Both ETA and ETB receptors are involved in mitogen-activated protein kinase activation and DNA synthesis of astrocytes:: study using ETB receptor-deficient rats (aganglionosis rats)

Endothelin (ET) is known to be a potent mitogen in astrocytes, However, the contribution and signalling pathway of ETA and/or ETB receptor to the proliferation of astrocytes remain unclear. We investigated ET-induced DNA synthesis in astrocytes using ETB receptor-deficient mutant rats (aganglionosis rats: sl/sl). Western blotting with anti-ET receptor subtype-specific antibodies and Scatchard analysis of binding revealed that ETB receptor expression in astrocytes depended on gene dosage (+/+: sl/+: sl/sl = 2:1:0), whereas ETA receptor expression was unchanged among the three genotypes. ET-1 (10 nM) stimulated [H-3]thymidine incorporation and mitogen-activated protein kinase (MAP kinase) activity not only in +/+ via both ETA and ETB receptors, but also in sl/sl astrocytes via ETA receptor with about half the extent of those observed in +/+ astrocytes. Treatment with pertussis toxin (PTX) suppressed the ET-1-induced increases in the incorporation and MAP kinase activity in +/+, but not sl/sl astrocytes, indicating that the ETB receptor-, but not the ETA receptor-, mediated pathway to DNA synthesis involves PTX-sensitive G proteins, e.g. G(i) and/or G(o) (G(i/o)). in +/+ astrocytes, ET-1 (1 nM) stimulated cAMP accumulation, and the ETB receptor-selective agonist IRL 1620 (1 nM) suppressed 10 mu M forskolin-induced cAMP accumulation, suggesting G(s) coupling to the ETA receptor and Oil, coupling to the ETB receptor. On the other hand, ET-1 did not increase cAMP accumulation in sl/sl astrocytes, although ET-1 (1 nM) suppressed the forskolin-induced response, suggesting G(i/o) coupling to the ETA receptor, Our results suggest the possibility that the selectivity of G protein for ETA receptor is changed from G(s) to G(i/o) in ETB receptor-deficient astrocytes.