Mechanism of bradykinin-induced Ca(2+) mobilization in MG63 human osteosarcoma cells.

BACKGROUND: The effect of bradykinin on intracellular free Ca(2+) levels (Ca(2+)](i)) in MG63 human osteosarcoma cells was explored using fura-2 as a Ca(2+) dye. METHODS/RESULTS: Bradykinin (0.1 nM-1 microM) increased Ca(2+)](i) in a concentration-dependent manner with an EC(50) value of 0.5 nM. The Ca(2+)](i) signal comprised an initial peak and a fast decay which returned to baseline in 2 min. Extracellular Ca(2+) removal inhibited the peak Ca(2+)](i )signals by 35 +/- 3%. Bradykinin (1 nM) failed to increase Ca(2+)](i) in the absence of extracellular Ca(2+ )after cells were pretreated with thapsigargin (an endoplasmic reticulum Ca(2+) pump inhibitor; 1 microM). Bradykinin (1 nM)-induced intracellular Ca(2+) release was nearly abolished by inhibiting phospholipase C with 2 microM 1-(6-((17 beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione (U73122). The Ca(2+)](i )increase induced by 1 nM bradykinin in Ca(2+)- free medium was abolished by 1 nM HOE 140 (a B2 bradykinin receptor antagonist) but was not altered by 100 nM Des-Arg-HOE 140 (a B1 bradykinin receptor antagonist). Pretreatment with 1 pM pertussis toxin for 5 h in Ca(2+) medium inhibited 30 +/- 3% of 1 nM bradykinin-induced peak Ca(2+)](i) increase. CONCLUSIONS: Together, this study shows that bradykinin induced Ca(2+)](i) increases in a concentration-dependent manner, by stimulating B2 bradykinin receptors leading to mobilization of Ca(2+) from the thapsigargin-sensitive stores in a manner dependent on inositol-1,4,5-trisphosphate, and also by inducing extracellular Ca(2+) influx. The bradykinin response was partly coupled to a pertussis toxin-sensitive G protein pathway.