Myelin-associated glycoprotein (MAG) has been implicated in inhibition of nerve regeneration in the CNS. This results from interactions between MAG and the Nogo receptor and gangliosides on the apposing axon, which generates intracellular inhibitory signals in the neuron. However, because myelin–axon signaling is bidirectional, we undertook an analysis of potential MAG-activated signaling in oligodendrocytes (OLs). In this study, we show that antibody cross-linking of MAG on the surface of OLs (to mimic axonal binding) leads to the redistribution of MAG into detergent (TX-100)-insoluble complexes, hyperphosphorylation of Fyn, dephosphorylation of serine and threonine residues in specific proteins, including lactate dehydrogenase and the β subunit of the trimeric G-protein-complex, and cleavage of α-fodrin followed by a transient depolymerization of actin. We propose that these changes are part of a signaling cascade in OLs associated with MAG function as a mediator of axon–glial communication which might have implications for the mutual regulation of the formation and stability of axons and myelin.