Aim:To characterize the functional and pharmacological features of agonist-induced hump currents in human α4β2-nicotinic acetylcholine receptors (nAChR).Methods:Whole-cell and outside-out patch recordings were performed using human α4β2-nAChR heterologously expressed in stably-transfected,native nAChR-null subclonal human epithelial 1 (SH-EP1) cells.RT-PCR was used to test the mRNA expression of transfected nAChR.Homology modeling and ace-tylcholine (Ach) docking were applied to show the possible Ach-binding site in the channel pore.Results:The rapid exposure of 10 mmol/L Ach induced an inward current with a decline from peak to steady-state.However,after the re-moval of Ach,an additional inward current,called "hump" current,reoccurred.The ability of agonists to produce these hump currents cannot be easily explained based on drug size,charge,acute potency,or actions as full or partial agonists.Hump currents were associated with a rebound increase in whole-cell conductance,and they had voltage dependence-like peak currents induced by agonist action.Hump currents blocked by the α4β2-nAChR antagonist dihydro-β-erythroidine were reduced when α4β2-nAChR were desensitized,and were more pronounced in the absence of exteal Ca2+.Outside-out single-channel recordings demon-strated that compared to 1 μmol/L nicotine,100 μmol/L nicotine reduced channel current amplitude,shortened the channel mean open time,and prolonged the channel mean closed time,supporting an agonist-induced open-channel block before hump current production.A docking model also simulated the agonist-binding site in the channel pore.Conclusion:These results support the hypoth-esis that hump currents reflect a rapid release of agonists from the α4β2-nAChR channel pore and a rapid recovery from desensitized α4β2-nAChR.