Abstract
Acetylcholine receptors (AChRs) are pentameric ligand-gated ion channels involved in the neurotransmission of both vertebrates and invertebrates. A number of anthelmintic compounds like levamisole and pyrantel target the AChRs of nematodes producing spastic paralysis of the worms. The muscle AChRs of nematode parasites fall into three pharmacological classes that are preferentially activated by the cholinergic agonists levamisole (L-type), nicotine (N-type) and bephenium (B-type), respectively. Despite a number of studies of the B-type AChR in parasitic species, this receptor remains to be characterized at the molecular level. Recently, we have reconstituted and functionally characterized two distinct L-AChR subtypes of the gastro-intestinal parasitic nematode Haemonchus contortus in the Xenopus laevis oocyte expression system by providing the cRNAs encoding the receptor subunits and three ancillary proteins (Boulin et al. in Br J Pharmacol 164(5):1421–1432, 2011). In the present study, the effect of the bephenium drug on Hco-L-AChR1 and Hco-L-AChR2 subtypes was examined using the two-microelectrode voltage-clamp technique. We demonstrate that bephenium selectively activates the Hco-L-AChR1 subtype made of Hco-UNC-29.1, Hco-UNC-38, Hco-UNC-63, Hco-ACR-8 subunits that is more sensitive to levamisole than acetylcholine. Removing the Hco-ACR-8 subunit produced the Hco-L-AChR2 subtype that is more sensitive to pyrantel than acetylcholine and partially activated by levamisole, but which was bephenium-insensitive indicating that the bephenium-binding site involves Hco-ACR-8. Attempts were made to modify the subunit stoichiometry of the Hco-L-AChR1 subtype by injecting five fold more cRNA of individual subunits. Increased Hco-unc-29.1 cRNA produced no functional receptor. Increasing Hco-unc-63, Hco-unc-38 or Hco-acr-8 cRNAs did not affect the pharmacological characteristics of Hco-L-AChR1 but reduced the currents elicited by acetylcholine and the other agonists. Here, we provide the first description of the molecular composition and functional characteristics of any invertebrate bephenium-sensitive receptor.
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Acknowledgments
This work was supported by the Institut National de la Recherche Agronomique (INRA). Claude Charvet, via a 2011 fellowship award under the OECD Co-operative Research Programme: Biological Resource Management for Sustainable Agricultural Systems to perform the present work at Richard Martin’s laboratory at Iowa State University. RJM and APR were funded by Grant Number R R56 AI047194–11 and R21AI092185-01, respectively, from the National Institute of Allergy and Infectious Diseases of the National Institutes of Health. We wish to acknowledge Samuel K. Buxton for technical assistance and training at electrophysiology. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of Allergy and Infectious Diseases of the National Institutes of Health.
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Charvet, C.L., Robertson, A.P., Cabaret, J. et al. Selective effect of the anthelmintic bephenium on Haemonchus contortus levamisole-sensitive acetylcholine receptors. Invert Neurosci 12, 43–51 (2012). https://doi.org/10.1007/s10158-012-0130-0
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DOI: https://doi.org/10.1007/s10158-012-0130-0