Predicting Mesoscopic Larmor Frequency Shifts in Ex Vivo Porcine Optic Nerve

PURPOSE: Larmor frequency shifts in white matter (WM) vary with fiber orientation due to anisotropic microstructure. Since clinical voxels are significantly larger than these microscopic frequency variations, the measured signal represents a bulk average of local shifts. Accurate estimation of magnetic susceptibility therefore requires accounting for these underlying frequency distributions that exist below the imaging resolution. METHODS: We evaluated whether Microstructure-informed Quantitative Susceptibility Mapping (μQSM) can predict orientation-dependent sub-voxel frequency shifts from orientationally dispersed hollow cylinders and spherical inclusions. Diffusion-weighted and multi-gradient-echo images were acquired from ex vivo pig optic nerves at multiple orientations relative to the main magnetic field using a 3T Siemens Connectom scanner. We also analyzed de-ironed optic nerves to try and separate the effects of myelin and iron on susceptibility. RESULTS: The estimated sub-voxel frequency shifts closely matched μQSM predictions, consistent with mesoscopic field perturbations generated by uniformly magnetized axons. The de-ironing experiments yielded different results depending on fixation protocol but had minimal effect on the frequency shift, indicating a negligible iron contribution to the bulk susceptibility in porcine optic nerve. CONCLUSION: μQSM accurately reproduces the orientation dependence of Larmor frequency shifts in optic nerve WM, providing insight into their microstructural origin and supporting improved estimation of tissue magnetic susceptibility in Quantitative Susceptibility Mapping.