Stimulatory Effect of Pituitary Adenylate Cyclase-Activating Polypeptide on Catecholamine Synthesis in Cultured Bovine Adrenal Chromaffin Cells: Involvements of Tyrosine Hydroxylase Phosphorylation Caused by Ca²⁺ Influx and cAMP

Abstract

In cultured bovine adrenal chromaffin cells, pituitary adenylate cylase-activating polypeptide (PACAP) stimulated [¹⁴C]catecholamine synthesis from [¹⁴C]tyrosine (but not from [¹⁴C]DOPA) in a concentration-dependent manner, causing maximal stimulation at 10^-7 M. The stimulatory action of PACAP was not affected by staurosporine (an inhibitor of protein kinase C) or in the cells in which protein kinase C was down-regulated by prolonged exposure to TPA (an activator of protein kinase C), whereas it was partially attenuated in Ca²⁺-free medium. PACAP (10^-7M) increased the formation of [³H]inositol phosphates, [Ca²⁺]_i and ⁴⁵Ca2+ uptake as well as cAMP. The peptide also stimulated the phosphorylation of tyrosine hydroxylase, the enzyme catalyzing the rate-limiting step in catecholamine synthesis. Catecholamine synthesis and tyrosine hydroxylase phosphorylation stimulated by the maximal effective concentration of dibutyryl cAMP or high K⁺, which activates Ca²⁺ uptake, were further enhanced by PACAP, suggesting that both cAMP- and Ca²⁺-dependent protein kinases may be involved in the stimulation of tyrosine hydroxylase phosphorylation and catecholamine synthesis caused by PACAP.