Deconstructing the hypothalmic ontogeny and plasticity via clonal analysis

Johns Hopkins University, Baltimore MD

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Abstract

? DESCRIPTION (provided by applicant): Mammalian brain function critically relies on sophisticated cytoarchitectonic organization during embryonic development. Cell generation, migration, synapse formation and circuit integration often follow highly stereotyped patterns to form complex laminated or nuclear structures in the brain. Recently developed clonal lineage analysis has revealed stem cell behavior giving rise to laminar structures, such as the cerebral cortex, cerebellum and retina, with unprecedented single-cell resolution. However, the formation of nuclear structures by neural stem cells (NSCs) remains unclear and has yet to be systematically investigated. The mammalian hypothalamus is a heterogeneous nuclear structure that is critical for the integration and homeostatic maintenance of endocrine, autonomic and behavioral functions. Reconstructing how hypothalamic neurons are generated from individual NSCs and organized into discrete nuclei during early development is essential to understand the structure-function relationship of different hypothalamic nuclei and the extent to which this can be modulated by environmental conditions. We have developed a genetically-based single-cell lineage tracing-technique that employs MADM (mosaic analysis of double marker) animals to label NSCs in the developing embryo and begin to address these outstanding questions of hypothalamic organization. The goal of the proposed research is to reconstruct and quantify the behavior of individually-labeled NSCs in vivo, decipher the general principles organizing hypothalamic nuclei, decode the ontogeny of individual hypothalamic nuclei and explore the ontogenetic plasticity of nuclear organization in the context of a maternal challenge. The complexity of the anatomical and molecular subdivisions of the hypothalamus, and lack of appropriate genetic tools, has thus far prevented a deep understanding of the organization and ontogeny of this nuclear structure. Successful completion of our study will result in a comprehensive map of single NSCs and their progeny at regional, zonal and nuclear levels, the clonal organization of sibling neurons in a three-dimensional context to determine migratory patterns of newly born neurons, and the capacity of single stem cells to contribute to functionally distinct nuclei. We will also have validated an experimental platform for future mechanistic investigations of hypothalamic dysregulation under pathological conditions, which can lead to targeted diagnostic and therapeutic strategies to preserve critical physiological functions.