UTILITY OF C6-GLIOMA CELLS FOR EXPLORATORY RISK ASSESSMENT OF MIXTURES OF INSECTICIDES

ROMERO, D.M; KOTLER, M.L; WOLANSKY, M.J.

Washington

Congreso; 50th Annual Meeting and Tox Expo of the Society of Toxicology; 2011

Society of Toxicology

Low levels of organophosphate (OP) and pyrethroid (PYR) insecticides may co-occur in real environments and food products. Considering each compound independently, these residue loadings may be considered safe for humans. However, there is a consensus on the need of considering joint neurotoxicity data to improve the accuracy of health risk estimates for these chemicals. We use a C6-glioma cell culture as an exploratory in vitro system derived from rat nervous system aimed to identify toxicologically relevant OP-PYR mixtures that may require cumulative risk analysis efforts in in vivo models. First, 4 OPs (chlorpyrifos; acephate; methidathion; diazinon) and 4 PYRs (type I: bifenthrin, tefluthrin; type II: alpha-cypermethrin, deltamethrin) were examined in single-compound assays carried out in C6-cells after 4-48 hr exposure to each compound (0.1-250 uM). Threshold doses for cell viability status were determined using optical microscopy and MTT assays. We generated time- and dose-effect data, and ED15 values were computed from assays conducted at t=24 hr. In general, micromolar exposures produced dose-related alterations in cell viability at 24-48 hr. ED15s were 5-100 uM for PYRs and 5-30 uM for OPs. Second, we evaluated the effects of each insecticide on nucleus morphology using Hoechst 33258 staining. We found dose-related increases in anomalous nuclei shapes from the low uM range up. Last, we examined a mixture of equitoxic levels of the 4 PYRs (1X mixture = ED15) using the MTT assay. This type-I/II mixture produced a dose-related, dual joint action at 24 hr: while addition of individual effects was observed after exposure to the 1X mixture, dose-addition was apparent at a 0.1X dose, i.e., well below the single-compound thresholds for mitochondrial impact. We plan to evaluate other mixtures of insecticides in order to select toxicologically relevant combinations for in vivo testing of cumulative neurotoxicity.