The use of differentiating N2a and C6 cell lines for studies of organophosphate toxicity

Abstract

A major goal of our cellular toxicology research has been the identification of novel targets of organophosphorous compounds (OPs), to which end we have studied the effects of sub-lethal concentrations of OPs on morphological and molecular end points in differentiating mammalian cell lines. This chapter describes the key practical approaches that we use to monitor OP toxicity in mouse N2a neuroblastoma and rat C6 glioma cell lines. These cell lines are easy to maintain and are useful for the study of short-term, potentially reversible, effects of OPs on the outgrowth and maintenance of neurites and associated regulatory proteins. The main topics covered include (a) the basic maintenance of cells and the induction and morphological assessment of cell differentiation in various experimental scenarios, (b) the measurements of key enzyme activities, and (c) the detection of molecular changes using a targeted proteomic approach focussed on immunofluorescence staining of fixed cells and western blotting analysis of cell extracts with antibodies to proteins known to be important in the regulation of cell differentiation. This approach can also be used for the study of other neurotoxins and is amenable to the measurement of a wide range of end points. © 2011 Springer Science+Business Media, LLC.