Gallamine interacts with an allosteric site on muscarinic acetylcholine receptor complexes in rat brain membranes, thereby slowing the dissociation of a radiolabelled ligand ([3H]N-methylscopolamine) from the receptor complex. This effect involves the elimination of the fast component of the biphasic dissociation curve. The allosteric effect of gallamine is equally prominent in membranes containing predominantly M1 (cerebral cortex) and M2 (brainstem) subtypes of muscarinic receptor. Gallamine's action is not affected by a variety of treatments which influence the conformational state of the receptor as reflected by agonist binding affinity, including treatments with heat, N-ethylmaleimide and trypsin. A guanine nucleotide (5'-guanylylimidodiphosphate), however, moderates the effects of gallamine on muscarinic receptors in brainstem, but not in cortical, membranes.