Quantification of anisotropy and orientation in 3D electron microscopy and diffusion tensor imaging in injured rat brain

Diffusion tensor imaging (DTI) reveals microstructural features of grey and white matter non-invasively. The contrast produced by DTI, however, is not fully understood and requires further validation. We used serial block-face scanning electron microscopy (SBEM) to acquire tissue metrics, i.e., anisotropy and orientation, using three-dimensional Fourier transform-based (3D-FT) analysis, to correlate with fractional anisotropy and orientation in DTI. SBEM produces high-resolution 3D data at the mesoscopic scale with good contrast of cellular membranes. We analysed selected samples from cingulum, corpus callosum, and perilesional cortex of sham-operated and traumatic brain injury (TBI) rats. Principal orientations produced by DTI and 3D-FT in all samples were in good agreement. Anisotropy values showed similar patterns of change in corresponding DTI and 3D-FT parameters in sham-operated and TBI rats. While DTI and 3D-FT anisotropy values were similar in grey matter, 3D-FT anisotropy values were consistently lower than fractional anisotropy values from DTI in white matter. We also evaluated the effect of resolution in 3D-FT analysis. Despite small angular differences in grey matter samples, lower resolution datasets provided reliable results, allowing for analysis of larger fields of view. Overall, 3D SBEM allows for more sophisticated validation studies of diffusion imaging contrast from a tissue microstructural perspective.