Intravital Imaging of Mouse Ear Vasculature

Intravital imaging of the mouse ear with intradermal injection has been described previously (Honkura et al., 2018 (link)). Briefly, following systemic administration of 2000 kDa FITC (SigmaAldrich, FD2000S) or TRITC Dextran (ThermoFischer Scientific, D7139) by tail-vein injection, mice were sedated by intraperitoneal injection of Ketamine-Xylazine (120 mg/kg Ketamine, 10 mg/kg Xylazine) and the ear secured to a solid support. Mice were maintained at a body temperature of 37 °C for the entire experiment, maximum 90 min. Time-lapse imaging was performed using single-photon microscopy (Leica SP8). For intradermal EC stimulation, a volume of approximately 0.1 μl histamine (SigmaAldrich, H7125), concentration 10 ng/μl, was injected using a sub-micrometer capillary needle. 10 kDa TRITC Dextran (ThermoFischer Scientific, D1817) was used as a tracer. Leakage sites were identified in time-lapse imaging as defined sites of concentrated dextran in the extravascular space.