Changes in actin cytoskeleton during volume regulation in C6 glial cells

Changes in actin cytoskeleton in. the C6 rat glial cell line were studied d uring decrease or increase (abrupt or gradual) of extracellular osmolality. Actin cytoskeleton was visualized by confocal microscopy after FITC-phallo idin labeling. G-actin, Triton-soluble F-actin and Triton-insoluble F-actin subfractions were determined by gel electrophoresis and scanning, and by D Nase I inhibition assays. In control conditions C6 glial cells exhibited we ll-defined stress fibers and a relatively smooth cortical network. Extracel lular anisosmotic changes induced a rapid actin cytoskeletal reorganization , which further progressed and was not reversed upon cell volume recovery. Hypotonic shock caused membrane ruffling and a shift towards polymerized ac tin, whereas hypertonicity (abrupt or gradual) led to a distinct morphologi cal appearance of abundant short actin microfilaments with, however, no det ectable alteration in actin subfractions. When anisosmotic cell volume regu lation was prevented, cytoskeleton reorganization depended on the osmotic c hange and the experimental protocol, but was not related to the absence of volume readjustment. Therefore, although involvement of cytoskeletal altera tions in transduction of volume regulatory responses cannot be excluded, it is likely that the observed changes in actin cytoskeleton in C6 glial cell s are linked with, but do not initiate, cell volume regulatory processes.