Elsevier

Neuropharmacology

Volume 45, Issue 1, July 2003, Pages 133-144
Neuropharmacology

Diverse actions of neonicotinoids on chicken α7, α4β2 and Drosophila–chicken SADβ2 and ALSβ2 hybrid nicotinic acetylcholine receptors expressed in Xenopus laevis oocytes

https://doi.org/10.1016/S0028-3908(03)00134-5Get rights and content

Abstract

The 2-nitroimino-imidazolidine and related moieties are structural features of neonicotinoid insecticides acting on nicotinic acetylcholine receptors (nicotinic AChRs). To evaluate these moieties in neonicotinoid interactions with nicotinic AChR α subunits, the actions of imidacloprid and related compounds on the chicken α7, α4β2 and Drosophila melanogaster-chicken hybrid (SADβ2 and ALSβ2) receptors expressed in Xenopus laevis oocytes were studied by voltage-clamp electrophysiology. Imidacloprid and nitenpyram were partial agonists and a nitromethylene analog of imidacloprid (CH-IMI) was a full agonist of the α7 receptor, whereas their agonist actions on the α4β2 receptor were very weak, contrasting with full agonist actions of DN-IMI, a desnitro derivative of imidacloprid. The neonicotinoids and DN-IMI were either full or partial agonists of the SADβ2 receptors. Nitenpyram and DN-IMI were partial agonists of the ALSβ2 receptor, whereas imidacloprid and CH-IMI scarcely activated the ALSβ2 receptor. Imidacloprid and CH-IMI in fact suppressed ACh-induced responses of the ALSβ2 receptor, whereas imidacloprid potentiated and CH-IMI suppressed ACh-induced responses of the α4β2 receptor. These results suggest that interactions with α subunits of the 2-nitroimino-imidazolidine moiety of imidacloprid play a role in determining not only agonist and antagonist actions on all four receptors, but also the potentiation of ACh-induced responses of the α4β2 receptor.

Introduction

Acetylcholine receptors (AChRs) of the nicotinic type mediate fast excitatory cholinergic synaptic transmission at neuromuscular junctions and in the nervous system of vertebrates. In insects, nicotinic AChRs are present at high concentrations in the nervous system (Breer and Sattelle, 1987) and are targeted by several classes of insecticides (Narahashi, 2000). Imidacloprid and related insecticides such as nitenpyram and acetamiprid (Fig. 1) are referred to as neonicotinoids and act on nicotinic AChRs (Matsuda et al., 2001). Evidence for the actions of neonicotinoids and their prototype compounds on nicotinic AChRs is based on electrophysiologial (Schroeder and Flattum, 1984, Sattelle et al., 1989, Bai et al., 1991, Zwart et al., 1994) and radioligand binding (Tomizawa and Yamamoto, 1992, Tomizawa and Yamamoto, 1993, Liu et al., 1993) assays. The selective toxicity of the neonicotinoids results, at least in part, from a higher affinity for insect nicotinic AChRs (Liu and Casida, 1993, Zwart et al., 1994, Yamamoto et al., 1995). There is evidence that the interaction of a part of the nitroguanidine moiety of imidacloprid with the α subunit of the nicotinic AChRs contributes to its partial agonist actions and its selectivity for insect nicotinic AChRs (Matsuda et al., 1998). However, it has been shown that [3H]-imidacloprid binding to the Drosophila SAD-rat β2 hybrid receptor expressed in Drosophila S2 cells is markedly reduced by replacement of the β2 subunit by the β4 subunit, suggesting a role of non-α subunits in interactions with imidacloprid (Lansdell and Millar, 2000).

A number of derivatives and analogues of imidacloprid have been generated to date. Using crude membrane preparations (Tomizawa and Yamamoto, 1992, Tomizawa and Yamamoto, 1993, Liu et al., 1993, Tomizawa and Casida, 1999, D’Amour and Casida, 1999, Zhang et al., 2000), binding affinities have been determined for a limited number of neonicotinoids. Preparations developed to generate membrane preparations include the Torpedo electric organ tissue, human SH-SY5Y cells, rodent brain tissues containing several combinations of nicotinic AChRs (Tomizawa and Casida, 1999) and insect Sf9 cells expressing the rat α4β2 nicotinic AChR (D’Amour and Casida, 1999). Despite the accumulation of such binding data, little is known about how variations in neonicotinoid structure affect their agonist or antagonist actions on nicotinic AChRs of known composition. The aim of this study is to investigate, using voltage-clamp electrophysiology, how the nitroguanidine and analogous moieties, as well as the imidazolidine ring structure, of neonicotinoids (Fig. 1) influence either agonist or antagonist actions of imidacloprid and related compounds on homomeric α7 and heteromeric α4β2 neuronal nicotinic AChRs and Drosophila-chicken SADβ2 and ALSβ2 hybrid heteromeric receptors (Bertrand et al., 1994). We demonstrate that responses to neonicotinoids of the four recombinant nicotinic AChRs tested (α7, α4β2, SADβ2 and ALSβ2) differ strikingly in response to structural changes in these ligands, indicating an important role for the α subunit. These results are of interest in the context of designing new and safer insecticides.

Section snippets

Expression of nicotinic AChRs in Xenopus oocytes by nuclear injection of cDNA

Oocytes at stage V or VI of development were removed under anesthetic (immersion in 1.5 g l−1 tricaine for 30–45 min) from adult female Xenopus laevis (see Matsuda et al., 2000) and then separated from the follicle cell layer by treatment with 2 mg ml−1 collagenase (Sigma-Aldrich Japan, Tokyo, Japan; Type IA) for 30 min at room temperature (19–25 °C). The authors made as much effort as possible to minimize animal suffering and reduce the number of animals used in accordance with the UK Animals

Agonist actions of neonicotinoids on the chicken α7 nicotinic AChR

Imidacloprid, CH-IMI and nitenpyram resulted in inward currents when bath-applied to oocytes expressing α7 nicotinic AChRs (Fig. 2a i). The currents were rapidly activated and also desensitized during the application. The maximum normalized current of the imidacloprid and nitenpyram-induced responses at saturating concentrations were 0.53 (n=7) and 0.25 (n=4) (Table 1). Thus, these two insecticides are partial agonists on the α7 receptor. The maximum current size of the response to a

Discussion

The nitroguanidine moiety of imidacloprid has been shown to contribute to its partial agonist action as well as its selectivity for insect nicotinic AChRs (Matsuda et al., 1998). Agonist profiles measured for the neonicotinoids on the recombinant nicotinic receptors revealed that structural changes in the nitroguanidine moiety of imidacloprid affect not only the affinity but also the efficacy of neonicotinoids on neuronal nicotinic AChRs (Fig. 4). It has been shown that neonicotinoids can have

Acknowledgements

This work was supported in part by a Grant-in-Aid for Scientific Research (C) from the Japan Society for the Promotion of Science (No. 13660113) and the Program for Promotion for Basic Research Activities for Innovative Biosciences (Bio-oriented Technology Research Advancement Institution) to K. Matsuda as well as the Medical Research Council of the UK (V. Raymond and D. B. Sattelle). The authors wish to thank Professor Mark Ballivet of The University of Geneva, Switzerland and Professor H.

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