PROJECT SUMMARY/ ABSTRACT The goal of this project is to characterize and develop the adult kidney of the novel zebrafish-related teleost fish, Danionella cerebrum, for optical-based kidney disease research. Visualizing cellular interactions within an intact live adult vertebrate kidney has led to discoveries of dynamic cell behaviors in their natural, unperturbed environment. Currently, the ability to perform this type of research remains limited to few investigators due to the high bar of entry mainly because of the costs associated with the sophisticated multiphoton microscopic equipment needed and the training required to perform complex surgical procedures. D. cerebrum possess many of the same amenable attributes seen in zebrafish, with the added benefits of adults remaining small and almost completely transparent in adulthood. These later characteristics allow for optical access to the mature, yet small mesonephric kidney of this species, ideal for optical-based studies using standard confocal techniques with no need for surgical interventions. Our recently published methods using custom 3D-printed chambers allow for prolonged and longitudinal imaging of the same adult fish over long periods of time. We have generated several kidney and innate immune cell-specific transgenic D. cerebrum lines for this project. In Aim 1, we will establish and standardize methods of inducing kidney injury in adult D. cerebrum using nephrotoxic chemical agents and laser ablation approaches. In vivo imaging of labeled kidney-specific cells combined with a fluorescence-based functional assay will allow us to observe and quantify the degree of renal damage. Traditional histological analyses will confirm and corroborate our imaging data. In Aim 2 we will demonstrate the utility of this new animal model by using imaging techniques to explore and characterize the innate immune system’s role in kidney injury and recovery. Combining new and available transgenic lines will allow us to explore the signaling mechanisms that regulate the innate immune response to kidney injury and determine what role innate immune cells play in kidney cell recovery and regeneration, or conversely, fibrosis and scar formation. This proposed study will provide the foundation for future kidney- related investigations. This model will facilitate important discoveries in the biology of kidney disease and have the potential to transform kidney research by providing a highly optically accessible and genetically tractable whole animal model system.

PROJECT NARRATIVE We aim to develop the adult Danionella cerebrum, a close relative to zebrafish (Danio rerio), for optical-based studies of kidney-related disease mechanisms. This novel model will provide advantages and serve as an attractive alternative to existing animal models because its small size, optical transparency, and ease of use. Development of this novel adult disease model will facilitate future studies allowing for the in vivo visualization of cell signaling events and cell-cell interactions responsible for kidney disease initiation and resolution, or disease progression and development of end stage renal disease.