Agonist-induced hump current production in heterologously-expressed human α4β2-nicotinic acetylcholine receptors  

作　　者：Qiang LIU Ke-wei YU Yong-chang CHANG Ronald J LUKAS Jie WU 

机构地区：[1]Divisions of Neurology , Barrow Neurological Institute, St Joseph's Hospital and Medical Center, Phoenix, Arizona 85013-4496, USA [2]Divisions of Neurobiology, Barrow Neurological Institute, St Joseph's Hospital and Medical Center, Phoenix, Arizona 85013-4496, USA

出　　处：《Acta Pharmacologica Sinica》2008年第3期305-319,共15页中国药理学报（英文版）

摘　　要：Aim： To characterize the functional and pharmacological features of agonistinduced 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 acetylcholine （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 removal 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 external Ca^2＋. Outside-out single-channel recordings demonstrated 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 agonistbinding site in the channel pore. Conclusion： These results support the hypothesis that hump currents reflect a rapid release of agonists from the α4β2-nAChR channel pore and a rapid recovery from desensitized α4β2-nAChR.Aim： To characterize the functional and pharmacological features of agonistinduced 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 acetylcholine （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 removal 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 external Ca^2＋. Outside-out single-channel recordings demonstrated 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 agonistbinding site in the channel pore. Conclusion： These results support the hypothesis that hump currents reflect a rapid release of agonists from the α4β2-nAChR channel pore and a rapid recovery from desensitized α4β2-nAChR.

关 键 词：ACETYLCHOLINE dihydro-β-erythroidine dimethyl-phenyl-piperazinium EPIBATIDINE nicotinic acetylcholine receptor 

分 类 号：R96[医药卫生—药理学]