Cranial Window Surgery for In Vivo Imaging

Cranial window surgery was performed as previously described (Mostany and Portera-Cailliau, 2008 (link); Mostany et al., 2013 (link); Voglewede et al., 2019 (link)). To prevent brain swelling and inflammation mice were injected with carprofen (5.0 mg/kg; s.c.; Zoetis Inc., Parsippany-Troy Hills, NJ, USA) and dexamethasone (0.2 mg/kg; s.c.; MWI, Boise, ID, USA) after anesthesia induction and before any incision was made. Mice were anesthetized with isoflurane (5.0% for induction, 1.5%–1.7% for maintenance), placed in a stereotaxic frame, and a 4-mm craniotomy was performed with a pneumatic dental drill over S1BF, centered at 3 mm lateral to the midline and 1.95 mm caudal to bregma. A 5-mm glass coverslip (#1; Electron Microscopy Sciences, Hatfield, PA, USA) was placed over the intact dura and secured using cyanoacrylate glue and dental acrylic (Lang Dental Mfg. Co, Inc., Wheeling, IL, USA) to the skull. A custom-made titanium bar (9.5 mm × 3.2 mm × 1.1 mm) was cemented within the dental acrylic for securing the mouse to the microscope’s stage. A 3-week recovery time was allowed before in vivo imaging.