The complex symptoms of chronic multiple sclerosis (MS) are due, in part, to widespread axonal pathology affecting lesioned and non-lesioned areas of the CNS. Here we describe an association between microglial activation and axon/ oligodendrocyte pathology at nodal and paranodal domains in normal appearing white matter (NAWM) of MS and experimental allergic encephalomyelitis (EAE). The extent of paranodal axo-glial (neurofascin-155+/Caspr1+) disruption correlated with the local degree of microglial inflammation and axonal injury (expression of nonphosphorylated neurofilaments) in MS NAWM. These changes were independent of demyelinating lesions and did not correlate with the density of infiltrating lymphocytes. Similar axo-glial alterations were seen in pre-symptomatic EAE, at a time-point characterised by microglia activation prior to the infiltration of immune cells. Disruption of the axo-glial unit in adjuvant immunised animals was reversible and coincided with the resolution of microglial inflammation, whereas paranodal damage and microglial inflammation persisted in chronic EAE. We were able to preserve axo-glial integrity by administering minocycline, which inhibited microglial activation, in actively immunised animals. Therefore, permanent disruption to axo-glial domains in an environment of microglial inflammation is an early indicator of axonal injury that would affect normal nerve conduction contributing to pathology outside of the demyelinated lesion.