Long-Term Potentiation Induces Extrasynaptic Exocytosis of Glun2A-containing NMDA Receptors that is Mainly Controlled by SNAP23

Abstract

NMDA receptors (NMDAR) are key players in the initiation of synaptic plasticity that underlies learning and memory. Long-term potentiation (LTP) of synapses require an increased calcium current via NMDA channels to trigger modifications in postsynaptic density (PSD). It is generally believed that the amount of NMDARs on the postsynaptic surface remains stationary, whereas their subunit composition is dynamically fluctuated during this plasticity process. However, the molecular machinery underlying this subunit-specific regulation remains largely elusive. Here, by detecting the time-lapse changes of surface GluN2A and GluN2B subunit levels using biochemical approaches, surface immunostaining, live-imaging and super-resolution microscopy, we uncovered a transient increase of surface GluN2A-type NMDARs shortly after the induction of chemical long term potentiation (cLTP). These augmented sub-diffraction-limited GluN2A clusters predominantly exist in extrasynaptic domains. We also showed that the spine-enriched SNARE associated protein SNAP-23, and to a minor extent its homologue SNAP-25, control both the basal and regulated surface level of GluN2A receptors. Using a total internal reflection fluorescence microscopy (TIRFM) based live-imaging assay, we resolved and analyzed individual exocytic events of NMDARs in live neurons and found that cLTP raised the frequency of NMDAR exocytosis at extrasynaptic regions, without altering the duration or the package size of these events. Our study thereby provides direct evidence that synaptic plasticity controls the postsynaptic exocytosis machinery, which induces the insertion of more GluN2A receptors into the extrasynaptic area.Significance StatementMemory formation involves the long-term modification of synapses, which is called synaptic plasticity. In the postsynaptic density (PSD) of excited neurons, this modification process occurs on a minute timescale, initiated by the opening of NMDARs that trigger downstream cascades to fix the potentiation (LTP) at specific synapses for longer timescales. Here, using a novel live-imaging assay we resolved the dynamic delivery of NMDARs to the cell surface, and found that only the insertion frequency, not the duration of individual insertion or number of GluN2A subunits each of these NMDAR vesicles contains, was altered during the synaptic potentiation process. We also identified SNAP-23 as the key molecule mediating this activity dependent NMDAR surface delivery. This study provides a novel mechanism of how NMDARs are regulated in the short window to initiate the long-lasting synaptic modifications.