The peripheral olfactory nervous system exhibits, uniquely, neuronal cell body replacement and reestablishment of central connections in adult mammals. The role of the olfactory nerve glia in these phenomena is unknown, but information might be provided by in vitro systems. This paper reports on the characterization of olfactory nerve glia in dissociated cell cultures of newborn rat nasal mucosal tissues. The predominant type of glial cell resembled Schwann cells and immunostained for the S-100 protein, found in all glial cell types; glial fibrillary acidic protein (GFAP), found in astrocytes and nonmyelinating Schwann cells; and showed binding of 217C, a monoclonal Schwann-cell marker that binds to the low-affinity NGF receptor in glioma cells. They were negative for A2B5. The Schwann-cell-like olfactory glia changed morphology upon culturing in serum-free medium, with further shape changes after plating on laminin. Plating on laminin increased cell numbers. A second population, found only after GFAP-immunostaining, was astrocyte-like in morphology and represented approximately 10 percent of all glial cells. These were S-100-, A2B5-, and 217C-negative, a unique glial cell immunological profile. At low dilutions of anti-GFAP (1/10,000), or with weak fluorescent secondary antibodies, astrocyte-like glia were immunostained but Schwann-cell-like glia were not detectable. Astrocyte-like glia were not an artifact of the dissection, since they were detectable in tissue sections of newborn-rat olfactory nerves immunostained with a low dilution of anti-GFAP. The presence of two types of glial cells in culture suggests similarities between olfactory glia and enteric glia.