The role of astrocytes in the process of demyelination has been controversial. A culture system in which explants of adult cerebral rabbit white matter were maintained on a Gelfoam matrix was used for evaluating the phagocytosis of myelin by astrocytes without an influx of phagocytic cells derived from actively circulating blood-borne cells. Adult neurons were not viable for more than a few days in these explant cultures, thus resulting in empty myelin sheaths following loss of their axons. After 7 days in vitro, astrocytes, recognized by positivity for glial fibrillary acidic protein by electron microscopic immunocytochemistry, contained numerous membrane-bound vesicles filled with myelin in various stages of degradation. Although the majority of macrophage-like cells were negative for glial fibrillary acidic protein, a minority were positive, in spite of the lack of bundles of intermediate filaments, and were interpreted as astrocytic. Astrocytes were also often positive for glutamine synthetase. This study presents evidence suggesting that astrocytes may actively participate in the phagocytosis and degradation of myelin, a function generally attributed to reactive macrophages.