Nerve extract induces increase and redistribution of acetylcholine receptors on cloned muscle cells

Abstract

The effect of rat spinal cord explants and cell-free nerve extract on acetylcholine receptor site density and distribution was studied using (125)I- and rhodamine-labeled alpha-bungarotoxin on L(6), a cloned rat muscle cell line. Control L(6) myotubes have a low and uniform distribution of acetylcholine receptors (20 +/- 3 sites per mum(2) in the present study). The addition of spinal cord explants caused an increase in average receptor site density of about 6 times on myotubes within 2 mm of the explant, while a smaller increase of 3 times was observed at distances greater than 5 mm. The formation of high-density patches of receptors was also stimulated. These observations suggested that a diffusible substance originating from the explant was responsible for these changes. Cell-free homogenates of the central nervous system were prepared and found to produce the same effects. The effect of the homogenate was not strongly dependent on the age of the fetus from which the tissue was isolated, and fetal liver had little or no effect. The active component(s) appears to be a protein(s) with a molecular weight of about 100,000. Because the nerve homogenates make the L(6) cells resemble primary muscle cultures, we suggest that a common factor is responsible for regulating the acetylcholine receptor in the two types of muscle culture. The normally acetylcholine receptor-poor L(6) cells may provide a more sensitive assay for these factors than do primary muscle cultures.