Intracerebral Infusion of Contrast Agent in Mice

The anesthetized mice were positioned in a stereotaxic frame and their neck slightly flexed with care to not compromise the airway and spontaneous breathing pattern. Through a midline incision, the atlanto-occipital membrane was exposed and a 34-ga shortened needle (Hamilton, US) connected via polyurethane tubing to a 50 µl Hamilton syringe (Hamilton, US) mounted in a micro-infusion pump (Legato 130, KD Scientific, Holliston, MA, USA) was inserted into the CM. Previous studies reporting on tracer delivery for GS transport studies in mice via the CM have used infusion rates of 0.1–2 µl/min for a total volume of 5–10 µl^{22 (link),37 (link)–40 (link)}. In this study we infused 7 µl of Gd-DOTA (Guerbet LLC, Princeton, NJ, US) prepared as a 1:20 dilution in sterile 0.9% NaCl or 7 µl of sterile 0.9% NaCl at an infusion rate of 1 µl/min. After the infusion, the needle was left in place for 2-min, after which it was removed, and the dura sealed with cyanoacrylate glue. Collectively, anesthesia, CM exposure and Gd-DOTA infusion required ~ 30 min.