AT-1001 Is a Partial Agonist with High Affinity and Selectivity at Human and Rat α3β4 Nicotinic Cholinergic Receptors

Abstract

AT-1001 [N-(2-bromophenyl)-9-methyl-9-azabicyclo[3.3.1] nonan-3-amine] is a high-affinity and highly selective ligand at a3b4 nicotinic cholinergic receptors (nAChRs) that was reported to decrease nicotine self-administration in rats. It was initially reported to be an antagonist at rat a3b4 nAChRs heterologously expressed in HEK293 cells. Here we compared AT-1001 actions at rat and human a3b4 and a4b2 nAChRs similarly expressed in HEK 293 cells. We found that, as originally reported, AT-1001 is highly selective for a3b4 receptors over a4b2 receptors, but its binding selectivity is much greater at human than at rat receptors, because of a higher affinity at human than at rat a3b4 nAChRs. Binding studies in human and rat brain and pineal gland confirmed the selectivity of AT-1001 for a3b4 nAChRs and its higher affinity for human compared with rat receptors. In patch-clamp electrophysiology studies, AT-1001 was a potent partial agonist with 65-70% efficacy at both human and rat a3b4 nAChRs. It was also a less potent and weaker (18%) partial agonist at a4b2 nAChRs. Both a3b4 and a4b2 nAChRs are upregulated by exposure of cells to AT-1001 for 3 days. Similarly, AT-1001 desensitized both receptor subtypes in a concentration-dependent manner, but it was 10 and 30 times more potent to desensitize human a3b4 receptors than rat a3b4 and human a4b2 receptors, respectively. After exposure to AT-1001, the time to recovery from desensitization was longest for the human a3b4 nAChR and shortest for the human a4b2 receptor, suggesting that recovery from desensitization is primarily related to the dissociation of the ligand from the receptor.

Agonist Selectivity and Ion Permeation in the α3β4 Ganglionic Nicotinic Receptor

“…AT-1001 is a partial agonist that acts functionally as an antagonist by causing rapid desensitization (Cippitelli et al, 2015) (Figure 3B). This compound displays sub-nanomolar affinity to the human a3b4 receptor and is $200-fold selective for this subtype over a4b2 (Tuan et al, 2015). Our structure revealed that AT-1001 binds at the orthosteric sites (Figure 3B).…”

Determinants for α4β2 vs. α3β4 Subtype Selectivity of Pyrrolidine-Based nAChRs Ligands: A Computational Perspective with Focus on Recent cryo-EM Receptor Structures

“…Competition binding experiments at hα3rβ4 and hα3hβ4 also demonstrated that these amino acids were responsible for a modest but significant (14-fold) difference in the affinity of binding of TMAQ to nAChRs containing the human and rat α4 subunit. Similarly, in 2015, Tuan et al reported AT-1001 (Figure 2) as a human selective α3β4 nAChR ligand with similar K i s at human and rat α4β2 nAChRs, while 20 times higher affinity at the human than rat α3β4 nAChRs [43].…”

Activation of nicotinic acetylcholine receptors induces potentiation and synchronization within in vitro hippocampal networks

“…High concentrations of the α4β2 antagonist DhβE that are needed to block their activation with 90 µM nicotine can also block α3β4 (Harvey, Maddox, & Luetje, ). Consequently, to distinguish the contributions of α4β2 and α3β4 nAChRs to network activity, we used sazatidine‐A (SAZ‐A), a partial agonist at rat α4β2 nAChRs (DeDominicis et al, ; Tuan et al, ) which potently desensitizes rat α4β2 but not rat α3β4 nAChRs (Xiao et al, ). We also used AT‐1001, a partial agonist at rat α3β4 nAChRs that is known to desensitize rat α3β4 and human α4β2 nAChRs (Tuan et al, ).…”

“…The concentrations of SAZ‐A and AT‐1001 were chosen based on the maximal desensitization each drug would cause at its target receptor (Tuan et al, ; Xiao et al, ), which coincides with the maximal activation of each receptor. As they are partial agonists, the long application time will desensitize the target nAChR receptor, essentially giving rise to a ‘time‐averaged antagonism’ (Hulihan‐Giblin et al, ).…”