title: A multiplexed assay for determination of neurotoxicant effects on spontaneous network activity and viability from microelectrode arrays.
abstract: Microelectrode array (MEA) recordings are increasingly being used as an in vitro method to detect and characterize the ability of drugs, chemicals and particles to cause neurotoxicity. While compound effects on spontaneous network activity are easily determined by MEA recordings, compound cytotoxicity is not routinely assessed, particularly within the same network from which recordings are collected. With the advent of higher-throughput 48 and 96 well MEA systems, rapid and simple methods to measure compound effects on cell health are required to facilitate efficient compound screening using MEAs. The present experiments sought to develop a multiplexed approach that allows measurement of network activity and cell health in the same MEA well. Primary cultures from rat cortex were exposed to six different compounds (glyphosate, β-cyfluthrin, domoic acid, tributyltin, lindane and fipronil). Effects of these compounds (0.03-100 μM) on spontaneous network activity (mean firing rate; MFR), cellular metabolic activity (Cell Titer Blue™ (CTB) assay) and lactate dehydrogenase (LDH) release were determined in the same well following a 60-min exposure. Glyphosate elicited no effect on MFR, LDH release or CTB reduction. Tributyltin caused concomitant decreases in MFR and CTB reduction and increases LDH release, while domoic acid and β-cyfluthrin decreased MFR in a concentration-dependent manner without altering either LDH release or CTB reduction. By contrast, lindane and fipronil did not alter LDH release or CTB reduction, but caused biphasic alterations in MFR, with increases in MFR at lower concentrations followed by decreases at higher concentrations. These results demonstrate a simple and rapid method for the simultaneous determination of test compound effects on spontaneous electrical activity and cell health from the same network, and will facilitate rapid screening of compounds for potential neurotoxicity.
authors.affiliation: Integrated Systems Toxicology Division, National Health and Environmental Effects Research Laboratory, Office of Research and Development, US Environmental Protection Agency, Research Triangle Park, NC, United States.
authors.forename: Kathleen
authors.initials: K
authors.lastname: Wallace
authors.affiliation: Axion Biosystems, Atlanta, GA, United States.
authors.forename: Jenna D
authors.initials: JD
authors.lastname: Strickland
authors.affiliation: Axion Biosystems, Atlanta, GA, United States.
authors.forename: Pablo
authors.initials: P
authors.lastname: Valdivia
authors.affiliation: Integrated Systems Toxicology Division, National Health and Environmental Effects Research Laboratory, Office of Research and Development, US Environmental Protection Agency, Research Triangle Park, NC, United States.
authors.forename: William R
authors.initials: WR
authors.lastname: Mundy
authors.affiliation: Integrated Systems Toxicology Division, National Health and Environmental Effects Research Laboratory, Office of Research and Development, US Environmental Protection Agency, Research Triangle Park, NC, United States. Electronic address: Shafer.tim@epa.gov.
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Shafer
mesh_terms: D000200:Action Potentials / Q000187:drug effects*; D000818:Animals; D000831:Animals, Newborn; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002525:Cerebellar Cortex / Q000166:cytology; D004305:Dose-Response Relationship, Drug; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008839:Microelectrodes; D009415:Nerve Net / Q000187:drug effects*; D009474:Neurons / Q000187:drug effects*; D009498:Neurotoxins / Q000494:pharmacology*; D051381:Rats
chemical_list: D009498:Neurotoxins; D007770:L-Lactate Dehydrogenase
keywords: Microelectrode array; Multiplexed assay; Screening