Ek. Dutton et al., ACETYLCHOLINE-RECEPTOR AGGREGATION AT NERVE-MUSCLE CONTACTS IN MAMMALIAN CULTURES - INDUCTION BY VENTRAL SPINAL-CORD NEURONS IS SPECIFIC TOAXONS, The Journal of neuroscience, 15(11), 1995, pp. 7401-7416
We used a novel mammalian coculture system to study ACh receptor (AChR
) redistribution and synaptic structure at nerve-muscle contacts, Vent
ral spinal cord (VSC) neurons were plated on cultures containing exten
sive myotubes but few fibroblasts, Neurite-induced redistribution of A
ChRs occurred within 6 hr after plating neurons and was maximal betwee
n 36-48 hr, This AChR redistribution appeared in two patterns: (1) ACh
R density at sites directly apposed to the neurite where neurites cros
sed preexisting AChR patches was sharply reduced, (2) Newly aggregated
AChRs formed swaths lateral to the neurite path. VSC neurons induced
more AChR aggregation than hippocampal, superior cervical ganglion and
dorsal root ganglion neurons. The 43 and 58 kDa postsynaptic proteins
were colocalized with AChR-enriched domains in all VSC neurite-induce
d aggregates whereas the colocalization of laminin was variable. Elect
ron microscopy of regions with neurite-induced AChR aggregation showed
postsynaptic membrane specializations characteristic of developing sy
napses and, in older cultures, features of more mature synaptic struct
ure, Thus, the coculture system is useful for studying early stages of
neuromuscular junction (NMJ) formation. Neurites in these cocultures
were identified as axons or dendrites by morphological criteria and by
their immunoreactivity for synaptophysin and phosphorylated heavy neu
rofilament subunits or for microtubule associated protein 2 (MAP2), re
spectively, Axons showed a 10-fold higher induction of AChR aggregatio
n than did dendrites, Thus, at least one essential signaling molecule
necessary for the induction-of AChR aggregation at sites of interactio
n with muscle appears to be expressed in a polarized fashion in develo
ping VSC neurons.