Mitochondrial and transcriptome responses in rat dopaminergic neuronal cells following exposure to the insecticide fipronil

• Published in: Neurotoxicology (Park Forest South) 2021-07, Vol.85, p.173-185
• Main Authors: Souders, Christopher L., Rushin, Anna, Sanchez, Christina L., Toth, Darby, Adamovsky, Ondrej, Martyniuk, Christopher J.
• Format: Article
• Language: English
• Subjects: Alzheimer's disease; Animals; Apoptosis; ATP; Bioenergetics; Butyric acid; Caspase-3; Cell Line, Transformed; Cell Survival - drug effects; Cell Survival - physiology; Central nervous system; Cytotoxicity; Degeneration; Depolarization; Disease; Dopamine; Dopamine receptors; Dopaminergic Neurons - drug effects; Dopaminergic Neurons - metabolism; Dose-Response Relationship, Drug; Exposure; Fipronil; Inflammatory diseases; Insecticides; Insecticides - toxicity; Ion channels (ligand-gated); Membrane permeability; Membrane potential; Membrane Potential, Mitochondrial - drug effects; Membrane Potential, Mitochondrial - physiology; Membranes; Mitochondria; Mitochondria - drug effects; Mitochondria - metabolism; Mitochondrial bioenergetics; Movement disorders; Neurodegeneration; Neurodegenerative diseases; Neurofibrillary tangles; Neurotoxicity; Neurotoxins; Oligomycin; Oxygen consumption; Parkinson's disease; Pathway analysis; Pesticides; Pyrazoles - toxicity; Rats; Reduction; Respiration; Toxicants; Toxicity; Toxins; Transcription; Transcriptome - drug effects; Transcriptome - physiology; Transcriptomics; γ-Aminobutyric acid
• ISSN: 0161-813X; 1872-9711
• DOI: 10.1016/j.neuro.2021.05.011

•Fipronil increased cytotoxicity and caspase 3/7 activity in dopamine neurons.•Fipronil impaired oxidative phosphorylation in dopamine neurons.•Mitochondrial membrane potential was significantly reduced by fipronil at 2.5 μM and above.•Apoptotic pathways, mitochondrial, and gene networks related to neurodegeneration were altered by fipronil.•Fipronil is a potent uncoupler of oxidative phosphorylation and is a mitotoxicant. The phenylpyrazole fipronil is an insecticide that inhibits γ -amino-butyric acid (GABA) ionotropic receptors in the central nervous system. Experimental evidence suggests that fipronil acts as a neurotoxin and it is implicated in neurodegenerative diseases; however, the mechanisms of neurotoxicity are not fully elucidated. The objective of this study was to quantify mechanisms of fipronil-induced neurotoxicity in dopamine cells. Rat primary immortalized mesencephalic dopaminergic cells (N27) were treated with fipronil (0.25 up to 500 μM depending on the assay). We measured endpoints related to mitochondrial bioenergetics, mitophagy, mitochondrial membrane potential, and ATP production in addition to discerning transcriptome responses to the pesticide. Fipronil reduced cell viability at 500 μM after 24 h exposure and caspase 3/7 activity was significant increased after 6 and 12 h by 250 and 500 μM fipronil. Subsequent endpoints were thus assessed at concentrations that were below cytotoxicity. We measured oxidative respiration of N27 cells following a 24 h exposure to one dose of either 0.25, 2.5, 25, or 50 μM fipronil. Oxygen consumption rates (OCR) were not different between vehicle-control and 0.25 or 2.5 μM fipronil treatments, but there was a ∼40–60 % reduction in basal respiration, as well as reduced oligomycin-induced ATP production at 50 μM. The reduction in OCR is hypothesized to be related to lower mitochondrial mass due to mitophagy. Mitochondrial membrane potential was also sensitive to fipronil, and it was compromised at concentrations of 2.5 μM and above. To further elucidate the mechanisms linked to neurotoxicity, we conducted transcriptomics in dopamine cells following treatment with 25 μM fipronil. Fipronil suppressed transcriptional networks associated with mitochondria (damage, depolarization, permeability, and fission), consistent with its effects on mitochondrial membrane potential. Altered gene networks also included those related to Alzheimer disease, inflammatory disease, nerve fiber degeneration, and neurofibrill