Internal and external differentiations of the postsynaptic membrane at the neuromuscular junction

Frog, snake and rat neuromuscular junctions were prepared for electron microscopy by the quick-freeze, deep-etch, rotary replication procedure. The postsynaptic membrane was exposed by treating muscles with 1 mg/ml collagenase to remove the basal lamina. Present on the apices of the postsynaptic folds are regular arrays of 8-9 nm protrusions. These are not seen in the depths of the folds nor elsewhere on the muscle surface, thus they presumably represent the heads of cholinergic receptor molecules. These protrusions tend to be arranged in parallel rows two-abreast. Their high concentration (10 000/microns2) and their orderly arrangement is basically similar to the receptors seen in Torpedo postsynaptic membrane. Their distribution did not appear to change after denervation. Efforts were made to expose possible anchoring structures of these receptors, by treating muscles with 0.1% Saponin immediately before and/or during fixation in 1% formaldehyde, or by homogenizing muscles after brief formaldehyde fixation. This washed most soluble protein out of the cytoplasm and exposed a submembraneous meshwork just beneath the postsynaptic membrane. This meshwork appears to connect the membrane to underlying bundles of intermediate filaments which course through the postsynaptic processes that border each fold. This meshwork is presumably equivalent to the postsynaptic 'density' seen in thin sections. Its three-dimensional structure suggests that it could anchor receptor molecules to underlying cytoskeletal elements and thus immobilize receptors in the plane of the postsynaptic membrane.