Early postsynaptic instability and acetylcholine receptor compartmentalization precede neuromuscular synapse dismantling

Abstract The neuromuscular junction (NMJ) is the peripheral synapse controlling muscle contraction. Although aging and neurodegeneration result in NMJ denervation and synaptic dismantling, early indicators of this process remain elusive. Here, we analyzed the organization and dynamics of postsynaptic nicotinic acetylcholine receptors (nAChR) following muscle denervation. Using fluorescent conjugates of α-bungarotoxin (BTX), we found that loss of nAChR stability preceded morphological disintegration. Early after denervation, the combined use of receptor labeling and lectin staining revealed a rearrangement of long-lasting or newly inserted receptors that resulted in a novel compartmentalized postsynaptic pattern in which stable, pre-existing nAChRs concentrated centrally, while newly inserted, dynamic receptors localized peripherally. Small ectopic, highly dynamic nAChR clusters emerged since early denervation. Additionally, intracellular ring-like nAChR aggregates emerged since early denervation stages and were distributed in perinuclear regions, co-localizing with the lysosomal marker LAMP1, consistent with a degradative fate. Altogether, specific combinations of nAChR dynamics and morphologies serve as early markers of NMJ dismantling. These novel criteria to assess NMJ integrity may help define therapeutic windows to promote reinnervation in degenerative neuromuscular conditions.