RESUMO
Alpha-neurexins are neuron-specific cell-surface molecules that are essential for the functional organization of presynaptic Ca2+ channels and release sites. We have now examined postsynaptic glutamate receptor function in alpha-neurexin knockout (KO) mice by using whole-cell recordings in cultured neocortical slices. Unexpectedly, we find that alpha-neurexins are required for normal activity of N-methyl-D-aspartate (NMDA)- but not alpha-amino-3-hydroxy-5-methyl-4-isoxyzolepropionic acid (AMPA)-type glutamate receptors. In alpha-neurexin-deficient mice, the ratio of NMDA- to AMPA-receptor currents, recorded as evoked synaptic responses, was diminished approximately 50%. Furthermore, the NMDA-receptor-dependent component of spontaneous synaptic miniature responses was reduced approximately 50%, whereas the AMPA-receptor-dependent component was unaffected. No alterations in the levels of NMDA- or AMPA-receptor proteins were detected. These results suggest that alpha-neurexins are required to maintain normal postsynaptic NMDA-receptor function. The decrease in NMDA-receptor activity in alpha-neurexin-deficient synapses could be due to a transsynaptic effect on the postsynaptic neuron (i.e., alpha-neurexins on the presynaptic inputs guide postsynaptic NMDA-receptor function) or to a cell-autonomous postsynaptic effect of alpha-neurexins on NMDA-receptor activity. To distinguish between these two possibilities, we cocultured WT GFP-labeled neurons with neocortical slices from alpha-neurexin-deficient or control mice. No difference was found between WT neurons innervated by inputs that contained or lacked alpha-neurexins, indicating that the absence of presynaptic alpha-neurexins alone does not depress postsynaptic NMDA-receptor function. Our data suggest that, in addition to the previously described presynaptic impairments, loss of alpha-neurexins induces postsynaptic changes by a cell-autonomous mechanism.
Assuntos
Neocórtex/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Neurônios/fisiologia , Receptores de N-Metil-D-Aspartato/fisiologia , Sinapses/fisiologia , Animais , Técnicas de Cultura de Células , Deleção de Genes , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Knockout , N-Metilaspartato/farmacologia , Proteínas do Tecido Nervoso/efeitos dos fármacos , Proteínas do Tecido Nervoso/genética , Neurônios/citologia , Receptores de AMPA/fisiologia , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/farmacologiaRESUMO
Central glutamatergic synapses are thought to initially form as immature, so-called silent synapses showing exclusively N-methyl-d-aspartate receptor-mediated synaptic transmission. Postsynaptic insertion of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors during further development leads to a conversion into functional, mature synapses. Here, we tested the hypothesis that, according to the "inside first-outside last" pattern of neocortical layer formation and synaptogenesis, pyramidal cells in the superficial layers might show a higher fraction of silent synapses compared with pyramidal cells in the deep layers. We performed an electrophysiological analysis of glutamatergic synapses in acute rat visual cortex slices during postnatal development. In layer VI pyramidal neurons the incidence of silent synapses was high during the first postnatal week and strongly declined during further development. Surprisingly, in superficial cortical plate pyramidal neurons (immature layers II/III), the fraction of silent synapses was initially very low and increased up to the second postnatal week. Thereafter, a similar decline as found in layer VI pyramidal neurons was observed. Thus the developmental regulation of silent synapses was clearly different in pyramidal neurons from different neocortical layers. The almost complete absence of silent synapses at early stages in layer II/III pyramidal neurons indicates that an initially formed subset of synapses is constitutively functional. This might be important to enable spontaneous activity and latter activity-dependent maturation of synapses.
Assuntos
Sinapses/fisiologia , Córtex Visual/crescimento & desenvolvimento , Córtex Visual/fisiologia , Animais , Animais Recém-Nascidos , Ratos , Ratos Wistar , Receptores de Glutamato/fisiologiaRESUMO
Synapses are specialized intercellular junctions in which cell adhesion molecules connect the presynaptic machinery for neurotransmitter release to the postsynaptic machinery for receptor signalling. Neurotransmitter release requires the presynaptic co-assembly of Ca2+ channels with the secretory apparatus, but little is known about how synaptic components are organized. Alpha-neurexins, a family of >1,000 presynaptic cell-surface proteins encoded by three genes, link the pre- and postsynaptic compartments of synapses by binding extracellularly to postsynaptic cell adhesion molecules and intracellularly to presynaptic PDZ domain proteins. Using triple-knockout mice, we show that alpha-neurexins are not required for synapse formation, but are essential for Ca2+-triggered neurotransmitter release. Neurotransmitter release is impaired because synaptic Ca2+ channel function is markedly reduced, although the number of cell-surface Ca2+ channels appears normal. These data suggest that alpha-neurexins organize presynaptic terminals by functionally coupling Ca2+ channels to the presynaptic machinery.