The formation of ethanolamine plasmalogen from labelled 1-alkyl-2-acyl-sn-glycero-3-phosphorylethanolamine was studied in neurons and glial cells of the developing rat brain. It was found that the conversion of the ether to the enol-ether bond of the 1-alkyl moiety by the neuronal and glial desaturase system requires unsaturated fatty acids at the 2 position of the substrate. There is almost no difference between the activity of the neuronal and glial desaturase during the period of active myelination, whereas the neuronal cell fraction of the adult rats displays a threefold higher enzyme activity as compared to the glial cells. Evidence for the involvement of a microsomal electron transport system in the enzymic conversion of alkylacyl-glycero-3-phosphorylethanolamine to ethanolamine plasmalogen was obtained by using specific antibodies against NADH-cytochrome b5 reductase. Cytochrome b5 stimulated the biosynthesis of ethanolamine plasmalogen.