RESUMO
Presynaptic GABA(B) receptors (GABA(B)R) control glutamate and GABA release at many synapses in the nervous system. In the present study we used whole-cell patch-clamp recordings of spontaneous excitatory and inhibitory synaptic currents in the presence of TTX to monitor glutamate and GABA release from synapses in layer II and V of the rat entorhinal cortex (EC)in vitro. In both layers the release of both transmitters was reduced by application of GABA(B)R agonists. Quantitatively, the depression of GABA release in layer II and layer V, and of glutamate release in layer V was similar, but glutamate release in layer II was depressed to a greater extent. The data suggest that the same GABA(B)R may be present on both GABA and glutamate terminals in the EC, but that the heteroreceptor may show a greater level of expression in layer II. Studies with GABA(B)R antagonists suggested that neither the auto- nor the heteroreceptor was consistently tonically activated by ambient GABA in the presence of TTX. Studies in EC slices from rats made chronically epileptic using a pilocarpine model of temporal lobe epilepsy revealed a reduced effectiveness of both auto- and heteroreceptor function in both layers. This could suggest that enhanced glutamate and GABA release in the EC may be associated with the development of the epileptic condition.
Assuntos
Córtex Entorrinal/metabolismo , Epilepsia do Lobo Temporal/metabolismo , Ácido Glutâmico/metabolismo , Terminações Pré-Sinápticas/metabolismo , Receptores de GABA-B/metabolismo , Ácido gama-Aminobutírico/metabolismo , Animais , Autorreceptores/efeitos dos fármacos , Autorreceptores/metabolismo , Doença Crônica , Convulsivantes/farmacologia , Modelos Animais de Doenças , Córtex Entorrinal/fisiopatologia , Epilepsia do Lobo Temporal/fisiopatologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Agonistas GABAérgicos/farmacologia , Agonistas dos Receptores de GABA-B , Potenciais Pós-Sinápticos Inibidores/efeitos dos fármacos , Potenciais Pós-Sinápticos Inibidores/fisiologia , Depressão Sináptica de Longo Prazo/fisiologia , Masculino , Inibição Neural/fisiologia , Técnicas de Cultura de Órgãos , Técnicas de Patch-Clamp , Ratos , Ratos Wistar , Bloqueadores dos Canais de Sódio/farmacologia , Transmissão Sináptica/fisiologiaRESUMO
We have previously shown that there are clear differences between spontaneous excitatory synaptic currents recorded in layers V and II of the rat entorhinal cortex (EC) in vitro, and have suggested that these might contribute to a more pronounced susceptibility of the deeper layer to epileptogenesis. In the present study, we have made a detailed comparison of spontaneous synaptic inhibition between the two layers by recording spontaneous inhibitory synaptic currents (sIPSCs) using whole-cell patch-clamp techniques in EC slices. Pharmacological studies indicated that sIPSCs were mediated exclusively by gamma-aminobutyric acid (GABA)(A) receptors. There was little difference in average amplitudes, rise or decay times of sIPSCs in layer II compared with layer V. However, in the former, events occurred at 4-5 times the frequency seen in the latter, and frequencies of =40 Hz were not uncommon. When activity-independent, miniature IPSCs were isolated in tetrodotoxin (TTX), the frequency in layer V was more than halved, but in layer II only a small reduction was seen, and the frequency remained very high. In terms of kinetics, while averaged sIPSCs in each layer were very similar, detailed comparison of individual sIPSCs within layers revealed distinct differences, possibly reflecting inputs from different subtypes of interneurons or inputs at different somatodendritic locations. In layer V, sIPSCs could be divided into three groups, one with slow rise and decay kinetics and a second with fast rise kinetics, further distinguished into two groups by either fast or slow decay kinetics. The distinction between events in layer II was simpler, one group having both fast rise and decay times and the second with both parameters much slower. Finally, IPSCs could occur in high-frequency bursts in both layers, although these were much more prevalent in layer II. The results are discussed in terms of the overall level of background inhibition in the two layers, as well as how this might relate to their susceptibilities to epileptogenesis.
Assuntos
Córtex Entorrinal/citologia , Córtex Entorrinal/fisiologia , Inibição Neural/fisiologia , Ácido gama-Aminobutírico/fisiologia , Anestésicos Locais/farmacologia , Animais , Cádmio/farmacologia , Antagonistas GABAérgicos/farmacologia , Cinética , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Piridazinas/farmacologia , Ratos , Ratos Wistar , Receptores de GABA-A/fisiologia , Estimulação Química , Sinapses/efeitos dos fármacos , Sinapses/fisiologia , Tetrodotoxina/farmacologiaRESUMO
We have recently shown that the anticonvulsant drugs phenytoin, lamotrigine and sodium valproate all reduce the release of glutamate at synapses in the entorhinal cortex in vitro. In the present investigation we determined whether this property was shared by gabapentin and pregabalin, using whole-cell patch-clamp recordings of excitatory postsynaptic currents (EPSCs) in layer V neurons in slices of rat entorhinal cortex. Both drugs reduced the amplitude and increased the paired-pulse ratio of EPSCs evoked by electrical stimulation of afferent inputs, suggesting a presynaptic effect to reduce glutamate release. The frequency of spontaneous EPSCs (sEPSCs) was concurrently reduced by GBP, further supporting a presynaptic action. There was no significant change in amplitude although a slight reduction was seen, particularly with gabapentin, which may reflect a reduction in the number of larger amplitude sEPSCs. When activity-independent miniature EPSCs were recorded in the presence of tetrodotoxin, both drugs continued to reduce the frequency of events with no change in amplitude. The reduction in frequency induced by gabapentin or pregabalin was blocked by application of the l-amino acid transporter substrate l-isoleucine. The results show that gabapentin and pregabalin, like other anticonvulsants, reduce glutamate release at cortical synapses. It is possible that this reduction is a combination of two effects: a reduction of activity-dependent release possibly via interaction with P/Q-type voltage-gated Ca channels, and a second action, as yet unidentified, occurring downstream of Ca influx into the presynaptic terminals.