Lethal effects of an insecticidal spider venom peptide involve positive allosteric modulation of insect nicotinic acetylcholine receptors

Abstract

Please cite this article as: Windley, M.J., Vetter, I., Lewis, R.J., Nicholson, G.M., Lethal effects of an insecticidal spider venom peptide involve positive allosteric modulation of insect nicotinic acetylcholine receptors, Neuropharmacology (2017), doi: 10.1016/j.neuropharm.2017.04.008. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Highlights• The previously characterized BK Ca channel block is not responsible for the lethal excitatory toxicity of κ-HXTX-Hv1c • κ-HXTX-Hv1c is a positive allosteric modulator of the insect nicotinic acetylcholine receptor which acts by prolonging current decay and reversing receptor desensitization • The likely lethal target of κ-HXTX-Hv1c is the nicotinic acetylcholine receptor, acting by a mechanism similar to that of the insecticide spinosyn A. A B S T R A C Tκ-Hexatoxins (κ-HXTXs) are a family of excitotoxic insect-selective neurotoxins from Australian funnel-web spiders that are lethal to a wide range of insects, but display no toxicity towards vertebrates. The prototypic κ-HXTX-Hv1c selectively blocks native and expressed cockroach largeconductance calcium-activated potassium (BK Ca or K Ca 1.1) channels, but not their mammalian orthologs. Despite this potent and selective action on insect K Ca 1.1 channels, we found that the classical K Ca 1.1 blockers paxilline, charybdotoxin and iberiotoxin, which all block insect K Ca 1.1 channels, are not lethal in crickets. We therefore used whole-cell patch-clamp analysis of cockroach dorsal unpaired median (DUM) neurons to study the effects of κ-HXTX-Hv1c on sodium-activated (K Na ), delayed-rectifier (K DR ) and 'A-type' transient (K A ) K + channels. 1 µM κ-HXTX-Hv1c failed to significantly inhibit cockroach K Na and K DR channels, but did cause a 30 ± 7% saturating inhibition of K A channel currents, possibly via a Kv4 (Shal-like) action. However, this modest action at such a high concentration of κ-HXTX-Hv1c would indicate a different lethal target. Accordingly, we assessed the actions of κ-HXTX-Hv1c on neurotransmitter-gated ion channels in cockroach DUM neurons. We found that κ-HXTX-Hv1c failed to produce any major effects on GABA A or glutamate-Cl receptors but dramatically slowed nicotine-evoked ACh receptor (nAChR) current decay and reversed nAChR desensitization. These actions occurred without any alterations to nAChR current amplitude or the nicotine concentration-response curve, and are consistent with a positive allosteric modulation of nAChRs. κ-HXTX-Hv1c therefore represents the first venom peptide that selectively modulates insect nAChRs with a mode of action similar to the excitotoxic insecticide spino...

Insecticidal spider toxins are high affinity positive allosteric modulators of the nicotinic acetylcholine receptor

“…Another consideration is that the nAChR subtype selectivity has only been explored for x-hexatoxin-Hv1a; so it remains possible that the hexatoxins can have diverse nAChR subtype selectivity, species selectivity or quality of effect when bound. In support of this, the study of Windley et al [21] shows that j-hexatoxin-Hv1c delays the decay time of slowly activating, non-desensitizing currents induced by nicotine application to cockroach neurons. These currents are mediated by the nAChN class of receptor [42] that is selective for spinosyn A and methyllycaconitine, so j-hexatoxin-Hv1c seems able to bind to more than one class of insect receptor.…”

“…In support of this, the study of Windley et al . shows that κ‐hexatoxin‐Hv1c delays the decay time of slowly activating, non‐desensitizing currents induced by nicotine application to cockroach neurons. These currents are mediated by the nAChN class of receptor that is selective for spinosyn A and methyllycaconitine, so κ‐hexatoxin‐Hv1c seems able to bind to more than one class of insect receptor.…”

“…Hexatoxin binding is also shown to allosterically modulate the binding of other ligands, supporting the electrophysiological studies of Windley et al . which concluded that κ‐hexatoxin‐Hv1c acts as a PAM of nAChRs. Our results suggest that all these hexatoxins primarily act in this way, since the potency of this effect is greater than that reported for any other molecular target.…”

Genomics, cys-loop ligand-gated ion channels and new targets for the control of insect pests and vectors

“…Several other classes of insecticides, such as spinosyns, sulfoxamines and butenolides, act on nAChRs [9]. Also, natural peptides found in venom, such as that of the funnel-web spider, can act on insect nAChRs, with a higher selectivity than on vertebrate nAChRs [10].…”

Identification and functional study of three nAChR regulators, ubiquilin‐1, PICK1, and CRELD2, in Locusta migratoria manilensis dorsal unpaired median neurons

“…; Windley et al . ). Numerous cys‐loop ligand‐gated ion channel genes have been annotated and characterized in the L. migratoria neuron transcriptome, including 39 putative nAChR genes (Wang et al .…”