Novel Single Cell Assay to Identify Genes Underlying Developmental Neurotoxicity

Abstract Traditional tests for developmental neurotoxicity have involved animal studies with behavioral or anatomical readouts that are integrative of toxicant effects on the whole organism However they are not informative of mechanism of action particularly with molecular detail Recently methods to produce D brain organoids from stem cells were developed These cortical organoids exhibit much more of the spatial and cell type organization of authentic cortical regions of the developing brain We propose to develop a novel genetic screen of neurotoxicant effects on brain organoid development The proposed project brings together several established as well as recent emerging technologies with a novel panel of over genetically diverse ES cell lines to solve a problem that has been a barrier to developmental neurotoxicity DNT testing Bulk harvesting of RNA required that expression studies use uniform cell populations However neurodevelopment is an inherently cell heterogenous process In this project we will use brain organoids produced in vitro from panel of genetically diverse ES cell lines in combination with single cell RNAseq to develop a scalable highly efficient and highly granular eQTL DNT screen The key innovations of our technology is combining complex trait genetics with single cell transcriptomes and defining subpopulations of differentiating neural cell types using single cell transcriptomes then assigning individual genetically diverse cells to the appropriate subpopulation In this way the muddle of data from heterogenous cell mixtures is avoided !The developing brain is highly vulnerable to chemicals that can cause damage to nervous system cell growth function and development We are creating a new test to understand that damage in deep molecular detail in individual cells of a minibrain model !