Jensen_2014_Eur.J.Pharmacol_740_703

The distribution and physiological functions of the alpha6 subunit-containing (alpha6()) nicotinic acetylcholine receptors in the central nervous system make them interesting putative therapeutic targets in several disorders. However, investigations into the receptors have been complicated by their inefficient functional expression in vitro. In the present study we have characterized and compared the pharmacological properties displayed by alpha6beta4 and alpha6beta4beta3 nicotinic acetylcholine receptors assembled in tsA201 cells from the classical alpha6/alpha3 chimera (C1) and two novel alpha6/alpha3 chimeras (C6(F223L) and C16(F223L)) identified in a recent study (Jensen et al., 2013). Whereas the Bmax values exhibited by [(3)H]epibatidine at wild-type alpha6beta4, C1beta4, C6(F223L)beta4 and C16(F223L)beta4 receptors differed substantially, the radioligand and seven orthosteric nicotinic agonists exhibited very similar KD and Ki values at the four receptors. In the FLIPR Membrane Potential Blue assay, the agonists exhibited the same rank order of potencies [(+/-)-epibatidine>sazetidine A>varenicline>(-)-cytisine~(S)-nicotine>acetylcholine>carbachol] at the C1beta4, C1beta4beta3, C6(F223L)beta4, C6(F223L)beta4beta3, C16(F223L)beta4 and C16(F223L)beta4beta3 receptors, albeit the absolute EC50 values differed somewhat between the receptors. Furthermore, four reference antagonists displayed the same rank order of inhibitory potencies at the six receptors [alpha-conotoxin PIA>2,2,6,6-tetramethylpiperidin-4-yl heptanoate>mecamylamine>dihydro-beta-erythroidine]. Although all interpretations based on these results should be made keeping the molecular modifications in the alpha6 surrogate subunits in mind, this study sheds light on the pharmacological properties of alpha6beta4() nicotinic acetylcholine receptors and demonstrates the applicability of the C6(F223L) and C16(F223L) chimeras for studies of these receptors.