Article

Pharmacological and functional comparisons of α6/ α3β2β3-nAChRs and α4β2-nAChRs heterologously expressed in the human epithelial SH-EP1 cell line

Authors: De-jie CHEN1,2, Fen-fei GAO2,3, Xiao-kuang MA2,3, Gang-gang SHI3, Yuan-bing HUANG1,2, Quang-xi SU1, Sterling SUD-WEEKS4, Ming GAO2, Turner DHARSHAUN2, Jason Brek EATON2, Yong-chang CHANG2, J Michael MCINTOSH5,6, Ronald J LUKAS2, Paul WHITEAKER2, Scott C STEFFENSEN7, Jie WU1,2,3
1 Department of Neurology, Yunfu People’s Hospital, Yunfu 527300, China
2 Department of Neurobiology, Barrow Neurological Institute, St Joseph’s Hospital and Medical Center, Phoenix, AZ 85013, USA
3 Department of Pharmacology, Shantou University Medical College, Shantou 515063, China
4 Departments of Psychology and Developmental Biology, Brigham Young University, Provo, UT 84602, USA
5 George E Wahlen Veterans Affairs Medical Center, Salt Lake City, UT 84108, USA
6 Departments of Psychiatry and Biology, University of Utah, Salt Lake City, UT 84112, USA
7 Department of Physiology and Neuroscience, Brigham Young University, Provo, UT 84602, USA
Correspondence to: Jie WU: jie.wu@DignityHealth.org,
DOI: 10.1038/aps.2017.209
Received: 30 September 2017
Accepted: 21 December 2017
Advance online: 24 May 2018

Abstract

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. Patch-clamp 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 subunit-selective antagonist α-conotoxin MII 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.
Keywords: nicotinic acetylcholine receptor; nicotine; acetylcholine; SH-EP1 cells; patch-clamp

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