Abstract
Drug development for nicotinic acetylcholine receptors (nAChR) is challenged by subtype diversity arising from variations in subunit composition. On-target activity for neuronal heteromeric receptors is typically associated with CNS receptors that contain α4 and other subunits, while off-target activity could be associated with ganglionic-type receptors containing α3β4 binding sites and other subunits, including β4, β2, α5, or α3 as a structural subunit in the pentamer. Additional interest in α3 β4 α5-containing receptors arises from genome-wide association studies linking these genes, and a single nucleotide polymorphism (SNP) in α5 in particular, to lung cancer and heavy smoking. While α3 and β4 readily form receptors in expression system such as the Xenopus oocyte, since α5 is not required for function, simple co-expression approaches may under-represent α5-containing receptors. We used a concatamer of human α3 and β4 subunits to form ligand-binding domains, and show that we can force the insertions of alternative structural subunits into the functional pentamers. These α3β4 variants differ in sensitivity to ACh, nicotine, varenicline, and cytisine. Our data indicated lower efficacy for varenicline and cytisine than expected for β4-containing receptors, based on previous studies of rodent receptors. We confirm that these therapeutically important α4 receptor partial agonists may present different autonomic-based side-effect profiles in humans than will be seen in rodent models, with varenicline being more potent for human than rat receptors and cytisine less potent. Our initial characterizations failed to find functional effects of the α5 SNP. However, our data validate this approach for further investigations.
Copyright © 2012 Elsevier Ltd. All rights reserved.
Publication types
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Comparative Study
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Acetylcholine / agonists
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Acetylcholine / antagonists & inhibitors
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Acetylcholine / metabolism*
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Alkaloids / metabolism
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Alkaloids / pharmacology
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Animals
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Azocines / metabolism
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Azocines / pharmacology
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Benzazepines / metabolism
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Benzazepines / pharmacology
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Drug Partial Agonism
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Evoked Potentials / drug effects
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Ganglia / metabolism*
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Humans
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Ligands
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Mutagenesis, Site-Directed
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Mutant Proteins / agonists
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Mutant Proteins / antagonists & inhibitors
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Mutant Proteins / genetics
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Mutant Proteins / metabolism
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Nerve Tissue Proteins / agonists
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Nerve Tissue Proteins / antagonists & inhibitors
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Nerve Tissue Proteins / genetics
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Nerve Tissue Proteins / metabolism*
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Nicotine / agonists
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Nicotine / antagonists & inhibitors
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Nicotine / metabolism
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Nicotinic Agonists / metabolism*
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Nicotinic Agonists / pharmacology
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Nicotinic Antagonists / metabolism*
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Nicotinic Antagonists / pharmacology
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Oocytes / drug effects
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Oocytes / metabolism
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Protein Subunits / agonists
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Protein Subunits / antagonists & inhibitors
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Protein Subunits / genetics
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Protein Subunits / metabolism*
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Quinolizines / metabolism
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Quinolizines / pharmacology
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Quinoxalines / metabolism
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Quinoxalines / pharmacology
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Rats
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Receptors, Nicotinic / chemistry
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Receptors, Nicotinic / genetics
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Receptors, Nicotinic / metabolism*
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Recombinant Fusion Proteins / agonists
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Recombinant Fusion Proteins / antagonists & inhibitors
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Recombinant Fusion Proteins / metabolism
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Species Specificity
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Varenicline
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Xenopus laevis
Substances
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Alkaloids
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Azocines
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Benzazepines
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Ligands
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Mutant Proteins
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Nerve Tissue Proteins
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Nicotinic Agonists
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Nicotinic Antagonists
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Protein Subunits
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Quinolizines
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Quinoxalines
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Receptors, Nicotinic
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Recombinant Fusion Proteins
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nicotinic receptor alpha3beta4
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cytisine
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Nicotine
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Acetylcholine
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Varenicline