UTP inhibits Na+ absorption in wild-type and DeltaF508 CFTR-expressing human bronchial epithelia.
Ca2+-mediated agonists, including UTP, are being developed for therapeutic use in cystic fibrosis (CF) based on their ability to modulate alternative Cl- conductances. As CF is also characterized by hyperabsorption of Na+, we determined the effect of mucosal UTP on transepithelial Na+ transport in primary cultures of human bronchial epithelia (HBE). In symmetrical NaCl, UTP induced an initial increase in short-circuit current (Isc) followed by a sustained inhibition. To differentiate between effects on Na+ absorption and Cl- secretion, Isc was measured in the absence of mucosal and serosal Cl- (INa). Again, mucosal UTP induced an initial increase and then a sustained decrease that reduced amiloride-sensitive INa by 73%. The Ca2+-dependent agonists histamine, bradykinin, serosal UTP, and thapsigargin similarly induced sustained inhibition (62-84%) of INa. Mucosal UTP induced similar sustained inhibition (half-maximal inhibitory concentration 296 nM) of INa in primary cultures of human CF airway homozygous for the DeltaF508 mutation. BAPTA-AM blunted UTP-dependent inhibition of INa, but inhibitors of protein kinase C (PKC) and phospholipase A2 had no effect. Indeed, direct activation of PKC by phorbol 12-myristate 13-acetate failed to inhibit Na+ absorption. Apyrase, a tri- and diphosphatase, did not reverse inhibitory effects of UTP on INa, suggesting a long-term inhibitory effect of UTP that is independent of receptor occupancy. After establishment of a mucosa-to-serosa K+ concentration gradient and permeabilization of the mucosal membrane with nystatin, mucosal UTP induced an initial increase in K+ current followed by a sustained inhibition. We conclude that increasing cellular Ca2+ induces a long-term inhibition of transepithelial Na+ transport across normal and CF HBE at least partly due to downregulation of a basolateral membrane K+ conductance. Thus UTP may have a dual therapeutic effect in CF airway: 1) stimulation of a Cl- secretory response and 2) inhibition of Na+ transport.
Devor
D C
DC
Department of Physiology and Cell Biology, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA. dd2+@pitt.edu
Pilewski
J M
JM
D000584:Amiloride / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects; D001980:Bronchi / Q000503:physiopathology*; D002034:Bumetanide; D002118:Calcium / Q000502:physiology; D002478:Cells, Cultured; D002712:Chlorides / Q000378:metabolism; D003550:Cystic Fibrosis / Q000235:genetics / Q000503:physiopathology*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics* / Q000502:physiology; D006632:Histamine / Q000494:pharmacology; D006720:Homozygote; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009092:Mucous Membrane / Q000187:drug effects / Q000503:physiopathology; D017384:Sequence Deletion; D012964:Sodium / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014544:Uridine Triphosphate / Q000494:pharmacology*
C505032:CFTR protein, human; D002712:Chlorides; D002034:Bumetanide; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000584:Amiloride; D006632:Histamine; D012964:Sodium; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D014544:Uridine Triphosphate