A ROLE FOR PROTEIN-TYROSINE KINASE IN THE STEROIDOGENIC PATHWAY OF ANGIOTENSIN-II IN BOVINE ZONA GLOMERULOSA CELLS

Stimulation of aldosterone synthesis in bovine adrenal zona glomerulosa (ZGB) cells by angiotensin II (AngII) is believed to be mediated by the phospholipase C (PLC) pathway that results in the increase of cytosolic free calcium concentration and in the activation of protein kinase C (PKC). However, the cell proliferation and contraction associated with AngII action are known to be mediated in part by protein tyrosine kinases (PTK). To assess the potential role of PTK in the stimulatory effect of AngII on adrenal steroidogenesis, the actions of a series of PTK inhibitors on this metabolic pathway were examined in isolated ZGB cells. Tyrphostin 23 (TP23) caused a dose-dependent inhibition of AngII-stimulated aldosterone production with an IC50 of 15 mu M and reached complete inhibition at 100 mu M Genistein (GS) was more potent with an IC50 of 35 nM and complete inhibition at 10 mu M. The stimulation of aldosterone production by the calcium-mobilizing agent thapsigargin (Thaps) was also dose-dependently inhibited by TP and GS with the same potency. A specific PKC inhibitor, calphostin C (0.1 mu M) caused only a 51.7% inhibition of AngII-stimulated aldosterone production. In the same way, a specific Ca2+/calmodulin-dependent protein kinase inhibitor, KN-62 (1 mu M), reduced aldosterone production stimulated by AngII by 64%. As expected, thapsigargin-stimulated aldosterone biosynthesis was not affected by calphostin C, but was completely inhibited by KN-62. These results demonstrate for the first time that protein tyrosine kinase activity is part of the angiotensin II signalling pathway in bovine zona glomerulosa cells. The activation of this PTK occurs subsequently to the mobilization of intracellular calcium. This calcium-dependent protein tyrosine kinase pathway is essential for the steroidogenic response to AngII in bovine zona glomerulosa cells.