Allosteric Antagonism of Insect Odorant Receptor Ion Channels

Abstract

BackgroundAt a molecular level, insects utilize members of several highly divergent and unrelated families of cell-surface chemosensory receptors for detection of volatile odorants. Most odors are detected via a family of odorant receptors (ORs), which form heteromeric complexes consisting of a well-conserved OR co-receptor (Orco) ion channel and a non-conserved tuning OR that provides coding specificity to each complex. Orco functions as a non-selective cation channel and is expressed in the majority of olfactory receptor neurons (ORNs). As the destructive behaviors of many insects are principally driven by olfaction, Orco represents a novel target for behavior-based control strategies. While many natural and synthetic odorants have been shown to agonize Orco/Or complexes, only a single direct Orco modulator, VUAA1, has been described. In an effort to identify additional Orco modulators, we have investigated the structure/activity relationships around VUAA1.ResultsA search of our compound library identified several VUAA1 analogs that were selected for evaluation against HEK cells expressing Orco from the malaria vector Anopheles gambiae (AgOrco). While the majority of compounds displayed no activity, many of these analogs possess no intrinsic efficacy, but instead, act as competitive VUAA1 antagonists. Using calcium mobilization assays, patch clamp electrophysiology, and single sensillum in vivo recording, we demonstrate that one such candidate, VU0183254, is a specific allosteric modulator of OR signaling, capable of broadly inhibiting odor-mediated OR complex activation.ConclusionsWe have described and characterized the first Orco antagonist, that is capable of non-competitively inhibiting odorant-evoked activation of OR complexes, thereby providing additional insight into the structure/function of this unique family of ligand-gated ion channels. While Orco antagonists are likely to have limited utility in insect control programs, they represent important pharmacological tools that will facilitate the investigation of the molecular mechanisms underlying insect olfactory signal transduction.

Detection of sweet tastants by a conserved group of insect gustatory receptors

“…Previous studies have shown that nonvolatile agonists or antagonists of the Or coreceptor, Orco, are able to act on olfactory neurons simply by their inclusion in the recording micropipette (28,29). Others have reported injected delivery of inhibitors into the sensillum lymph that mimics the effects of in vitro studies (30)(31)(32).…”

Odorant receptor-mediated sperm activation in disease vector mosquitoes

“…8-Bromo-cAMP was obtained from Sigma-Aldrich, Inc. (Cat# B5386). VUAA-class compounds were prepared as previously described (8,31,32). All other compounds were obtained from Sigma-Aldrich at the highest purity available.…”

“…Responses were scored by post hoc examination of video clips in a double-blinded fashion (Movie S1). The stimuli used in these assays were comprised of a range of unitary odorants as well as highly specific Orco modulators that have been recently characterized (8,31,32). Flagellar beating responses were significantly elevated in the presence of two Orco agonists, VUAA1 and VUAA4, but not in the presence of identical concentrations of a nonpotent structural analog, VUAA0 (32) (Fig.…”

Mutational Analysis of Cysteine Residues of the Insect Odorant Co-receptor (Orco) from Drosophila melanogaster Reveals Differential Effects on Agonist- and Odorant-tuning Receptor-dependent Activation

“…Screening of cell lines expressing these homomeric channels led to the discovery of VUAA1 as a novel allosteric agonist of insect ORs (16). Further exploration of the structure-* This work was supported by Grant PAF-09-01 from the Marsden Fund activity relationships of VUAA1 identified several more potent Orco allosteric agonists (17)(18)(19). A number of compounds structurally related to VUAA1 that are inactive as agonists act as competitive inhibitors of VUAA1 (17,18,20).…”