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Establishing sample-preparation protocols for X-ray phase-contrast CT of rodent spinal cords: Aldehyde fixations and osmium impregnation
- Source :
- Journal of Neuroscience Methods. 339:108744
- Publication Year :
- 2020
- Publisher :
- Elsevier BV, 2020.
-
Abstract
- Background Dense and unbiased cellular-resolution representations of extended volumetric central nervous system soft-tissue anatomy are difficult to obtain, even in experimental post-mortem settings. Interestingly, X-ray phase-contrast computed tomography (X-PCI-CT), an emerging soft-tissue-sensitive volumetric imaging technique, can provide multiscale organ- to cellular-level morphological visualizations of neuroanatomical structure. New Method Here, we tested different nervous-tissue fixation procedures, conventionally used for transmission electron microscopy, to better establish X-PCI-CT-specific sample-preparation protocols. Extracted rat spinal medullas were alternatively fixed with a standard paraformaldehyde-only aldehyde-based protocol, or in combination with glutaraldehyde. Some specimens were additionally post-fixed with osmium tetroxide. Multiscale X-PCI-CT datasets were collected at several synchrotron radiation facilities, using state-of-the-art setups with effective image voxel sizes of 3.03 to 0.33 μm3, and compared to high-field magnetic resonance imaging, histology and vascular fluorescence microscopy data. Results Multiscale X-PCI-CT of aldehyde-fixed spinal cord specimens resulted in dense histology-like volumetric representations and quantifications of extended deep spinal micro-vascular networks and of intra-medullary cell populations. Osmium post-fixation increased intra-medullary contrast between white and gray-matter tissues, and enhanced delineation of intra-medullary cellular structure, e.g. axon fibers and motor neuron perikarya. Comparison with Existing Methods Volumetric X-PCI-CT provides complementary contrast and higher spatial resolution compared to 9.4 T MRI. X-PCI-CT’s advantage over planar histology is the volumetric nature of the cellular-level data obtained, using samples much larger than those fit for volumetric vascular fluorescence microscopy. Conclusions Deliberately choosing (post-)fixation protocols tailored for optimal nervous-tissue structural preservation is of paramount importance in achieving effective and targeted neuroimaging via the X-PCI-CT technique.
- Subjects :
- Micro-CT
X-ray phase-contrast
0301 basic medicine
Materials science
Rodentia
computer.software_genre
03 medical and health sciences
chemistry.chemical_compound
Percutaneous Coronary Intervention
0302 clinical medicine
Neuroimaging
Voxel
medicine
Fluorescence microscope
Animals
Image resolution
Aldehydes
Synchrotron radiation
medicine.diagnostic_test
X-Rays
Soft-tissue fixation
General Neuroscience
Spinal cord imaging
Magnetic resonance imaging
X-Ray Microtomography
Osmium
Spinal cord
Rats
030104 developmental biology
medicine.anatomical_structure
Spinal Cord
Multiscale neuroimaging
chemistry
Osmium tetroxide
Glutaraldehyde
computer
030217 neurology & neurosurgery
Biomedical engineering
Subjects
Details
- ISSN :
- 01650270
- Volume :
- 339
- Database :
- OpenAIRE
- Journal :
- Journal of Neuroscience Methods
- Accession number :
- edsair.doi.dedup.....15cd202d504cab614c885db480f50428