IPPA08 allosterically enhances the action of imidacloprid on nicotinic acetylcholine receptors

Our previous study showed that IPPA08, a cis-configuration neonicotinoid compound with unique oxabridged substructure, acted as a specific synergist to neonicotinoid insecticides targeting nicotinic acetylcholine receptors (nAChRs). Heteropentamer nAChRs have diverse characteristics and can form canonical and noncanonical subunit interfaces. While canonical interfaces have been exploited as targets of many drugs, noncanonical interfaces have received less attention. In this study, the mechanism of IPPA08 synergism was evaluated on hybrid nAChRs consisting of three alpha 1 subunits from the brown planthopper and two rat beta 1 subunits (Nl alpha 1/r beta 2) expressed in Xenopus oocytes. IPPA08 alone evoked inward currents, but only at very high concentrations, greater than 1 mM. However, at concentrations below 200 mu M, IPPA08 slowed the decay of inward currents evoked by imidacloprid, but not by acetylcholine, and also increased the sensitivity of Nl alpha 1/r beta 2 to imidacloprid. Both modulations by IPPA08 were concentration-dependent in the same concentration range of 10-150 mu M. Experimentally induced mutations in canonical (alpha+/beta-) and noncanonical (beta+/alpha-) interfaces of NI alpha 1/r beta 2 receptors were also examined to evaluate the presence of possible binding sites for IPPA08 on the receptors. Our results showed that mutations in the canonical interfaces affected only the potency of IPPA08 as an agonist, while mutations in the noncanonical interfaces affected only the synergistic action of IPPA08. Based on these results, we propose that at low concentrations IPPA08 can act as a positive allosteric modulator of noncanonical interfaces, and likely slow the decay of currents through stabilizing the open-channel state caused by the action of imidacloprid on canonical interfaces. (C) 2016 Elsevier Ltd. All rights reserved.