Whole-brain three-dimensional morphometry of the mouse choroid plexus at subcellular resolution

In this repository for our publication, we provide supplementary information, code, and tables with results data to enhance the reproducibility and facilitate broader adoption of post mortem synchrotron radiation-based hard X-ray micro computed tomography (SRµCT). Documentation and data are published under Creative Commons Attribution 4.0 International, code under GNU General Public License v3.0 or later.

Reconstructed 3D SRμCT datasets (each around 4.5 TB) are available under a Creative Commons Attribution 4.0 International license via EBRAINS (Sample: mouse4) and at BioImage Archive under the accession number S-BIAD2598 together with their respective annotations (each around 30 GB, all brains).

Information on the mice, scan details and other available datasets from our Consortium can be accessed via the FABRIC4 Portal.

Abstract

Background

The choroid plexus (ChP) produces cerebrospinal fluid (CSF) and its epithelial cells form the blood–CSF barrier that regulates exchange between the blood and CSF. Despite its physiological importance, the precise size and surface area of the ChP are not well know and recent stereological analyses have indicated that the ChP surface area may approach that of the brain microvasculature. In the mouse, a principal model organism in neurobiology, the complete three-dimensional morphology of the ChP has not yet been quantified at cellular resolution across the entire brain.

Methods

We here performed whole-brain synchrotron radiation-based microtomography at 0.65 µm isotropic voxel size in three ex vivo healthy adult mouse brains stored in ethanol. A convolutional neural network was trained to segment the ChP epithelium and stroma across all ventricles, enabling direct measurement of ChP epithelial thickness, surface areas, and compartmental volumes in all ventricles without extrapolation from partial volumes data. Primary results are given for the two brains that were stored in ethanol for one day (Brain 1 and Brain 2). To assess the influence of prolonged ethanol storage, these were compared with a third brain stored for 600 days.

Results

In both Brain 1 and Brain 2, the total ChP epithelium volume was 0.48 mm³, with stroma accounting for approximately 30% of total ChP volume. The apical epithelial surface area, after accounting for microvillar amplification, was estimated at 526 to 986 mm². Without microvillar amplification, the measured epithelial surface area (63 mm² and 68 mm²) was comparable to the total ventricular wall surface (67 mm² and 75 mm²). Mean epithelial thickness was 7.1 µm and 7.8 µm for Brain 1 and Brain 2, respectively.

Conclusions

Our results confirm the ChP as a major interface for blood-CSF exchange, smaller but comparable in scale to the endothelial blood-brain barrier. Using synchrotron radiation-based microtomography and deep-learning-based segmentation, we provide the first whole-brain, three-dimensional representations of the ChP at micrometer-scale resolution. The open dataset and analysis pipeline provide a foundation for future studies of ChP remodeling in aging and disease.