Anatoxin-a interactions with cholinergic synaptic molecules.

Abstract

Anatoxin-a, a bicyclic amine isolated from blue-green alga, binds to the nicotinic acetylcholine receptor of Torpedo ocellata electric tissue, thereby inducing conformational changes in the postsynaptic receptor-ion channel complex as evidenced by alterations in the binding of radiolabeled ligands to the complex. Anatoxin-a binds to the acetylcholine recognition site (Kd = 0.1-0.2 .mu.M) as indicated by its competitive inhibition of specific [3H]acetylcholine and d-[3H]tubocurarine binding. Anatoxin-a stimulates the binding of 3 physiologically identified ion channel blockers, [3H]perhydrohistrionicotoxin, [3H]phencyclidine and [3H]phencyclidine methiodide. The 50% effective doses for these effects range from 0.14-0.28 .mu.M. Incubation of Torpedo membranes with anatoxin-a before addition of a radiolabeled channel probe produced a time- and concentration-dependent attenuation of the binding compared to the situation in which anatoxin-a and the probe are added simultaneously. The time course for the elaboration of this decrease corresponds to electrophysiological measurements of anatoxin-a-induced desensitization of neuromuscular junction responses. In these nicotinic actions, anatoxin-a is about as potent as acetylcholine. Anatoxin-a has relatively low affinity for the muscarinic acetylcholine receptors of rat brain, inhibiting 3-[3H]quinuclidinyl benzilate binding (10-10 M) by 50% at concentrations between 10-20 .mu.M. In contrast to classical muscarinic agonists, anatoxin-a displays little regional selectivity in its binding, and its unaltered by alkylation of the neural membranes with N-ethylmaleimide.