3D visualization and quantification of microvessels in the whole ischemic mouse brain using solvent-based clearing...

Area of Science:

• Neuroscience
• Biomedical Engineering
• Medical Imaging

Background:

• Cerebral microvessel visualization is crucial for understanding stroke-induced brain remodeling.
• Current methods like dye injection and 2D histochemistry have limitations in accuracy and detail.
• Interpreting complex vascular circuits in 2D poses challenges for stroke research.

Purpose of the Study:

• To develop a straightforward technique for comprehensive 3D visualization of microvasculature in the entire ischemic mouse brain.
• To quantify microvessel changes and assess the impact of focal cerebral ischemia on vascular networks.
• To provide a robust method for studying microvascular injury and remodeling post-stroke.

Main Methods:

• Developed a novel protocol combining fluorescent hydrogel conjugate injection, 3D solvent clearing, and automated light sheet microscopy.
• Utilized transient middle cerebral artery occlusion in C57Bl/6j mice to induce focal cerebral ischemia.
• Employed advanced image processing and filament modeling to calculate vessel length density.

Main Results:

• Achieved detailed 3D vasculature imaging of whole ischemic mouse brains.
• Calculated vessel length density, showing a significant reduction in ischemic tissue (0.329 ± 0.131 m/mm³) compared to healthy tissue (0.922 ± 0.176 m/mm³).
• Demonstrated a marked loss of capillaries (≤10 µm) and moderate loss of small microvessels (>10–≤20 µm), with larger vessels (>20 µm) remaining unaffected.

Conclusions:

• The developed technique offers a powerful tool for high-resolution 3D visualization of cerebral microvasculature.
• The findings highlight the significant impact of focal cerebral ischemia on microvessel density, particularly affecting smaller caliber vessels.
• This protocol is highly suitable for advancing the study of microvascular injury and remodeling in stroke research.