Immunofluorescence study on the organization of actin in astroglial cells in primary cultures

Abstract

Actin antibodies were purified by affinity chromatography from the serum of rabbits immunized with actin isolated from bovine skeletal myofibrils. By the indirect immunofluorescence technique the pattern of organization of actin was studied in cultured astroglial cells prepared from cerebral hemispheres of newborn rats. Labelling with monospecifc actin antibody revealed that the flat epitheloid astrocytes contained bundles of actin fibres arranged in characteristic patterns. Actin fibres disaggregated in round cells produced by trypsinization and reorganized when cells returned to their elongate or polygonal form. The retraction of cell bodies produced by treatment with cytochalasin B was associated with the disappearance of actin filament bundles. Reversion to the former flattened morphology and organization of fibres occurred after removal of the drug. In response to either dibutyryl adenosine 3'5'-monophosphate, 3-isobutyl-l-methylxanthine, rat brain extract or to lesser extent to norepinephrine, flat epitheloid cells took on a stellate appearance with extensive processes, resembling more closely mature astrocytes. The conversion of flat epitheloid cells into stellate cells was associated with the disappearance of immunofluorescent fibres and the appearance of diffuse immunofluorescence indicating the structural reorganization of actin from a highly organized fibrillar state into a loosely organized actin network. It is concluded that the respect to actin patterns, the behaviour of flat epithelioid astrocytes in culture does not differ from that of culture non-muscle cells of mesenchymal origin. The pattern of organization of actin in stellate process-bearing astrocytes is compatible with the involvement of actin in typical astroglial motility and in establishment of characteristic stellate shape.