We have examained the specificity and the mechanism of acetylcholine receptor (AChR) accumulation at embryonic chick nerve-muscle contacts that form in culture. Spinal cord motoneurons were identified in vitro after labelling them in vivo with Lucifer Yellow-wheat germ agglutinin conjugates. All of their processes induced receptor clusters on contacted myotubes; after 24 to 48 hr of co-culture, the incidence of neurite-associated receptor patches (NARPs) was ~ 1.2/100 μm of contact. In contrast, NARPs were rarely associated with spinal cord interneurons (~ 0.1/100 μm of contact). Neurons dissociated from ciliary ganglia induce NARPs to the same extent as motoneurons. The relative contribution to NARPs of AChRs present in the membrane prior to plating ciliary ganglion neurons and of 'new' AChRs inserted 8, 11, or 17 hr after addition of neurons was assessed with two fluorescent receptor probes. Rhodamine-conjugated α-bungarotoxin was used to label either old or new receptors; a monoclonal, anti-receptor antibody visualized with fluorescein-second antibody was used to label all (new and old) receptors. Analysis of digitized fluorescence images showed that NARPs contained both new and old receptors but that within the first 24 hr of co-culture the majority (60 to 80%) were new. We estimate that cholinergic neurites increase the rate of receptor insertion 4- to 5-fold during the first 8 hr of NARP formation. The contribution of new receptors to NARPs declines with time. After 3 days of co-culture, receptors inserted over an 8-hr interval comprised only 20% of the total NARP complement. Receptor clusters that appear beneath or adjacent to neuron cell-myotube contacts are apparently different from NARPs in that the contribution of newly inserted clusters never exceeded ~ 20% of the total even during the first 24 hr of co-culture. Thus, the relative importance for NARP formation of receptor migration within the membrane and of receptor insertion varies with time and position along the cell body-neurite axis.
|Original language||English (US)|
|Number of pages||8|
|Journal||Journal of Neuroscience|
|State||Published - 1985|
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