Î±-Conotoxins Identify The Î±3Î²4Âˆ— Subtype As The Predominant Nicotinic Acetylcholine Receptor Expressed In Human Adrenal Chromaffin Cells
Ligands that selectively inhibit human Î±3Î²2 and Î±6Î²2 nicotinic acetylcholine receptor (nAChRs) and not the closely related Î±3Î²4 and Î±6Î²4 subtypes are lacking. Current Î±-conotoxins (Î±-Ctxs) that discriminate among these nAChR subtypes in rat fail to discriminate among the human receptor homologs. In this study, we describe the development of Î±-Ctx LvIA(N9R,V10A) that is 3000-fold more potent on oocyte-expressed human Î±3b2 than Î±3b4 and 165-fold more potent on human a6/a3Î²2Î²3 than a6/ a3Î²4 nAChRs. This analog was used in conjuction with three other Î±-Ctx analogs and patch-clamp electrophysiology to characterize the nAChR subtypes expressed by human adrenal chromaffin cells. LvIA(N9R,V10A) showed little effect on the acetylcholine-evoked currents in these cells at concentrations expected to inhibit nAChRs with Î²2 ligand-binding sites. In contrast, the b4-selective Î±-Ctx BuIA(T5A,P6O) inhibited .98% of the acetylcholine-evoked current, indicating that most of the heteromeric receptors contained b4 ligand-binding sites. Additional studies using the Î±6-selective Î±-Ctx PeIA(A7V,S9H,V10A, N11R,E14A) indicated that the predominant heteromeric nAChR expressed by human adrenal chromaffin cells is the a3b4âˆ— subtype (asterisk indicates the possible presence of additional subunits). This conclusion was supported by polymerase chain reaction experiments of human adrenal medulla gland and of cultured human adrenal chromaffin cells that demonstrated prominent expression of RNAs for Î±3, Î±5, Î±7, Î²2, and Î²4 subunits and a low abundance of RNAs for Î±2, Î±4, Î±6, and Î±10 subunits.
Digital Object Identifier (DOI)
Hone, Arik J.; McIntosh, J. Michael; Azam, Layla; Lindstrom, Jon; Lucero, Linda; Whiteaker, Paul; Passas, Juan; BlÃ¡zquez, JesÃºs; and Albillos, Almudena, "Î±-Conotoxins Identify The Î±3Î²4Âˆ— Subtype As The Predominant Nicotinic Acetylcholine Receptor Expressed In Human Adrenal Chromaffin Cells" (2015). Neurobiology. 389.