Neuritis of the optic nerve is one of the most frequent early symptoms of multiple sclerosis. There are only scarce data correlating magnetic resonance imaging (MRI) contrast alterations with the underlying pathology, that is inflammation, demyelination, and axonal damage. Here we studied optic neuritis in a rat model of experimental autoimmune encephalomyelitis by comparing in vivo MRI findings from multiple techniques (T1, T2, proton density, magnetization transfer) to histopathology. We further assessed a breakdown of the blood-brain barrier by using Gd-DTPA and indirectly estimated the intracellular accumulation of calcium as a consequence of axonal damage by using manganese-enhanced MRI. Hyperintensity on T2-weighted images and signal enhancement after Gd-DTPA were highly sensitive to lesions of the optic nerve but did not differentiate between mild, moderate, and severe damage. Signal reduction on T1-weighted images was less sensitive but correlated well with the severity of tissue damage. No significant changes in magnetization transfer ratio were observed. Manganese ions tended to accumulate in the central parts of the inflamed optic nerve. The resulting signal enhancement at 24 h after administration positively correlated with the severity of axonal loss. Thus, manganese might be an indicator of intracellular calcium accumulation that is known to be associated with axon damage. Although none of the methods alone distinguished between inflammation, demyelination, and reduced axon density, their specific capabilities should prove useful for future in vivo MRI studies of optic neuritis in both animal models and humans.