Metabolic properties of human acetylcholine receptors can be characterized on cultured human muscle.
Experiments examining acetylcholine receptor (AChR) metabolism in tissue culture have hitherto been limited to animal systems. For many studies, the human AChR on human skeletal muscle provides a more physiologic target. However, previous studies suggested that the levels of AChR produced on cultured human muscle were inadequate for metabolic studies. We demonstrate here that the metabolism of human acetylcholine receptors can be analysed on pure human muscle fibers that develop in tissue culture. Degradation of AChR follows first-order kinetics and is inhibited 85% by leupeptin, demonstrating that proteolysis of human AChR occurs in the lysosome. New AChR continue to appear on the cell surface for 3 h in the presence of cycloheximide, indicating the existence of a pool of intracellular AChR destined for the cell membrane. This pool is equivalent to approximately one-third of the AChR present on the surface of the cell. At any given time, the rate of AChR accumulation on the cell surface can be quantitatively accounted for by the rates of synthesis and degradation. Our results demonstrate that studies on the effects of hormones, neurotoxins or antibodies from patients with autoimmune neuromuscular diseases are now possible with human AChR which develop on intact human muscle myotubes formed in tissue culture.
D002454:Cell Differentiation; D002455:Cell Division; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D007976:Leupeptins / Q000494:pharmacology; D008247:Lysosomes / Q000378:metabolism; D009132:Muscles / Q000166:cytology / Q000378:metabolism*; D011950:Receptors, Cholinergic / Q000096:biosynthesis / Q000378:metabolism*
D007976:Leupeptins; D011950:Receptors, Cholinergic; D003513:Cycloheximide