Repetitive electrical activity produces microstructural alteration in myelinated axons, which may afford the opportunity to noninvasively monitor function of myelinated fibers in peripheral nervous system (PNS)/CNS pathways. Microstructural changes were assessed via two different magnetic-resonance-based approaches: diffusion fMRI and dynamic T2 spectroscopy in the ex vivo perfused bullfrog sciatic nerves. Using this robust, classical model as a platform for testing, we demonstrate that noninvasive diffusion fMRI, based on standard diffusion tensor imaging (DTI), can clearly localize the sites of axonal conduction blockage as might be encountered in neurotrauma or other lesion types. It is also shown that the diffusion fMRI response is graded in proportion to the total number of electrical impulses carried through a given locus. Dynamic T2 spectroscopy of the perfused frog nerves point to an electrical-activity-induced redistribution of tissue water and myelin structural changes. Diffusion basis spectrum imaging (DBSI) reveals a reversible shift of tissue water into a restricted isotropic diffusion signal component. Submyelinic vacuoles are observed in electron-microscopy images of tissue fixed during electrical stimulation. A slowing of the compound action potential conduction velocity accompanies repetitive electrical activity. Correlations between electrophysiology and MRI parameters during and immediately after stimulation are presented. Potential mechanisms and interpretations of these results are discussed. © 2018 National Academy of Sciences. All rights reserved.