Background: We conducted a multi-modal optical coherence tomography (OCT) and magnetoencephalography (MEG) study to test whether there is a relationship between retinal layer integrity and electrophysiological activity and connectivity (FC) in the visual network influenced by optic neuritis (ON) in patients with multiple sclerosis (MS). Methods: One hundred and two MS patients were included in this MEG/OCT study. Retinal OCT data were collected from the optic discs, macular region, and segmented. Neuronal activity and FC in the visual cortex was estimated from source-reconstructed resting-state MEG data by computing relative power and the phase lag index (PLI). Generalized estimating equations (GEE) were used to account for intereye within-patient dependencies. Results: There was a significant relationship for both relative power and FC in the visual cortex with retinal layer thicknesses. The findings were influenced by the presence of MSON, particularly for connectivity in the alpha bands and the outer macular layers. In the absence of MSON, this relationship was dominated by the lower frequency bands (theta, delta) and inner and outer retinal layers. Conclusion: These results suggest that visual cortex FC more than activity alters in the presence of MSON, which may guide the understanding of FC plasticity effects following MSON.