Acetylcholine receptor alpha-subunit mRNA is increased by ascorbic acid in cloned L5 muscle cells: Northern blot analysis and in situ hybridization

Ascorbic acid is the major factor in brain extract responsible for increasing the average acetylcholine receptor (AChR) site density on the cloned muscle cell line L5. In the present study, we show that this effect of ascorbic acid requires mRNA synthesis, and that the mRNA level for the AChR alpha-subunit is increased to about the same level as are the surface receptors. We have found no increase in the mRNA levels of the beta-, gamma-, and delta-subunits, or in the mRNAs of other muscle-specific proteins, such as that of light chain myosin 2, alpha-actin, and creatine kinase. By in situ hybridization, we further show that the increase in alpha-mRNA in response to ascorbic acid is exclusively in myotubes and is located near clusters of nuclei. mRNA levels for the alpha-subunit in mononucleated cells are very low and do not significantly increase in response to ascorbic acid. The mononucleated cells are thus excluded as a possible source for the increase in alpha-subunit mRNA detected by Northern blot analysis. Our results indicate that there is a very specific action of ascorbic acid on the regulation of AChR alpha-mRNA in the L5 muscle cells, and that the expression of surface receptors in these cells is limited by the amount of AChR alpha-subunit mRNA.