methods
Functional MRI Processing for Mouse Micturition Studies
Functional MRI (fMRI) studies in rodents is somewhat uncommon as it requires some cognitive or physiological stimulus that causes measurable changes in brain activity. This project however, lends itself well to fMRI studies as voiding induced by cystometry has a cyclical rhythm that is regulated by Barrington’s nucleus (aka PMC) and the periaqueductal gray regions of the brain; and these studies can be done while the animal is anesthetized with urethane.
Image Acquisition
Cystometry Catheter. Mice are anesthetized with 1.2g/kg body weight urethane. Surgery to implant the catheter is carried out using continuous flow 3% isoflurane. A description of the procedure can be found here.
MRI scanning. Magnetic Resonance Imaging (BOLD-MRI) is carried out in a Bruker 9.4T magnet with a 4 element mouse brain phased array coil. A series of 2D multislice gradient echo-planar images are acquired every 2s while the mouse undergoes cystometry. 25 slices are acquired with slice 500µm, interslice distance 600µm, and in-plane resolution 250µm. Imaging occurs over a 40 minute time frame during which the mouse undergoes 10-15 voiding cycles (CMG).
Data Processing
Processing is mostly performed within the program Statistical Parametric Mapping (SPM) from the Wellcome Institute for Cognitive Neurology. It is free software that runs within MATLAB. In addition, a toolbox called spmmouse has been downloaded for display on mouse atlases.
fMRI processing of our mouse micturition studies is still in development. Particular outstanding issues to resolve include:
1 Motion reduction and correction
2 Accurate registration of echoplanar images to high resolution anatomic images
3 Identifying or defining anatomic location of smaller nuclei (PMC, …)
4 Converting pressure curves to key features for linear model analysis.
Links to software used
MRICRON: http://www.cabiatl.com/mricro/mricron/dcm2nii.html
Statistical Parameter Mapping (SPM): http://www.fil.ion.ucl.ac.uk/spm/software/spm12/
MatLab (note this is commercial software): https://www.mathworks.com/?s_tid=gn_logo
SPM_Mouse “Link unreliable, you may need to search for this”: http://spmmouse.org
The mouse brain template: https://www.nitrc.org/projects/tpm_mouse
We use the C57BL/6J template: https://www.nitrc.org/frs/?group_id=935&release_id=3299
The Allen Brain Atlas – much fun if you are interested in exploring either human or mouse brains. Hint: find Barrington’s nucleus also called the Pontine Micturition Center (PMC). This is the part of the hindbrain that triggers voiding: https://portal.brain-map.org
Step by Step Methods
* Convert Images. Our Bruker scanner produces images in DICOM format. SPM12 requires NifTi images for analysis. Images can be converted in SPM12 or separately using MRICRON.
* Converting images using MRICRON (link to separate pdf for this method).
* All steps in SPM12 (link to a pdf file containing detailed descriptions of the Realignment).
* Realignment and re-slicing creates an averaged (mean) image. This step will also show how much movement occurred during the scan.
* Mask Image. Masking is done to smooth processing and remove any regions that are not brain.
* Reorientation. Image Rotation or Reorientation is done so that the coronal, sagittal and axial scan images are in the same orientation as the mouse brain template.
* Normalization. Normalization fits the scanned image to a template brain so that anatomical structures can be identified.
* Correlation to Cystometry Data. Here the cystometry data are matched to the scanned images using Excel.
* Level 1 Analysis.
* Estimate.
* Review.
Do you want to try analysis of a dataset? Here is a link that will take you to a zipped file containing anatomical and BOLD MRI (fMRI) scans from seven male C57BL/6J mice. Note this is a large file of about 2.16GB.