In vitro approach for organophosphorous compounds-induced neurotoxicity (OP Neurotox Assay)

In vitro test system based on in SH-SY5Y human neuroblastoma cell line was proposed for evaluation of organophosphorus compounds (OPs) induced neurotoxicity. The assay is based on  the measurements of acetylcholineesterase (AChE) and neuropathy target esterase (NTE) activity since OPs exert neurotoxicities following inhibition of these two critical esterases: AChE for acute toxicity and neurotoxic esterase NTE for delayed neuropathy.  To be effective insecticides, AChE needs to be inhibited, but NTE does not.  The relationship between AChE and NTE inhibition provides information on which OP compounds could be expected to be effective as insecticides without them causing progressive, irreversible delayed neuropathy in humans and susceptible animals.  Concentration-responses of AChE and NTE activities can be measured in cultured neuronal cells, plotted against the log of their concentrations, and a ratio of the concentrations responsible for 50% inhibition (IC50) determined by dividing the NTE IC50 by the AChE IC50.  Ratios >100 suggest that NTE inhibition and subsequent delayed neuropathy are unlikely because test compound concentrations would need to be 100-fold higher than those causing acute toxicity. 

Protoxicants can be tested in this system following incubation with 0.05% bromine solution or rat liver microsomes.  NTE and AChE inhibition can be measured following either a single exposure or exposures for up to 28 days.  Use of this in vitro system would decrease the need to dose animals and collect brain and spinal cord for esterase measurements. Evaluated organophosphorus compounds were chosen based on animal studies that identified them as active toxicants capable of producing delayed neuropathy (n=6), protoxicants capable of producing delayed neuropathy (n=3), active AChE inhibitors not producing delayed neuropathy in unprotected animals (n=5), and protoxicants not producing delayed neuropathy in unprotected animals (n=5).  The in vitro test comparing the NTE inhibition with the AChE inhibition produced by the 19 compounds tested matched the in vivo results 100% of the time.