Liquid chromatography tandem mass spectrometry of AMPA receptor containing vesicles

Regulated delivery of AMPA receptors (AMPARs) to the postsynaptic membrane
is an essential step in synaptic strength modification, and in particular,
long-term potentiation (LTP). While LTP has been extensively studied using
electrophysiology and light microscopy, several questions regarding the
molecular mechanisms of AMPAR delivery via trafficking vesicles remain
outstanding, including the gross molecular make up of AMPAR trafficking
organelles and identification and location of calcium sensors required for
SNARE complex-dependent membrane fusion of such trafficking vesicles with
the plasma membrane. Here, we isolated AMPAR containing vesicles (ACVs)
from whole mouse brains via immunoisolation and characterized them using
immunoelectron microscopy, immunoblotting, and liquid chromatography
tandem mass spectrometry (LC-MS/MS). We identified several proteins on
ACVs that were previously found to play a role in AMPAR trafficking,
including synaptobrevin-2, Rabs, the SM protein Munc18-1, the
calcium-sensor synaptotagmin-1, as well as several new candidates,
including synaptophysin and synaptogyrin on ACV membranes. Here,
we present three biological replicates of liquid chromatography tandem
mass spectrometry of AMPA receptor containing vesicles.