Pituitary adenylate cyclase-activating polypeptide triggers dual transduction signaling in CATH.a cells and transcriptionally activates tyrosine hydroxylase and c-fos expression

Abstract

We used a catecholaminergic neuron-like cell line (CATH.a cells) as a model system to investigate the likelihood that pituitary adenylate cyclase- activating polypeptide (PACAP) may participate in the regulation of specific gene expression in catecholaminergic neurons. Analysis by reverse transcriptase-PCR amplification revealed the presence in these cells of type I PACAP receptors, with a short isoform, together with a heavier so-called Hop splice variant. PACAP38 and PACAP27 enhanced, in a dose-dependent manner, both cyclic AMP formation and phosphoinositide breakdown, with EC50 values of, respectively, 0.6 x 10-10 and 2 x 10-9 M. These peptides, in addition, also elevated [Ca2+](i) by mobilizing intracellular calcium pools. Vasoactive intestinal peptide (VIP) was ~1,000-fold less potent in stimulating cyclic AMP (with EC50 = 2 x 10-7 M) and failed to change the turnover of phosphoinositides and to alter [Ca2+](i). Both forms of PACAP, as well as forskolin, stimulated transcriptional induction of tyrosine hydroxylase (TH) and c-fos promoters fused to a chloramphenicol acetyltransferase (CAT) reporter gene in transiently transfected cells (p < 0.91 vs. controls). Induction of CAT activity linked to both TH and c-fos promoters was obliterated upon coexpression of a dominant inhibitory mutant (Mt-RAB) of cyclic AMP-dependent protein kinase. We conclude that CATH.a cells do express functional PACAP type I receptors, the activation of which impinges on TH and c-fos transcription according to a process that is primarily dependent on the cyclic AMP-PKA pathway.