ENDOTHELINS ARE EXTRACELLULAR SIGNALS MODULATING CYTOSKELETAL ACTIN ORGANIZATION IN RAT CULTURED ASTROCYTES

Effects of endothelin-3 on rapid morphological changes and cytoskeletal actin organization of rat cortical cultured astrocytes were examined. In serum-free medium, treatments with 1 mM dibutyryl cAMP and 5 mu M cytochalasin B, an inhibitor of actin polymerization, caused astrocytic morphological changes with cytoplasmic retraction (stellation). Concurrent addition of 1 nM endothelin-3 prevented astrocytic stellation by dibutyryl cAMP and cytochalasin B. The inhibition of endothelin-3 on the astrocytic stellation was dose-dependent, where IC50 and maximal effective dose were about 50 pM and 0.1 nM, respectively. Endothelin-1 and sarafotoxin S6b prevented the cytochalasin B-induced stellation with similar potencies to endothelin-3. Endothelin-3 reversed the stellate morphology of cytochalasin B-treated cells. Sixty minutes after addition of endothelin-3, most cytochalasin B-treated astrocytes lost their apparent distinction between cell body and processes. Treatment with dibutyryl cAMP and cytochalasin B decreased actin content in a 0.5% Triton X-100-insoluble fraction (cytoskeletal fraction) of cultured astrocytes. Subsequent treatments with endothelin-3 for 2 h restored the decreased cytoskeletal actin to that of non-treated cells. Rhodamine-phalloidin staining showed that a prominent structure of organized filamentous actin in protoplasmic astrocytes is stress fibers. The astrocytic stress fibers disappeared after treatment with dibutyryl cAMP and cytochalasin B. Endothelin-3 stimulated reorganization of stress fibers both in the dibutyryl cAMP- and the cytochalasin B-treated astrocytes. These results suggest that endothelins are extracellular signals to regulate cytoskeletal actin organization of astrocytes.