The energy providing substrate ATP can be released from various cells and act extracellularly to regulate the same cells or neighboring cells. However, the pathway for ATP release and the eliciting physiological stimulus are unclear. Recently, we showed that ATP activates P2X and P2Y purinergic receptors on pancreatic ducts. Thus, it was relevant to ask whether the upstream acini could be the source of releasable ATP and what the stimulus might be. We used freshly prepared rat pancreatic acini and applied conventional luminescence measurements of luciferin/luciferase reaction. As a new application of this reaction in confocal microscopy, we monitored luciferin fluorescence as a sign of ATP release by single acini. In addition we used quinacrine to mark ATP stores, which were similar to those marked with fluorescent ATP, 2′-(or-3′)-O-(N-methylanthraniloyl) adenosine 5′-triphosphate, but only partially overlapping with those marked by acridine orange and LysoTracker Red. In functional studies we show that native pancreatic acini release ATP in response to various stimuli but most importantly to cholinergic stimulation, a very likely physiological stimulus in this epithelium. In a close vicinity of acini we detect about 9 μm ATP after cholinergic stimulation. Thus, ATP is poised as the paracrine mediator between pancreatic acini and ducts.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers and new therapeutic targets are urgently needed. One of the hallmarks of cancer is changed pH-homeostasis and potentially pH-sensors may play an important role in cancer cell behavior. Two-pore potassium channels (K2P) are pH-regulated channels that conduct a background K(+) current, which is involved in setting the plasma membrane potential (Vm). Some members of the K2P superfamily were reported as crucial players in driving tumor progression. The aim of this study was to investigate pH-regulated K(+) currents in PDAC cells and determine possible effects on their pathological phenotype. Using a planar high-throughput patch-clamp system (SyncroPatch 384PE) we identified a pH-regulated K(+) current in the PDAC cell line BxPC-3. The current was inhibited by extracellular acidification and intracellular alkalization. Exposure to a set of different K(+) channel inhibitors, and the TREK-1 (K2P2.1)-specific activator BL1249, TREK-1 was identified as the main component of pH-regulated current. A voltage-sensor dye (VF2.1.Cl) was used to monitor effects of pH and BL1249 on Vm in more physiological conditions and TREK-1-mediated current was found as critical player in setting Vm. We assessed a possible role of TREK-1 in PDAC progression using cell proliferation and migration assays and observed similar trends with attenuated proliferation/migration rates in acidic (pH7.4) conditions. Notably, BL1249 inhibited both PDAC cell proliferation and migration indicating that hyperpolarization of Vm attenuates cancer cell behavior. TREK-1 may therefore be a promising novel target for PDAC therapy.

The mechanism by which pancreas secretes high HCO₃^- has not been fully resolved. This alkaline secretion, formed in pancreatic ducts, can be achieved by transporting HCO₃^- from serosa to mucosa or by moving H⁺ in the opposite direction. The aim of the present study was to determine whether H⁺/K⁺-ATPases are expressed and functional in human pancreatic ducts and whether proton pump inhibitors (PPIs) have effect on those. Here we show that the gastric HKα1 and HKβ subunits (ATP4A; ATP4B) and non-gastric HKα2 subunits (ATP12A) of H⁺/K⁺-ATPases are expressed in human pancreatic cells. Pumps have similar localizations in duct cell monolayers (Capan-1) and human pancreas, and notably the gastric pumps are localized on the luminal membranes. In Capan-1 cells, PPIs inhibited recovery of intracellular pH from acidosis. Furthermore, in rats treated with PPIs, pancreatic secretion was inhibited but concentrations of major ions in secretion follow similar excretory curves in control and PPI treated animals. In addition to HCO₃^-, pancreas also secretes K⁺. In conclusion, this study calls for a revision of the basic model for HCO₃^- secretion. We propose that proton transport is driving secretion, and that in addition it may provide a protective pH buffer zone and K⁺ recirculation. Furthermore, it seems relevant to re-evaluate whether PPIs should be used in treatment therapies where pancreatic functions are already compromised.

Introduction: Peripheral correlates of age-associated cognitive decline are important tools in the screening for potentially abnormal courses of cognitive aging. Since salivary gland function is controlled by the autonomic and central nervous system, associations between cognitive changes and salivary gland hypofunction were tested in two groups of middle-aged men in late midlife, who differed substantially with respect to their midlife performance in verbal intelligence when compared with their performance in young adulthood. Materials and Methods: Participants (n = 193) were recruited from the Danish Metropolit Cohort of men born in 1953. Based on their individual change in performance in two previously administered intelligence tests, they were allocated to one group of positive and one group of negative outliers in midlife cognition scores, indicating no decline versus decline in test performance. All participants underwent a clinical oral examination including assessments of their dental, periodontal, and mucosal conditions. Whole and parotid saliva flow rates were measured, and the number of systemic diseases and medication intake as well as daytime and nocturnal xerostomia were registered. Results: Participants with decline in cognitive test performance in midlife had significantly lower unstimulated whole saliva flow rates, higher prevalence of hyposalivation and daytime xerostomia and a higher caries experience than participants with no decline in midlife performance. Daytime and nocturnal xerostomia were associated with daily intake of medication and alcohol. Discussion: Overall, hyposalivation, xerostomia and poor dental status distinguished a group of men displaying relative decline in cognitive performance from a group of men without evidence of cognitive decline. Thus, hyposalivation and poor dental health status may represent potential correlates of age-related cognitive decline in late midlife, provided that other causes can be excluded.

Pancreatic duct cells are equipped with acid/base transporters important for exocrine secretion. Pancreatic ductal adenocarcinoma (PDAC) cells may utilize such transporters to acidify extracellular tumor microenvironment, creating a niche favoring cell proliferation, fibrosis and resistance to chemotherapy—all contributing to the notoriously bad prognosis of this disease. Here, we report that gastric and non-gastric H⁺, K⁺-ATPases (coded by ATP4A and ATP12A) are overexpressed in human and murine pancreatic cancer and that we can target them specifically with proton pump inhibitors (PPIs) and potassium-competitive acid blockers (P-CABs) in in vitro models of PDAC. Focusing on pantoprazole, we show that it significantly reduced human cancer cell proliferation by inhibiting cellular H⁺ extrusion, increasing K⁺ conductance and promoting cyclin D1-dependent cell cycle arrest and preventing STAT3 activation. Pantoprazole also decreased collagen secretion from pancreatic stellate cells. Importantly, in vivo studies show that pantoprazole treatment of tumor-bearing mice reduced tumor size, fibrosis and expression of angiogenic markers. This work provides the first evidence that H⁺, K⁺-ATPases contribute to PDAC progression and that these can be targeted by inhibitors of these pumps, thus proving a promising therapeutic strategy.