Ligand-gated ion channels as targets of neuroactive insecticides

The Cys-loop superfamily of ligand-gated ion channels (Cys-loop receptors) is one of the most ubiquitous ion channel families in vertebrates and invertebrates. Despite their ubiquity, they are targeted by several classes of pesticides, including neonicotinoids, phenylpyrazols, and macrolides such as ivermectins. The current commercialized compounds have high target site selectivity, which contributes to the safety of insecticide use. Structural analyses have accelerated progress in this field; notably, the X-ray crystal structures of acetylcholine binding protein and glutamate-gated Cl channels revealed the details of the molecular interactions between insecticides and their targets. Recently, the functional expression of the insect nicotinic acetylcholine receptor (nAChR) has been described, and detailed evaluations using the insect nAChR have emerged. This review discusses the basic concepts and the current insights into the molecular mechanisms of neuroactive insecticides targeting the ligand-gated ion channels, particularly Cys-loop receptors, and presents insights into target-based selectivity, resistance, and future drug design.

A Novel and Functionally Diverse Class of Acetylcholine-gated Ion Channels

Fast cholinergic neurotransmission is mediated by pentameric acetylcholine-gated ion channels; in particular, cationic nicotinic acetylcholine receptors play well-established roles in virtually all nervous systems. Acetylcholine-gated anion channels have also been identified in some invertebrate phyla, yet their roles in the nervous system are less well-understood. Here we describe the functional properties of five previously-uncharacterized acetylcholine-gated anion channels from C. elegans, including four from a novel nematode specific subfamily known as the diverse group. In addition to their activation by acetylcholine, these diverse group channels are activated at physiological concentrations by other ligands; three, encoded by the lgc-40, lgc-57 and lgc-58 genes, are activated by choline, while lgc-39 encoded channels are activated by octopamine and tyramine. Intriguingly, these and other acetylcholine-gated anion channels show extensive co-expression with cation-selective nicotinic receptors, implying that many cholinergic synapses may have both excitatory and inhibitory potential. Thus, the evolutionary expansion of cholinergic ligand-gated ion channels may enable complex synaptic signalling in an anatomically compact nervous system.

Structural dynamics determine voltage and pH gating in human voltage-gated proton channel

Voltage-gated ion channels are key players of electrical signaling in cells. As a unique subfamily, voltage-gated proton (Hv) channels are standalone voltage sensors without separate ion conductive pores. They are gated by both voltage and transmembrane proton gradient (i.e ΔpH), serving as acid extruders in most cells. Amongst their many functions, Hv channels are known to regulate the intracellular pH of human spermatozoa and compensate for the charge and pH imbalances caused by NADPH oxidases in phagocytes. Like the canonical voltage sensors, the Hv channel is a bundle of 4 helices (named S1 through S4), with the S4 segment carrying 3 positively charged Arg residues. Extensive structural and electrophysiological studies on voltage-gated ion channels generally agree on an outwards movement of the S4 segment upon activating voltage, but the real time conformational transitions are still unattainable. With purified human voltage-gated proton (hHv1) channel reconstituted in liposomes, we have examined its conformational dynamics at different voltage and pHs using the single molecule fluorescence resonance energy transfer (smFRET). Here we provided the first glimpse of real time conformational trajectories of the hHv1 voltage sensor and showed that both voltage and pH gradient shift the conformational dynamics of the S4 segment to control channel gating. Our results suggested the biological gating is determined by the conformational distributions of the hHv1 voltage sensor, rather than the conformational transitions between the presumptive ‘resting’ and ‘activated’ conformations. We further identified H140 as the key residue sensing extracellular pH and showed that both the intracellular and extracellular pH sensors act on the voltage sensing S4 segment to enrich the resting conformations. Taken together, we proposed a model that explains the mechanisms underlying voltage and pH gating in Hv channels, which may also serve as a general framework to understand the voltage sensing and gating in other voltage-gated ion channels.