DOI: 10.5281/zenodo.15690357
title: Synchrotron radiation-based micro computed tomography dataset - in vivo mouse brain - CA019 - Choroid plexus motion

description: Imaging choroid plexus motion at CLS BMIT beamline

This datasets features a synchrotron radiation-based micro computed tomography dataset of a C57BL/6J mouse injected with barium-based contrast agent into the lateral ventricle. Data was recorded at the beamline BMIT of the Canadian Light Source in November 2023. For more information, see the associated publication, repository or the FABRIC4 portal.



subjCA019_scan002_slice490_time-steps.zip2D slice z = 490, all time points except 32, from start of contrast agent infusion

subjCA019_scan002_slice597_time-steps.zip2D slice z = 597, all time points except 32, from start of contrast agent infusion

subjCA019_scan002_time-steps_036_scan_000.zipFull 3D-stack, time point 36, 36 min after start of contrast agent infusion

subjCA019_scan002_time-steps_038_scan_000.zipFull 3D-stack, time point 38, 38 min after start of contrast agent infusion

Supplementary_Video_6.mp42D slice z = 490, time points 14-31 and 33-50, starting 13 min after start of contrast agent infusion


Methods

C57BL/6J (stock 000664) mice were supplied by Jackson Labs. To ensure proper acclimatization, the animals were housed within the local animal facility one week before the experiments. The experimental protocols were reviewed and approved by the responsible ethics committee of the University of Saskatchewan, approval number AUP 20230072 MOD#1 

For the dataset shown, a male mouse (SubjectID: CA019) twelve weeks of age and 28.5 g body weight was first injected subcutaneously with buprenorphine (0.1 mg/kg) for analgesia. Anesthesia was induced after onset of analgesia 30 min later, via intraperitoneal injection of a cocktail of ketamine (50 mg/kg) and medetomidine (1 mg/kg). The animals were kept warm using heating pads. The depth of anesthesia was monitored by testing reflexes, and additional intraperitoneal injections were given as needed. Eye ointment was applied and the skull, neck, and upper thoracic region of the mouse were shaved to avoid potential artefacts during X-ray imaging. 2 × 0.5 ml glucose 10% was administered in two separate subcutaneous injections, and tracheotomy was performed using a custom, radiotransparent, 3D-printed plastic tracheal cannula with Y-adapter.

The mouse was then secured in a stereotactic frame and ventilated with a MiniVent, providing 96% oxygen from an oxygen accumulator device and 0.5 – 2% isoflurane. The exhaled air was routed through a water column to maintain 2 cmH2O positive end-expiratory pressure. Stroke volume was set to 150 µl, respiration rate to 140 bpm.For cannula implantation, excess periosteum from the bone was removed and the bregma was identified. A small hole of about 1 mm diameter was drilled through the parietal bone at the injection coordinates. An MRI-compatible PEEK infusion cannula of 2.3 mm length was filled with standard concentration contrast agent 320 mg Ba/ml and implanted into the right lateral ventricle. Cannula coordinates were 1.2 mm lateral and 0.03 mm caudal of the bregma.

The animal was then transferred to the radiation hutch containing the SRµCT imaging setup, where it was imaged at a photon energy of 37.8 keV, provided by the beamline’s double bent Laue crystal monochromator. 2 000 radiographs over a rotation range of 180° were acquired with a pco.edge 4.2 camera coupled with a tandem lens system (105 mm f/2.4 and 85 mm f/1.4) and a 200 µm LuAG:Ce scintillator for 8 µm effective pixel size. The field of view was reduced to 2048 × 1200 pixels due to the limited size of the X-ray beam, and recorded with 15 ms exposure time and 62 frames per second, meaning 1 ms overhead time. Acquisition time per scan was 32 s. Sample-detector distance was 0.6 m, source-sample distance was 57.8 m.

One tomographic scan was acquired every minute for 50 min, while the animal was infused with contrast agent. Each scan took 32 s, the rotation stage returned to the start position in 18 s and the next scan was started after a 10 s delay. Contrast agent flow rate was ramped up linearly from 0 to 0.25 µl/min within the first minute, then kept until 5 µl of contrast agent were infused. To avoid backflow of CSF into the infusion cannula, a very low flow rate of 0.02 µl/min was maintained thereafter. Flat-field images were only acquired at the beginning and the end of the 50 scans.

Tomograms were reconstructed using the image processing toolkit tofu, which employs the UFO framework as a back-end. Center-of-rotation was determined automatically. Output image intensity values correspond to µΔx, the product of the linear attenuation coefficients µ and the voxel length Δx.In order to isolate potential motion of the ventricles from the effect of whole animal motion, data volumes of the different time points used for comparison were registered to the first reconstruction of the series (called reference image) via rigid registration. An extended bone mask without ventricular spaces was created for the reference image to ensure that the contrast enhancement of the ventricles would not affect registration. The registration was driven by maximizing normalized cross-correlation as an image similarity measure within the extended bone mask. Automatic image registration used the open-source software elastix (version 4.9). Images were rescaled to the intensity range in the ROI of the reference image to reduce quantification errors when using 256 bins for covering the intensity range during registration. Images of the original intensities were afterwards transformed based on the registration result.

To visualize examples of the movement in 2D occurring during 1 min and 2 min, respectively, coronal and transversal planes were calculated between three timepoints of a time series (36, 37 and 38 min after infusion start) were subtracted. The 3D quantification of choroid plexus motion was based on datasets with a time difference of 2 min (36 and 38 min after infusion start).