Retrieving neuronal orientations using 3 D 1 scanning SAXS and comparison with 2 diffusion MRI 3

While diffusion MRI (dMRI) is currently the method of choice to non-invasively probe tissue 14 microstructure and study structural connectivity in the brain, its spatial resolution is limited and its 15 results need structural validation. Current ex vivo methods employed to provide 3D fiber orientations 16 have limitations, including tissue-distorting sample preparation, small field of view or inability to 17 quantify 3D fiber orientation distributions. 3D fiber orientation in tissue sections can be obtained 18 from 3D scanning small-angle X-ray scattering (3D sSAXS) by analyzing the anisotropy of scattering 19 signals. Here we adapt the 3D sSAXS method for use in brain tissue, exploiting the high sensitivity 20 of the SAXS signal to the ordered molecular structure of myelin. We extend the characterization of 21 anisotropy from vectors to tensors, employ the Funk-Radon-Transform for converting scattering 22 information to real space fiber orientations, and demonstrate the feasibility of the method in thin 23 sections of mouse brain with minimal sample preparation. We obtain a second rank tensor 24 representing the fiber orientation distribution function (fODF) for every voxel, thereby generating 25 fODF maps. Finally, we illustrate the potential of 3D sSAXS by comparing the result with diffusion 26 MRI fiber orientations in the same mouse brain. We show a remarkably good correspondence, 27 considering the orthogonality of the two methods, i.e. the different physical processes underlying 28 the two signals. 3D sSAXS can serve as validation method for microstructural MRI, and can provide 29 novel microstructural insights for the nervous system, given the method’s orthogonality to dMRI, 30 high sensitivity to myelin sheath’s orientation and abundance, and the possibility to extract myelin31 specific signal and to perform micrometer-resolution scanning. 32