Immunocytochemical localization of GD3 ganglioside to astrocytes in murine cerebellar mutants.

Biochemical analysis of the murine mutants, Purkinje cell degeneration (pcd/pcd), staggerer (sg/sg) and lurcher (Lc/+), which are characterized by neuronal degeneration in the cerebellar cortex, have revealed substantially elevated levels of GD3 ganglioside (ceramide-Glu-Gal-NANA-NANA). Ultrastructural studies on pcd/pcd and sg/sg have shown astrocytes elaborating slender sheet-like processes which wrap around neuronal processes. Seyfried et al. hypothesized that the elevation in GD3 seen in these mutants is attributed to its expression by altered astrocytes. Using a monoclonal antibody to GD3 and a polyclonal antibody to GFAP we have explored the cellular localization of GD3. Positive immunofluorescence was observed in sg/sg, pcd/pcd and Lc/+ cerebella, but not in age-matched normal littermates or in weaver (wv/wv) a fourth cerebellar murine mutation which destroys granule cells prior to their migration across the molecular layer. In wv/wv cerebella, astrocytes do not elaborate sheets of processes and no significant elevations of GD3 are observed biochemically. The positive GD3 staining in pcd/pcd and Lc/+ was confined to the granule cell layer and appeared as many punctate or short, fine profiles, suggestive of binding to thin cytoplasmic processes. No GD3 positive staining was seen in the Bergmann glia or astrocytes of the white matter. GD3-positive staining was seen throughout the cortex in sg/sg which displayed severe disruption of its histoarchitecture with no clear delineation between the molecular and granule cell layers. Ultrastructural localization of GD3 was performed using pre-embedding immunocytochemistry with a PAP technique in sg/sg mice. The cytoplasmic processes and cell bodies of astrocytes displayed positive membrane staining. Our results suggest that astrocytes undergo important changes in membrane composition during pathological reaction caused by neuronal degeneration.