Mechanisms of bradykinin-mediated Ca2+ signalling in canine cultured corneal epithelial cells

Samuel C.M. Huang, Chin Sung Chien, Li Der Hsiao, Chuan Chwan Wang, Chi Tso Chiu, Kao Yi Liang, Chuen Mao Yang*

*Corresponding author for this work

Research output: Contribution to journalJournal Article peer-review

6 Scopus citations

Abstract

Experiments were designed to differentiate the mechanisms of bradykinin receptors mediating the changes in intracellular Ca2+ concentration ([Ca2+]i) in canine cultured corneal epithelial cells (CECs). Bradykinin and Lys-bradykinin caused an initial transient peak of [Ca2+]i in a concentration-dependent manner, with half-maximal stimulation (pEC50) obtained at 6.9 and 7.1, respectively. Pretreatment of CECs with pertussis toxin (PTX) or cholera toxin (CTX) for 24 h did not affect the bradykinin-induced [Ca2+]i changes. Application of Ca2+ channel blockers, diltiazem and Ni2+, inhibited the bradykinin-induced Ca2+ mobilization, indicating that Ca2+ influx was required for the bradykinin-induced responses. 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 readded to this buffer. Pretreatment of CECs with TG completely abolished bradykinin-induced initial transient [Ca2+]i, but had slight effect on bradykinin-induced Ca2+ influx. Pretreatment of CECs with 1-[β-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole (SKF96365) and 1-(6-((17β-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H- pyrrole-2,5-dione (U73122) inhibited the bradykinin-induced Ca2+ release and Ca2+ influx, consistent with the inhibition of receptor-gated Ca2+ channels and phospholipase C (PLC) in CECs, respectively. These results demonstrate that bradykinin directly stimulates B2 receptors and subsequently Ca2+ mobilization via a PTX-insensitive G protein in canine CECs. These results suggest that bradykinin-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 bradykinin-induced Ca2+ influx that is dependent on extracellular Ca2+ in CECs.

Original languageEnglish
Pages (from-to)565-574
Number of pages10
JournalCellular Signalling
Volume13
Issue number8
DOIs
StatePublished - 2001

Keywords

  • Bradykinin receptor
  • Ca
  • Corneal epithelial cells
  • Thapsigargin

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