In this study, we report the effects of the quinoline derivatives quinine, its optical isomer quinidine, and chloroquine on alpha9alpha10-containing nicotinic acetylcholine receptors (nAChRs). The compounds blocked acetylcholine (ACh)-evoked responses in alpha9alpha10-injected Xenopus laevis oocytes in a concentration-dependent manner, with a rank order of potency of chloroquine (IC50 = 0.39 microM) > quinine (IC50 = 0.97 microM) approximately quinidine (IC50= 1.37 microM). Moreover, chloroquine blocked ACh-evoked responses on rat cochlear inner hair cells with an IC50 value of 0.13 microM, which is within the same range as that observed for recombinant receptors. Block by chloroquine was purely competitive, whereas quinine inhibited ACh currents in a mixed competitive and noncompetitive manner. The competitive nature of the blockage produced by the three compounds was confirmed by equilibrium binding experiments using [3H]methyllycaconitine. Binding affinities (Ki values) were 2.3, 5.5, and 13.0 microM for chloroquine, quinine, and quinidine, respectively. Block by quinine was found to be only slightly voltage-dependent, thus precluding open-channel block as the main mechanism of interaction of quinine with alpha9alpha10 nAChRs. The present results add to the pharmacological characterization of alpha9alpha10-containing nicotinic receptors and indicate that the efferent olivocochlear system that innervates the cochlear hair cells is a target of these ototoxic antimalarial compounds.