Structure and Dynamics of Nicotinic Acetylcholine Receptor at the Cell Membrane

A combination of ensemble averaging methods (confocal FRAP and FCS) and single molecule experimental techniques (single-particle tracking, high-resolution fluorescence microscopy, patch-clamp) was used to study the supramolecular organization of the acetylcholine receptor (AChR), receptor dynamics at the cell surface, and the kinetics of receptor internalization. Chol depletion produced gain-of-function of single-channel dwell time. Submicron-sized particles could be resolved into AChR "nano-clusters" with a peak size distribution of ∼55 nm by superresolution STED and GSDIM microscopies. Chol depletion reduced the number of nanoclusters, increasing their size, and changed their supramolecular organization on larger scales (0.5-3.5 microns). FRAP, FCS and SPT experiments provided information on the dynamics of AChR nanoclusters, disclosing the dependence of their mobility on Chol content and cortical cytoskeleton. Chol content at the plasmalemma may thus modulate cell-surface organization and dynamics of receptor nanoclusters, and fine-tune receptor channel function to temporarily compensate for acute AChR loss from the cell surface.