TRANSDUCTION MECHANISMS INVOLVED IN THROMBIN RECEPTOR-INDUCED NERVE GROWTH-FACTOR SECRETION AND CELL-DIVISION IN PRIMARY CULTURES OF ASTROCYTES

In astrocytes, thrombin and thrombin receptor-activating peptide (TRAP -14), a 14-amino-acid agonist of the proteolytic activating receptor f or thrombin (PART), significantly increased cell division as assessed by [H-3]-thymidine incorporation into DNA (EC(50) = 1 nM and + 650% at 100 nM for thrombin; EC(50) = 3 mu M and + 600% at 100 mu M for TRAP- 14) and nerve growth factor (NGF) secretion (approximately twofold at 100 nM thrombin or 100 mu M TRAP-14). The [H-3]thymidine incorporation was prevented by protein kinase C inhibitors (staurosporine and H7) o r by down-regulation of this enzyme by chronic exposure of astrocytes to phorbol 12-myristate 13-acetate (PMA). Thrombin-induced NGF secreti on was completely inhibited by protein kinase C inhibitors. Treatment with PMA stimulated NGF secretion 19-fold, and this effect was not fur ther enhanced by thrombin. These data suggest an absolute requirement of protein kinase C activity for thrombin-induced NGF secretion and ce ll division. Pretreatment of astrocytes with pertussis toxin (PTX) red uced thrombin- and TRAP-14-induced DNA synthesis. PART activation caus ed a decrease in forskolin-stimulated cyclic AMP accumulation. PTX tre atment prevented the inhibitory effect of PART activation on cyclic AM P accumulation, suggesting that a PTX-sensitive G protein, such as G(i ) or G(o), is involved in thrombin-induced cell division. In contrast, thrombin-induced NGF secretion was not inhibited by PTX. Finally, the protein tyrosine kinase inhibitor herbimycin A partially but signific antly prevented thrombin- and TRAP-14-induced cell division but was wi thout effect on NGF secretion. Taken together, these results demonstra te that, in astrocytes, PART(s)-triggered cell division or NGF secreti on is mediated by distinct transduction mechanisms.