title: Inhibition of Voltage-Gated Calcium Channels After Subchronic and Repeated Exposure of PC12 Cells to Different Classes of Insecticides.
abstract: We previously demonstrated that acute inhibition of voltage-gated calcium channels (VGCCs) is a common mode of action for (sub)micromolar concentrations of chemicals, including insecticides. However, because human exposure to chemicals is usually chronic and repeated, we investigated if selected insecticides from different chemical classes (organochlorines, organophosphates, pyrethroids, carbamates, and neonicotinoids) also disturb calcium homeostasis after subchronic (24 h) exposure and after a subsequent (repeated) acute exposure. Effects on calcium homeostasis were investigated with single-cell fluorescence (Fura-2) imaging of PC12 cells. Cells were depolarized with high-K(+) saline to study effects of subchronic or repeated exposure on VGCC-mediated Ca(2+) influx. The results demonstrate that except for carbaryl and imidacloprid, all selected insecticides inhibited depolarization (K(+))-evoked Ca(2+) influx after subchronic exposure (IC50's: approximately 1-10 µM) in PC12 cells. These inhibitory effects were not or only slowly reversible. Moreover, repeated exposure augmented the inhibition of the K(+)-evoked increase in intracellular calcium concentration induced by subchronic exposure to cypermethrin, chlorpyrifos, chlorpyrifos-oxon, and endosulfan (IC50's: approximately 0.1-4 µM). In rat primary cortical cultures, acute and repeated chlorpyrifos exposure also augmented inhibition of VGCCs compared with subchronic exposure. In conclusion, compared with subchronic exposure, repeated exposure increases the potency of insecticides to inhibit VGCCs. However, the potency of insecticides to inhibit VGCCs upon repeated exposure was comparable with the inhibition previously observed following acute exposure, with the exception of chlorpyrifos. The data suggest that an acute exposure paradigm is sufficient for screening chemicals for effects on VGCCs and that PC12 cells are a sensitive model for detection of effects on VGCCs.
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Faculty of Veterinary Medicine, Institute for Risk Assessment Sciences (IRAS), Utrecht University, NL-3508 TD Utrecht, The Netherlands.
authors.forename: Marieke
authors.initials: M
authors.lastname: Meijer
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Faculty of Veterinary Medicine, Institute for Risk Assessment Sciences (IRAS), Utrecht University, NL-3508 TD Utrecht, The Netherlands.
authors.forename: Joske A R
authors.initials: JA
authors.lastname: Brandsema
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Faculty of Veterinary Medicine, Institute for Risk Assessment Sciences (IRAS), Utrecht University, NL-3508 TD Utrecht, The Netherlands.
authors.forename: Desirée
authors.initials: D
authors.lastname: Nieuwenhuis
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Faculty of Veterinary Medicine, Institute for Risk Assessment Sciences (IRAS), Utrecht University, NL-3508 TD Utrecht, The Netherlands.
authors.forename: Fiona M J
authors.initials: FM
authors.lastname: Wijnolts
authors.affiliation: Neurotoxicology Research Group, Toxicology Division, Faculty of Veterinary Medicine, Institute for Risk Assessment Sciences (IRAS), 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, Faculty of Veterinary Medicine, Institute for Risk Assessment Sciences (IRAS), 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: D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D001921:Brain / Q000166:cytology / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000008:administration & dosage / Q000633:toxicity*; D015220:Calcium Channels / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D007306:Insecticides / Q000008:administration & dosage / Q000633:toxicity*; D016716:PC12 Cells / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar; D059550:Toxicity Tests, Subchronic / Q000379:methods
chemical_list: D002121:Calcium Channel Blockers; D015220:Calcium Channels; D007306:Insecticides; D002118:Calcium
keywords: calcium homeostasis; insecticides; in vitro neurotoxicology; repeated exposure; subchronic exposure; voltage-gated calcium channels