Astrocytes are implicated in the function of nodes of Ranvier because their perinodal processes form contacts with the axonal membrane at nodes. We have filled astrocytes iontophoretically with horseradish peroxidase in the intact mouse optic nerve to resolve the precise relationship between perinodal processes and astrocyte three dimensional structure. We confirm that nodal contacts were formed either by single processes which almost completely enveloped nodes, or by delicate, finger-like projections from larger processes which made discrete nodal contacts. A single perinodal process can form multiple contacts with a node and nodes were contacted by processes from more than one astrocyte. Perinodal processes emanated from larger processes, which terminated as end-feet on blood vessels and at the pia, as well as collateral branches which subsequently ended at nodes; these latter may specifically subserve nodes. Perinodal contacts were also formed directly by the soma and cytoplasmic expansions of the cell body. Both primary processes and collateral branches formed multiple associations with nodes which often appeared in clusters. Thus, all astrocytes formed multiple contacts with nodes, blood vessels and the subpial glia limitans. We conclude that perinodal processes are not formed by a specialized astrocyte in the mouse optic nerve.