Collaborative Research: Specification of Excitatory Fates in the Spinal Cord

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This project examines how appropriately-functioning neuronal circuits are made as embryos develop and grow. The work focuses on circuits in the spinal cord of zebrafish embryos, which develop early in life and are simpler and more experimentally-tractable than neural circuits in the brain. The goal is to identify key regulatory genes that instruct cells to grow into one particular population of nerve cells (interneurons) with specific functions in controlling precise body movements. The outcomes of this project will be widely applicable, because these types of nerve cell exist in all vertebrates (including humans). The genes identified in this study and the ways that they influence nerve cell and circuit development are likely to be similar in many different species. This project will significantly increase understanding of how genes instruct cells to develop with particular characteristics, and the roles that specific genes play in the construction of neuronal circuitry. The principal investigators will incorporate this research into their university teaching and presentations made to the general public. This research will contribute to the diversification of the scientific workforce through the active participation of both women and under-represented minorities. The principal investigators will also perform outreach to local city high schools and develop teaching modules for both 9th/10th grade and advanced placement high school Biology students. The aim of these activities is to significantly impact the lives of traditionally under-served high school students and enthuse under represented minority high school students about science and research.An essential first step in understanding how neuronal networks are generated is determining how distinct interneurons (INs) are specified with particular functional properties. One key property that helps to define INs is which neurotransmitter they use to communicate with other cells. Neurotransmitters can be excitatory or inhibitory and INs utilizing inappropriate neurotransmitters form malfunctioning circuits. Therefore, determining how IN neurotransmitter properties are specified is crucial for understanding both IN and circuit development. Relatively little is known about how excitatory spinal IN properties are specified. This proposal will characterize a gene regulatory network (GRN) that specifies spinal IN excitatory properties, using zebrafish as a model system. The investigators have identified genes that encode for 11 different DNA-binding proteins as candidate members of a GRN that specifies V0v IN excitatory (glutamatergic) properties. V0v INs reside in the middle of the spinal cord and have crucial roles in locomotor circuitry. The research team has already shown that four of these genes, evx1/2 and lmx1ba/b are required for correct excitatory fates of V0v INs. In addition, in evx1/2 double mutants V0v INs lose expression of both lmx1ba/b and the other 7 genes, suggesting that all 9 of these genes may act downstream of Evx1/2 in specifying V0v glutamatergic properties. The specific objectives of this proposal are to test if any of these 7 other genes are required for correct V0v neurotransmitter properties and determine the epistatic relationships among all of these Evx1/2-regulated genes. Single and compound zebrafish mutants will be used to answer these questions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.