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Non-competitive Inhibition of Nicotinic Acetylcholine Receptors by Ladybird Beetle Alkaloids

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Abstract

Ladybird beetles (Family Coccinellidae) secrete an alkaloid rich venom from their leg joints that protects them from predators. Coccinellines, the major venom constituents, are alkaloids composed of three fused piperidine rings that share a common nitrogen atom. Although many coccinellines have been isolated and chemically characterized, their pharmacological properties are essentially unknown. Using radioligand binding and functional assays we investigated the actions of several coccinellines on skeletal muscle and α7 nicotinic acetylcholine receptors (nAChRs). The alkaloids were shown to displace the specific binding of tritiated piperidyl-N-(1-(2-thienyl)cyclohexyl)-3,4-piperidine ([3H]-TCP), which has been shown to bind deep within the ion channel of the electric fish (Torpedo) muscle nAChR. The stereoisomers precoccinelline and hippodamine (whose nitrogens are predicted to be ionized at physiological pH) and their respective analogs N-methyl-precoccinelline and N-methyl-hippodamine (whose quaternary nitrogens are permanently charged) displayed similar IC50s for inhibition of [3H]-TCP binding. However, the corresponding precoccinelline and hippodamine N-oxides, coccinelline and convergine (which have an electronegative oxygen bonded to an electropositive nitrogen) displayed significantly higher binding IC50s. Finally, exochomine, a dimeric coccinelline containing the hippodamine structure, displayed the highest IC50 (lowest affinity) for displacing specific [3H]-TCP binding. The presence of a desensitizing concentration (10−3 M) of carbachol (CCh) had little or no effect on the affinity of the Torpedo nAChR for the three coccinellines tested. High concentrations of the coccinellid alkaloids did not affect binding of [3H]-cytisine to Torpedo receptor ACh binding sites. Inhibition of the alpha7 nAChR with pre-equilibrated precoccinelline was insurmountable with respect to ACh concentration. We conclude that the coccinellines bind to one or more allosteric sites rather than to the ACh binding sites, and inhibit nAChR responses to ACh through a non-competitive mechanism. Future chemical and pharmacological investigations of other ladybird beetle alkaloids are likely to reveal other interesting alkaloids affecting ligand-gated receptors.

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Abbreviations

ACh:

Acetylcholine

CCh:

Carbamylcholine

nAChR:

Nicotinic acetylcholine receptor

NCA:

Non-competitive antagonist

[3H]-TCP:

[Piperidyl-3,4-3H(N)]-N-(1-(2-thienyl)cyclohexyl)-3,4-piperidine

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Authors and Affiliations

  1. Department of Pharmacology and Therapeutics, University of Florida, Gainesville, FL, 32610, USA

    Ron L. Leong, Hong Xing & William R. Kem

  2. Laboratory of Bioorganic Chemistry, Faculty of Sciences, University of Brussels, 1050, Brussels, Belgium

    Jean-Claude Braekman

Authors
  1. Ron L. Leong
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  2. Hong Xing
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  3. Jean-Claude Braekman
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  4. William R. Kem
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Corresponding author

Correspondence to William R. Kem.

Additional information

Ron L. Leong and Hong Xing have contributed equally to this study.

Special Issue: In Honor of Lynn Wecker.

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Leong, R.L., Xing, H., Braekman, JC. et al. Non-competitive Inhibition of Nicotinic Acetylcholine Receptors by Ladybird Beetle Alkaloids. Neurochem Res 40, 2078–2086 (2015). https://doi.org/10.1007/s11064-014-1466-0

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