title: Effects of organophosphorus compounds on ATP production and mitochondrial integrity in cultured cells.
abstract: Recent studies in vivo and in vitro suggested that mitochondrial dysfunction follows exposure to organophosphorus (OP) esters. As mitochondrial ATP production is important for cellular integrity, ATP production in the presence of OP neurotoxicants was examined in a human neuronal cell line (SH-SY5Y neuroblastoma cells) and primary dorsal root ganglia (DRG) cells isolated from chick embryos and subsequently cultured to achieve maturation with axons. These cell culture systems were chosen to evaluate toxic effects on the mitochondrial respiratory chain associated with exposure to OP compounds that do and do not cause OP-induced delayed neuropathy (OPIDN), a disorder preceded by inhibition of neurotoxic esterase (NTE). Concentration- and time-response studies were done in neuroblastoma cells exposed to phenyl saligenin phosphate (PSP) and mipafox, both compounds that readily induce delayed neuropathy in hens, or paraoxon, which does not. Phenylmethylsulfonyl fluoride (PMSF) was included as a non-neuropathic inhibitor of NTE. Purified neuronal cultures from 9 day-old chick embryo DRG were treated for 12 h with 1 microM PSP, mipafox, or paraoxon. In situ evaluation of ATP production measured by bioluminescence assay demonstrated decreased ATP concentrations both in neuroblastoma cells and chick DRG neurons treated with PSP. Mipafox decreased ATP production in DRG but not in SH-SY5Y cells. This low energy state was present at several levels of the mitochondrial respiratory chain, including Complexes I, II, III, and IV, although Complex I was the most severely affected. Paraoxon and PMSF were not effective at all complexes, and, when effective, required higher concentrations than needed for PSP. Results suggest that mitochondria are an important early target for OP compounds, with exposure resulting in depletion of ATP production. The targeting of neuronal, rather than Schwann cell mitochondria in DRG following exposure to PSP and mipafox was verified by loss of the mitochondrial-specific dye, tetramethylrhodamine, in these cells. No such loss was seen in paraoxon exposed neurons isolated from DRG or in Schwann cells treated with any of the test compounds.
authors.affiliation: Virginia-Maryland Regional College of Veterinary Medicine, 1 Duck Pond Drive, Blacksburg, VA 24061-0442, USA.
authors.forename: C
authors.initials: C
authors.lastname: Massicotte
authors.forename: K
authors.initials: K
authors.lastname: Knight
authors.forename: C J
authors.initials: CJ
authors.lastname: Van der Schyf
authors.forename: B S
authors.initials: BS
authors.lastname: Jortner
authors.forename: M
authors.initials: M
authors.lastname: Ehrich
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D002642:Chick Embryo; D002800:Cholinesterase Inhibitors / Q000633:toxicity*; D004305:Dose-Response Relationship, Drug; D005727:Ganglia, Spinal / Q000166:cytology; D006801:Humans; D007531:Isoflurophate / Q000031:analogs & derivatives*; D018613:Microscopy, Confocal / Q000379:methods; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D009097:Multienzyme Complexes / Q000378:metabolism; D009447:Neuroblastoma; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D009943:Organophosphorus Compounds / Q000633:toxicity*; D010261:Paraoxon; D010664:Phenylmethylsulfonyl Fluoride; D012583:Schwann Cells / Q000187:drug effects; D013997:Time Factors
chemical_list: D002800:Cholinesterase Inhibitors; D009097:Multienzyme Complexes; D009943:Organophosphorus Compounds; D007531:Isoflurophate; C005238:mipafox; C046357:phenylsaligenin cyclic phosphate; D010664:Phenylmethylsulfonyl Fluoride; D000255:Adenosine Triphosphate; D010261:Paraoxon