Perhydrohistrionicotoxin: a potential ligand for the ion conductance modulator of the acetylcholine receptor.

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Abstract
Histrinicotoxin from the Colombian frog Dendrobates histrionicus and its perhydro derivative reversibly block the acetylcholine-sensitive ion conductance system in frog neuromuscular preparations. The perhydro derivative and [3H]perhydrohistrionicotoxin, like histrionicotoxin, caused a significant decrease in the peak amplitude of the end-plate current and shortened its rise time and half-decay time. In membrane preparations from Torpedo [ocellata] electroplax, [3H]perhydrohistrionicotoxin bound reversibly to a limited number of high-affinity sites [dissociation constant, (KD) = 0.4 .mu.M]. The ratio of perhydrohistrionicotoxin to acetylcholine binding sites in these membrane preparations approached 2. Histrionicotoxins, local anesthetics and certain cholinergic agonists inhibited binding of perhydrohistrionicotoxin. Binding of perhydrohistrionicotoxin to membranes was decreased by heat or treatment with proteases. Treatment of membranes with Triton X-100 solubilized acetylcholine binding proteins and apparently also perhydrohistrionicotoxin-binding proteins. The detergent Triton X-100 also bound [3H]perhydrohistrionicotoxin. This nonspecific binding was not saturable and complicated studies on the antagonism by drugs of binding of [3H]perhydrohistrionicotoxin. In solubilized preparations the binding protein for acetylcholine could be removed by affinity chromatography or immunoprecipitation without affecting binding of perhydrohistrionicotoxin. Sephadex chromatography also separated acetylcholine- from perhydrohistrionicotoxin-binding proteins. Perhydrohistrionicotoxin did not bind significantly to purified acetylcholine-receptor protein but presumably bound to an ion conductance modulator protein that was associated with the acetylcholine-receptor in intact membrane and readily separable from the receptor protein after solubilization.