Effects of extracellular matrix macromolecules on the differentiation of plasma membrane structure in cultured astrocytes.

Orthogonal aggregates of small intramembrane particles, termed "assemblies," are concentrated especially in the cell membranes of astrocytic processes that form the glia limitans at the outer surface of the brain and the perivascular sheath surrounding the parenchymal blood vessels. As an initial step to clarifying the totally unknown biochemical nature of this intramembrane structure, we have devised a culture system which enhances the differentiation of assemblies in secondary cultures of astrocytes derived from neonatal mouse neopallium. Since assemblies are most concentrated in the plasma membranes attaching to the basement membrane, we expected that extracellular matrix molecules constituting the basement membrane would be suitable candidates for our aim. We report here that a mixture of type IV collagen, laminin, and fibronectin, major components of the extracellular matrix, has the potency to increase assembly density in cultured astrocytes. We also report that, in freeze-fracture electron microscopy of cultured cells, one can satisfactorily preserve membrane structure and reliably obtain large replicas by inoculating cells on aluminium foil and peeling it from the cells in a freeze-fracture apparatus.