Craniula: A cranial window technique for prolonged imaging of brain surface vasculature with simultaneous adjacent intracerebral injection

Abstract

BackgroundImaging of the brain surface vasculature following inflammatory insults is critical to study structural and functional changes in the living brain under normal and pathological conditions. Although there have been published reports relating to the changes that occur in the blood brain barrier (BBB) during the inflammatory process, the ability to visualize and track such changes in vivo and over time has proven to be problematic. Different techniques have been used to achieve visualization of pial vessels, but the approach has limits, which can jeopardize the well-being of the animals. Development of the cranial window technique provided a major advance in the acquisition of live images of the brain vasculature and its response to different insults and treatments.MethodsWe describe in detail a protocol for delivery of a localized inflammatory insult to the mouse brain via a craniula (cranial window and adjacent cannula) and subsequent imaging of the mouse brain vasculature by intravital microscopy and two-photon laser scanning microscopy. The surgical implantation of the craniula can be completed in 30-45 min and images can be acquired immediately and for several months thereafter. The technique is minimally invasive and permits serial injections directly to the brain, thereby allowing longitudinal imaging studies. The craniula technique permits the study of structural and functional changes of the BBB following inflammatory insult and as such has wide application to neuroscience research.

Secoisolariciresinol diglucoside is a blood-brain barrier protective and anti-inflammatory agent: implications for neuroinflammation

“…Intravital video microscopy (IVM) (via cranial window) was used to quantify in vivo leukocyte adhesion in the presence or absence of SDG treatment and inflammatory insult [ 31 ]. 5 days after implantation of the cranial window with adjacent cannula [ 31 ], mice were injected with rhodamine 6G (0.1%).…”

PARP inhibition in leukocytes diminishes inflammation via effects on integrins/cytoskeleton and protects the blood-brain barrier

“…Prior to IVM, mice were treated with LPS (6 mg/kg) i.v. or TNFα (0.5 μg/mouse) by IC injection [ 9 , 35 , 42 , 43 ]. Two hours post-injection of TNFα, leukocytes were labeled in vivo with Vybrant® DiI Cell-Labeling Solution (DiI) (Life Technologies, Carlsbad, CA) introduced i.v.…”

Removable cranial window for sustained wide-field optical imaging in mouse neocortex

“…With this system, however, the methods used to remove and replace are faster and simpler and carry less risk of tissue damage compared. Additionally, the described methods of facilitating tissue access can be advantageous over a fixed access port by providing full access without compromising the image field (12)(13)(14).…”

“…Using a wide-field fluorescence microscope with a scientific-CMOS camera, we can record activity in hundreds of distinct neurons across wide areas of the brain of awake behaving mice (7)(8)(9). Additionally, when combined with optical or pharmacological manipulation and/or electrophysiological recording, optical imaging provides a sophisticated approach to investigate neuronal function within the larger neural network (10)(11)(12)(13)(14). One of the key advantages of in vivo imaging regards the ability to observe and record from the same brain region for extended periods to track long-term changes (1,9,15,16).…”

“…The ability to access and manipulate tissue during real-time imaging opens the door in experimental designs to an expansive toolbox of neuromanipulation possibilities allowing exploration of uncharted connectivity and dynamic processes of the brain (17). Several strategies have been reported to gain access to regions below glass cranial windows by incorporating features such as an access port sealed with elastomer (12,13), infusion cannula (14,32), or the use of microfluidic channels (25). Nonetheless, the approaches limit the tissue accessibility to a single designated site predetermined before an experiment begins and do not offer uniform access over the imaging area.…”