Clusters of intramembranous particles on cultured myotubes at sites that are highly sensitive to acetylcholine.

Abstract

Electrophysiological and autoradiographic studies have shown that distribution of acetylcholine (AcCho) receptors on uninnervated cultured chicken muscle cells is not uniform. Regions of high receptor density (hot spots) 10-40 times more sensitive than surrounding areas are localized as discrete patches or clusters about 10 .mu.m in diameter on myotube muscle membranes. Hot spots were also found on fusion-arrested mononucleated myoblasts. A method for freeze-fracturing monolayer cultures that allows unambiguous reidentification of membrane patches previously assayed for AcCho sensitivity was developed.Freeze-fractured membranes at physiologically defined hot spots contain aggregates of many (10-20) small clusters of large (10-19 nm in diameter) intramembranous particles. Clusters are found on both fracture faces, but particle density is much greater on the protoplasmic (P) face than on the extracellular (E) face (about 2000/.mu.m2 vs. 700/.mu.m2). Some of the particles may be composed of 5 or 6 subunits arranged cylindrically around a central dark dot. Because the aggregates are present at sites of high AcCho sensitivity, intramembranous particles may in some way be related to the AcCho receptor molecule.