The Effects of Acute and Repeated Exposure/s to the Organophosphate Pesticide Chlorpyrifos at Subthreshold Doses on Brain Structural Integrity

• Wistar Rats weighing 250 – 300g were treated with Chloropyrifos (CPF) either a large subtoxic dose acutely and sacrificed after 6 or 24 hours, or they were given varying doses over a 30 day period and given a washout. • After the rats were sacrificed, the prefrontal cortex was isolated and processed using a subcellular fractionation protocol. • Total protein was quantified for each subcellular fraction (cytosolic (soluble), nuclear and membrane) using a detergent-compatible protein assay reagent (Bio-Rad) according to manufacturer’s instructions. • Cytosolic (Soluble) extracts were prepared in Laemmli buffer containing 2-mercaptoethanol for SDS-polyacrylabmide gel electrophoresis (PAGE). • Resolved extracted proteins were electroblotted to PVDF and probed for the expression of acetylated (lysine 40) and total -tubulin (SigmaAldrich mouse monoclonal antibodies T7451 and 9026, respectively) • Anti-tubulin antibodies were detected using horseradish peroxidase and enhanced chemiluminescence on film. • Protein band optical densities were analyzed from scanned film images using ImageJ software. Dr. Alvin V. Terry, Jr., for kindly permitting use of lab bench space. GRU’s Small Animal Behavioral Core Facility’s Research Associates (Samantha Sinha, Kristy Bouchard and Leah Vandenhuerk) for assistance with injecting rats. Kristina Skinner (Hope College) for assistance with regent preparation. Ms. Abigail Drescher and CURS program for this opportunity and stipend support. Repeated Exposures