Intravital Imaging of Ultrasound-induced Blood Brain Barrier Opening Using Transgenic Mice with Two-Photon Microscopy

Delivery of therapeutic agents to the brain is limited by the blood-brain barrier (BBB), which prevents intravenously-administered drugs from reaching the central nervous system (CNS). Increasing the permeability of the BBB temporarily with focused ultrasound combined with microbubbles (FUS/MB) has great potential in non-invasive treatment of CNS disorders. Thus, it is important to understand the short-term or long-term effect in vivo of FUS/MB on cerebrovascular health and the pharmacokinetics of drugs. However, there is a spatial and temporal gap in our understanding of the drug delivery mechanism. In this study, by using Tie2-Cre::Ai14 transgenic mice that robustly express fluorescence in the brain vasculature endothelium, the dynamic effect of FUS/MB on cerebral vessels was visualized by two-photon fluorescence microscopy. We established a viable approach to investigate the interactions of FUS/MB on the brain endothelium in vivo. The dynamic process of macromolecular extravasation across the BBB could be observed in the scale of micrometers. We found the endothelial cells defect after FUS/MB treatment at the site of leakage. Moreover, it was observed that discrete punctae of the dextran attached to the outside of the blood vessel wall induced by FUS/MB, suggesting another pathway of the BBB opening.