Fig. 4: The Aqp4-mRuby3 knock-in reporter mouse allows for in vivo imaging of the glia limitans.
From: In vivo imaging of the barrier properties of the glia limitans during health and neuroinflammation
a, c, e Schemes showing localization and the respective surgical preparations of the cranial window (a), skull thinning (c) and spinal cord window (e) for 2P-IVM imaging of the Aqp4-mRuby3 knock-in reporter mouse. 2P excitation wavelengths of 920 nm and 1045 nm were used simultaneously. Created in BioRender. Kloster, F. (2025) https://BioRender.com/dh3qfvt. b Representative images of 2P-IVM of the brain surface through a cranial window of Aqp4-mRuby3 mice after infusion of 10 kDa-FITC dextran (green) via a carotid artery catheter. Second harmonic generation (SHG) is visible in green at this excitation wavelength, superposing the collagen type I fibers (green) over the FITC dextran-filled vascular lumens (green). XY maximum intensity projections (MIP) from the whole Z-stack (top panel) and from the cortical surface excluding the dura mater (middle panel) are shown. Bottom panel shows a 70° angle YZ MIP from the cortical surface, with the AQP4-mRuby3 signal coating the vessels that are diving into the parenchyma (white arrowheads). d Representative images of 2P-IVM of the brain surface of Aqp4-mRuby3 mice through a thinned skull. XY and YZ MIPs from the surface of the brain are shown. White arrows point to AQP4-mRuby3 signal following vessels within the brain cortex. f Representative images of 2P-IVM of the spinal cord surface of Aqp4-mRuby3 mice through a cervical spinal cord window. YZ and XY MIPs from the surface of the spinal cord are shown. Data are representative of 4 independent mice for each preparation. DV dorsal vein, GL glia limitans, MIP maximum intensity projection, i.a. intra-arterial, SHG second harmonic generation.
