithium in the excitotoxic process induced by KA in mouse hippocampus was demonstrated via modulation of calcium entry and the subsequent inhibition of the calpain pathway. These mechanisms may act in an additive way with other mechanisms previously described for lithium, suggesting that it may be useful as a possible therapeutic strategy for Alzheimer's disease.
Assuntos
Calpaína/metabolismo , Hipocampo , Cloreto de Lítio/farmacologia , Doenças Neurodegenerativas/tratamento farmacológico , Fármacos Neuroprotetores/farmacologia , Animais , Cálcio/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Quinase 5 Dependente de Ciclina/metabolismo , Modelos Animais de Doenças , Quinases da Glicogênio Sintase/metabolismo , Hipocampo/metabolismo , Hipocampo/patologia , Ácido Caínico/toxicidade , Masculino , Camundongos , Camundongos Endogâmicos , Doenças Neurodegenerativas/induzido quimicamente , Fosforilação , Proteínas tau/metabolismoRESUMO
Lithium is a simple cation that has been used clinically since 1950 for the treatment of bipolar disorder. However in the last decade numerous studies either using animal models or human trials suggest that this cation may delay progression of neurodegenerative diseases. One of the main challenges facing researchers in the neurosciences is to identify key molecules in neuronal apoptosis. This would facilitate the identification of targets in order to design drugs for the treatment of Alzheimer's disease, Parkinson's disease and other neurological disorders. Although enormous effort has been made in the past few years and it has been demonstrated that the mitochondria comprise a key component of the neuronal apoptotic route, it seems that in addition to the mitochondria other intracellular components are implicated in this process. It has been proposed that DNA damage and re-entry into the cell cycle or the activation of different proteases, such as calpain, could constitute a common pathway in the apoptotic process and thus death processes in neurological diseases. The hypothesis about the implication of calpain in neuronal cell death is supported by existing data on neurodegenerative disorders in the brains of patients who show an increase in proteolytic activity of calpain compared with control brains. Indeed, studies performed in neuronal cell preparations suggest that activation of this protease is accompanied by other features such as structural modifications of the cytoskeleton, cleavage of several receptors, activation of kinases, such as cdk5 or GSK3ss, etc. Here, we summarize the potential routes involved in neurodegenerative disorders related to calpain activation, mainly those connected with changes in calcium homeostasis machinery, activation of kinase pathways, transcription factors, and the cell cycle.
Assuntos
Calpaína/metabolismo , Sistemas de Liberação de Medicamentos , Doenças Neurodegenerativas/tratamento farmacológico , Animais , Antimaníacos/farmacologia , Antimaníacos/uso terapêutico , Apoptose , Ensaios Clínicos como Assunto , Humanos , Compostos de Lítio/farmacologia , Compostos de Lítio/uso terapêutico , Doenças Neurodegenerativas/fisiopatologia , Neurônios/metabolismoRESUMO
3-Nitropropionic acid (3-NP) is a neurotoxin that inhibits mitochondrial complex II and is used in an experimental model of Huntington's disease. Treatment of rats with 3-NP 30mgkg(-1) i.p. once a day for 5 days induced an increase in calpain activation in rat striatum, measured by the formation of 145kDa fragment of alpha-spectrin breakdown and by an increase in enzymatic calpain activity. In this neurotoxic model, Western blot studies revealed that calpain activity increase was followed by changes in cyclin-dependent kinase 5 (cdk5) and its activator p25. Our results indicated, after 10 days of treatment with 3-NP, a decrease in myocyte enhancer factor phosphorylation, a neuronal prosurvival factor. Thus, a decrease in its expression indicates a new potential mechanism of neuronal cell death mediated by the neurotoxin 3-NP. Accordingly, in our study we demonstrated in rat striatum the activation of the calpain/cdk5/p25 pathway in the 3-NP model. Previous studies have linked the deregulation of cdk5 with neurodegenerative diseases, such as Alzheimer's and Parkinson's. We suggest that calpain/cdk5 activation could also be a common pathway activated in other neurodegenerative diseases, which is liable to be targeted.
Assuntos
Calpaína/metabolismo , Convulsivantes/farmacologia , Corpo Estriado/efeitos dos fármacos , Quinase 5 Dependente de Ciclina/metabolismo , Nitrocompostos/farmacologia , Propionatos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Análise de Variância , Animais , Ativação Enzimática/efeitos dos fármacos , Masculino , Ratos , Ratos Sprague-Dawley , Fatores de TempoRESUMO
Certain effects of melatonin on senescence were investigated. The experimental model used was 10-month-old senescence-accelerated mouse prone 8 (SAMP8). The mice in the experiment were administered melatonin (10 mg/kg) from the age of 1 month. Results showed that chronic administration of melatonin decreased cell loss in the cerebral cortex and reduced oxidative damage in protein and lipids. There are several studies suggesting that the activation of the cdk5/p35 pathway at its cleavage to cdk5/p25 may play a role in hyperphosphorylation of tau during aging and neurodegenerative diseases. Melatonin not only reduced the cerebral aging disturbances, but also prevented tau hyperphosphorylation present in the experimental model used in this study. Melatonin reduced cdk5 expression, as well as the cleavage of p35 to p25. The other tau kinase studied, GSK3beta, showed a reduction in this activity in comparison with SAMP8 nontreated SAMP8. These data indicate that melatonin possesses neuroprotective properties against cerebral damage gated to senescence. Moreover, these data suggest that the cdk5/GSKbeta signaling cascade has a potential role as a target for neurodegenerative diseases related to aging.
Assuntos
Lesão Encefálica Crônica/tratamento farmacológico , Lesão Encefálica Crônica/metabolismo , Melatonina/administração & dosagem , Fármacos Neuroprotetores/administração & dosagem , Animais , Biomarcadores/metabolismo , Lesão Encefálica Crônica/patologia , Catalase/metabolismo , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Córtex Cerebral/patologia , Quinase 5 Dependente de Ciclina/metabolismo , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Mutantes Neurológicos , Proteínas do Tecido Nervoso/metabolismo , Fosforilação , Fosfotransferases/metabolismoRESUMO
Tau is a neuronal microtubule-associated protein found predominantly on axons. Tau phosphorylation regulates both normal and pathological functions of this protein. Hyperphosphorylation impairs the microtubule binding function of tau, resulting in the destabilization of microtubules in brain, ultimately leading to the degeneration of the affected neurons. Numerous serine/threonine kinases, including GSK-3beta and Cdk5 can phosphorylate tau. SAMR1 and SAMP8 are murine strains of senescence. We show an increase in hyperphosphorylated forms of tau in SAMP8 (senescent mice) in comparison with resistant strain SAMR1. Moreover, an increase in Cdk5 expression and activation is described but analysis of GSK3beta isoforms failed to show differences in SAMP8 in comparison to age-matched SAMR1. In conclusion, tau hyperphosphorylation occurs in SAMP-8 (early senescent) mice, indicating a link between aging and tau modifications in this murine model.
Assuntos
Microtúbulos/metabolismo , Proteínas tau/metabolismo , Envelhecimento , Animais , Axônios/metabolismo , Western Blotting , Encéfalo/metabolismo , Encéfalo/patologia , Membrana Celular/metabolismo , Senescência Celular , Quinase 5 Dependente de Ciclina/metabolismo , Camundongos , Camundongos Transgênicos , Neurônios/metabolismo , Fosforilação , Isoformas de Proteínas , Distribuição Tecidual , Tubulina (Proteína)/metabolismo , Proteínas tau/químicaRESUMO
Injury to the central nervous system causes atrophy or death of connecting neurons and can modify the expression of neurotrophic factors. We observed transneuronal upregulation of brain-derived neurotrophic factor (BDNF) expression in the rat ipsilateral substantia nigra pars compacta after a striatal lesion induced by kainate. This effect is developmentally regulated because the enhancement of nigral BDNF expression was only observed when striatal lesion was performed on postnatal day (P) 15 and in adulthood, but not at P7. Interestingly, the lack of regulation of BDNF was coincident with the transynaptic degeneration of nigral neurons after striatal excitotoxic injury. Hence, the number of tyrosine hydroxylase-positive neurons in the substantia nigra pars compacta decreased when the lesion was performed at P7, but not at P15 or at P30. The analysis of the functional significance of this BDNF upregulation was done using trkB-IgG fusion proteins. After striatal injury, blockade of endogenous BDNF by trkB fusion proteins induced an atrophy of the dopaminergic neurons of the pars compacta. The injection of trkB-IgG fusion proteins did not modify the effects of kainate in the substantia nigra pars reticulata. Thus, our results show that BDNF exerts an autocrine/paracrine protective effect selectively on dopaminergic neurons against the loss of trophic support from the target striatum.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Corpo Estriado/fisiopatologia , Dopamina/metabolismo , Degeneração Neural/prevenção & controle , Neurônios/metabolismo , Substância Negra/citologia , Fatores Etários , Análise de Variância , Animais , Animais Recém-Nascidos , Northern Blotting/métodos , Lesões Encefálicas/induzido quimicamente , Lesões Encefálicas/complicações , Lesões Encefálicas/prevenção & controle , Contagem de Células , Corpo Estriado/efeitos dos fármacos , Diagnóstico por Imagem/métodos , Ensaio de Desvio de Mobilidade Eletroforética/métodos , Lateralidade Funcional/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Imunoglobulina G/farmacologia , Imuno-Histoquímica/métodos , Hibridização In Situ/métodos , Ácido Caínico , Masculino , Degeneração Neural/etiologia , Neurônios/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Receptor trkB/imunologia , Fatores de Tempo , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
The neurotoxic action of the abuse drugs methamphetamine (METH) and 3,4-methylenedioxymethamphetamine (MDMA) on cerebellar granule neurones (CGNs) culture was examined. Treatment for 48 h with METH or MDMA (1-5 mM) induced a higher decrease in viability than 24 h treatment. z.VAD.fmk (100 microM) but not MK-801 nor NBQX recovered control viability values. In both cases, cell death was characterised as apoptotic rather than necrotic by morphology cell observation. Apoptosis measured by flow cytometry indicated an increase in the hypodiploid population after 48 h treatment with METH and MDMA. Apoptosis was reverted by the presence of z.VAD.fmk (100 microM) but not by 10 microM MK-801 or NBQX. Similar results were obtained by analysing nuclear chromatine condensation. These results ruled out excitotoxic participation in amphetamine derivative-induced neurotoxicity in CGNs. Participation of radical oxygen species (ROS) was evaluated using alpha-tocopherol (1-15 microM) and cytometric studies. The co-treatment with 4 mM METH or MDMA for 48 h partially reverted neurotoxic action and apoptotic features, indicating ROS implication in CGNs death by amphetamine derivatives. Alteration of mitochondrial function induced cytochrome C (Cyt C) release after 48-h treatment with METH and MDMA (4 mM). There was also indication of caspase-3-like activation, measured by immunoanalysis and biochemically. Finally, neurodegenerative action caused by amphetamine derivatives may be prevented by using caspase inhibitors.
Assuntos
Clorometilcetonas de Aminoácidos/farmacologia , Caspases/metabolismo , Cerebelo/efeitos dos fármacos , Cerebelo/enzimologia , Metanfetamina/antagonistas & inibidores , Metanfetamina/toxicidade , N-Metil-3,4-Metilenodioxianfetamina/antagonistas & inibidores , N-Metil-3,4-Metilenodioxianfetamina/toxicidade , Fármacos Neuroprotetores/farmacologia , Animais , Apoptose/efeitos dos fármacos , Caspase 3 , Cerebelo/citologia , Cerebelo/metabolismo , Citocromos c/metabolismo , Maleato de Dizocilpina/farmacologia , Interações Medicamentosas , Ativação Enzimática , Citometria de Fluxo , Microscopia de Fluorescência , Microscopia de Contraste de Fase , Mitocôndrias/enzimologia , Mitocôndrias/metabolismo , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Neurônios/metabolismo , Quinoxalinas/farmacologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , alfa-Tocoferol/farmacologiaRESUMO
Kainic acid (KA)-induced neuronal damage and the protective effects of flavopiridol were studied in primary cultures of rat cerebellar granule cells (CGNs). When neurons were treated with 500 microM KA, the percentage of cells with condensed nuclei measured by nuclear counting increased by up to 55%. After flavopiridol treatment, an antitumoral drug that is a broad inhibitor of cyclin-dependent kinases, the percentage of condensed nuclei decreased by up to 26%. Furthermore, this KA-mediated cell death was only partially dependent on the activation of the initiator caspase-9 and the effector caspases-3 and -6. This argues for a minor role of caspases in the intracellular pathway leading to KA-induced programmed cell death in CGNs. We examined the possible implication of cell cycle proteins in KA-induced neurotoxicity. We found an increase in the expression of proliferating cell nuclear antigen and E2F-1, two proteins implicated in S-phase, by Western blot. KA increased bromodeoxyuridine incorporation in CGNs, a marker of cell proliferation, and flavopiridol attenuated this effect. These results indicated that flavopiridol decreased the expression of cell cycle markers in CGNs after KA treatment. Flavopiridol might thus be used as a preventive agent against neurodegenerative diseases associated with cell cycle activation.
Assuntos
Proteínas de Ciclo Celular , Proteínas de Ligação a DNA , Flavonoides/farmacologia , Ácido Caínico/farmacologia , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Piperidinas/farmacologia , Animais , Bromodesoxiuridina , Caspases/metabolismo , Ciclo Celular , Núcleo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Cerebelo/citologia , Interações Medicamentosas , Fatores de Transcrição E2F , Fator de Transcrição E2F1 , Ativação Enzimática , Inibidores Enzimáticos/farmacologia , Agonistas de Aminoácidos Excitatórios , Neurônios/citologia , Antígeno Nuclear de Célula em Proliferação/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de AMPA/metabolismo , Fatores de Transcrição/metabolismo , Regulação para CimaRESUMO
Neuronal apoptosis may be partly due to inappropriate control of the cell cycle. We used serum deprivation as stimulus and reduced potassium from 25 to 5mM (S/K deprivation), which induces apoptosis in cerebellar granule neurons (CGNs), to evaluate the direct correlation between re-entry in the cell cycle and apoptosis. Roscovitine (10 microM), an antitumoral drug that inhibits cyclin-dependent kinase 1 (cdk1), cdk2 and cdk5, showed a significant neuroprotective effect on CGNs deprived of S/K. S/K deprivation induced the expression of cell cycle proteins such as cyclin E, cyclin A, cdk2, cdk4 and E2F-1. It also caused CGNs to enter the S phase of the cell cycle, measured by a significant incorporation of BrdU (30% increase over control cells), which was reduced in the presence of roscovitine (10 microM). On the other hand, roscovitine modified the expression of cytochrome c (Cyt c), Bcl-2 and Bax, which are involved in the apoptotic intrinsic pathway induced by S/K deprivation. We suggest that the antiapoptotic effects of roscovitine on CGNs are due to its anti-proliferative efficacy and to an action on the mitochondrial apoptotic mechanism.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Ciclo Celular , Cerebelo/efeitos dos fármacos , Potássio/administração & dosagem , Purinas/farmacologia , Animais , Bromodesoxiuridina , Caspase 3 , Inibidores de Caspase , Caspases/metabolismo , Cerebelo/citologia , Meios de Cultura Livres de Soro , Ativação Enzimática , Imuno-Histoquímica , Ratos , Ratos Sprague-Dawley , RoscovitinaRESUMO
Flavopiridol was developed as a drug for cancer therapy due to its ability to inhibit cell cycle progression by targeting cyclin-dependent kinases (CDKs). In this study, we show that flavopiridol may also have a neuroprotective action. We show that at therapeutic dosage (or at micromolar range), flavopiridol almost completely prevents colchicine-induced apoptosis in cerebellar granule neurones. In agreement with this, flavopiridol inhibits both the release of cyt c and the activation of caspase-3 induced in response to colchicine treatment. We demonstrate that in this cellular model for neurotoxicity, neither re-entry in the cell cycle nor activation of stress-activated protein kinases, such as c-Jun N-terminal kinase (JNK) or p38 MAP kinase, is involved. In contrast, we show that colchicine-induced apoptosis correlates with a substantial increase in the expression of cdk5 and Par-4, which is efficiently prevented by flavopiridol. Accordingly, a cdk5 inhibitor such as roscovitine, but not a cdk4 inhibitor such as 3-ATA, was also able to protect neurons from apoptosis as well as prevent accumulation of cdk5 and Par-4 in response to colchicine. Our data suggest a potential therapeutic use of flavopiridol in disorders of the central nervous system in which cytoskeleton alteration mediated by cdk5 activation and Par-4 expression has been demonstrated, such as Alzheimer's disease.
Assuntos
Apoptose , Colchicina/farmacologia , Flavonoides/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular , Proteínas Quinases JNK Ativadas por Mitógeno , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Piperidinas/farmacologia , Clorometilcetonas de Aminoácidos/farmacologia , Animais , Animais Recém-Nascidos , Antracenos/farmacologia , Antibacterianos/farmacologia , Proteínas Reguladoras de Apoptose , Western Blotting , Bromodesoxiuridina/metabolismo , Quinases relacionadas a CDC2 e CDC28/metabolismo , Proteínas de Transporte/metabolismo , Caspase 3 , Caspases/metabolismo , Contagem de Células , Sobrevivência Celular , Células Cultivadas , Cerebelo/citologia , Cerebelo/efeitos dos fármacos , Cerebelo/fisiologia , Cromatina/metabolismo , Ciclina E/metabolismo , Quinase 2 Dependente de Ciclina , Quinase 5 Dependente de Ciclina , Quinases Ciclina-Dependentes/metabolismo , Citocromos c/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Agonistas de Aminoácidos Excitatórios/farmacologia , Citometria de Fluxo , Imuno-Histoquímica , Ácido Caínico/farmacologia , MAP Quinase Quinase 4 , Microtúbulos/metabolismo , Minociclina/farmacologia , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Neurônios/fisiologia , Purinas/farmacologia , Ratos , Ratos Sprague-Dawley , Roscovitina , Fatores de Tempo , Tubulina (Proteína)/metabolismoRESUMO
We have investigated the neuroprotective effects of (+/-)-huprine Y on excitotoxic lesions in rat cerebellar granule cells (CGCs). (+/-)-Huprine Y prevented cell death induced by 100 microM glutamate, as well as, 10 microM MK-801, a NMDA receptor antagonist, in a significant manner. On the other hand, intracellular calcium increase induced by NMDA (200 microM), measured by fura-2 fluorescence, was prevented by (+/-)-huprine Y with an EC(50) of 12.44 microM, which evidences the modulatory action of this compound on NMDA-induced calcium currents. In vivo, we have studied (+/-)-huprine Y neuroprotective effects on striatal lesions induced by the subacute administration of the mitochondrial toxin 3-nitropropionic acid (3-NP, 30 mg/kg, ip, for 10 days). We have assessed that both the behavioral and the morphological consequences of the lesion were prevented by pretreatment with (+/-)-huprine Y (2.5 mg/kg/twice a day, ip). Striatal gliosis induced by 3-NP treatment was prevented by (+/-)-huprine Y pretreatment, as demonstrated by the attenuation of both the increase in [(3)H]PK 11195 specific binding indicative of microgliosis and the expression of hsp27 kDa, a chaperone expressed mainly in astrocytes. In conclusion, (+/-)-huprine Y attenuated excitotoxic-induced lesions, both in vitro and in vivo, and further evidence is provided for the potential use of this compound in the prevention of neurodegenerative disorders.
Assuntos
Aminoquinolinas/farmacologia , Proteínas de Choque Térmico , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Doenças Neurodegenerativas/prevenção & controle , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Neurotoxinas/toxicidade , Animais , Ligação Competitiva/efeitos dos fármacos , Cálcio/metabolismo , Morte Celular/efeitos dos fármacos , Células Cultivadas , Cerebelo/citologia , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/patologia , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Feminino , Gliose/induzido quimicamente , Gliose/prevenção & controle , Ácido Glutâmico/toxicidade , Proteínas de Choque Térmico HSP27 , Isoquinolinas/farmacocinética , Camundongos , Chaperonas Moleculares , Atividade Motora/efeitos dos fármacos , N-Metilaspartato/farmacologia , Proteínas de Neoplasias/biossíntese , Doenças Neurodegenerativas/induzido quimicamente , Doenças Neurodegenerativas/patologia , Neurônios/citologia , Nitrocompostos , Propionatos , RatosRESUMO
The consequences of the neurotoxic insult induced by 3,4-methylenedioxymethamphetamine (MDMA, an amphetamine derivative with specific action on the serotonergic system) were compared with those of methamphetamine (a derivative with specific action on dopaminergic system) in rats. Both drugs induced a very similar loss of body weight, especially evident 24 h after treatment. Their hyperthermic profile was also very similar and was dependent on ambient temperature, corroborating the thermo-dysregulatory effect of both substances. Methamphetamine (four injections of 10 mg kg(-1) s.c. at 2-h intervals) induced the loss of dopaminergic (35%) but not of serotonergic, terminals in the rat striatum and, simultaneously, a significant increase in striatal peripheral-type benzodiazepine receptor density, pointing to a glial reaction. Evidence for this drug-induced astrogliosis was the increased heat shock protein 27 (HSP27) expression in striatum, cortex and hippocampus. MDMA (20 mg kg(-1) s.c. b.i.d. for 4 days) induced a similar dopaminergic lesion in the striatum 3 days post-treatment, which reversed 4 days later. An important neurotoxic effect on serotonergic terminals was also observed in the cortex, striatum and hippocampus 3 days post-treatment, which partially reversed 4 days later in the striatum and hippocampus. No microglial activation was noticeable at either 3 or 7 days after MDMA treatment. This lack of effect on microglial cells was assessed by [(3)H]PK 11195 binding and OX-6 immunostaining, which were unchanged in the striatum and cortex after MDMA treatment. A non-significant tendency to increase was noted in the hippocampus 3 days after MDMA treatment. Furthermore, in MDMA-treated rats, neither HSP27 expression nor an increase in HSP27 immunoreactivity were detected. This result, together with the lack of increase in glial fibrilliary acidic protein (GFAP) immunoreactivity, indicate no astroglial activation at either 3 or 7 days post-treatment. Without microglial activation, an inflammatory process would not accompany the lesion induced by MDMA. The differences in glial activation between methamphetamine and MDMA observed in the present study could have implications for the prognosis of the injury induced by these drugs.
Assuntos
Doenças do Sistema Nervoso Central/induzido quimicamente , Glicoproteínas de Membrana , Metanfetamina/toxicidade , N-Metil-3,4-Metilenodioxianfetamina/toxicidade , Proteínas do Tecido Nervoso , Neuroglia/efeitos dos fármacos , Animais , Sítios de Ligação/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/ultraestrutura , Proteínas de Transporte/efeitos dos fármacos , Proteínas de Transporte/metabolismo , Dopamina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Dopamina , Esquema de Medicação , Febre/induzido quimicamente , Proteína Glial Fibrilar Ácida/efeitos dos fármacos , Proteína Glial Fibrilar Ácida/ultraestrutura , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/imunologia , Masculino , Proteínas de Membrana Transportadoras/efeitos dos fármacos , Proteínas de Membrana Transportadoras/metabolismo , Metanfetamina/administração & dosagem , Metanfetamina/metabolismo , N-Metil-3,4-Metilenodioxianfetamina/administração & dosagem , N-Metil-3,4-Metilenodioxianfetamina/metabolismo , Neuroglia/patologia , Neuroglia/ultraestrutura , Ratos , Ratos Sprague-Dawley , Serotonina/metabolismo , Fatores de TempoRESUMO
Stimulation of ionotropic glutamate receptors are implicated in neurodegenerative diseases such as Alzheimer's and Parkinson's diseases. Recently this has been demonstrated in the expression of cell cycle proteins in vulnerable neurons in Alzheimer's disease. Thus, the aim of the present study was to evaluate the expression of cell cycle proteins in cerebellar granule cells after stimulation of AMPA/KA receptors and likewise to study the neuroprotective effects of CDK inhibitors. Our results demonstrated that after a treatment with CDK inhibitors, a significant decrease in apoptotic nuclei induced by kainic acid was found in the presence of flavopiridol and 3-ATA. We concluded that CDK activation is involved, at least, in part, in the pro-apoptotic effects of kainic acid.
Assuntos
Apoptose/efeitos dos fármacos , Quinases Ciclina-Dependentes/antagonistas & inibidores , Ácido Caínico/toxicidade , Neurônios/efeitos dos fármacos , Animais , Proteínas de Ciclo Celular/metabolismo , Inibidores Enzimáticos/farmacologia , Flavonoides/farmacologia , Neurônios/citologia , Neurônios/fisiologia , Piperidinas/farmacologia , RatosRESUMO
This study was undertaken to investigate whether kainic acid (KA) may regulate the expression of several proteins which plays an important role in cell-cycle progression in cerebellar granule neurons (CGNs). KA induced decrease in MTT values in a concentration dependent way. Flow cytometric analysis showed that KA was able to induce 30% apoptosis in CGNs. Apoptotic nuclear condensation were detected 24 h of exposure to KA (200 microM). An associated marked increase in DNA synthesis, measured by BrdU incorporation, was observed. Western blot analysis showed that KA induced an increase in the expression of Cdk2, cyclin E and E2F-1. It is proposed that, in post-mitotic cells like CGNs, re-entry cell cycle could be responsible for the apoptotic effect of KA.
Assuntos
Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Cerebelo/efeitos dos fármacos , Ácido Caínico/farmacologia , Neurônios/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Apoptose/fisiologia , Ciclo Celular/fisiologia , Proteínas de Ciclo Celular/biossíntese , Células Cultivadas , Cerebelo/citologia , Cerebelo/fisiologia , Relação Dose-Resposta a Droga , Agonistas de Aminoácidos Excitatórios/farmacologia , Neurônios/citologia , Neurônios/fisiologia , Ratos , Ratos Sprague-DawleyRESUMO
Glial cell line-derived neurotrophic factor (GDNF) family ligands are important regulators of neuronal development and maintenance of the connectivity in the basal ganglia and show neuroprotective activities in several paradigms of brain injury. The mRNAs of two members of this family, GDNF and neurturin, and also their receptors have been detected in the basal ganglia. In the present work, we analyzed the time course changes in the expression of these neurotrophic factors and receptors in the adult rat striatum, induced by quinolinate or kainate excitotoxicity. Our results show that stimulation of NMDA or non-NMDA receptors induced different effects on the mRNA levels analyzed. Expression of GDNF and its preferred receptor, GDNF family receptor-alpha1 (GFRalpha1), was transiently up-regulated by quinolinate and kainate, but with differing intensity and temporal pattern. Immunohistochemical analysis showed that, although GDNF and GFRalpha1 were initially localized in neurons, excitotoxicity induced the expression of these proteins in astrocyte-like cells. Neurturin mRNA levels were only up-regulated after quinolinate injection, whereas quinolinate or kainate injection did not modify GFRalpha2 mRNA. The mRNA for the common receptor, c-Ret, was up-regulated by both agonists with similar temporal pattern but with differing intensity. Immunohistochemical analysis showed that c-Ret protein was located on neurons. These changes in mRNA levels and protein localization of GDNF family components could reflect an endogenous trophic response of striatal cells to different excitotoxic insults.
Assuntos
Corpo Estriado/metabolismo , Proteínas de Drosophila , Aminoácidos Excitatórios/metabolismo , Fatores de Crescimento Neural/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Animais , Corpo Estriado/citologia , Corpo Estriado/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Fator Neurotrófico Derivado de Linhagem de Célula Glial , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial , Imuno-Histoquímica , Ácido Caínico/toxicidade , Ligantes , Masculino , Fatores de Crescimento Neural/genética , Proteínas do Tecido Nervoso/genética , Neurturina , Ensaios de Proteção de Nucleases , Proteínas Proto-Oncogênicas c-ret , Ácido Quinolínico/toxicidade , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Regulação para Cima/efeitos dos fármacosRESUMO
1. We examined the role of non-NMDA receptors in kainic acid (KA)-induced apoptosis in cultures of rat cerebellar granule cells (CGCs). KA (1 - 500 microM) induced cell death in a concentration-dependent manner, which was prevented by NBQX and GYKI 52466, non-NMDA receptor antagonists. Moreover, AMPA blocked KA-induced excitotoxicity, through desensitization of AMPA receptors. 2. Similarly, KA raised the intracellular calcium concentration of CGCs, which was inhibited by NBQX and GYKI 52466. Again, AMPA (100 microM) abolished the KA (100 microM)-induced increase in intracellular calcium concentration. 3. KA-induced cell death in CGCs had apoptotic features, which were determined morphologically, by DNA fragmentation, and by expression of the prostate apoptosis response-4 protein (Par-4). 5. KA (500 microM) slightly (18%) increased caspase-3 activity, which was strongly enhanced by colchicine (1 microM), an apoptotic stimulus. However, neither Z-VAD.fmk, a pan-caspase inhibitor, nor the more specific caspase-3 inhibitor, Ac-DEVD-CHO, prevented KA-induced cell death or apoptosis. In contrast, both drugs inhibited colchicine-induced apoptosis. 5. The calpain inhibitor ALLN had no effect on KA or colchicine-induced neurotoxicity. 6. Our findings indicate that colchicine-induced apoptosis in CGCs is mediated by caspase-3 activation, unlike KA-induced apoptosis.