Extrasynaptic Membrane Trafficking Regulated by GluR1 Serine 845 Phosphorylation Primes AMPA Receptors for Long-term Potentiation

Eric S Guire,Victor A Derkach,Michael C Oh,Thomas R Soderling

doi:10.1074/jbc.m509677200

Eric S Guire, Victor A Derkach + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m509677200

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Abstract

Enhancement of synaptic transmission, as occurs in long-term potentiation (LTP), can result from several mechanisms that are regulated by phosphorylation of the AMPA-type glutamate receptor (AMPAR). Using a quantitative assay of net serine 845 (Ser-845) phosphorylation in the GluR1 subunit of AMPARs, we investigated the relationship between phospho-Ser-845, GluR1 surface expression, and synaptic strength in hippocampal neurons. About 15% of surface AMPARs in cultured neurons were phosphorylated at Ser-845 basally, whereas chemical potentiation (forskolin/rolipram treatment) persistently increased this to 60% and chemical depression (N-methyl-D-aspartate treatment) decreased it to 10%. These changes in Ser-845 phosphorylation were paralleled by corresponding changes in the surface expression of AMPARs in both cultured neurons and hippocampal slices. For every 1% increase in net phospho-Ser-845, there was 0.75% increase in the surface fraction of GluR1. Phosphorylation of Ser-845 correlated with a selective delivery of AMPARs to extrasynaptic sites, and their synaptic localization required coincident synaptic activity. Furthermore, increasing the extrasynaptic pool of AMPA receptors resulted in stronger theta burst LTP. Our results support a two-step model for delivery of GluR1-containing AMPARs to synapses during activity-dependent LTP, where Ser-845 phosphorylation can traffic AMPARs to extrasynaptic sites for subsequent delivery to synapses during LTP.

Highlights

Expression of AMPA-type glutamate receptor (AMPAR) by increasing the pool of GluR1 recycled back to the surface after their endocytosis [13]
These in vitro phospho-serine 845 (Ser-845) calibration standards were run on every SDS-PAGE (Fig. 2a) with extracts of cultured neurons or hippocampal slices and used for Western blotting with phospho-specific antibody to quantify the net Ser-845 phosphorylation levels of endogenous GluR1 in cultured hippocampal neurons
There was no detectable presynaptic effect, as the pair-pulse facilitation (PPF) ratio remained at 1.5 during baseline and 40 min after forskolin plus rolipram (F/R) treatment. These findings revealed that in the absence of basal stimulation, F/R treatment was sufficient to robustly increase surface AMPA receptors (Fig. 4c) but not synaptic strength (Fig. 5a), indicating that AMPA receptors phosphorylated at Ser-845 were probably trafficked to extrasynaptic sites

Summary

Introduction

Expression of AMPARs by increasing the pool of GluR1 recycled back to the surface after their endocytosis [13]. CaM-KII activity is critical for synaptic incorporation of AMPARs but requires a serine at position 845, which is not phosphorylated by CaM-KII, in GluR1 [14], suggesting an important role of Ser-845 phosphorylation for trafficking of AMPARs in an activity-dependent manner. These studies indicate that AMPAR phosphorylation and trafficking are coupled [13, 14], exactly how Ser-845 phosphorylation contributes to trafficking is unclear. We quantified the net Ser-845 phosphorylation and surface expression of GluR1 in hippocampal neurons during bidirectional synaptic plasticity. Ser-845 phosphorylation serves as a “priming” step in enhancing synaptic strength during LTP by enhancing the extrasynaptic delivery of AMPARs

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Conclusion