Title : The unique alpha4(+)\/(-)alpha4 agonist binding site in (alpha4)3(beta2)2 subtype nicotinic acetylcholine receptors permits differential agonist desensitization pharmacology versus the (alpha4)2(beta2)3 subtype - Eaton_2014_J.Pharmacol.Exp.Ther_348_46 |
Author(s) : Eaton JB , Lucero LM , Stratton H , Chang Y , Cooper JF , Lindstrom JM , Lukas RJ , Whiteaker P |
Ref : Journal of Pharmacology & Experimental Therapeutics , 348 :46 , 2014 |
Abstract :
Selected nicotinic agonists were used to activate and desensitize high-sensitivity (HS) (alpha4)2(beta2)3) or low-sensitivity (LS) (alpha4)3(beta2)2) isoforms of human alpha4beta2-nicotinic acetylcholine receptors (nAChRs). Function was assessed using (86)Rb(+) efflux in a stably transfected SH-EP1-halpha4beta2 human epithelial cell line, and two-electrode voltage-clamp electrophysiology in Xenopus laevis oocytes expressing concatenated pentameric HS or LS alpha4beta2-nAChR constructs (HSP and LSP). Unlike previously studied agonists, desensitization by the highly selective agonists A-85380 [3-(2(S)-azetidinylmethoxy)pyridine] and sazetidine-A (Saz-A) preferentially reduced alpha4beta2-nAChR HS-phase versus LS-phase responses. The concatenated-nAChR experiments confirmed that approximately 20% of LS-isoform acetylcholine-induced function occurs in an HS-like phase, which is abolished by Saz-A preincubation. Six mutant LSPs were generated, each targeting a conserved agonist binding residue within the LS-isoform-only alpha4(+)/(-)alpha4 interface agonist binding site. Every mutation reduced the percentage of LS-phase function, demonstrating that this site underpins LS-phase function. Oocyte-surface expression of the HSP and each of the LSP constructs was statistically indistinguishable, as measured using beta2-subunit-specific [(125)I]mAb295 labeling. However, maximum function is approximately five times greater on a "per-receptor" basis for unmodified LSP versus HSP alpha4beta2-nAChRs. Thus, recruitment of the alpha4(+)/(-)alpha4 site at higher agonist concentrations appears to augment otherwise-similar function mediated by the pair of alpha4(+)/(-)beta2 sites shared by both isoforms. These studies elucidate the receptor-level differences underlying the differential pharmacology of the two alpha4beta2-nAChR isoforms, and demonstrate that HS versus LS alpha4beta2-nAChR activity can be selectively manipulated using pharmacological approaches. Since alpha4beta2 nAChRs are the predominant neuronal subtype, these discoveries likely have significant functional implications, and may provide important insights for drug discovery and development. |
PubMedSearch : Eaton_2014_J.Pharmacol.Exp.Ther_348_46 |
PubMedID: 24190916 |
Eaton JB, Lucero LM, Stratton H, Chang Y, Cooper JF, Lindstrom JM, Lukas RJ, Whiteaker P (2014)
The unique alpha4(+)\/(-)alpha4 agonist binding site in (alpha4)3(beta2)2 subtype nicotinic acetylcholine receptors permits differential agonist desensitization pharmacology versus the (alpha4)2(beta2)3 subtype
Journal of Pharmacology & Experimental Therapeutics
348 :46
Eaton JB, Lucero LM, Stratton H, Chang Y, Cooper JF, Lindstrom JM, Lukas RJ, Whiteaker P (2014)
Journal of Pharmacology & Experimental Therapeutics
348 :46