Abstract
Scientific work showed that 70 to 80% of flying insects had disappeared in the last 30 years. Several studies extended these results to fish and birds, species being at the top of the food chain, and to several places on the surface of the globe. The main cause, even if it is not exclusive, had been attributed to pesticides in general, and more particularly to insecticides. These molecules, most often neurotoxic, are used by spreading in fields, in seed coating, and in veterinary or domestic products in the form of sprays against mosquitoes, ants, and hornets... They are now present in any place, such as fields, rivers, or even in the air. As soon as they do not quickly kill insects via massive doses, they can, via lower doses, permanently affect flight, locomotion, olfaction, and memory of insects that are exposed; even in the case of eusocial insects such as Apis mellifera bees, affecting the colony, by modifying the production or detection of the pheromones which ensure its proper functioning, going as far as death. Their main targets in insects are synaptic ion channels (voltage-gated channels: sodium and calcium, channels opened by ligands: GABAergic or nicotinic), which can be overstimulated or blocked, quickly becoming lethal... Scientific studies on the disturbances caused by these molecules are most often interested in their direct molecular targets, and relatively little is known about the effect on the other actors of the physiological functions described above, often because these other actors are poorly described in the literature.The study carried out during these three years of thesis was mainly focused on two of these actors: octopamine receptors (OctR), which are receptors coupled to G proteins and which play important roles in cardiac function, olfaction and/or memory; and olfactory receptors (OR), which are the primary step in odor detection. In both cases, the cloning or recloning of these receptors in the bee Apis mellifera and/or the Varroa destructor, which is an important parasite of the hive, was carried out, with the development of the experimental conditions for their expression. in Xenopus oocyte. Voltage-clamp experiments with two-electrode then made it possible to analyze their biophysical and pharmacological properties. The results of their characterization highlight several intracellular signaling pathways stimulated by the activation of OctRs via different G proteins, including the production of cAMP and/or IP3, but also the G proteins independant activation of a conductance that remains to be determined. The specificity of the stimulation of these pathways is dependent on the receptor, but also on the ligand used.The analysis of 10 ORs (including 9 being orphans), structurally representative of the diversity of the ORs of the 9-exon clade (overexpressed in males and workers), shows that the constitutive subunit Orco is not highly sensitive to VUAA1 in the bee Apis mellifera, unlike that of the fruit fly Drosophila melanogaster. The Orco/OR heterotetramer has a different selectivity, and thus different pore structure than the Orco homotetramer, suggesting the actual formation of the heterotetramer. Apart from AmOR11, which was studied in the presence of 9-ODA, a pheromone specific to this OR and secreted by the queen's mandibles, the 9 other ORs seem to form heterotetramers with Orco, due to their sensitivity to VUUA1, however, none of them could be deorphanized with the molecules of interest that we had been chosen in this study.