HCO(3)(-)-dependent soluble adenylyl cyclase activates cystic fibrosis transmembrane conductance regulator in corneal endothelium.
cAMP-dependent activation of the cystic fibrosis transmembrane conductance regulator (CFTR) regulates fluid transport in many tissues. Secretion by the corneal endothelium is stimulated by cAMP and dependent on HCO(3)(-). We asked whether HCO(3)(-) can secondarily increase CFTR permeability in bovine corneal endothelial cells (BCEC) by activating soluble adenylyl cyclase (sAC). Immunofluorescence suggests that sAC is distributed throughout the cytoplasm. HCO(3)(-) (40 mM) increased cAMP concentration 42% in the presence of 50 microM rolipram (a phosphodiesterase 4 inhibitor), and a standard HCO(3)(-) Ringer solution (28.5 mM) increased apical Cl(-) permeability by 78% relative to HCO(3)(-)-free solution. The HCO(3)(-)-dependent increase in Cl(-) permeability was reduced 60% by 20 mM NaHSO(3) (a weak agonist of sAC). NaHSO(3) alone increased apical Cl(-) permeability by only 13%. The HCO(3)(-)-dependent increase in Cl(-) permeability was reduced 57% in the presence of 50 microM Rp-adenosine 3',5'-cyclic monophosphorothioate, and 86% by 50 microM 5-nitro-2-(3-phenylpropyl-amino)benzoic acid but unaffected by 200 microM apical H(2)DIDS. CFTR phosphorylation was increased 23, 150, and 32% by 20 mM HSO(3)(-), 28.5 mM HCO(3)(-), and 28.5 mM HCO(3)(-) + 20 mM HSO(3)(-), respectively. Activation of apical Cl(-) permeability by 5 microM genistein was increased synergistically by HCO(3)(-) over that due to genistein and HCO(3)(-) alone. We conclude that HCO(3)(-)-stimulated sAC is a form of autocrine signaling that contributes to baseline cAMP production, thereby affecting baseline CFTR activity in BCEC. This form of autocrine signaling may be important in tissues that express sAC and exhibit robust HCO(3)(-) influx (e.g., ocular ciliary epithelium, choroid plexus, and airway epithelium).
Sun
Xing Cai
XC
School of Optometry, Indiana University, Bloomington, Indiana 47405, USA.
Zhai
Chang-Bin
CB
Cui
Miao
M
Chen
Yanqiu
Y
Levin
Lonny R
LR
Buck
Jochen
J
Bonanno
Joseph A
JA
D000262:Adenylyl Cyclases / Q000502:physiology*; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism* / Q000494:pharmacology; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002712:Chlorides / Q000378:metabolism; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000502:physiology*; D004728:Endothelium, Corneal / Q000378:metabolism*; D010539:Permeability / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D012995:Solubility; D013447:Sulfites / Q000494:pharmacology
D001639:Bicarbonates; D002712:Chlorides; D013447:Sulfites; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D000242:Cyclic AMP; D000262:Adenylyl Cyclases