The specificity of action of extrinsic acetylcholine on extrajunctional and junctional receptors in the chick biventer cervicis muscle was studied by determining its ability to protect the responses evoked by acetylcholine and by tetanic nerve stimulation from the blockade by alpha-bungarotoxin, an irreversible binding agent of acetylcholine receptors. At concentrations of 50-100 mug/ml, acetylcholine caused a desensitization to extrinsic acetylcholine but not to nerve stimulation and protected only the contractile response to extrinsic acetylcholine from the toxin blockade whereas neither the response to tetanic nerve stimulation nor the endplate potentials were protected. For the protection of the latter, higher concentrations of acetylcholine were needed. In the presence of physostigmine, a concentration of acetylcholine as low as 10 mug/ml protected the endplate potentials from the toxin blockade. By contrast, d-tubocurarine protected the tetanic contraction and the endplate potentials induced by nerve stimulation at a concentration which produced the same protection of acetylcholine-induced contraction as that produced by 50-100 mug/ml acetylcholine. These results indicate that in contrast to d-tubocurarine, extrinsic acetylcholine at low concentrations acts preferentially on the extrajunctional receptors in the absence of an anticholinesterase.