Effect of endothelin-3 (ET-3) on dibutyryl cAMP (DBcAMP)-induced stellation of rat cerebral cultured astrocytes was examined. Treatment with 1 mM DBcAMP, 10 microM forskolin, 100 microM isoproterenol and 500 nM phorbol 12-myristate 13-acetate changed protoplasmic cultured astrocytes into process-bearing ones. ET-3 (1 nM) completely prevented the astrocytic stellation induced by these agents. The effect of ET-3 showed a dose-dependence, where IC50 value and maximal effective dose were 49 pM and about 0.1 nM, respectively. ET-1 and sarafotoxin (SRTX) S6b prevented the DBcAMP-induced astrocytic stellation with potencies similar to that of ET-3. ET-3 (1 nM) did not affect the cAMP accumulation after DBcAMP treatment in cultured astrocytes. Stellate astrocytes were reversed to the protoplasmic type cells by addition of 1 nM ET-3 in the presence of DBcAMP. ET-1 and SRTX similarly reversed the astrocytic stellation. ET-3 reversed the astrocytic stellation in the absence of extracellular Ca2+. Pre-loading of BAPTA-AM, a permeable Ca2+ chelator, on stellate astrocytes had no effect on the reversal by ET-3. ET-3 did not increase intracellular free Ca2+ concentration ([Ca2+]i) of most astrocytes tested at 0.1 nM. A high concentration (100 nM) of ET-3 increased astrocytic [Ca2+]i which was negated by Ca(2+)-free and BAPTA-AM loading. These results suggest that ETs modulate morphological changes in astrocytes through cAMP- and Ca(2+)-independent mechanisms.