Apis mellifera ion channels and tools to study agroecosystems contaminants

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The dramatic decline and disappearance of insect populations, and of many other species, around the world is now a scientific fact, with more than one million species on the endangered list. Insect populations have been harder hit, with a 75% to 98% decline in insect biomass in the last 25-40 years. Although there are many anthropogenic causes, chronic exposure to pesticides is central to the conflict between agricultural imperatives and environmental issues, which is front and centre in the political arena and most media. Despite the fact that the pesticides currently used in modern agriculture are relatively safe for humans, they can severely affect both target pests and beneficial insects. Pesticides include insecticides, herbicides, and fungicides. The most widely used insecticides are neurotoxic. The most active substances belong to 4 major families and target synaptic ion channels: GABA and nicotinic receptors (nAChR), which are targeted by fipronil and neonicotinoids, Nav channels, which are targeted by pyrethroids and other Nav blockers such as metaflumizone, and Choline esterase, which is targeted by organophosphates and methylcarbamates. Their ecotoxicological effects include a significant increase in the loss of honey bee colonies. We will perform in vitro analyses of a collection of commonly used pesticides on several ion channels expressed in the synapses of the honey bee (Apis mellifera) brain and will clone and characterize these channels in an expression system. We hypothesize that honey bee ion channels are sensitive to insecticides, which may explain their deleterious effect on these needed pollinators. Our research program hypothesis will be tested via 3 Aims: Aim #1: Identify, clone, and sequence GABA receptor subunit genes expressed at honeybee synapses and analyze their expression patterns in tissues from adult honey bees and larvae. Characterize, after expression in Xenopus oocytes, the biophysical and pharmacological properties of these pore-forming subunits as well as the effects of a panel of pesticides currently used for agricultural or domestic purposes. Aim #2: Identify, clone, and sequence nAChR receptor subunit genes expressed at honeybee synapses, characterize their expression in Xenopus oocytes, and study their biophysical and pharmacological properties well as the effects of pesticides currently used for agricultural or domestic purposes, including neonic pesticides. Aim #3: Produce a molecular model of each channel type onto which effective molecules can be docked to better understand the molecular mechanisms of ion channel regulation and identify possible species specificities through structural/sequence variations. Impact: This project will in fine provide an invaluable tool for providing an in-depth understanding of the toxicity of the main phytopharmaceutical products and will constitute the first comprehensive molecular and functional characterization of a complete synaptic channelome.