Extracellular vesicles (EVs) are biological vectors that can modulate the metabolism of target cells by conveying signalling proteins and genomic material. The level of EVs in plasma is significantly increased in cardiometabolic diseases associated with obesity, suggesting their possible participation in the development of metabolic dysfunction. With regard to the poor definition of adipocyte‐derived EVs, the purpose of this study was to characterise both qualitatively and quantitatively EVs subpopulations secreted by fat cells. Adipocyte‐derived EVs were isolated by differential centrifugation of conditioned media collected from 3T3‐L1 adipocytes cultured for 24 h in serum‐free conditions. Based on morphological and biochemical properties, as well as quantification of secreted EVs, we distinguished two subpopulations of adipocyte‐derived EVs, namely small extracellular vesicles (sEVs) and large extracellular vesicles (lEVs). Proteomic analyses revealed that lEVs and sEVs exhibit specific protein signatures, allowing us not only to define novel markers of each population, but also to predict their biological functions. Despite similar phospholipid patterns, the comparative lipidomic analysis performed on these EV subclasses revealed a specific cholesterol enrichment of the sEV population, whereas lEVs were characterised by high amounts of externalised phosphatidylserine. Enhanced secretion of lEVs and sEVs is achievable following exposure to different biological stimuli related to the chronic low‐grade inflammation state associated with obesity. Finally, we demonstrate the ability of primary murine adipocytes to secrete sEVs and lEVs, which display physical and biological characteristics similar to those described for 3T3‐L1. Our study provides additional information and elements to define EV subtypes based on the characterisation of adipocyte‐derived EV populations. It also underscores the need to distinguish EV subpopulations, through a combination of multiple approaches and markers, since their specific composition may cause distinct metabolic responses in recipient cells and tissues.

Nicotinic acetylcholine receptors (nAChRs) are the main target of neonicotinoid insecticides, which are widely used in crop protection against insect pests. Electrophysiological and molecular approaches have demonstrated the presence of several nAChR subtypes with different affinities for neonicotinoid insecticides. However, the precise mode of action of neonicotinoids on insect nAChRs remains to be elucidated. Radioligand binding studies with [3H]-α-bungarotoxin and [3H]-imidacloprid have proved instructive in understanding ligand binding interactions between insect nAChRs and neonicotinoid insecticides. The precise binding site interactions have been established using membranes from whole body and specific tissues. In this review, we discuss findings concerning the number of nAChR binding sites against neonicotinoid insecticides from radioligand binding studies on native tissues. We summarize the data available in the literature and compare the binding properties of the most commonly used neonicotinoid insecticides in several insect species. Finally, we demonstrate that neonicotinoid-nAChR binding sites are also linked to biological samples used and insect species.

Neonicotinoid insecticides are used worldwide and have been demonstrated as toxic to beneficial insects such as honeybees. Their effectiveness is predominantly attributed to their high affinity for insect neuronal nicotinic acetylcholine receptors (nAChRs). Mammalian neuronal nAChRs are of major importance because cholinergic synaptic transmission plays a key role in rapid neurotransmission, learning and memory processes, and neurodegenerative diseases. Because of the low agonist effects of neonicotinoid insecticides on mammalian neuronal nAChRs, it has been suggested that they are relatively safe for mammals, including humans. However, several lines of evidence have demonstrated that neonicotinoid insecticides can modulate cholinergic functions through neuronal nAChRs. Major studies on the influence of neonicotinoid insecticides on cholinergic functions have been conducted using nicotine low-affinity homomeric α7 and high-affinity heteromeric α4β2 receptors, as they are the most abundant in the nervous system. It has been found that the neonicotinoids thiamethoxam and clothianidin can activate the release of dopamine in rat striatum. In some contexts, such as neurodegenerative diseases, they can disturb the neuronal distribution or induce oxidative stress, leading to neurotoxicity. This review highlights recent studies on the mode of action of neonicotinoid insecticides on mammalian neuronal nAChRs and cholinergic functions.

Neonicotinoid insecticides act on nicotinic acetylcholine receptor and are particularly effective against sucking pests. They are widely used in crops protection to fight against aphids, which cause severe damage. In the present study we evaluated the susceptibility of the pea aphid Acyrthosiphon pisum to the commonly used neonicotinoid insecticides imidacloprid (IMI), thiamethoxam (TMX) and clothianidin (CLT). Binding studies on aphid membrane preparations revealed the existence of high and low-affinity binding sites for [³H]-IMI (Kd of 0.16±0.04 nM and 41.7±5.9 nM) and for the nicotinic antagonist [¹²⁵I]-α-bungarotoxin (Kd of 0.008±0.002 nM and 1.135±0.213 nM). Competitive binding experiments demonstrated that TMX displayed a higher affinity than IMI for [¹²⁵I]-α-bungarotoxin binding sites while CLT affinity was similar for both [¹²⁵I]-α-bungarotoxin and [³H]-IMI binding sites. Interestingly, toxicological studies revealed that at 48 h, IMI (LC₅₀ = 0.038 µg/ml) and TMX (LC₅₀ = 0.034 µg/ml) were more toxic than CLT (LC₅₀ = 0.118 µg/ml). The effect of TMX could be associated to its metabolite CLT as demonstrated by HPLC/MS analysis. In addition, we found that aphid larvae treated either with IMI, TMX or CLT showed a strong variation of nAChR subunit expression. Using semi-quantitative PCR experiments, we detected for all insecticides an increase of Apisumα10 and Apisumβ1 expressions levels, whereas Apisumβ2 expression decreased. Moreover, some other receptor subunits seemed to be differently regulated according to the insecticide used. Finally, we also demonstrated that nAChR subunit expression differed during pea aphid development. Altogether these results highlight species specificity that should be taken into account in pest management strategies.