title: Inhibition of voltage-gated calcium channels as common mode of action for (mixtures of) distinct classes of insecticides.
abstract: Humans are exposed to distinct structural classes of insecticides with different neurotoxic modes of action. Because calcium homeostasis is essential for proper neuronal function and development, we investigated the effects of insecticides from different classes (pyrethroid: (α-)cypermethrin; organophosphate: chlorpyrifos; organochlorine: endosulfan; neonicotinoid: imidacloprid) and mixtures thereof on the intracellular calcium concentration ([Ca(2+)]i). Effects of acute (20 min) exposure to (mixtures of) insecticides on basal and depolarization-evoked [Ca(2+)]i were studied in vitro with Fura-2-loaded PC12 cells and high resolution single-cell fluorescence microscopy. The data demonstrate that cypermethrin, α-cypermethrin, endosulfan, and chlorpyrifos concentration-dependently decreased depolarization-evoked [Ca(2+)]i, with 50% (IC50) at 78nM, 239nM, 250nM, and 899nM, respectively. Additionally, acute exposure to chlorpyrifos or endosulfan (10μM) induced a modest increase in basal [Ca(2+)]i, amounting to 68 ± 8nM and 53 ± 8nM, respectively. Imidacloprid did not disturb basal or depolarization-evoked [Ca(2+)]i at 10μM. Following exposure to binary mixtures, effects on depolarization-evoked [Ca(2+)]i were within the expected effect additivity range, whereas the effect of the tertiary mixture was less than this expected additivity effect range. These results demonstrate that different types of insecticides inhibit depolarization-evoked [Ca(2+)]i in PC12 cells by inhibiting voltage-gated calcium channels (VGCCs) in vitro at concentrations comparable with human occupational exposure levels. Moreover, the effective concentrations in this study are below those for earlier described modes of action. Because inhibition of VGCCs appears to be a common and potentially additive mode of action of several classes of insecticides, this target should be considered in neurotoxicity risk assessment studies.
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Institute for Risk Assessment Sciences (IRAS), Faculty of Veterinary Medicine, Utrecht University, NL-3508 TD Utrecht, The Netherlands.
authors.forename: Marieke
authors.initials: M
authors.lastname: Meijer
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Institute for Risk Assessment Sciences (IRAS), Faculty of Veterinary Medicine, Utrecht University, NL-3508 TD Utrecht, The Netherlands.
authors.forename: Milou M L
authors.initials: MM
authors.lastname: Dingemans
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Institute for Risk Assessment Sciences (IRAS), Faculty of Veterinary Medicine, Utrecht University, NL-3508 TD Utrecht, The Netherlands.
authors.forename: Martin
authors.initials: M
authors.lastname: van den Berg
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Institute for Risk Assessment Sciences (IRAS), Faculty of Veterinary Medicine, Utrecht University, NL-3508 TD Utrecht, The Netherlands r.westerink@uu.nl.
authors.forename: Remco H S
authors.initials: RH
authors.lastname: Westerink
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000737:chemistry / Q000633:toxicity*; D015220:Calcium Channels / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D004785:Environmental Pollutants / Q000737:chemistry / Q000633:toxicity*; D006801:Humans; D007306:Insecticides / Q000737:chemistry / Q000633:toxicity*; D020258:Neurotoxicity Syndromes / Q000378:metabolism; D016716:PC12 Cells; D051381:Rats; D013329:Structure-Activity Relationship
chemical_list: D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004785:Environmental Pollutants; D007306:Insecticides; D002118:Calcium
keywords: PC12 cells; calcium homeostasis; in vitro neurotoxicity; insecticides; mixture toxicity; voltage-gated calcium channels