Computational protocol: Disruption of astrocyte-vascular coupling and the blood-brain barrier by invading glioma cells

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Protocol publication

[…] Animals were anesthetized with 2-5% isoflurane. The hair and skin of the skull was removed. A metal bar was secured to the skull behind lambda by dental cement. Removal of the skull anterior to lambda and posterior to bregma was performed on one hemisphere. Animals were injected into the tail vein with 200 μL of 5 mg ml-1 70,000 MW tetramethylrhodamine dextran (Invitrogen) with a 28 gauge, 0.5 inch long needle (BD Biosciences). Animals were placed on a custom built apparatus with a heating pad and the metal bar was secured to immobilize the animal's skull. Arteries and arterioles were identified by their size (<7 μm) and direction of blood flow or by retro-orbitally administered Alexa Fluor 633 Hydrazide (Invitrogen). Using a multiphoton laser scanning fluorescence microscope (Prairie Technologies) equipped with a X40/0.8 N.A. W water-immersion objective (Olympus) Z stacks of perivascular glioma cells were acquired. Channels were unmixed using ImageJ software and projection images created using Imaris x64 7.5.2 Software (Bitplane Scientific Software). […]

Pipeline specifications

Software tools ImageJ, Imaris
Applications Laser scanning microscopy, Microscopic phenotype analysis
Organisms Mus musculus
Diseases Glioma, Neoplasms