Involvement of calpains in the destabilization of the acetylcholine receptor clusters in rat myotubes.
The effects of calpain inhibitors on the total number of acetylcholine receptors (AChRs) on cultured rat myotubes and on the stability of AChR clusters in these myotubes were investigated. The degradation rate of total AChRs labeled with (125)I-alpha-bungarotoxin was assessed from radioactivity remaining in the myotubes as a function of time. Treatment with calpain inhibitors resulted in a two- to three-fold increase in the half-life of total AChRs. Incubation with these inhibitors produced 40% increases in intracellular AChRs but no major changes in surface AChRs, indicating that the increased AChR half-life is due to intracellular accumulation. The rate loss of AChRs from the clusters was assessed by measuring the loss of fluorescence intensity in rhodaminated-alpha-bungarotoxin-labeled clusters with time. Treatment with calpain inhibitors resulted in twofold increases in cluster half-life. Thus, there was generally no change in total surface receptors with the calpain inhibitors, whereas cluster half-life was substantially increased. Furthermore, with a low dose of calpeptin there was no change in turnover of total cellular AChRs, whereas cluster half-life was doubled. Taken together, these results suggest that the increased half-life of clusters produced by the calpain inhibitors may be due to retardation of the lateral movement from AChRs in the clusters.
Kim
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Laboratory of Developmental Neurobiology, Bldg. 49, Rm. 5A 38, National Institute of Child Health and Human Development, National Institutes of Health, 9000 Rockville Pike, Bethesda, Maryland 20892, USA.
Nelson
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PG
D000818:Animals; D000831:Animals, Newborn; D002038:Bungarotoxins / Q000378:metabolism; D002154:Calpain / Q000037:antagonists & inhibitors*; D002478:Cells, Cultured; D004151:Dipeptides / Q000494:pharmacology*; D009210:Myofibrils / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D011950:Receptors, Cholinergic / Q000187:drug effects* / Q000378:metabolism
D002038:Bungarotoxins; D004151:Dipeptides; D011950:Receptors, Cholinergic; C071482:calpeptin; D002154:Calpain