RESUMO
N-methyl-D-aspartate (NMDA) receptors (NMDARs) are implicated in synaptic plasticity and modulation of glutamatergic excitatory transmission. Effect of NMDAR activation on inhibitory GABAergic transmission remains largely unknown. Here, we report that a brief application of NMDA could induce two distinct actions in CA1 pyramidal neurons in mouse hippocampal slices: 1) an inward current attributed to activation of postsynaptic NMDARs; and 2) fast phasic synaptic currents, namely spontaneous inhibitory postsynaptic currents (sIPSCs), mediated by GABA(A) receptors in pyramidal neurons. The mean amplitude of sIPSCs was also increased by NMDA. This profound increase in the sIPSC frequency and amplitude was markedly suppressed by the sodium channel blocker TTX, whereas the frequency and mean amplitude of miniature IPSCs were not significantly affected by NMDA, suggesting that NMDA elicits repetitive firing in GABAergic interneurons, thereby leading to GABA release from multiple synaptic sites of single GABAergic axons. We found that the NMDAR open-channel blocker MK-801 injected into recorded pyramidal neurons suppressed the NMDA-induced increase of sIPSCs, which raises the possibility that the firing of interneurons may not be the sole factor and certain retrograde messengers may also be involved in the NMDA-mediated enhancement of GABAergic transmission. Our results from pharmacological tests suggest that the nitric oxide signaling pathway is mobilized by NMDAR activation in CA1 pyramidal neurons, which in turn retrogradely facilitates GABA release from the presynaptic terminals. Thus NMDARs at glutamatergic synapses on both CA1 pyramidal neurons and interneurons appear to exert feedback and feedforward inhibition for determining the spike timing of the hippocampal microcircuit.
Assuntos
Hipocampo/citologia , Terminações Pré-Sinápticas/fisiologia , Células Piramidais/citologia , Receptores de N-Metil-D-Aspartato/metabolismo , Transmissão Sináptica/fisiologia , Ácido gama-Aminobutírico/metabolismo , Agatoxinas , Animais , Animais Recém-Nascidos , Bloqueadores dos Canais de Cálcio/farmacologia , Óxidos N-Cíclicos/farmacologia , Interações Medicamentosas , Inibidores Enzimáticos/farmacologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Sequestradores de Radicais Livres/farmacologia , GABAérgicos/farmacologia , Imidazóis/farmacologia , Técnicas In Vitro , Potenciais Pós-Sinápticos Inibidores/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , NG-Nitroarginina Metil Éster/farmacologia , Inibição Neural/efeitos dos fármacos , Inibição Neural/fisiologia , Técnicas de Patch-Clamp , Terminações Pré-Sinápticas/efeitos dos fármacos , Venenos de Aranha/farmacologia , Transmissão Sináptica/efeitos dos fármacos , Fatores de Tempo , ômega-Conotoxina GVIA/farmacologiaRESUMO
With Flammulina velutipes material,an improved method was developed for extracting total RNA from domestic fungus that are rich in RNase,polyphenols, polymeric carbohydrates and proteoglycans. Phenol-chloroform-isoamyl alcohol were used twice to clear DNA and protein under higher concentration of denaturing solution and isopentanol, sodium acetate were used to precipitate RNA selectively. Pure and intact RNA can be effectively prepared by this method.
