Activation of the α7 nicotinic acetylcholine receptor typically occurs upon binding of agonists at the receptor orthosteric binding site and is characterized by intrinsically low open probability. Positive allosteric modulators (PAMs) reverse this limitation by binding in the transmembrane domain, which promotes prolonged receptor activity. While PAM effects are linked to the presence of an orthosteric agonist, ago-PAMs are able to evoke α7 ion channel currents in the absence of orthosteric ligands via a direct allosteric activation (DAA) site in the extracellular domain. At present, allosteric agonists displaying DAA but not PAM activity have been not reported. We here identify 1,1-diethyl-4-(naphthalene-2-yl)piperazin-1-ium (2NDEP) as the first α7 allosteric agonist that selectively activates α7 via DAA with no intrinsic PAM component, as confirmed by the requirement of PAM co-application. Despite being formerly reported as an α7 partial agonist potentiated by type II PAM co-application, our present investigation highlighted favorable docking and MMPBSA binding energies at the α7 DAA site for 2NDEP. We therefore hypothesized that part of the PAM-potentiated responses observed for 2NDEP are ascribable to the coupling between the PAM with the DAA site rather than the orthosteric site. To verify this hypothesis, we tested 2NDEP on the α7 Y93C and C190A mutants, which are known to be insensitive to orthosteric agonists but effectively activated by ago-PAMs like GAT107. 2NDEP acted as an antagonist of GAT107 on Y93C, and as an allosteric agonist of C190A when co-applied with the type II PAM PNU-120596. Moreover, co-application with the DAA site-selective antagonist 4-(2,3,5,6-tetramethylphenyl)-3a,4,5,9btetrahydro-3H-cyclopenta[c]quinoline-8-sulfonamide (2,3,5,6MP-TQS) nearly abolished the 2NDEP allosteric activity. These results were consistent with involvement of the DAA site and were further investigated in computational analyses. Overall, our findings reveal a novel mode for α7 agonist activation via the DAA site and offer new structural insights to develop purely allosteric α7 ligands.

2NDEP highlights allosteric activation of the α7 nicotinic acetylcholine receptor

2019 Spring National Meeting

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.