Coincident elevation of cAMP and calcium influx by PACAP-27 synergistically regulates vasoactive intestinal polypeptide gene transcription through a novel PKA-independent signaling pathway.
Pituitary adenylate cyclase-activating polypeptide (PACAP) causes calcium influx, intracellular calcium release, and elevation of cAMP in chromaffin cells. Calcium influx is required for PACAP-stimulated secretion of catecholamines and neuropeptides. The role of cAMP elevation in the action of PACAP at either sympathetic or adrenomedullary synapses, however, is unknown. Here, we show that PACAP-27-induced calcium influx through voltage-sensitive calcium channels (VSCCs), together with elevation of intracellular cAMP, was sufficient to stimulate vasoactive intestinal polypeptide (VIP) biosynthesis at least 40-fold. Combined treatment of chromaffin cells with 40 mm KCl, which elevates intracellular calcium, and 25 micrometer forskolin, which elevates intracellular cAMP, caused an increase in VIP peptide and mRNA much greater than that elicited by either agent alone, and comparable to the increase caused by 10-100 nm PACAP-27. Elevation of VIP mRNA by either KCl plus forskolin, or PACAP, (1) was independent of new protein synthesis, (2) was blocked by inhibition of calcium influx through voltage-sensitive calcium channels, (3) was calcineurin dependent, and (4) was dependent on MAP kinase activation but not activation of protein kinase A. The degree of activation of two different second-messenger pathways, calcium influx and cAMP elevation, appears to determine the magnitude of transcriptional activation of the VIP gene in chromaffin cells. Maximal stimulation of VIP biosynthesis by PACAP appears to require the coincident activation of both of these pathways.
Hamelink
Carol
C
Section on Molecular Neuroscience, Laboratory of Cellular and Molecular Regulation, National Institute of Mental Health, National Institutes of Health, Bethesda, Maryland 20892, USA.
Lee
Hyeon-Woo
HW
Chen
Yun
Y
Grimaldi
Maurizio
M
Eiden
Lee E
LE
D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020013:Calcium Signaling; D002417:Cattle; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000187:drug effects / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000096:biosynthesis*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004357:Drug Synergism; D006801:Humans; D020935:MAP Kinase Signaling System; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009479:Neuropeptides / Q000494:pharmacology*; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D011188:Potassium / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D015290:Second Messenger Systems; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured; D014660:Vasoactive Intestinal Peptide / Q000378:metabolism*
C495401:ADCYAP1 protein, human; D002121:Calcium Channel Blockers; D009479:Neuropeptides; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D012333:RNA, Messenger; D005576:Colforsin; D014660:Vasoactive Intestinal Peptide; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D011188:Potassium