ACTIVATION OF TYROSINE-HYDROXYLASE BY PITUITARY ADENYLATE CYCLASE-ACTIVATING POLYPEPTIDE (PACAP-27) IN BOVINE ADRENAL CHROMAFFIN CELLS

The effect of pituitary adenylate cyclase-activating polypeptide (PACA P-27) on tyrosine hydroxylase activity has been studied in intact, cul tured, bovine adrenal chromaffin cells. Tyrosine hydroxylase activity was determined in situ by measuring the production of (CO2)-C-14 follo wing the hydroxylation and rapid decarboxylation of [C-14]tyr offered to the cells. PACAP-27 increased tyrosine hydroxylase activity 3-fold over 10 min, with an EC(50) of 10-20 nM. PACAP-38 was approximately 2- fold less potent. Removing extracellular Ca2+ reduced basal tyrosine h ydroxylase activity and the activation produced by both PACAP-27 and f orskolin by about 20%. In the absence of extracellular Ca2+, chelation of intracellular Ca2+ by treating cells with BAPTA-AM (50 mu M) cause d a consistent 40-50% reduction in basal tyrosine hydroxylase activity and in the responses to forskolin and PACAP-27. The tyrosine hydroxyl ase activation produced by PACAP-27 was unaffected by the protein kina se C inhibitor Ro 31-8220 (3 mu M), but was reduced by 85% by the prot ein kinase A inhibitor H89 (10 mu M). PACAP-27 increased cellular cycl ic AMP levels 3-fold at 100 nM. The results suggest that PACAP-27 acti vates tyrosine hydroxylase in bovine chromaffin cells through cyclic A MP formation and protein kinase A activation, and that both extracellu lar and intracellular Ca2+ modulate the effect of the adenylate cyclas e/cyclic AMP/protein kinase A signalling pathway on tyrosine hydroxyla se activity.