Advances in studying whole mouse brain vasculature using high-resolution 3D light microscopy imaging

Abstract

Significance: The cerebrovasculature has become increasingly recognized as a major player in overall brain health and many brain disorders. Although there have been several landmark studies to understand details of these crucially important structures in an anatomically defined area, brain-wide examination of the whole cerebrovasculature, including microvessels, has been challenging. However, emerging techniques, including tissue processing and three-dimensional (3D) microscopy imaging, enable neuroscientists to examine the total vasculature in the entire mouse brain. Aim: Here, we aim to highlight advances in these high-resolution 3D mapping methods including block-face imaging and light sheet fluorescent microscopy. Approach: We summarize latest mapping tools to understand detailed anatomical arrangement of the cerebrovascular network and the organizing principles of the neurovascular unit (NVU) as a whole. Results: We discuss biological insights gained from studies using these imaging methods and how these tools can be used to advance our understanding of the cerebrovascular network and related cell types in the entire brain. Conclusions: This review article will help to understand recent advance in high-resolution NVU mapping in mice and provide perspective on future studies.

Keywords: block-face imaging; brain; cerebrovasculature; light sheet fluorescence microscopy; neurovascular unit; three-dimensional light microscopy.

Fig. 1

Overview of block-face imaging versus LSFM. (a–d) STPT is used as an example of the block-face image. (a) STPT uses a built-in vibratome to physically remove brain slices after block-face imaging. (b) Results produce 3D stack of 2D high-resolution images with 1 to 2 TB in size for the whole mouse brain. (c) Maximum intensity projection (MIP) of coronal sections ($1000\text{-}\mu \mathrm{m}$ thick) with fluorescein isothiocyanate (FITC) filled vascular signals. (d) Physical serial section with optical scans in between physical sectioning can provide sufficient high resolution in 3D to resolve the entire cerebrovasculature. (e) LSFM to visualize the entire cerebrovasculature from an intact and cleared mouse brain. (f) LSFM produces about 1 to 2 TB data to visualize the whole mouse brain vasculature. (g) MIP of horizontal sections ($6600\text{-}\mu \mathrm{m}$ thick) with artery-specific antibody labeling.

Fig. 2

Summary of cerebrovascular network organization in the mouse brain. (a) The spatial arrangement of cerebrovasculature is variable across different animals. (b) Robustness of the cerebrovascular network. Cerebrovasculature from young mouse brains showed high redundancy and relatively straight vasculature with triad branching features. (c) The primary sensory cortices showed higher vascular length density than association cortices. (d) Brain stem areas show a high degree of anisotropy, with preferential directionality in a vascular arrangement. D, dorsal; V, ventral; R, rostral; C, caudal.