RESUMO
The accumulation of intracellular ionic zinc and pharmaceutical compounds, like the antibiotic sulfamethoxazole, may contribute to various neuropathologies. Sulfamethoxazole and the drug trimethoprim, are inhibitors of enzymes involved in the synthesis of tetrahydrofolate and also of carbonic anhydrases. The inhibition of the latter enzymes, which are localized both intra- and extracellularly and have a key role in pH regulation, causes alkalinization that is associated with higher spontaneous transmitter release. Intense synaptic stimulation causes the entry of released zinc into postsynaptic neurons, through glutamate receptor channels or voltage dependent calcium channels. The aim of this study was to evaluate the effect of sulfamethoxazole (180 µM) on basal postsynaptic zinc and to compare it with that caused by two depolarizing media, containing high potassium or tetraethylammonium, which may induce long term synaptic plasticity. The studies were performed in brain slices from gestating rats, at the mossy fiber synapses from hippocampal CA3 area, using the zinc indicator Newport Green. In the presence of KCl (20 mM) and sulfamethoxazole (180 µM) the zinc signals were enhanced, unlike in tetraethylammonium (25 mM). After sulfamethoxazole the tetraethylammonium evoked zinc signal had reduced amplitude. Thus, the data suggests that sulfamethoxazole enhances transmitter release affecting synaptic zinc physiology.
Assuntos
Anti-Infecciosos/toxicidade , Fibras Musgosas Hipocampais/efeitos dos fármacos , Sulfametoxazol/toxicidade , Sinapses/efeitos dos fármacos , Zinco/metabolismo , Animais , Feminino , Fibras Musgosas Hipocampais/metabolismo , Técnicas de Cultura de Órgãos , Gravidez , Ratos , Ratos WistarRESUMO
The application of tetraethylammonium (TEA), a blocker of voltage-dependent potassium channels, can induce long-term potentiation (LTP) in the synaptic systems CA3-CA1 and mossy fiber-CA3 pyramidal cells of the hippocampus. In the mossy fibers, the depolarization evoked by extracellular TEA induces a large amount of glutamate and also of zinc release. It is considered that zinc has a neuromodulatory role at the mossy fiber synapses, which can, at least in part, be due to the activation of presynaptic ATP-dependent potassium (KATP) channels. The aim of this work was to study properties of TEA-induced zinc signals, detected at the mossy fiber region, using the permeant form of the zinc indicator Newport Green. The application of TEA caused a depression of those signals that was partially blocked by the KATP channel inhibitor tolbutamide. After the removal of TEA, the signals usually increased to a level above baseline. These results are in agreement with the idea that intense zinc release during strong synaptic events triggers a negative feedback action. The zinc depression, caused by the LTP-evoking chemical stimulation, turns into potentiation after TEA washout, suggesting the existence of a correspondence between the observed zinc potentiation and TEA-evoked mossy fiber LTP.
Assuntos
Região CA3 Hipocampal/citologia , Fibras Musgosas Hipocampais/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Sinapses/efeitos dos fármacos , Tetraetilamônio/farmacologia , Tolbutamida/farmacologia , Zinco/metabolismo , Animais , Região CA3 Hipocampal/efeitos dos fármacos , Feminino , Canais KATP/metabolismo , Potenciação de Longa Duração/efeitos dos fármacos , Bloqueadores dos Canais de Potássio/farmacologia , Gravidez , Ratos , Ratos Wistar , Sinapses/metabolismoRESUMO
The hippocampal mossy fibers contain a substantial quantity of loosely-bound zinc in their glutamatergic presynaptic vesicles, which is released in synaptic transmission processes. Despite the large number of studies about this issue, the zinc changes related to short and long-term forms of potentiation are not totally understood. This work focus on zinc signals associated with chemically-induced mossy fiber synaptic plasticity, in particular on postsynaptic zinc signals evoked by KCl depolarization. The signals were detected using the medium affinity fluorescent zinc indicator Newport Green. The application of large concentrations of KCl, 20 mM and 60 mM, in the extracellular medium evoked zinc potentiations that decreased and remained stable after washout of the first and the second media, respectively. These short and long-lasting enhancements are considered to be due to zinc entry into postsynaptic neurons. We have also observed that following established zinc potentiation, another application of 60 mM KCl only elicited further enhancement when combined with external zinc. These facts support the idea that the KCl-evoked presynaptic depolarization causes higher zinc release leading to zinc influx into the postsynaptic region.
Assuntos
Potenciais da Membrana/fisiologia , Fibras Musgosas Hipocampais/fisiologia , Plasticidade Neuronal/fisiologia , Sinapses/fisiologia , Transmissão Sináptica/fisiologia , Zinco/metabolismo , Animais , Células Cultivadas , Feminino , Potenciação de Longa Duração/efeitos dos fármacos , Potenciação de Longa Duração/fisiologia , Potenciais da Membrana/efeitos dos fármacos , Fibras Musgosas Hipocampais/efeitos dos fármacos , Plasticidade Neuronal/efeitos dos fármacos , Cloreto de Potássio/administração & dosagem , Ratos , Ratos Wistar , Sinapses/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacosRESUMO
The aims of the present work were to assess the application of a chemical process to degrade a mixture of parabens and determine the influence of a natural river water matrix on toxicity. Model effluents containing either a single compound, namely methylparaben, ethylparaben, propylparaben, butylparaben, benzylparaben or p-hydroxybenzoic acid, or to mimic realistic conditions a mixture of the six compounds was used. Fenton process was applied to reduce the organic charge and toxic properties of the model effluents. The efficiency of the decontamination has been investigated using a chemical as well as a toxicological approach. The potential reduction of the effluents' toxicity after Fenton treatment was evaluated by assessing (i) Vibrio fischeri luminescence inhibition, (ii) lethal effects amongst freshwater Asian clams (Corbicula fluminea), and (iii) the impact on mammalian neuronal activity using brain slices. From the environmental point of view such a broad toxicity analysis has been performed for the first time. The results indicate that Fenton reaction is an effective method for the reduction of chemical oxygen demand of a mixture of parabens and their toxicity to V. fischeri and C. fluminea. However, no important differences were found between raw and treated samples in regard to mammalian neuronal activity.
Assuntos
Parabenos/química , Eliminação de Resíduos Líquidos/métodos , Poluentes Químicos da Água/química , Aliivibrio fischeri/efeitos dos fármacos , Animais , Encéfalo/efeitos dos fármacos , Corbicula/efeitos dos fármacos , Feminino , Oxirredução , Parabenos/toxicidade , Ratos , Ratos Wistar , Poluentes Químicos da Água/toxicidadeRESUMO
Zinc is a transition metal that is particularly abundant in the mossy fibers of the hippocampal CA3 area. Despite the large number of studies about the zinc role in mossy fibers, the action of zinc in synaptic mechanisms is only partly known. The use of computational models can be a useful tool for this study. In a previous work, a model was developed to evaluate zinc dynamics at the mossy fiber synaptic cleft, following weak stimulation, insufficient to evoke zinc entry into postsynaptic neurons. For intense stimulation, cleft zinc effluxes must be considered. Therefore, the initial model was extended to include postsynaptic zinc effluxes based on the Goldman-Hodgkin-Katz current equation combined with Hodgkin and Huxley conductance changes. These effluxes occur through different postsynaptic escape routes, namely L- and N-types voltage-dependent calcium channels and NMDA receptors. For that purpose, various stimulations were assumed to induce high concentrations of cleft free zinc, named as intense (10 µM), very intense (100 µM) and extreme (500 µM). It was observed that the main postsynaptic escape routes of cleft zinc are the L-type calcium channels, followed by the NMDA receptor channels and by N-type calcium channels. However, their relative contribution for cleft zinc clearance was relatively small and decreased for higher amounts of zinc, most likely due to the blockade action of zinc in postsynaptic receptors and channels. Therefore, it can be concluded that the larger the zinc release, the more predominant the zinc uptake process will be in the cleft zinc clearance.
Assuntos
Fibras Musgosas Hipocampais , Zinco , Zinco/metabolismo , Sinapses/fisiologia , Hipocampo/metabolismo , Receptores de N-Metil-D-Aspartato/fisiologia , Transmissão Sináptica/fisiologiaRESUMO
Fura-2 is widely used as a fluorescent probe to monitor dynamic changes in cytosolic free calcium in cells, where Ca(2+) can enter through several types of voltage-operated or ligand-gated channels. However, Fura-2 is also sensitive to other metal ions, such as zinc, which may be involved in ionic channels and receptors. There is interest, in particular, in studying the synapses between mossy fibers and CA3 pyramidal cells which contain both calcium and high quantities of free or loosely bound zinc. We have found, through fluorescence probing, that endogenous zinc inhibits mossy fiber calcium transients. However, since these results might be explained by an effect of the zinc chelator N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) on the spectral properties of Fura-2, we have carried out a validation of the method through fluorescence excitation spectra of the complex Fura-2/calcium, and show that TPEN does not affect these spectra. This supports the idea that the observed calcium enhancement is related to a zinc inhibition of presynaptic calcium mechanisms, and confirms the use of the chelator TPEN as a general procedure for the biophysical study of Ca(II) in the presence of Zn(II) using Fura-2.
Assuntos
Cálcio/metabolismo , Células/metabolismo , Quelantes/química , Etilenodiaminas/química , Corantes Fluorescentes/química , Fura-2/química , Zinco/química , Cálcio/química , Reprodutibilidade dos Testes , Soluções , Espectrometria de FluorescênciaRESUMO
The electrochemical oxidation (EO) of phenolic wastewaters mimicking olive oil mill effluents was carried out in a batch stirring reactor using Ti/IrO2 anodes, varying the nature (NaCl and Na2SO4) and electrolyte concentration (1.8-20 g L-1), current density (57-119 mA cm-2) and initial pH (3.4-9). Phenolic content (TPh) and chemical oxygen demand (COD) removals were monitored as a function of applied charge and over time. The nature of the electrolyte greatly affected the efficiency of the system, followed by the influence of the current density. The NaCl concentration and the initial pH influenced the process in a lesser extent. The best operating conditions achieved were 10 g L-1 of NaCl, current density of 119 mA cm-2 and initial pH of 3.4. These parameters led to 100 and 84.8% of TPh and COD removal, respectively. Under these conditions, some morphological differences were observed by SEM on the surface of the anode after treatment. To study the potential toxicity of the synthetic effluent in neuronal activity, this mixture was applied to rat brain slices prior to and after EO. The results indicate that although the treated effluent causes a smaller depression of the neuronal reactive oxygen species (ROS) signal than the untreated one, it leads to a potentiation instead of recovery, upon washout. Furthermore, the purification of a real olive mill wastewater (OMW), with the organic load of the synthetic effluent, using the same optimised operating conditions, achieved total phenolic compounds abatement and 62.8% of COD removal.This study demonstrates the applicability of this EO as a pre-treatment process of a real effluent, in order to achieve the legal limit values to be discharged into natural streams regarding its organic load.
Assuntos
Técnicas Eletroquímicas/métodos , Irídio/química , Fenóis , Titânio/química , Poluentes Químicos da Água , Purificação da Água/métodos , Eletrodos , Oxirredução , Fenóis/análise , Fenóis/química , Fenóis/isolamento & purificação , Cloreto de Sódio/química , Sulfatos/química , Poluentes Químicos da Água/análise , Poluentes Químicos da Água/química , Poluentes Químicos da Água/isolamento & purificaçãoRESUMO
The involvement of type I metabotropic glutamate receptors in hippocampal CA1 long-term potentiation (LTP) depends on the applied tetanic stimulation protocol. Activation of these receptors may cause an elevation of intracellular calcium via the formation of the second messenger inositol triphosphate (IP3) and subsequent intracellular calcium release. It has been shown that the type I metabotropic receptors antagonist L-2-amino-3-phosphonopropionate (L-AP3) blocks CA1 LTP. Combining dendritic calcium and field potential measurements in CA1 hippocampal area, we found that L-AP3 did not affect single calcium transients but reduced the calcium changes evoked by a single tetanus, preventing the long-lasting calcium enhancements associated with CA1 LTP. These findings suggest that the formation of this type of LTP requires calcium release from IP3-sensitive stores.
Assuntos
Alanina/análogos & derivados , Alanina/farmacologia , Sinalização do Cálcio/fisiologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Hipocampo/metabolismo , Líquido Intracelular/metabolismo , Potenciação de Longa Duração/fisiologia , Receptores de Glutamato Metabotrópico/metabolismo , 2-Amino-5-fosfonovalerato/farmacologia , 6-Ciano-7-nitroquinoxalina-2,3-diona/farmacologia , Animais , Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Estimulação Elétrica , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Corantes Fluorescentes , Fura-2 , Hipocampo/citologia , Hipocampo/efeitos dos fármacos , Inositol 1,4,5-Trifosfato/metabolismo , Líquido Intracelular/efeitos dos fármacos , Potenciação de Longa Duração/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Ratos , Ratos Wistar , Receptores de Glutamato Metabotrópico/antagonistas & inibidores , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Receptores de N-Metil-D-Aspartato/metabolismo , Transmissão Sináptica/efeitos dos fármacos , Transmissão Sináptica/fisiologia , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologiaRESUMO
Multiple calcium signaling pathways, including intracellular calcium release that is mediated by inositol triphosphate (IP3) or ryanodine calcium store receptors, seem to be involved in CA1 hippocampal synaptic plasticity. We have addressed the role of dendritic calcium release in short- and long-term potentiation (STP and LTP) using thapsigargin, which depletes intracellular calcium stores. Measuring Fura-2 calcium signals and extracellular field potentials, we have found that thapsigargin did not affect single pre-tetanus calcium transients but reduced tetanically evoked calcium changes. The latter effect prevented the formation of short- and long-lasting calcium enhancements. These results are consistent with the idea that intracellular calcium release is not involved in baseline synaptic transmission but is essential for those forms of synaptic plasticity.
Assuntos
Cálcio/metabolismo , Inibidores Enzimáticos/farmacologia , Hipocampo/metabolismo , Potenciação de Longa Duração/efeitos dos fármacos , Tapsigargina/farmacologia , Animais , Sinalização do Cálcio/efeitos dos fármacos , Hipocampo/efeitos dos fármacos , Ratos , Ratos WistarRESUMO
An important pool of chelatable zinc is present in the synaptic vesicles of mossy fiber terminals from hippocampal CA3 area, being zinc released following single or repetitive electrical stimulation. Previous studies have suggested different synaptic roles for released mossy fiber zinc, including the inhibition of presynaptic calcium and of postsynaptic N-methyl-D-aspartate (NMDA) and gamma amino-butyric acid (GABAA) receptors. The effect of endogenously released zinc on mossy fiber long-term potentiation (LTP) induction also is not yet established. We have investigated the effect of the permeant zinc chelator N,N,N',N'-tetrakis(2-pyridylmethyl) ethylenediamine (TPEN) on mossy fiber calcium and on synaptic transmission, before and during the application of LTP-inducing stimulation. We have found, using the calcium indicator Fura-2, that single and tetanically-evoked mossy fiber calcium signals are both enhanced in the presence of 20 microM TPEN, while the single field potentials are unaffected. As expected, no effect was observed on the single calcium signals or field potentials obtained at the CA3-CA1 synapses, from the CA1 area, which has a lower concentration of vesicular zinc. These results support the idea that at the hippocampal mossy fiber synapses, released zinc inhibits presynaptic calcium mechanisms. A higher concentration of TPEN (100 microM) significantly reduced mossy fiber synaptic transmission but did not prevent the induction of mossy fiber LTP, suggesting that zinc is not required for the formation of this form of LTP.
Assuntos
Sinalização do Cálcio/efeitos dos fármacos , Quelantes/farmacologia , Etilenodiaminas/farmacologia , Fibras Musgosas Hipocampais/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Animais , Sinalização do Cálcio/fisiologia , Estimulação Elétrica , Potenciação de Longa Duração , Ratos , Ratos Wistar , Transmissão Sináptica/fisiologiaRESUMO
An important pool of chelatable zinc is present in the synaptic vesicles of mossy fiber terminals from hippocampal CA3 area, being zinc released following single or repetitive electrical stimulation. Previous studies have suggested different synaptic roles for released mossy fiber zinc, including the inhibition of presynaptic calcium and of postsynaptic N-methyl-D-aspartate (NMDA) and gamma amino-butiric acid (GABA A) receptors. The effect of endogenously released zinc on mossy fiber long-term potentiation (LTP) induction also is not yet established. We have investigated the effect of the permeant zinc chelator N,N,N',N'-tetrakis(2-pyridylmethyl) ethylenediamine (TPEN) on mossy fiber calcium and on synaptic transmission, before and during the application of LTP-inducing stimulation. We have found, using the calcium indicator Fura-2, that single and tetanically-evoked mossy fiber calcium signals are both enhanced in the presence of 20 ìM TPEN, while the single field potentials are unaffected. As expected, no effect was observed on the single calcium signals or field potentials obtained at the CA3-CA1 synapses, from the CA1 area, which has a lower concentration of vesicular zinc. These results support the idea that at the hippocampal mossy fiber synapses, released zinc inhibits presynaptic calcium mechanisms. A higher concentration of TPEN (100 ìM) significantly reduced mossy fiber synaptic transmission but did not prevent the induction of mossy fiber LTP, suggesting that zinc is not required for the formation of this form of LTP.