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Barbiturates bind to an allosteric regulatory site on nicotinic acetylcholine receptor-rich membranes.

Authors
  • Dodson, B A
  • Braswell, L M
  • Miller, K W
Type
Published Article
Journal
Molecular pharmacology
Publication Date
Jul 01, 1987
Volume
32
Issue
1
Pages
119–126
Identifiers
PMID: 3600612
Source
Medline
License
Unknown

Abstract

The ability of barbiturates to bind to acetylcholine receptor-rich membranes purified from the electroplaques of Torpedo nobiliana was examined by centrifugation assay. [14C]Amobarbital both partitioned into the membrane and bound displaceably to a site with an equilibrium dissociation constant of 12 microM. This low affinity made the stoichiometry difficult to obtain despite the high specific activity of acetylcholine receptors in this membrane preparation. However, the data are not inconsistent with a stoichiometry of one barbiturate-binding site per acetylcholine-binding site. Displaceable [14C]amobarbital binding was completely inhibited by barbiturates (IC50: amobarbital, 28 microM; secobarbital, 110 microM; pentobarbital, 400 microM; phenobarbital, 690 microM; butabarbital, 690 microM; and barbital, 5.1 mM. alpha-Bungarotoxin had no effect, but cholinergic ligands that convert the acetylcholine receptor to the desensitized state (acetylcholine, carbamylcholine, and, to a lesser extent, d-tubocurarine) partially inhibited displaceable [14C]amobarbital binding. This cholinergic inhibition was prevented by preincubation with alpha-bungarotoxin, implying an allosteric mediation through the classical cholinergic site. This negative interaction between the cholinergic and the barbiturate sites was mutual with barbiturates partially decreasing equilibrium [3H]acetylcholine binding in a saturable fashion with relative affinities that parallel those for inhibiting [14C]amobarbital binding (IC50). These data establish a mutual negative heterotropic interaction between barbiturate-binding sites and cholinergic binding sites on the nicotinic acetylcholine receptor from Torpedo.

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