RESUMO
Starting with cholesterol homeostasis, the first part of the review addresses various aspects of cholesterol metabolism in neuronal and glial cells and the mutual crosstalk between the two cell types, particularly the transport of cholesterol from its site of synthesis to its target loci in neuronal cells, discussing the multiple mechanistic aspects and transporter systems involved. Statins are next analyzed from the point of view of their chemical structure and its impingement on their pharmacological properties and permeability through cell membranes and the blood-brain barrier in particular. The following section then discusses the transcriptional effects of statins and the changes they induce in brain cell genes associated with a variety of processes, including cell growth, signaling and trafficking, uptake and synthesis of cholesterol. We review the effects of statins at the cellular level, analyzing their impact on the cholesterol composition of the nerve and glial cell plasmalemma, neurotransmitter receptor mobilization, myelination, dendritic arborization of neurons, synaptic vesicle release, and cell viability. Finally, the role of statins in disease is exemplified by Alzheimer and Parkinson diseases and some forms of epilepsy, both in animal models and in the human form of these pathologies.
Assuntos
Encéfalo/metabolismo , Colesterol/metabolismo , Inibidores de Hidroximetilglutaril-CoA Redutases/química , Neurônios/metabolismo , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/patologia , Encéfalo/patologia , Permeabilidade da Membrana Celular/efeitos dos fármacos , Homeostase/genética , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Neuroglia/metabolismo , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/patologia , Vesículas Sinápticas/efeitos dos fármacos , Vesículas Sinápticas/genéticaRESUMO
Acute ethanol treatment induces neurodegeneration in cultured neurons and can lead to brain damage in animal models. Neuronal cells exposed to ethanol showed an increase in reactive oxygen species (ROS), oxidative damage and mitochondrial impairment contributing to synaptic failure. However, the underlying mechanisms of these events are not well understood. Here, we studied the contribution of NADPH oxidase, as a relevant source of ROS production in the brain, to mitochondrial impairment and oxidative stress induced by ethanol. We used primary hippocampal neurons subjected to an acute treatment of ethanol at increasing concentrations (25, 50, and 75 mM, 24 h), and we evaluated ROS production, mitochondrial function, and synaptic vesicle activity. Our studies showed that after ethanol administration, hippocampal neurons presented an increase in ROS levels, mitochondrial dysfunction, calcium handling defects, and synaptic impairment. Interestingly, treatment with the NADPH inhibitor, apocynin, significantly prevented oxidative stress, mitochondrial dysfunction, and the impairment of synaptic vesicle activity induced by ethanol treatment. These results indicate that NADPH oxidase could be a key participant in the molecular mechanism by which alcohol affects the brain.
Assuntos
Intoxicação Alcoólica/enzimologia , Intoxicação Alcoólica/patologia , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Mitocôndrias/efeitos dos fármacos , NADPH Oxidases , Neurônios/efeitos dos fármacos , Neurônios/patologia , Estresse Oxidativo , Trifosfato de Adenosina/metabolismo , Animais , Sinalização do Cálcio/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Feminino , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Gravidez , Cultura Primária de Células , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio , Sinapses/efeitos dos fármacos , Vesículas Sinápticas/efeitos dos fármacosRESUMO
The signal transmission in the nervous system operates through a sensitive balance between excitatory (E) inputs and inhibitory (I) responses. Imbalances in this system contribute to the development of pathologies such as seizures. In Caenorhabditis elegans, the locomotor circuit operates via the coordinated activity of cholinergic excitatory (E) and GABAergic inhibitory (I) transmission. Changes in E/I inputs can cause uncontrolled electrical discharges, mimicking the physiology of seizures. Molecules derived from 1,3,4-oxadiazole have been found to exhibit diverse biological activities, including anticonvulsant effect. In this work, we study the activity of the compound 2-[(4-methoxyphenylselenyl)methylthio]-5-phenyl-1,3,4-oxadiazole (MPMT-OX) in the GABAergic and cholinergic systems. We demonstrate that MPMT-OX reduced the locomotor activity of C. elegans with a normal balance between the E/I systems and increased the resistance to paralysis in worms exposed to pentylenetetrazol and aldicarb. MPMT-OX increased seizure resistance and assisted in the recovery of locomotor activity in worms with deletions in the genes unc-46, which regulates the transport of GABA into vesicles, and unc-49, which encodes the GABAA receptor. C. elegans with deletions in the unc-25 and unc-47 genes did not respond to treatment. Therefore, we suggest that the compound MPMT-OX upregulates GABAergic signaling in a manner dependent on the unc-25 gene, which is responsible for GABA synthesis, and unc-47, which encodes the vesicular GABA transporter.
Assuntos
Comportamento Animal/efeitos dos fármacos , Caenorhabditis elegans , Agonistas GABAérgicos/farmacologia , Oxidiazóis/farmacologia , Convulsões/prevenção & controle , Transmissão Sináptica/efeitos dos fármacos , Animais , Proteínas de Caenorhabditis elegans/biossíntese , Proteínas de Caenorhabditis elegans/genética , Fenômenos Eletrofisiológicos/efeitos dos fármacos , Locomoção/efeitos dos fármacos , Sistema Nervoso Parassimpático/efeitos dos fármacos , Convulsões/induzido quimicamente , Convulsões/psicologia , Vesículas Sinápticas/efeitos dos fármacos , Ácido gama-Aminobutírico/fisiologiaRESUMO
Acetazolamide (AZ), a molecule frequently used to treat different neurological syndromes, is an inhibitor of the carbonic anhydrase (CA), an enzyme that regulates pH inside and outside cells. We combined fluorescent FM styryl dyes and electrophysiological techniques at ex vivo levator auris longus neuromuscular junctions (NMJs) from mice to investigate the modulation of synaptic transmission and vesicle recycling by AZ. Transmitter release was minimally affected by AZ, as evidenced by evoked and spontaneous end-plate potential measurements. However, optical evaluation with FM-styryl dyes of vesicle exocytosis elicited by 50 Hz stimuli showed a strong reduction in fluorescence loss in AZ treated NMJ, an effect that was abolished by bathing the NMJ in Hepes. The remaining dye was quenched by bromophenol, a small molecule capable of diffusing inside vesicles. Furthermore, in transgenic mice expressing Synaptophysin-pHluorin (SypHy), the fluorescence responses of motor nerve terminals to a 50 Hz train of stimuli was decrease to a 50% of controls in the presence of AZ. Immunohistochemistry experiments to evaluate the state of the Myosin light chain kinase (MLCK), an enzyme involved in vesicle recycling, demonstrated that MLCK phosphorylation was much stronger in the presence than AZ than in its absence in 50 Hz stimulated NMJs. We postulate that AZ, via cytosol acidification and activation of MLCK, shifts synaptic vesicle recycling to a fast (kiss-and-run) mode, which changes synaptic performance. These changes may contribute to the therapeutic action reported in many neurological syndromes like ataxia, epilepsy, and migraine.
Assuntos
Acetazolamida/farmacologia , Inibidores da Anidrase Carbônica/farmacologia , Fármacos Neuromusculares/farmacologia , Junção Neuromuscular/efeitos dos fármacos , Vesículas Sinápticas/efeitos dos fármacos , Animais , Miosinas Cardíacas/metabolismo , Citosol/efeitos dos fármacos , Citosol/metabolismo , Exocitose/efeitos dos fármacos , Exocitose/fisiologia , Concentração de Íons de Hidrogênio , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Cadeias Leves de Miosina/metabolismo , Quinase de Cadeia Leve de Miosina/metabolismo , Junção Neuromuscular/citologia , Junção Neuromuscular/metabolismo , Fosforilação/efeitos dos fármacos , Vesículas Sinápticas/metabolismoRESUMO
JM-20 (3-ethoxycarbonyl-2-methyl-4-(2-nitrophenyl)-4,11-dihydro-1H-pyrido[2,3-b][1,5]benzodiazepine) is a novel benzodiazepine dihydropyridine hybrid molecule, which has been shown to be a neuroprotective agent in brain disorders involving glutamate receptors. However, the effect of JM-20 on the functionality of the glutamatergic system has not been investigated. In this study, by using different in vitro preparations, we investigated the effects of JM-20 on (i) rat brain synaptic vesicles (L-[(3)H]-glutamate uptake, proton gradient built-up and bafilomycin-sensitive H(+)-ATPase activity), (ii) rat brain synaptosomes (glutamate release) and (iii) primary cultures of rat cortical neurons, astrocytes and astrocyte-neuron co-cultures (L-[(3)H]-glutamate uptake and glutamate release). We observed here that JM-20 impairs H(+)-ATPase activity and consequently reduces vesicular glutamate uptake. This molecule also inhibits glutamate release from brain synaptosomes and markedly increases glutamate uptake in astrocytes alone, and co-cultured neurons and astrocytes. The impairment of vesicular glutamate uptake by inhibition of the H(+)-ATPase caused by JM-20 could decrease the amount of the transmitter stored in synaptic vesicles, increase the cytosolic levels of glutamate, and will thus down-regulate neurotransmitter release. Together, these results contribute to explain the anti-excitotoxic effect of JM-20 and its strong neuroprotective effect observed in different in vitro and in vivo models of brain ischemia.
Assuntos
Benzodiazepinas/farmacologia , Encéfalo/efeitos dos fármacos , Ácido Glutâmico/metabolismo , Neurônios/efeitos dos fármacos , Niacina/análogos & derivados , Vesículas Sinápticas/efeitos dos fármacos , Sinaptossomos/efeitos dos fármacos , Animais , Encéfalo/citologia , Encéfalo/metabolismo , Células Cultivadas , Masculino , Neurônios/metabolismo , Niacina/farmacologia , Ratos , Ratos Wistar , Vesículas Sinápticas/metabolismo , Sinaptossomos/metabolismoRESUMO
BACKGROUND: The hippocampal CA3 area contains large amounts of vesicular zinc in the mossy fiber terminals which is released during synaptic activity, depending on presynaptic calcium. Another characteristic of these synapses is the presynaptic localization of high concentrations of group II metabotropic glutamate receptors, specifically activated by DCG-IV. Previous work has shown that DCG-IV affects only mossy fiber-evoked responses but not the signals from associational-commissural afferents, blocking mossy fiber synaptic transmission. Since zinc is released from mossy fibers even for single stimuli and it is generally assumed to be co-released with glutamate, the aim of the work was to investigate the effect of DCG-IV on mossy fiber zinc signals. RESULTS: Studies were performed using the membrane-permeant fluorescent zinc probe TSQ, and indicate that DCG-IV almost completely abolishes mossy fiber zinc changes as it does with synaptic transmission. CONCLUSIONS: Zinc signaling is regulated by the activation of type II metabotropic receptors, as it has been previously shown for glutamate, further supporting the corelease of glutamate and zinc from mossy fibers.
Assuntos
Anticonvulsivantes/farmacologia , Ciclopropanos/farmacologia , Glicina/análogos & derivados , Fibras Musgosas Hipocampais/efeitos dos fármacos , Receptores de Glutamato Metabotrópico/metabolismo , Zinco/metabolismo , 6-Ciano-7-nitroquinoxalina-2,3-diona/farmacologia , Animais , Antagonistas de Aminoácidos Excitatórios/farmacologia , Ácido Glutâmico/metabolismo , Glicina/farmacologia , Hipocampo/efeitos dos fármacos , Fibras Musgosas Hipocampais/metabolismo , Terminações Pré-Sinápticas/efeitos dos fármacos , Terminações Pré-Sinápticas/metabolismo , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Estatísticas não Paramétricas , Transmissão Sináptica/efeitos dos fármacos , Vesículas Sinápticas/efeitos dos fármacos , Vesículas Sinápticas/metabolismoRESUMO
BACKGROUND: The hippocampal CA3 area contains large amounts of vesicular zinc in the mossy fiber terminals which is released during synaptic activity, depending on presynaptic calcium. Another characteristic of these synapses is the presynaptic localization of high concentrations of group II metabotropic glutamate receptors, specifically activated by DCG-IV. Previous work has shown that DCG-IV affects only mossy fiber-evoked responses but not the signals from associational-commissural afferents, blocking mossy fiber synaptic transmission. Since zinc is released from mossy fibers even for single stimuli and it is generally assumed to be co-released with glutamate, the aim of the work was to investigate the effect of DCG-IV on mossy fiber zinc signals. RESULTS: Studies were performed using the membrane-permeant fluorescent zinc probe TSQ, and indicate that DCG-IV almost completely abolishes mossy fiber zinc changes as it does with synaptic transmission. CONCLUSIONS: Zinc signaling is regulated by the activation of type II metabotropic receptors, as it has been previously shown for glutamate, further supporting the corelease of glutamate and zinc from mossy fibers.
Assuntos
Animais , Ratos , Zinco/metabolismo , Receptores de Glutamato Metabotrópico/metabolismo , Fibras Musgosas Hipocampais/efeitos dos fármacos , Ciclopropanos/farmacologia , Glicina/análogos & derivados , Anticonvulsivantes/farmacologia , Vesículas Sinápticas/efeitos dos fármacos , Vesículas Sinápticas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ratos Wistar , Terminações Pré-Sinápticas/efeitos dos fármacos , Terminações Pré-Sinápticas/metabolismo , Transmissão Sináptica/efeitos dos fármacos , 6-Ciano-7-nitroquinoxalina-2,3-diona/farmacologia , Estatísticas não Paramétricas , Ácido Glutâmico/metabolismo , Antagonistas de Aminoácidos Excitatórios/farmacologia , Fibras Musgosas Hipocampais/metabolismo , Glicina/farmacologia , Hipocampo/efeitos dos fármacosRESUMO
Etomidate is an intravenous anesthetic used during anesthesia induction. This agent induces spontaneous movements, especially myoclonus after its administration suggesting a putative primary effect at the central nervous system or the periphery. Therefore, the aim of this study was to investigate the presynaptic and postsynaptic effects of etomidate at the mouse neuromuscular junction (NMJ). Diaphragm nerve muscle preparations were isolated and stained with the styryl dye FM1-43, a fluorescent tool that tracks synaptic vesicles exo-endocytosis that are key steps for neurotransmission. We observed that etomidate induced synaptic vesicle exocytosis in a dose-dependent fashion, an effect that was independent of voltage-gated Na(+) channels. By contrast, etomidate-evoked exocytosis was dependent on extracellular Ca(2+) because its effect was abolished in Ca(2+)-free medium and also inhibited by omega-Agatoxin IVA (30 and 200nM) suggesting the participation of P/Q-subtype Ca(2+) channels. Interestingly, even though etomidate induced synaptic vesicle exocytosis, we did not observe any significant difference in the frequency and amplitude of miniature end-plate potentials (MEPPs) in the presence of the anesthetic. We therefore investigated whether etomidate could act on nicotinic acetylcholine receptors labeled with α-bungarotoxin-Alexa 594 and we observed less fluorescence in preparations exposed to the anesthetic. In conclusion, our results suggest that etomidate exerts a presynaptic effect at the NMJ inducing synaptic vesicle exocytosis, likely through the activation of P-subtype voltage gated Ca(2+) channels without interfering with MEPPs frequency. The present data contribute to a better understanding about the effect of etomidate at the neuromuscular synapse and may help to explain some clinical effects of this agent.
Assuntos
Etomidato/farmacologia , Potenciais Evocados/efeitos dos fármacos , Exocitose/efeitos dos fármacos , Hipnóticos e Sedativos/farmacologia , Placa Motora/efeitos dos fármacos , Junção Neuromuscular/efeitos dos fármacos , Vesículas Sinápticas/efeitos dos fármacos , Animais , Canais de Cálcio Tipo P/efeitos dos fármacos , Canais de Cálcio Tipo P/metabolismo , Canais de Cálcio Tipo Q/efeitos dos fármacos , Canais de Cálcio Tipo Q/metabolismo , Diafragma/efeitos dos fármacos , Diafragma/inervação , Relação Dose-Resposta a Droga , Feminino , Camundongos , Receptores Nicotínicos/efeitos dos fármacosRESUMO
We investigated the effects of cholesterol removal on spontaneous and KCl-evoked synaptic vesicle recycling at the frog neuromuscular junction. Cholesterol removal by methyl-ß-cyclodextrin (MßCD) induced an increase in the frequency of miniature end-plate potentials (MEPPs) and spontaneous destaining of synaptic vesicles labeled with the styryl dye FM1-43. Treatment with MßCD also increased the size of MEPPs without causing significant changes in nicotinic receptor clustering. At the ultrastructural level, synaptic vesicles from nerve terminals treated with MßCD were larger than those from control. In addition, treatment with MßCD reduced the fusion of synaptic vesicles that are mobilized during KCl-evoked stimulation, but induced recycling of those vesicles that fuse spontaneously. We therefore suggest that MßCD might favor the release of vesicles that belong to a pool that is different from that involved in the KCl-evoked release. These results reveal fundamental differences in the synaptic vesicle cycle for spontaneous and evoked release, and suggest that deregulation of cholesterol affects synaptic vesicle biogenesis and increases transmitter packing.
Assuntos
Membrana Celular/fisiologia , Colesterol/metabolismo , Junção Neuromuscular/fisiologia , Vesículas Sinápticas/fisiologia , Animais , Membrana Celular/efeitos dos fármacos , Exocitose/efeitos dos fármacos , Exocitose/fisiologia , Microeletrodos , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , Potenciais Pós-Sinápticos em Miniatura/efeitos dos fármacos , Potenciais Pós-Sinápticos em Miniatura/fisiologia , Fármacos Neuromusculares/farmacologia , Junção Neuromuscular/efeitos dos fármacos , Junção Neuromuscular/ultraestrutura , Cloreto de Potássio/farmacologia , Compostos de Piridínio , Compostos de Amônio Quaternário , Rana catesbeiana , Receptores Nicotínicos/metabolismo , Vesículas Sinápticas/efeitos dos fármacos , Vesículas Sinápticas/ultraestrutura , Técnicas de Cultura de Tecidos , beta-Ciclodextrinas/farmacologiaRESUMO
AIM: To investigate the presynaptic effects of propofol, a short-acting intravenous anesthetic, in the frog neuromuscular junction. METHODS: Frog cutaneous pectoris nerve muscle preparations were prepared. A fluorescent tool (FM1-43) was used to visualize the effect of propofol on synaptic vesicle exocytosos in the frog neuromuscular junction. RESULTS: Low concentrations of propofol, ranging from 10 to 25 µmol/L, enhanced spontaneous vesicle exocytosis monitored by FM1-43 in a Ca(2+)-dependent and Na(+)-independent fashion. Higher concentrations of propofol (50, 100, and 200 µmol/L) had no effect on spontaneous exocytosis. By contrast, higher concentrations of propofol inhibited the Na(+)-dependent exocytosis evoked by 4-aminopyridine but did not affect the Na(+)-independent exocytosis evoked by KCl. This action was similar and non-additive with that observed by tetrodotoxin, a Na(+) channel blocker. CONCLUSION: Our data suggest that propofol has a dose-dependent presynaptic effect at the neuromuscular transmission which may help to understand some of the clinical effects of this agent on neuromuscular function.