Jing Wang:
Mechanisms underlying ATP regulated ion transport in human pancreatic duct cells

Date: 30-03-2012    Supervisor: Ivana Novak

Adenosine 5’-triphosphate (ATP) is a ubiquitous and fundamentally important biological molecule that acts both as an essential intracellular energy source and as an extracellular signaling molecule for purinergic receptors. ATP takes part in a wide spectrum of cellular processes in the human body. With regard to pancreas, acinar cells of exocrine pancreas secrete digestive enzymes and ATP-rich fluid into the lumen of pancreatic duct, where various purinergic P1 and P2 receptors have been identified, which can then be stimulated by ATP or its break-down product adenosine. The aim of the project is to identify the physiological roles of ATP in human pancreatic duct epithelia formed by pancreatic duct cell lines.

The pancreatic duct epithelium, generated by CAPAN-1 cells grown on permeable membranes, shows electrical parameters and polarized morphology, as well as functional receptors for secretin, vasoactive intestinal peptide (VIP), acetylcholine, adenosine, and purinergic P2 receptors. Thus, it is comparable to rat, mouse, guinea-pig and dog pancreatic ducts. Furthermore, the agonists stimulate net anion transport as detected by transepithelial electrophysiological measurements in open-circuit Ussing chambers. In particular, luminal nucleotides were the most potent activators of secretory responses. Therefore, purinergic receptors are probably significant players in regulating secretion in human pancreatic ducts.

The effect of ATP in CAPAN-1 monolayers was further analyzed in conjunction with Cl- and K+ channel modulators. Apically applied ATP opens both cystic fibrosis transmembrane conductance regulator (CFTR) and Ca2+ activated Cl- channels (CaCC), while basolateral ATP regulates secretion predominantly through CFTR. ATP also regulates K+ (most likely the intermediate-conductance K+ channel, IK) channels, which provide the driving force for Cl- secretion. Expression study confirms CFTR and IK in CAPAN-1 cells, as well as TMEM16A (the candidate for CaCC). Luminal P2 receptors activate Cl- and K+ channels leading to electrogenic secretory events, which will have to be followed by other transporters and exchangers on both plasma membranes to eventuate in HCO3--rich secretion. This scenario underpins the physiological function of luminal ATP that is secreted by acini.

Additionally, human pancreatic duct cells express both gastric and non-gastric H+/K+ ATPases on both luminal and lateral membranes, and ATP will thus function as an intracellular energy source for the pumps. Moreover, intracellular pH of CAPAN-1 cells is regulated by the H+/K+ ATPases, indicating their role in pancreatic acid-base homeostasis. Laterally or basolaterally expressed H+/K+ ATPases could fulfill the model for pancreatic HCO3- secretion by pumping H+ to the basolateral side. Function of the luminal expressed H+/K+ ATPases is unclear, but we postulate that they may The human pancreatic duct epithelium formed by CAPAN-1 cells resembles the function of native ducts, and has ATP regulated Cl- and K+ conductance as well as H+/K+ ATPases. The ATP regulated and IK channel potentiated Cl- channels could possibly be therapeutic targets for some pancreatic diseases including cystic fibrosis.