Enhanced cAMP production mediates the stimulatory action of pituitary adenylate cyclase activating polypeptide (PACAP) on in vitro catecholamine secretion from bovine adrenal chromaffin cells.
The 38 amino acid peptide pituitary adenylate cyclase activating polypeptide (PACAP) induced a dose dependent increase of catecholamine secretion in cultures of bovine chromaffin cells. This secretagogue activity of PACAP was strictly dependent on the presence of calcium in the culture medium. If calcium was omitted from the medium no effect of PACAP on catecholamine secretion could be detected during an incubation of 20 min. Preincubation of cells with 1 nM PACAP for 5 min facilitated the subsequent nicotine stimulated catecholamine secretion during a 20 min incubation without addition of the peptide. PACAP induced catecholamine secretion was clearly accompanied by a dose dependent increase of intracellular cAMP concentrations. The percentage of cells responding to PACAP with increased catecholamine secretion was assessed by immunocytochemistry of the transient appearance of dopamine-beta-hydroxylase, associated with the membranes of the chromaffin granules on the cell surface during the secretory process. About 70% of adrenal medullary cells responded to 100 nM PACAP with enhanced secretory activity. Though PACAP stimulated catecholamine secretion, we did not observed major effects on intracellular free calcium concentrations ([Ca2+]i) as determined with fura-2 by single cell fluorescence microscopy. In maximally 20% of the cells a rise in [Ca2+]i in response to a challenge with 500 nM PACAP was observed. Lower concentrations of PACAP were without an effect on [Ca2+]i. These data indicate that the stimulatory action of PACAP on in vitro catecholamine secretion from bovine chromaffin cells is linked to a rise of intracellular cAMP.
Perrin
D
D
Centre de Recherche Pierre Fabre, Department of Cancerology, Castres, France.
Germeshausen
A
A
Söling
H D
HD
Wuttke
W
W
Jarry
H
H
D000313:Adrenal Medulla / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002395:Catecholamines / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D003470:Culture Media; D000242:Cyclic AMP / Q000096:biosynthesis / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000494:pharmacology; D007150:Immunohistochemistry; D009479:Neuropeptides / Q000494:pharmacology*; D018377:Neurotransmitter Agents / Q000494:pharmacology*; D009538:Nicotine / Q000494:pharmacology; D018722:Nicotinic Agonists / Q000494:pharmacology; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D013997:Time Factors
D002395:Catecholamines; D002614:Chelating Agents; D003470:Culture Media; D009479:Neuropeptides; D018377:Neurotransmitter Agents; D018722:Nicotinic Agonists; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D004533:Egtazic Acid; D009538:Nicotine; D000242:Cyclic AMP; D002118:Calcium