The Mechanism Of Activity-dependent Membrane-trafficking Of GluN2A-NMDA Receptors In Neuron

Background N-Methyl-D-aspartate (NMDA) receptors are important ionotropic glutamate receptors, and they take pivotal parts in the processing of long term potentiation (LTP) which is believed to be highly related to learning and memory. Previous studies have shown that the number of NMDA receptors increased during the NMDA-mediated LTP process in hippocampal neurons, but the particular change on surface expression and the resource of robust inserted GluN2A subunits are not clear.Objectives Research on the activity-dependent surface expression and the resource of fast inserted GluN2A subunits in order to have a better understanding of the formation of learning and memory.Methods and Results At this study we established two LTP stimulation models-chemical LTP and high K+induced depolarization. We focused our research on the change of surface expression and transportation of NMDA receptor GluN2A subunits in cultured rat cortical and hippocampel neurons under these two models. Using the methods of surface biotinylation, postsynaptic density fraction extraction and western-blot, we found the surface expression of GluN2A increased during the two models of stimulation. And the observations under microscopy of fixed GFP labeled GluN2A subunits or the live cell imaging of SEP labeled GluN2A subunits agreed with our previous results. By applying rat cortical neurons with microtubules depolymerization agent Nocodazole or actin depolymerization agent Cytochalasin D, we figured that Cytochalasin D could abolish the increasing surface expression of GluN2A, but Nocodazole plays no effect on GluN2A insertion. Meanwhile, the treatment of translation inhibitor cycloheximide over0.5h could also attenuate the surface expression of GluN2A, but applied cycloheximide in a short time(<0.5h) had no effect on the robust insertion of GluN2A. At the same time we found the colocalization of GluN2A and dendritic endoplasmic reticulum decreased while the colocalization of GluN2A and PSD increased under the two LTP stimulation models.Conclusion Herein we show that the activity-dependent fast inserted GluN2A-NMDA receptors are not transported in a canonical way by which proteins were packed in vesicles and transported along microtubles to dendrites but transported from the dendritic part of endoplasmic reticulum, suggesting that the dendrtic endoplasmic reticulum may be a pool of GluN2A-NMDAs\recetpors and it could release NMDA receptors in response to the activity of excitotary synaptic plasticity.