Characterizing 3D trajectories of immune cells in live larval zebrafish*

Immune responses involve complex chemical and physical dynamics. Innate immune cells are the first responders to a variety of disturbances such as infected cells, tumors, and wounds. They also maintain microbial homeostasis, especially in the gut, where they show remarkable tolerance towards commensal microbes while preserving strong reactions to pathogens. Immune response in-vivo is typically assessed in the context of chemical signaling pathways. Physical characterization is typically limited to total cell count, but in-vitro studies show cellular motility and morphology are also altered upon stimulation. How these physical behaviors are manifested inside a living host remains unclear. We examined innate immune cells - neutrophils and macrophages - in larval zebrafish, a model vertebrate. Using light sheet fluorescence microscopy to obtain 3D images over several hours, we tracked innate immune cells in wound assays and in the presence of opportunistic gut pathogens. Characterizing speeds, measures of random-walk motion, and cellular morphology, we were able to identify a subpopulation of cells that perform directed motion towards stimuli while most cells show random or largely stationary trajectories. These observations suggest that heterogeneity is a key part of innate immune response.