RESUMO
An electrophysiological study was performed with mice lacking complexin II, a presynaptic protein. The long-term potentiation (LTP) by high-frequency stimulation, recorded in the hippocampal CA1 area, was decreased in complexin II-lacking mice (CPXII KO mice). The overall postsynaptic currents elicited by low frequency stimulation on the Schaffer collateral/commissural fibers in the hippocampal CA1 pyramidal cells were not different between wild-type and mutant mice. Excitatory postsynaptic currents (EPSCs) recorded in the presence of 50 microM bicuculline and inhibitory postsynaptic currents (IPSCs) recorded in the presence of 50 microM AP-5 (DL-2-amino-5-phosphonopentanoic acid) + 30 microM CNQX (6-cyano-7-nitroquinoxaline-2,3-dione) were also identical between wild-types and mutants. Furthermore, the EPSCs following repetitive stimulation (10 Hz) in CPXII KO mice did not show any difference with wild-types. These findings suggest that complexin II does not play a crucial role in ordinary neural transmission, short-term synaptic plasticity or synaptic transmission during high-frequency repetitive stimulation. Therefore, the protein is thought to be involved in the LTP process following tetanic stimulation, including the induction and/or maintenance of the LTP.
Assuntos
Hipocampo/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Plasticidade Neuronal/fisiologia , Proteínas Adaptadoras de Transporte Vesicular , Animais , Eletrofisiologia , Feminino , Hipocampo/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/deficiência , Neurônios/metabolismoRESUMO
We evaluated the association between distress and expressed emotion (EE) in family members of patients with schizophrenia by the General Health Questionnaire (GHQ), the Camberwell Family Interview (CFI), and the Five-Minute Speech Sample (FMSS). The GHQ score was higher in the high-EE group determined by both the CFI and FMSS. The difference in the GHQ score between high-EE and low-EE groups was more marked for the CFI. Family distress is closely associated with the EE classification, but the EE classification by the CFI more markedly reflected family distress versus the FMSS. Even in relatives with low EE, distress was marked, and therefore, coping with mental health in family members is important.
Assuntos
Afeto , Depressão/psicologia , Família/psicologia , Esquizofrenia , Comportamento Verbal , Adaptação Psicológica , Adulto , Depressão/diagnóstico , Saúde da Família , Feminino , Humanos , Japão , Masculino , Escalas de Graduação Psiquiátrica , Psicologia do EsquizofrênicoRESUMO
BACKGROUND: Both in vivo and in vitro studies suggest that excess stimulation of non-NMDA receptors can result in massive neuronal death in the retina. In particular, murine amacrine neurons have been known to show marked susceptibility to the toxic effects of kainate. PURPOSE: This study was designed to examine and characterize the role of N-methyl-D-aspartate (NMDA) receptor vs non-NMDA receptor in glutamate-induced neurotoxicity in the retina. METHODS: Primary cultures obtained from fetal rat retina (gestation day 16-19) were used for the experiment. The neurotoxicity was assessed quantitatively using the trypan blue exclusion method. Electrophysiological studies using patch-clamp techniques were performed to record whole-cell currents evoked by these excitatory amino acids. RESULTS: Removal of extracellular Ca2+ from the medium or application of MK-801 reduced the extent of cell death induced by the brief exposure to glutamate, NMDA, and kainate. By contrast, cell death induced by a 60-min exposure to kainate was not affected by MK-801. The electrophysiological study demonstrated that MK-801 abolished the whole-cell currents evoked by NMDA but had no effect on those induced by kainate or AMPA. CONCLUSION: These findings demonstrate that brief exposure to kainate induces cell death by way of activating NMDA receptors in cultured fetal retinal neurons and that NMDA receptors are the predominant route of fetal retinal neurotoxicity induced by brief glutamate exposure.
Assuntos
Agonistas de Aminoácidos Excitatórios/toxicidade , Ácido Caínico/toxicidade , Neurônios/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/metabolismo , Retina/efeitos dos fármacos , Animais , Cálcio/metabolismo , Morte Celular , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Maleato de Dizocilpina/farmacologia , Ácido Glutâmico/toxicidade , Potenciais da Membrana , N-Metilaspartato/toxicidade , Neurônios/metabolismo , Fármacos Neuroprotetores/farmacologia , Técnicas de Patch-Clamp , Ratos , Retina/embriologia , Retina/metabolismo , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/toxicidadeRESUMO
The SNAP receptor (SNARE) complex is a core complex specialized for synaptic vesicle exocytosis, and the binding of SNAPs to the complex is an essential step for neurotransmitter release. Complexin I and II have been identified as SNARE-complex-associated proteins. Importantly, complexins compete with alpha-SNAP for binding to the complex, suggesting that complexins may modulate neurotransmitter release process. To examine this possibility and to understand the physiological function of complexins, we generated complexin II knockout mice. The complexin-II-deficient mice (-/-) were viable and fertile, and appeared normal. Electrophysiological recordings in the mutant hippocampus showed that ordinary synaptic transmission and paired-pulse facilitation, a form of short-term synaptic plasticity, were normal. However, long-term potentiation (LTP) in both CA1 and CA3 regions was impaired, suggesting that complexin II may not be essential for synaptic vesicle exocytosis, but it does have a role in the establishment of hippocampal LTP.
Assuntos
Hipocampo/fisiologia , Potenciação de Longa Duração/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Proteínas Adaptadoras de Transporte Vesicular , Animais , Eletrofisiologia , Hipocampo/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout/genética , Proteínas do Tecido Nervoso/genética , Terminações Pré-Sinápticas/metabolismoRESUMO
Effects of ethanol on acutely dissociated medial vestibular nucleus (MVN) neurons were examined using whole-cell patch clamp technique to elucidate the mechanism underling the inhibitory effects of this drug on the neurons observed in in vivo studies. Dissociated MVN neurons obtained from male Wistar rats were superfused with extracellular solution continuously at a flow rate of 1-3 ml/min. Whole-cell patch clamp recording was performed according to standard procedures. GABA was applied by pressure from a pipette placed near the neuron recorded. Ethanol was applied via pipette by pressure or through bath perfusion. Acutely dissociated MVN neurons regularly showed spontaneous firing. Under current-clamp conditions, bath application of ethanol at 0.1% caused hyperpolarization and reduced spontaneous firing in MVN neurons, while 0.1% ethanol did not affect spontaneous firing. Pulse application of higher concentrations of ethanol (0.1-1%) caused similar hyperpolarization. Under voltage-clamp conditions at a holding potential of -30 mV, GABA induced outward currents in a concentration-dependent manner. GABA-induced currents were potentiated in the presence of 0.01% ethanol. These results indicate that high concentrations of ethanol (0.1-1%) directly induce inhibition of spontaneous firing and low concentrations (0.01%) enhance GABA-induced inhibition in the MVN neurons.
Assuntos
Etanol/farmacologia , Núcleos Vestibulares/efeitos dos fármacos , Ácido gama-Aminobutírico/farmacologia , Potenciais de Ação/efeitos dos fármacos , Animais , Depressão Química , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Técnicas In Vitro , Masculino , Neurônios/efeitos dos fármacos , Técnicas de Patch-Clamp , Ratos , Ratos WistarRESUMO
The effects of muscimol, a gamma-aminobutyric acid (GABA)A-receptor agonist, and aminooxy-acetic acid (AOAA), an inhibitor of GABA-converting enzyme, on tonic and absence-like seizures in spontaneously epileptic rats (SER: zi/zi, tm/tm) were investigated to elucidate whether GABAergic function operates normally in these animals. Muscimol at doses of 1 and 3 mg/kg (i.p.) induced high-voltage slow waves in the cortical and hippocampal EEG of SER, although the behavioral observation suggested inhibition of absence-like seizures. Similar high-voltage slow waves were also observed in the cortical and hippocampal EEG of normal rats with muscimol (1 and 3 mg/kg). Tonic convulsions in SER were dose-dependently inhibited by muscimol. AOAA (3 and 10 mg/kg, i.v.) inhibited both tonic and absence-like seizures in SER, although there were no obvious changes in EEG pattern. The inhibitory effects of AOAA on tonic convulsions appeared more slowly and lasted longer than those on absence-like seizures. Cerebral, hippocampal and cerebellar GABA levels were significantly higher in SER than the normal Kyo:Wistar and zitter rat (zi/zi), which were both the parent strains. These findings suggest that GABA receptors and GABAergic neurons are functional in SER and that the GABA system is involved in the inhibition of both seizures.
Assuntos
Ácido Amino-Oxiacético/farmacologia , Epilepsia/fisiopatologia , Agonistas de Receptores de GABA-A , Muscimol/farmacologia , 4-Aminobutirato Transaminase/antagonistas & inibidores , Animais , Encéfalo/metabolismo , Eletroencefalografia , Inibidores Enzimáticos/farmacologia , Feminino , Masculino , Ratos , Ácido gama-Aminobutírico/metabolismoRESUMO
Topiramate, a novel antiepileptic drug, inhibits the seizures of spontaneously epileptic rat (SER), a double mutant (zi/zi, tm/tm) which exhibits both tonic convulsion and absence-like seizures from the age of 8-weeks. Hippocampal CA3 pyramidal neurons in SER show a long-lasting depolarization shift with accompanying repetitive firing when a single electrostimulation is delivered to the mossy fibers in vitro. The effects of topiramate on the excitability of CA3 pyramidal neurons in SER were examined to elucidate the mechanism underlying the antiepileptic action. Intracellular recordings were performed in 23 hippocampal slice preparations of 16 SER aged 8-17 weeks. Topiramate (10-100 microM) dose-dependently inhibited the depolarizing shifts with repetitive firing induced by mossy fiber stimulation without affecting the first spike and resting membrane potentials in hippocampal CA3 neurons of SER. Higher dose of topiramate (100 microM) sometimes inhibited the first spike, and decreased excitatory postsynaptic potentials in the SER CA3 neurons. However, topiramate up to 100 microM did not affect the single action potential elicited by the stimulation in the hippocampal CA3 neurons of age-matched Wistar rat devoid of the seizure. Application of topiramate (100 microM) did not significantly affect the firing induced by depolarizing pulse applied in the CA3 neurons of the SER. In addition, topiramate (100 microM) had no effects on the Ca2+ spike induced by intracellularly applied depolarizing pulse in the presence of tetrodotoxin and tetraethylammonium. In contrast, a dose-dependent inhibition of depolarization and repetitive firing induced by bath application of glutamate in CA3 pyramidal neurons was obtained with topiramate (10-100 microM). Furthermore, topiramate (100 microM) decreased the number of miniature postsynaptic potential of CA3 pyramidal neurons of SER. In patch clamp whole cell recording using acutely dissociated hippocampal CA3 neurons from SER aged 8-weeks and age-matched normal Wistar rats, there were no remarkable effects on voltage dependent Ca2+ current with topiramate up to 300 microM in either animal; the current was completely blocked by Cd2+ at a concentration of 1 mM. These findings suggest that topiramate inhibits release of glutamate from the nerve terminals and/or abnormal firing of the CA3 pyramidal neurons of SER by mainly blocking glutamate receptors in the neurons.
Assuntos
Anticonvulsivantes/farmacologia , Epilepsia/fisiopatologia , Frutose/análogos & derivados , Hipocampo/efeitos dos fármacos , Hipocampo/fisiopatologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Animais , Cálcio/fisiologia , Eletrofisiologia , Epilepsia/genética , Feminino , Frutose/farmacologia , Ácido Glutâmico/farmacologia , Masculino , Técnicas de Patch-Clamp , Células Piramidais/efeitos dos fármacos , Células Piramidais/fisiologia , Ratos , Ratos Mutantes/genética , Transmissão Sináptica/efeitos dos fármacos , TopiramatoRESUMO
1. Patch-clamp whole cell recording was performed to elucidate whether or not ethanol, at low concentration, has an effect on the GABAA receptor in cultured rat cortical neurons as compared with flunitrazepam. 2. Bath application of ethanol (0.01%) or flunitrazepam (1 mM) potentiated the peak amplitude of GABA-induced (10 microM) current without affecting the equilibrium potential. 3. The decay time constant and time to peak of GABA-induced current were shortened in the presence of ethanol or flunitrazepam. 4. These findings indicate that a low concentration of ethanol and flunitrazepam potentiates the GABA-induced current concomitantly with acceleration of desensitization to the drug.
Assuntos
Depressores do Sistema Nervoso Central/farmacologia , Córtex Cerebral/efeitos dos fármacos , Etanol/farmacologia , Flunitrazepam/farmacologia , Moduladores GABAérgicos/farmacologia , Ácido gama-Aminobutírico/farmacologia , Animais , Células Cultivadas , Córtex Cerebral/fisiologia , Sinergismo Farmacológico , Neurônios/efeitos dos fármacos , Técnicas de Patch-Clamp , Ratos , Ratos Wistar , Receptores de GABA-A/efeitos dos fármacos , Receptores de GABA-A/fisiologiaRESUMO
A patch clamp study was performed to determine which subtype of ionotropic glutamate receptors is involved in the glutamate-induced excitation of the medial vestibular nucleus (MVN) neurons. Whole cell recording was performed on MVN neurons that were acutely dissociated by enzymatic and mechanical treatments. Application of glutamate at a concentration of 100 microM produced a current with a reversal potential of approximately 0 mV. The glutamate-induced current was completely blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 microM), a non-N-methyl-D-aspartate (NMDA)-receptor antagonist. Application of alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA) and kainic acid (KA), non-NMDA-receptor agonists, at concentrations of 30 and 100 microM produced a concentration-dependent depolarization concomitantly with an increase in firing rates during current clamp recording. During voltage clamp recording, glutamate, AMPA and KA elicited a concentration-dependent current with an equilibrium potential of approximately 0 mV. To clarify whether NMDA receptors are present in MVN neurons, the effects of glycine on the glutamate- and NMDA-induced current were examined. Two types of NMDA receptor-mediated current (types 1 and 2) were obtained in terms of the difference in sensitivity to both magnesium ion and MK-801, which act on the NMDA-receptor channel. In the type 1 neurons, the NMDA-induced current was not apparently blocked by magnesium ion or MK-801, although a larger current was obtained in the absence of magnesium ion. In the type 2 neurons, marked blockade of the NMDA-induced current was seen in the presence of magnesium ion and MK-801, as previously reported in other neurons of the central nervous system. These findings indicate the presence of both non-NMDA and NMDA receptors, which are involved in primary afferent transmission, in the MVN neuron, and two distinct types of NMDA receptors.
Assuntos
Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Receptores de Glutamato/efeitos dos fármacos , Receptores de Glutamato/fisiologia , Núcleos Vestibulares/efeitos dos fármacos , Núcleos Vestibulares/fisiologia , 6-Ciano-7-nitroquinoxalina-2,3-diona/farmacologia , Animais , Maleato de Dizocilpina/farmacologia , Eletrofisiologia , Agonistas de Aminoácidos Excitatórios/farmacologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Masculino , Potenciais da Membrana/fisiologia , Microscopia de Polarização , Técnicas de Patch-Clamp , Ratos , Ratos Wistar , Núcleos Vestibulares/citologia , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/farmacologiaRESUMO
We compared the effects of Pb2+ on four types of NMDA receptors expressed in Xenopus oocytes. Pb2+ reduced the currents evoked by glutamate and glycine. The Ki values of the receptors, epsilon 1/zeta 1, epsilon 2/zeta 1, epsilon 3/zeta 1 and epsilon 4/zeta 1, were 39, 34, 54 and 42 microM, respectively, and their Hill coefficients were 0.53, 4.6, 0.52 and 0.37, respectively. The epsilon 2/zeta 1 receptor that was inhibited in the presence of over 30 microM Pb2+ was not recovered to the control level after a Pb2+ washout for over 30 min, suggesting that epsilon 2/zeta 1 is responsible for the chronic Pb2+ intoxication in the nervous system.
Assuntos
Chumbo/farmacologia , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Condutividade Elétrica , Camundongos , Oócitos , Receptores de N-Metil-D-Aspartato/metabolismo , Proteínas Recombinantes/efeitos dos fármacos , Proteínas Recombinantes/metabolismo , XenopusRESUMO
PURPOSE: To examine the effects of Zn2+ on glutamate-induced neurotoxicity in cultured retinal neurons. METHODS: Primary cultures obtained from fetal rat retinas (16 to 19 days gestation) were used. The neurotoxic effects of excitatory amino acids were quantitatively assessed using the trypan blue exclusion method. RESULTS: A brief exposure of retinal cultures to glutamate or N-methyl-D-aspartate (NMDA) induced delayed cell death. Zn2+ at concentrations of 3 to 30 microM ameliorated glutamate- and NMDA-induced neurotoxicity in a dose-dependent manner. By contrast, neurotoxicity induced by a 1-hour exposure to kainate was not affected by Zn2+. CONCLUSIONS: These findings demonstrate that Zn2+ protects retinal neurons from NMDA receptor-mediated glutamate neurotoxicity.
Assuntos
Cloretos/farmacologia , Ácido Glutâmico/toxicidade , Neurônios/efeitos dos fármacos , Retina/efeitos dos fármacos , Compostos de Zinco/farmacologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Feto , Ácido Caínico/toxicidade , N-Metilaspartato/toxicidade , Ratos , Receptores de Glutamato/metabolismo , Retina/citologiaRESUMO
We examined the effects of conventional antiepileptic drugs (AEDs) on absence-like seizures in homozygous tremor rats (tm/tm) to determine if they corresponded pharmacologically to human absence seizures and absence-like seizures in spontaneously epileptic rats (SER: zi/zi, tm/tm) with both tonic convulsive and absence-like seizures. Cortical and hippocampal EEG activity was recorded with chronically implanted electrodes. The effects of AEDS on seizures of the tremor rat showed profiles similar to those observed in human absence seizures and also in absence-like seizures of SER. The absence-like seizures, associated with paroxysmal bursts of 5-7-Hz spike-wave complexes, were inhibited by trimethadione (TMO 200 mg/kg intraperitoneally, i.p.), ethosuximide (ESM 100 and 200 mg/kg, i.p.), valproate (VPA 100 mg/kg, i.p.), and phenobarbital (PB 10 and 20 mg/kg, i.p.). Phenytoin (PHT 20 mg/kg, i.p.) was ineffective. These results are consistent with the conclusion that the tremor rat is a useful model for evaluating new AEDS for human absence seizures.
Assuntos
Anticonvulsivantes/uso terapêutico , Epilepsia Tipo Ausência/tratamento farmacológico , Animais , Interpretação Estatística de Dados , Quimioterapia Combinada , Eletroencefalografia , Etossuximida/administração & dosagem , Etossuximida/uso terapêutico , Feminino , Injeções Intraperitoneais , Masculino , Fenobarbital/administração & dosagem , Fenobarbital/uso terapêutico , Fenitoína/administração & dosagem , Fenitoína/uso terapêutico , Ratos , Ratos Endogâmicos WKY , Ratos Mutantes , Convulsões/tratamento farmacológico , Fatores de Tempo , Trimetadiona/administração & dosagem , Trimetadiona/uso terapêutico , Ácido Valproico/administração & dosagem , Ácido Valproico/uso terapêuticoRESUMO
Effects of 20-hydroxyecdysone (20-HE), a neurosteroid, on cultured rat cortical neurons were examined using the whole cell recording technique. Under the voltage and current clamp conditions, brief application (5 sec) of 20-HE alone did not produce current changes nor any changes in the membrane potential. However, the chemical dose-dependently potentiated the GABA-induced current and hyperpolarization, which were blocked by bicuculline. These results suggest that 20-HE acts on the modulatory site of the GABAA receptor and potentiates GABAergic inhibition in rat cortical neurons.
Assuntos
Ecdisterona/farmacologia , Neurônios/efeitos dos fármacos , Ácido gama-Aminobutírico/farmacologia , Animais , Bicuculina/farmacologia , Células Cultivadas , Córtex Cerebral/citologia , Sinergismo Farmacológico , Potenciais da Membrana , Neurônios/fisiologia , Ratos , Ratos Wistar , Receptores de GABA/efeitos dos fármacosRESUMO
The effects of lead ions (Pb2+) on neuronal calcium channels were examined in cultured hippocampal neurons. Pb2+ blocked calcium channels in a concentration-dependent manner. The current-voltage relationship of the inhibition suggested a selective blockade of high-threshold calcium channels by Pb2+ up to a concentration of 3 microM. Pb2+ (3 microM) preferentially suppressed the omega-agatoxin IVA-sensitive components of the calcium channels. These findings suggest that Pb2+ mainly affects the P-type calcium channels at low concentrations.
Assuntos
Bloqueadores dos Canais de Cálcio/farmacologia , Hipocampo/metabolismo , Chumbo/farmacologia , Neurônios/metabolismo , Animais , Bário/metabolismo , Células Cultivadas , Hipocampo/citologia , Ativação do Canal Iônico/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Técnicas de Patch-Clamp , Ratos , Ratos Wistar , Venenos de Aranha/antagonistas & inibidores , Venenos de Aranha/farmacologia , ômega-Agatoxina IVARESUMO
The effects of dopamine on glutamate-induced cytotoxicity were examined using the primary cultures of rat striatal neurons. Cell viability was significantly reduced by exposure of cultures to glutamate or kainate for 24 h. In contrast, similar application of N-methyl-D-aspartate (NMDA) or alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA) did not induce cytotoxicity. Kainate-induced cytotoxicity was significantly inhibited by kynurenate but not by MK-801. Dopamine at concentrations of 1-100 microM dose-dependently reduced kainate-induced cytotoxicity. Forskolin also significantly reduced kainate cytotoxicity. The neuroprotective effect of dopamine was antagonized by SCH 23390, a D1 receptor antagonist, but not by domperidone, a D2 receptor antagonist. Moreover, kainate-induced cytotoxicity was prevented by SKF 38393, a D1 receptor agonist, or forskolin but not by quinpirole, a D2 receptor agonist. The patch clamp study revealed that the same striatal neurons responded to both kainate and NMDA. During voltage clamp recording, neither kainate-induced currents nor NMDA-induced currents were affected by dopamine. Moreover, dopamine did not affect glutamate- or kainate-induced Ca2+ influx measured with fura-2. These findings indicate that dopamine prevents kainate receptor-mediated cytotoxicity without affecting the kainate receptor activities and intracellular Ca2+ movement. Dopamine-induced neuroprotection may be mediated by an increased intracellular cAMP formed following activation of D1 receptors.
Assuntos
Dopamina/farmacologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Ácido Glutâmico/toxicidade , Neostriado/citologia , Neurônios/efeitos dos fármacos , Receptores de Ácido Caínico/antagonistas & inibidores , Animais , Cálcio/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Citosol/efeitos dos fármacos , Citosol/metabolismo , Agonistas de Dopamina/farmacologia , Antagonistas de Dopamina/farmacologia , Potenciais Evocados/efeitos dos fármacos , N-Metilaspartato/toxicidade , Neostriado/efeitos dos fármacos , Técnicas de Patch-Clamp , Ratos , Receptores de Ácido Caínico/efeitos dos fármacosRESUMO
PURPOSE: The electrophysiologic study using patch-clamp techniques demonstrated that NMDA-induced currents had properties similar to those recorded in the brain. METHODS: Primary cultures obtained from the fetal rat retina (gestation days 16 to 19) were used for the experiment. Immunocytochemical and electrophysiologic studies were done to identify the cultured cells. The neurotoxic effects of glutamate or N-methyl-D-aspartate (NMDA) on the retinal cultures were quantitatively assessed using the trypan blue exclusion method. RESULTS: The immunocytochemical study revealed that the major component of the rat retinal cultures was neurons including amacrine cells. The electrophysiologic study using patch-clamp techniques demonstrated that exposure to NMDA-induced currents with properties characteristic of those recorded in the brain. Brief exposure of these neurons to glutamate or NMDA induced delayed cell death. Glutamate neurotoxicity was prevented by the application of dopamine and forskolin. The protective action of dopamine was antagonized by a D1 receptor antagonist (SCH 23390) but not by D2 receptor antagonists (domperidone and sulpiride). A D1 receptor agonist (SKF 38393) protected glutamate-induced neurotoxicity in a concentration-dependent manner, whereas a D2 receptor agonist (quinpirole) did not affect it. CONCLUSIONS: These findings demonstrate that dopamine protects retinal neuronal cells against NMDA receptor-mediated glutamate neurotoxicity via D1 receptors.
Assuntos
Dopamina/farmacologia , Antagonistas de Aminoácidos Excitatórios , Glutamatos/toxicidade , Retina/efeitos dos fármacos , 2,3,4,5-Tetra-Hidro-7,8-Di-Hidroxi-1-Fenil-1H-3-Benzazepina/farmacologia , Animais , Benzazepinas/farmacologia , Morte Celular/efeitos dos fármacos , Células Cultivadas , Antagonistas de Dopamina , Relação Dose-Resposta a Droga , Eletrofisiologia , Feto , Imunofluorescência , Ácido Glutâmico , N-Metilaspartato/antagonistas & inibidores , N-Metilaspartato/toxicidade , Proteínas do Tecido Nervoso/metabolismo , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Ratos , Receptores de N-Metil-D-Aspartato/metabolismo , Retina/citologia , Retina/fisiologiaRESUMO
Effects of a nitric oxide (NO)-producing agent, sodium nitroprusside, on N-methyl-D-aspartate (NMDA) receptor activation in the cultured retinal neurons of rats were examined. NMDA in a Mg(2+)-free medium evoked inward currents at the resting membrane potential. Inward currents were also evoked by kainate. Sodium nitroprusside markedly reduced the NMDA-induced currents without affecting those induced by kainate. These results suggest the possible existence of a negative feed back system of NO which serves to regulate the activation of NMDA receptors in retinal neurons.
Assuntos
Neurônios/metabolismo , Óxido Nítrico/metabolismo , Nitroprussiato/farmacologia , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Retina/metabolismo , Animais , Feminino , Ácido Caínico/farmacologia , Potenciais da Membrana/efeitos dos fármacos , N-Metilaspartato/farmacologia , Neurônios/efeitos dos fármacos , Gravidez , Ratos , Receptores de N-Metil-D-Aspartato/metabolismo , Retina/citologia , Retina/efeitos dos fármacosRESUMO
The spontaneously epileptic rat (SER:zi/zi, tm/tm), a double mutant, shows both tonic convulsions and absence-like seizures characterized by low-voltage fast waves and by 5-7 Hz spike and wave-like complexes in the cerebral cortical and hippocampal EEG, respectively. Characteristics of hippocampal CA3 pyramidal neurons were examined to determine whether these neurons are abnormally excitable. When a single stimulus was given to the mossy fiber, there was repetitive firing and a depolarization shift in neurons of mature SER (over 12 weeks old), in which epileptic seizures had fully developed. However, in young SER (7-8 weeks old) and littermates (zi/zi, tm/+), which did not show any seizures, only a single spike was elicited with each single stimulation of the mossy fiber. Intracellular recording showed that the resting membrane potential was not significantly different among young and mature SER and littermates, but a long-lasting (100-200 ms) depolarizing shift accompanied by repetitive firing was observed following a single stimulation of the mossy fiber in half of the CA3 neurons of mature SER. Furthermore, the input impedance of the CA3 neurons in mature SER was lower than that in young SER and in littermates. These results indicate that SER hippocampal CA3 neurons become abnormally excitable in conjunction with the development of epileptic seizures.
Assuntos
Epilepsia Tipo Ausência/fisiopatologia , Epilepsia/fisiopatologia , Hipocampo/fisiopatologia , Animais , Eletrofisiologia , Espaço Extracelular/fisiologia , Hipocampo/patologia , Técnicas In Vitro , Membranas Intracelulares/fisiologia , Neurônios/fisiologia , Ratos , Ratos Mutantes , Ratos WistarRESUMO
The role of protein kinase C (PKC) in the muscarinic excitation of chromaffin cells freshly isolated from rat adrenal medullae was examined by the patch-clamp recording method. Acetylcholine and McN-A-343, a M1-receptor agonist, depolarized the cell and induced action potentials. Phorbol 12,13-dibutyrate (PDBu), an activator of PKC, increased acetylcholine-induced firing concomitant with a persistent depolarization. Under voltage-clamp recording, both McN-A-343 and PDBu decreased the cesium-sensitive K+ current, which was induced by shifting the membrane potential between -140 mV and -40 mV. These results suggested that the stimulation of muscarinic M1-receptors by cholinergic drugs activated phospholipase C to degrade phosphoinositide, consequently producing diacylglycerol, and diacylglycerol activates PKC to induce excitation of adrenal chromaffin cells.
Assuntos
Medula Suprarrenal/fisiologia , Células Enterocromafins/fisiologia , Proteína Quinase C/fisiologia , Receptores Muscarínicos/efeitos dos fármacos , Cloreto de (4-(m-Clorofenilcarbamoiloxi)-2-butinil)trimetilamônio/farmacologia , Acetilcolina/farmacologia , Medula Suprarrenal/citologia , Medula Suprarrenal/efeitos dos fármacos , Animais , Células Enterocromafins/efeitos dos fármacos , Técnicas In Vitro , Masculino , Microeletrodos , Dibutirato de 12,13-Forbol/farmacologia , Canais de Potássio/efeitos dos fármacos , Ratos , Ratos WistarRESUMO
The ontogeny of the N-methyl-D-aspartate (NMDA) subtype of glutamatergic receptor/ion channel was studied by examining whole cell currents evoked by NMDA in cultured hippocampal neurons 1 to 30 days after plating of cells from 18- to 20-day-gestation rat fetuses. We observed a maturation-dependent increase in conductance, compatible with an increased density of NMDA receptors, which is in agreement with previous binding data. The whole cell currents evoked by NMDA (10-100 microM) in the presence of glycine (1-100 microM) had two components that contributed to the peak amplitude. The first was a rapidly decaying current (fast component) and the second a slowly decaying current (slow component), their ratio depending upon glycine concentration. The EC50 values for glycine were 1.8 and 0.3 microM for the fast and slow components of the current, respectively. The quantitative analysis of these components indicated the existence of two distinct glycine sites, which differ in their affinity for glycine. The fast component originates from the action of glycine at the site with lower affinity. Moreover, the ratio of the fast to the slow component was also dependent on the time lapsed after plating of the fetal hippocampal neurons. The slow component became more predominant and the fast component less predominant along with cell maturation in culture, a phenomenon which reflects a change in the ratio of high- to low-affinity glycine binding sites. In addition, studies on Zn2+ gave further evidence of a change in the NMDA receptor/channel properties related to maturation of the cultured neurons.(ABSTRACT TRUNCATED AT 250 WORDS)