title: Bradykinin-induced phosphoinositide hydrolysis and Ca2+ mobilization in canine cultured tracheal epithelial cells.
abstract: 1. Experiments were designed to differentiate the mechanisms and subtype of kinin receptors mediating the changes in intracellular Ca2+ concentration ([Ca2+]i) induced by bradykinin (BK) in canine cultured tracheal epithelial cells (TECs). 2. BK and Lys-BK caused an initial transient peak of [Ca2+]i in a concentration-dependent manner, with half-maximal stimulation (pEC50) obtained at 7.70 and 7.23, respectively. 3. Kinin B2 antagonists Hoe 140 (10 nM) and [D-Arg0, Hyp3, Thi5,8, D-Phe7]-BK (1 microM) had high affinity in antagonizing BK-induced Ca2+ response with pKB values of 8.90 and 6.99, respectively. 4. Pretreatment of TECs with pertussis toxin (100 ng ml(-1)) or cholera toxin (10 microg ml(-1)) for 24 h did not affect the BK-induced IP accumulation and [Ca2+]i changes in TECs. 5. Removal of Ca2+ by the addition of EGTA or application of Ca2+-channel blockers, verapamil, diltiazem, and Ni2+, inhibited the BK-induced IP accumulation and Ca2+ mobilization, indicating that Ca2+ influx was required for the BK-induced responses. 6. Addition of thapsigargin (TG), which is known to deplete intracellular Ca2+ stores, transiently increased [Ca2+]i in Ca2+-free buffer and subsequently induced Ca2+ influx when Ca2+ was re-added to this buffer. Pretreatment of TECs with TG completely abolished BK-induced initial transient [Ca2+]i, but had slight effect on BK-induced Ca2+ influx. 7. Pretreatment of TECs with SKF96365 and U73122 inhibited the BK-induced Ca2+ influx and Ca2+ release, consistent with the inhibition of receptor-gated Ca2+-channels and phospholipase C in TECs, respectively. 8. These results demonstrate that BK directly stimulates kinin B2 receptors and subsequently phospholipase C-mediated IP accumulation and Ca2+ mobilization via a pertussis toxin-insensitive G protein in canine TECs. These results also suggest that BK-induced Ca2+ influx into the cells is not due to depletion of these Ca2+ stores, as prior depletion of these pools by TG has no effect on the BK-induced Ca2+ influx that is dependent on extracellular Ca2+ in TECs.
authors.affiliation: Department of Internal Medicine, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan.
authors.forename: S F
authors.initials: SF
authors.lastname: Luo
authors.forename: S L
authors.initials: SL
authors.lastname: Pan
authors.forename: W B
authors.initials: WB
authors.lastname: Wu
authors.forename: C C
authors.initials: CC
authors.lastname: Wang
authors.forename: C T
authors.initials: CT
authors.lastname: Chiu
authors.forename: Y J
authors.initials: YJ
authors.lastname: Tsai
authors.forename: C M
authors.initials: CM
authors.lastname: Yang
mesh_terms: D000818:Animals; D001920:Bradykinin / Q000031:analogs & derivatives / Q000494:pharmacology*; D065168:Bradykinin Receptor Antagonists; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000494:pharmacology; D004285:Dogs; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004963:Estrenes / Q000494:pharmacology; D005260:Female; D006868:Hydrolysis / Q000187:drug effects; D007093:Imidazoles / Q000494:pharmacology; D008297:Male; D037342:Pertussis Toxin; D010716:Phosphatidylinositols / Q000378:metabolism*; D011760:Pyrrolidinones / Q000494:pharmacology; D043782:Receptor, Bradykinin B2; D019284:Thapsigargin / Q000494:pharmacology; D014132:Trachea / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
chemical_list: D065168:Bradykinin Receptor Antagonists; D002121:Calcium Channel Blockers; D004791:Enzyme Inhibitors; D004963:Estrenes; D007093:Imidazoles; D010716:Phosphatidylinositols; D011760:Pyrrolidinones; D043782:Receptor, Bradykinin B2; D010566:Virulence Factors, Bordetella; C058532:HOE k86-4321; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D019284:Thapsigargin; C065679:icatibant; D002772:Cholera Toxin; D037342:Pertussis Toxin; D010738:Type C Phospholipases; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole; D001920:Bradykinin; D002118:Calcium