Microglial contributions to the development of the mammalian optic stalk

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Project Summary Ocular development is marked by a key embryonic process, the closure of the ventral optic fissure. Failure in the proper closure of the optic fissure results in coloboma, a cause of significant visual impairment. Microglia, the resident innate immune phagocyte of the central nervous system, are found to populate the retina prior to the closure of the optic fissure. Microglia are specialized to recognize and eliminate apoptotic cells, and critically, cell death is associated with optic fissure closure in mouse and human. It is unknown, however, whether microglia participate in the proper closure of the optic fissure in mammals. To address this, Aim1 will determine whether microglia facilitate optic fissure closure through elimination of apoptotic cells. I will collect tissue from Bax-/-, MerTK-/-, and Pu.1-/- embryos, and their littermate controls, at onset of optic fissure closure, during optic fissure closure, and following optic fissure closure. These mouse lines will allow me to test whether blocking developmental apoptosis, blocking microglial recognition of apoptotic cells, or eliminating microglia completely alters optic fissure closure. Aim2 will determine the expression profile of optic fissure-associated microglia during closure. I will collect tissue from Cx3CR1GFP/+ mice during optic fissure closure and determine the gene expression profile of optic fissure-associated microglia versus adjacent ventral retina microglia using in situ hybridization chain reaction (HCR). While I have a strong background and laboratory experience in neuroscience, both in the context of development and disease, I require additional training to develop as an independent researcher. My goals as a graduate student are to build strength in neuroimmunology and techniques and tools to study contributions of immunology to neurodevelopment. This foundation will prepare me to pursue my long-term career goal of becoming a research scientist. I anticipate that my preliminary data plus the aims described here will produce at least one scientific manuscript while also supplying preliminary data for future grant applications. Under the training plan provided by this fellowship, will refine technical skills, deepen my expertise in neuroimmunology, and become independent in experimental choice, design, analysis, and communication. These skills will propel me on my trajectory towards a career as an independent researcher. Completion of this work will define how microglia shape the closure of the optic fissure and contribute to the embryonic development of the retina and optic nerve. This proposal aims to address the role of microglia in mammalian optic fissure closure, yielding insights into pathways involved in optic fissure closure, and more broadly, revealing how microglia regulate tissue morphogenesis.