The effects of injecting the calcium-selective ionophore, ionomycin, into myelinated tracts in the dorsal columns of adult rat spinal cords were examined electron microscopically. In vivo, ionomycin induced a primary vesicular demyelination, together with a variable degree of axonal degeneration, in a dose-dependent manner. The results are consistent with previous demonstrations that mature oligodendrocytes are more vulnerable to alterations in levels of [Ca2+]i than other glial cells. We speculate that demyelination induced by ionomycin in vivo occurs as a result of direct activation of endogenous Ca(2+)-dependent enzymes and/or as a consequence of oligodendrocyte injury mediated via astrocytes.