Adenylyl cyclase 6 mediates the action of cyclic AMP‐dependent secretagogues in mouse pancreatic exocrine cells via protein kinase A pathway activation

Abstract

Key points• Cyclic AMP (cAMP), produced from ATP and the enzyme adenylyl cyclase (AC), plays an important role in the regulation of pancreatic exocrine cells.• We identified five AC isoforms in pancreatic exocrine cells. AC3, AC4, AC6 and AC9 are expressed in both pancreatic acini and duct fragments, whereas AC7 is expressed only in duct fragments.• In mice deficient in AC6, cAMP formation and protein kinase A activation were impaired. As a consequence, a reduction in amylase secretion and pancreatic fluid production was observed.• These results indicate that AC6 plays a regulatory role in pancreatic exocrine cells.Abstract Both secretin and vasoactive intestinal polypeptide (VIP) receptors are responsible for the activation of adenylyl cyclases (ACs), which increase intracellular cyclic AMP (cAMP) levels in the exocrine pancreas. There are nine membrane-associated isoforms, each with its own pattern of expression and regulation. In this study we sought to establish which AC isoforms play a regulatory role in pancreatic exocrine cells. Using RT-PCR, AC3, AC4, AC6, AC7 and AC9 were found to be expressed in the pancreas. AC3, AC4, AC6 and AC9 were expressed in both pancreatic acini and ducts, whereas AC7 was expressed only in pancreatic ducts. Based on known regulation by intracellular signals, selective inhibitors and stimulators were used to suggest which isoforms play an important role in the induction of cAMP formation. AC6 appeared to be an important isoform because protein kinase A (PKA), PKC and calcium all inhibited VIP-induced cAMP formation, whereas calcineurin or calmodulin did not modify the response to VIP. Mice with genetically deleted AC6 were studied and showed reduced cAMP formation and PKA activation in both isolated pancreatic acini and duct fragments. The absence of AC6 reduced cAMP-dependent secretagogue-stimulated amylase secretion, and abolished fluid secretion in both in vivo and isolated duct fragments. In conclusion, several AC isoforms are expressed in pancreatic acini and ducts. AC6 mediates a significant part of pancreatic amylase and fluid secretion in response to secretin, VIP and forskolin through cAMP/PKA pathway activation.

International Union of Basic and Clinical Pharmacology. CI. Structures and Small Molecule Modulators of Mammalian Adenylyl Cyclases

“…The distribution of AC isoforms in the digestive system was also reviewed (Sabbatini et al, 2014). Similar to the other organs/cells reported, mRNA for multiple ACs were found in hepatocytes and cholangiocytes (Strazzabosco et al, 2009), as well as in the pancreatic acini and duct (Sabbatini et al, 2013). Another paper describing the cAMP signaling elements in lung reported the AC protein/mRNA expression patterns in several tissues and cells including bronchial epithelium, vascular endothelium, airway smooth muscle cells, vascular smooth muscle cells, pulmonary fibroblasts (WI-38 cells), and inflammatory cells (Oldenburger et al, 2012).…”

“…The localization of AC6 in osteocyte cilia likely contributes to its role in loading-induced bone adaptation (Lee et al, 2014). Other functions of AC6 include the regulation of basal sympathetic nerve discharge or sympathetic tone within the brain stem (Chien et al, 2013) and regulation of cAMP-dependent pancreatic amylase and fluid secretion (Sabbatini et al, 2013). In humans, the ADCY6 S674 variant results in a hyperfunctional AC6 protein (Gros et al, 2007), but follow up studies are required.…”

Adenylyl cyclases in the digestive system

“…Soluble AC plays a role as a metabolic and intracellular pH sensor because soluble AC can be activated by ${\mathrm{HCO}}_3^{-}$, Ca 2+ and ATP [35]. Like transmembrane AC isoforms [36–39], an increase in cAMP induced by soluble AC activates both PKA [40] and Epac/Rap1 pathways [41,42]. …”

“…Recently, the expression profile of the transmembrane AC isoforms in intact mouse pancreas, isolated pancreatic acini and sealed duct fragments was establishsed [36]. Using reverse transcriptase-PCR analysis, five different transmembrane AC isoforms were able to be identified in pancreatic exocrine cells: AC3, AC4, AC6, AC9 mRNAs were expressed in isolated pancreatic acini and sealed duct fragments, whereas AC7 mRNAs was only expressed in sealed duct fragments.…”

“…Using a number of chemical stimulators and inhibitors, as well as mice deficient in AC6 [60], the study established that AC6 plays a regulatory role in the functions of pancreatic exocrine cells [36]. The effect of VIP on cAMP levels was enhanced by PKA and PKC inhibitors, as well as by a depletion of Ca 2+ , and not modified by stimulants including the Ca 2+ agonist A23187, phorbol ester, and the cholinergic agonist carbachol, indicating that Ca 2+ /PKC/PKA-inhibited AC6 is likely to mediate the action of VIP.…”