RESUMO
Under pathological conditions, nitric oxide can become a mediator of oxidative cellular damage, generating an unbalance between oxidant and antioxidant systems. The participation of neuronal nitric oxide synthase (nNOS) in the neurodegeneration mechanism has been reported; the activation of N-methyl-D-aspartate (NMDA) receptors by agonist quinolinic acid (QUIN) triggers an increase in nNOS function and promotes oxidative stress. The aim of the present work was to elucidate the participation of nNOS in QUIN-induced oxidative stress in knock-out mice (nNOS-/-). To do so, we microinjected saline solution or QUIN in the striatum of wild-type (nNOS +/+), heterozygote (nNOS+/-), and knock-out (nNOS-/-) mice, and measured circling behavior, GABA content levels, oxidative stress, and NOS expression and activity. We found that the absence of nNOS provides a protection against striatal oxidative damage induced by QUIN, resulting in decreased circling behavior, oxidative stress, and a partial protection reflected in GABA depletion. We have shown that nNOS-derived NO is involved in neurological damage induced by oxidative stress in a QUIN-excitotoxic model.
Assuntos
Corpo Estriado/efeitos dos fármacos , Óxido Nítrico Sintase Tipo I/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Ácido Quinolínico/farmacologia , Animais , Antioxidantes/farmacologia , Corpo Estriado/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Camundongos Knockout , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase/efeitos dos fármacos , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Ácido Quinolínico/metabolismo , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/metabolismoRESUMO
Parkinson's disease is a neurodegenerative disorder characterized by movement alterations caused by reduced dopaminergic neurotransmission in the nigrostriatal pathway, presumably by oxidative stress (OS). MPP(+) intrastriatal injection leads to the overproduction of free radicals (FR). The increasing formation of FR produces OS, a decline in dopamine (DA) content, and behavioral disorders. Epicatechin (EC) has shown the ability to be FR scavenger, an antioxidant enzyme inductor, a redox state modulator, and transition metal chelator. Acute administration of 100 mg/kg of EC significantly prevented (P < 0.05) the circling MPP(+)-induced behavior (10 µg/8 µL). Likewise, EC significantly (P < 0.05) reduced the formation of fluorescent lipid products caused by MPP(+). MPP(+) injection produced (P < 0.05) increased enzymatic activity of the constitutive nitric oxide synthase (cNOS). This effect was blocked with acute EC pretreatment. Cu/Zn-dependent superoxide dismutase (Cu/Zn-SOD) activity was significantly (P < 0.05) reduced as a consequence of MPP(+) damage. EC produced a slight increase (≈20%) in Cu/Zn-SOD activity in the control group. Such effects persisted in animals injured with MPP(+). The results show that EC is effective against MPP(+)-induced biochemical and behavioral damage, which is possible by an increase in Cu/Zn-SOD activity.
Assuntos
1-Metil-4-fenilpiridínio/toxicidade , Catequina/metabolismo , Cobre/metabolismo , Zinco/metabolismo , Animais , Catequina/farmacologia , Modelos Animais de Doenças , Masculino , Ratos , Ratos WistarRESUMO
BACKGROUND: l-Kynurenine has antinociceptive effects in acute and inflammatory pain. This study determined the effect of l-kynurenine and its metabolite (kynurenic acid) on rats subjected to neuropathic pain. METHODS: L5/L6 spinal nerve ligation induced tactile allodynia as measured with von Frey filaments using the up-down method. High-performance liquid chromatography and Western blot analysis determined kynurenic acid levels and expression of kynurenine amino transferase II (KAT II), respectively. RESULTS: l-Kynurenine (50-200 mg/kg, i.p.) or probenecid (100 mg/kg, i.p.) did not affect allodynia in neuropathic rats. In contrast, l-kynurenine (50-200 mg/kg, i.p.) in combination with probenecid (100 mg/kg, i.p.), an inhibitor of organic anion transport, reversed allodynia. Furthermore, intrathecal kynurenic acid (1-30 µg) reversed allodynia. Probenecid (100 mg/kg, i.p.) supplementation enhanced the maximal antiallodynic effect of intrathecal kynurenic acid (10 µg). Only the combined administration of l-kynurenine (200 mg/kg)/probenecid (100 mg/kg) increased the kynurenic acid concentration in cerebrospinal fluid. KAT II is expressed in dorsal root ganglia and dorsal spinal cord. KAT II expression was unchanged by the spinal nerve ligation or l-kynurenine/probenecid combination. The kynurenine/probenecid combination did not affect motor activity. CONCLUSIONS: l-Kynurenine produces its antiallodynic effect in the central nervous system through kynurenic acid. This effect may result from blockade of N-methyl-d-aspartate receptors. KAT II is expressed in dorsal root ganglion and dorsal spinal cord. Combined l-kynurenine and probenecid therapy has the potential to reduce neuropathic pain in humans.
Assuntos
Hiperalgesia/tratamento farmacológico , Cinurenina/uso terapêutico , Neuralgia/tratamento farmacológico , Probenecid/uso terapêutico , Animais , Quimioterapia Combinada , Feminino , Cinurenina/farmacologia , Atividade Motora/efeitos dos fármacos , Medição da Dor , Probenecid/farmacologia , Ratos , Ratos Wistar , Medula Espinal/efeitos dos fármacosRESUMO
Striatal administration of 1-methyl-4-phenylpyridinium (MPP(+)), the active metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), causes nigrostriatal dopaminergic pathway damage similar to that observed in Parkinson's disease. Copper acts as a prosthetic group of several antioxidant enzymes and recent data show that copper attenuated MPP(+)-evoked neurotoxicity. We evaluated the effect of copper (as a supplement) upon proteins nitration (60 kDa) and tyrosine hydroxylase (TH) inactivation induced by MPP(+) (10 microg/8 microL) injection into the rat striatum. Copper pretreatment (10 micromol/kg i.p.) prevented both MPP(+)-induced proteins nitration and TH inactivation. Copper treatment also prevented the dopamine-depleting effect of MPP(+) injection. Those results were accompanied by a significant reduction of enzymatic activity of the constitutive nitric oxide synthase (cNOS), whereas, the protein levels of the three isoforms of NOS remained unchanged. Results indicate that the effect of copper against MPP(+)-induced proteins nitration and TH inactivation in the striatum of rat may be mediated by a reduction of cNOS activity.
Assuntos
Sulfato de Cobre/farmacologia , Intoxicação por MPTP/prevenção & controle , Neostriado/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Nitratos/metabolismo , Tirosina 3-Mono-Oxigenase/antagonistas & inibidores , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina , Animais , Western Blotting , Dopamina/metabolismo , Dopaminérgicos/toxicidade , Eletroforese em Gel de Poliacrilamida , Masculino , Neostriado/efeitos dos fármacos , Óxido Nítrico Sintase Tipo I/biossíntese , Doença de Parkinson Secundária/prevenção & controle , Ratos , Ratos WistarRESUMO
Epidemiological studies demonstrate an association between chronic consumption of arsenic contaminated water and cognitive deficits, especially when the exposure takes place during childhood. This study documents structural changes and nitrergic deficits in the striatum of adult female Wistar rats exposed to arsenic in drinking water (3 ppm, approximately 0.4 mg/kg per day) from gestation, throughout lactation and development until the age of 4 months. Kainic acid injected animals (10mg/kg, i.p.) were also analyzed as positive controls of neural cell damage. Morphological characteristics of cells, fiber tracts and axons were analyzed by means of light microscopy as well as immunoreactivity to neuronal nitric oxide synthase (nNOS). As nitrergic markers, nitrite/nitrate concentrations, nNOS levels and expression of nNOS-mRNA were quantified in striatal tissue. Reactive oxygen species (ROS) and lipid peroxidation (LPx) were determined as oxidative stress markers. Arsenic exposure resulted in moderate to severe alterations of thickness, organization, surrounding space and shape of fiber tracts and axons, while cell bodies remained healthy. These anomalies were not accompanied by ROS and/or LPx increases. By contrast, except the expression of nNOS-mRNA, all nitrergic markers including striatal nNOS immunoreactivity presented a significant decrease. These results indicate that arsenic targets the central nitrergic system and disturbs brain structural organization at low exposure levels.
Assuntos
Arsenitos/toxicidade , Corpo Estriado/efeitos dos fármacos , Neurônios Nitrérgicos/efeitos dos fármacos , Óxido Nítrico/metabolismo , Efeitos Tardios da Exposição Pré-Natal , Compostos de Sódio/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Corpo Estriado/enzimologia , Corpo Estriado/patologia , Agonistas de Aminoácidos Excitatórios/toxicidade , Feminino , Idade Gestacional , Ácido Caínico/toxicidade , Lactação , Peroxidação de Lipídeos/efeitos dos fármacos , Nitratos/metabolismo , Neurônios Nitrérgicos/enzimologia , Neurônios Nitrérgicos/patologia , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo I , Nitritos/metabolismo , Gravidez , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismoRESUMO
Oxidative/nitrosative stress is involved in NMDA receptor-mediated excitotoxic brain damage produced by the glutamate analog quinolinic acid. The purpose of this work was to study a possible role of peroxynitrite, a reactive oxygen/nitrogen species, in the course of excitotoxic events evoked by quinolinic acid in the brain. The effects of Fe(TPPS) (5,10,15,20-tetrakis (4-sulfonatophenyl)porphyrinate iron (III)), an iron porphyrinate and putative peroxynitrite decomposition catalyst, were tested on lipid peroxidation and mitochondrial function in brain synaptic vesicles exposed to quinolinic acid, as well as on peroxynitrite formation, nitric oxide synthase and superoxide dismutase activities, lipid peroxidation, caspase-3-like activation, DNA fragmentation, and GABA levels in striatal tissue from rats lesioned by quinolinic acid. Circling behavior was also evaluated. Increasing concentrations of Fe(TPPS) reduced lipid peroxidation and mitochondrial dysfunction induced by quinolinic acid (100 microM) in synaptic vesicles in a concentration-dependent manner (10-800 microM). In addition, Fe(TPPS) (10 mg/kg, i.p.) administered 2 h before the striatal lesions, prevented the formation of peroxynitrite, the increased nitric oxide synthase activity, the decreased superoxide dismutase activity and the increased lipid peroxidation induced by quinolinic acid (240 nmol/microl) 120 min after the toxin infusion. Enhanced caspase-3-like activity and DNA fragmentation were also reduced by the porphyrinate 24 h after the injection of the excitotoxin. Circling behavior from quinolinic acid-treated rats was abolished by Fe(TPPS) six days after quinolinic acid injection, while the striatal levels of GABA, measured one day later, were partially recovered. The protective effects that Fe(TPPS) exerted on quinolinic acid-induced lipid peroxidation and mitochondrial dysfunction in synaptic vesicles suggest a primary action of the porphyrinate as an antioxidant molecule. In vivo findings suggest that the early production of peroxynitrite, altogether with the enhanced risk of superoxide anion (O2*-) and nitric oxide formation (its precursors) induced by quinolinic acid in the striatum, are attenuated by Fe(TPPS) through a recovery in the basal activities of nitric oxide synthase and superoxide dismutase. The porphyrinate-mediated reduction in DNA fragmentation simultaneous to the decrease in caspase-3-like activation from quinolinic acid-lesioned rats suggests a prevention in the risk of peroxynitrite-mediated apoptotic events during the course of excitotoxic damage in the striatum. In summary, the protective effects that Fe(TPPS) exhibited both under in vitro and in vivo conditions support an active role of peroxynitrite and its precursors in the pattern of brain damage elicited by excitotoxic events in the experimental model of Huntington's disease. The neuroprotective mechanisms of Fe(TPPS) are discussed.
Assuntos
Lesões Encefálicas/metabolismo , Doença de Huntington/prevenção & controle , Ácido Peroxinitroso/metabolismo , Porfirinas/uso terapêutico , Radiossensibilizantes/uso terapêutico , Análise de Variância , Animais , Comportamento Animal , Southern Blotting/métodos , Lesões Encefálicas/complicações , Caspase 3 , Caspases/metabolismo , Cromatografia Líquida de Alta Pressão/métodos , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Interações Medicamentosas , Eletroforese em Gel de Ágar/métodos , Doença de Huntington/etiologia , Peroxidação de Lipídeos/efeitos dos fármacos , Peroxidação de Lipídeos/fisiologia , Masculino , Mitocôndrias/fisiologia , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Óxido Nítrico Sintase/metabolismo , Ácidos Quinolínicos/farmacologia , Ratos , Ratos Wistar , Teste de Desempenho do Rota-Rod/métodos , Superóxido Dismutase/metabolismo , Vesículas Sinápticas/metabolismo , Sinaptossomos/fisiologia , Tirosina/análogos & derivados , Tirosina/metabolismo , Ácido gama-Aminobutírico/metabolismoRESUMO
We examined the degree of oxidative damage to the brain of mice transgenic for the mutation responsible for Huntington's disease. We found that there is a progressive increase in striatal lipid peroxidation (LP), that parallels the worsening of the neurological phenotype. We consider that these transgenic mice may provide an interesting system to test treatments aimed at protecting cells from damage induced by free radicals.
Assuntos
Corpo Estriado/metabolismo , Modelos Animais de Doenças , Doença de Huntington/metabolismo , Camundongos Transgênicos , Estresse Oxidativo/genética , Animais , Cerebelo/citologia , Cerebelo/metabolismo , Córtex Cerebral/citologia , Córtex Cerebral/metabolismo , Corpo Estriado/citologia , Metabolismo Energético/genética , Feminino , Humanos , Doença de Huntington/genética , Peroxidação de Lipídeos/genética , Masculino , Camundongos , Camundongos Endogâmicos CBA , Neurônios Motores/metabolismo , Mutação/fisiologia , FenótipoRESUMO
The toxicity of lead (Pb) is of concern to public health due to its persistence in the environment. Brain is one of the major target organs where severe neurologic alterations may be triggered after exposure. The primary effects of lead on brain functions are thought to be a damage to the nervous system microvasculature. However, the mechanism of this toxicity is poorly understood. Nitric oxide synthase (NOS) may be a target for lead and changes in its function can result in a cascade of pathophysiological effects that may be observed in isolated capillaries and synaptosomes. We have determined the concentration of lead in blood, capillaries and synaptosomes in brain from mice receiving 0, 250, 500, and 1000 ppm of lead for 14 days, through the drinking water. NOS activity was determined in the capillaries and synaptosomes by following the conversion of 3H-L-arginine to 3H-L-citrulline. The results show that blood lead levels were dose-dependent. Brain capillaries showed a preferential accumulation of lead as compared to synaptosomes. With all Pb treatments, synaptosomal constitutive NOS was inhibited (about 50% of control) while the inducible NOS activity in capillaries was enhanced. These data suggest that inhibition of cNOS activity and increase in iNOS may contribute to the Pb effects on the CNS.