Neuronal nicotinic acetylcholine receptors containing α6 subunits (α6*-nAChRs) show highly restricted distribution in midbrain neurons associated with pleasure,reward,and mood control,suggesting an important impact of α6*-nAChRs in modulating mesolimbic functions.However,the function and pharmacology of α6*-nAChRs remain poorly understood because of the lack of selective agonists for α6*-nAChRs and the challenging heterologous expression of functional α6*-nAChRs in mammalian cell lines.In particular,the α6 subunit is commonly co-expressed with α4*-nAChRs in the midbrain,which masks α6*-nAChR (without α4) function and pharmacology.In this study,we systematically profiled the pharmacology and function of α6*-nAChRs and compared these properties with those of α4β2 nAChRs expressed in the same cell line.Heterologously expressed human α6/α3 chimeric subunits (α6 N-terminal domain joined with α3 trans-membrane domains and intracellular loops) with β2 and β3 subunits in the human SH-EP1 cell line (α6*-nAChRs) were used.Patchclamp whole-cell recordings were performed to measure these receptor-mediated currents.Functionally,the heterologously expressed α6*-nAChRs exhibited excellent function and showed distinct nicotine-induced current responses,such as kinetics,inward rectification and recovery from desensitization,compared with α4β2-nAChRs.Pharmacologically,α6*-nAChR was highly sensitive to the α6 subunitselective antagonist α-conotoxin MIl but had lower sensitivity to mecamylamine and dihydro-β-erythroidine.Nicotine and acetylcholine were found to be full agonists for α6*-nAChRs,whereas epibatidine and cytisine were determined to be partial agonists.Heterologously expressed α6*-nAChRs exhibited pharmacology and function distinct from those of α4β2-nAChRs,suggesting that α6*-nAChRs may mediate different cholinergic signals.Our α6*-nAChR expression system can be used as an excellent cell model for future investigations of α6*-nAChR function and pharmacology.