Direct blood flow velocity measurements of zebrafish cerebral vessels with a dual-mode light-sheet microscope

Characterization of cerebral blood flow is critical to studying brain function and disorders. We report a method for directly measuring zebrafish cerebral blood flow velocity in 3D. A dual-mode light sheet microscopy combined with structured illumination was developed to image zebrafish 3D brain vasculature in structured illumination mode and to trace the blood cells in fast imaging mode at the same region. Evans blue (EB) was injected into the heart of zebrafish to visualize the vessels, while the blood cells appeared to be dark in the vessels. Our approach significantly improves imaging speed to 100 Hz. Algorithms were developed to trace the blood cells and match the flow velocity to vascular structures. The blood flow velocity in the cerebral microvessels at different depths was obtained for zebrafish up to 8 days post-fertilization (dpf). The flow velocity and flux in "Y" shaped and curved vessels were analyzed.