Formation of neuromuscular junctions in adult rats: accumulation of acetylcholine receptors, acetylcholinesterase, and components of synaptic basal lamina

Abstract

We describe the appearance and accumulation of four specialized molecular components of the adult neuromuscular junction during ectopic endplate formation in adult rat soleus muscles. One component, the acetylcholine receptor (AChR), is a major constituent of the postsynaptic muscle membrane. The other three, acetylcholinesterase (AChE) and two extracellular synapse-specific antigens, are associated, at least in part, with the basal lamina in the synaptic cleft. The accumulation of each component was studied by immunocytochemistry. In addition, the accumulation of AChRs was measured by autoradiography after reaction with 125I-alpha-bungarotoxin, and AChE was examined by histochemistry and by sedimentation analysis. Endplate formation was initiated by cutting the original nerve to a muscle in which a foreign nerve had been previously implanted. Within 2 days, clusters of AChRs appeared in the new endplate zone. The density of AChRs in these clusters increased from 1 x 10(4) sites/microm2 to nearly the final value of 2 x 10(4) sites/ microm2 by 4 days. The cluster continued to grow in size and receptor number over the next month. AChE was not detected on the surface of the muscle fiber until after 1 week, when it was present at some, but not all ectopic endplates. Its appearance coincided with a rapid accumulation of the endplate-specific, 16 S form of AChE (A12) in the portion of the muscle containing new endplates. By 2 weeks virtually all endplates stained for AChE and by 1 month both immunochemical and histochemical staining resembled that of normal adult endplates. The synapse-specific basal lamina antigens that we studied were detected at some endplates by Day 6, and their further appearance followed a time course similar to, or slightly ahead of, that of AChE. Thus maturation of the synaptic basal lamina occurs after the AChRs have formed clusters and achieved nearly their final density.