Detection of Neurotoxicity Using In Vitro Neuronal Cell Systems

Abstract

Implementation of the EC White Paper, "Strategy for a future chemicals policy" (2001), is estimated to require the testing of approximately 30,000 'existing' chemicals by 2012. In vitro tests were recommended for use where possible. This study evaluated a novel in vitro testing strategy for neurotoxicity, as neurotoxic data is a requisite of the White Paper (2001). Sensitivities of undifferentiated and differentiated PC12 cells and primary CGC cultures to identify neurotoxins were compared. Cytotoxicants and neurotoxicants used in other neurotoxicity studies and covering a range of mechanisms and potencies were used. Undifferentiated PC12 cells were used to indicate basal cytotoxicity to which sensitivities of neuronal-like models were compared. Collectively, the tested toxins failed to significantly distinguish cell system responses as measured using the following endpoints; cell viability/activity, ATP depletion, MMP depolarisation, ROS production and cytoskeleton changes. As p53 is involved in mechanisms of neurotoxicity, including neurodegeneration and neurodisease, a p53 transfected PC12 cell line was characterised. The general functionality of the transfected cells was found to be questionable and use of the transfected p53 PC12 cell line was not recommended. All cell system responses produced IC50 values, some with at least 3x differences in cell system responses, due to differentiated cell systems being consistently less sensitive to both neurotoxicants and cytotoxicants. Comparisons between IC50 values and rat acute oral LD50 values did not give significant differences. This was hypothesised to be due to the differentiated cell systems post-mitotic state and the presence of elevated defence mechanisms. More neurospecific endpoints were recommended for inclusion in a testing strategy in order to elucidate the greater sensitivity of the neuronal-like cell systems.JRC.I.2-Validation of biomedical testing method