Identification of the Vascular Source of Vasogenic Brain Edema following Traumatic Brain Injury Using In Vivo 2-Photon Microscopy in Mice

Vasogenic brain edema due to vascular leakage is one of the most important factors determining the clinical outcome of patients following acute brain injury. To date, performing a detailed in vivo quantification of vascular leakage has not been possible. Here, we used in vivo 2-photon microscopy (2-PM) to determine the spatial (3D) and temporal development of vasogenic brain edema following traumatic brain injury (TBI) in mice; in addition, we identified the vessel types involved in vascular leakage. Thirteen male Tie2-GFP mice (6-8 weeks old) were subjected to controlled cortical impact (CCI) or a sham operation; subsequently, a cranial window was prepared adjacent to the injury site, and tetramethylrhodamine-dextran (TMRM, 40mg/kg, MW 40,000) was injected intravenously to visualize blood plasma leakage. Parenchymal fluorescence intensity was monitored in three regions for 2-4h post-CCI, reaching from the surface of the brain to a depth of 300m, and TMRM leakage was measured as an increase in TMRM fluorescence intensity outside the vessel lumen and in the parenchyma. In the CCI group, vascular leakage was detected in all investigated regions as early as 2.5h post-injury. This leakage increased over time and was more pronounced proximal to the primary contusion. Both arterioles and venules contributed similarly to brain edema formation and their contribution was independent of vessel size; however, capillaries were the major contributor to leakage. In summary, using 2-PM to perform in vivo 3D deep-brain imaging, we found that TBI induces vascular leakage from capillaries, venules, and arterioles. Thus, all three vessel types are involved in trauma-induced brain edema and should be considered when developing novel therapies for preventing vasogenic brain edema.