Postsynaptic differentiation can be experimentally induced in cultured Xenopus myotomal muscle cells by polyornithine-coated latex beads (Peng, H. B., and P.-C. Cheng, 1982, J. Neurosci., 2:1760-1774). In this study, we examined the time course of this process. Small, punctate acetylcholine receptor (AChR) clusters were detectable as early as 1.5 h after the addition of the beads. Subsequently, both the size and the number of the clusters increased with time until a saturation level was reached between 8-24 h. Because the onset and the site of the AChR clustering could be precisely marked, we were able to examine the early structural specializations associated with presumptive AChR clusters. At 1 h, when less than 20% bead-muscle contacts displayed AChR clusters, 70% of the contacts already exhibited a meshwork of 5-6-nm filaments, which were of the same size as the thin filaments within the myofibrils and thus may contain actin. A system of cisternae similar to the smooth endoplasmic reticulum was suspended within this meshwork, but other organelles were excluded from it. This meshwork, being the earliest cytoplasmic specialization at the presumptive AChR clusters and appearing before the clusters, may be a mechanism for the clustering process.

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