Bradykinin induces formation of inositol phosphates and causes an increase in cytoplasmic Ca 2+ in the osteoblastic cell line MC3T3‐E1

Recordings of fura‐2 fluorescence from single osteoblastic MC3T3‐E1 cells showed that bradykinin (BK, 1 μM) induced a rapid increase in cytoplasmic free Ca 2+ (Ca i 2+ , from 114 + 13 to 239 + 17 nM, mean + SEM). Following this initial transient (<1 minute) increase there was a second slow increase in Ca i 2+ (from 117 + 11 to 151 + 12 nM). Incubation in buffer with no Ca 2+ did not affect the first rapid BK‐induced increase in Ca i 2+ but eliminated the second slow increase. Addition of indomethacin or hydrocortisone to the incubation buffer did not inhibit the effect of BK on Ca i 2+ . BK caused a dose‐dependent initial rapid increase in Ca i 2+ with threshold at 1 nM and a maximal effect (241 + 30% of basal Ca i 2+ concentration) at 0.1 μM. The B1 BK receptor agonist des‐Arg 9 ‐BK (1 μM) caused only a small increase in Ca i 2+ in MC3T3‐E1 cells (from 101 + 20 to 140 + 4 nM). BK dose and time dependently stimulated the formation of inositol phosphates in MC3T3‐E1 cells with EC 50 at 2.4 nM and a significant increase in inositol trisphosphate already seen after 15 s. The Ca 2+ ionophore ionomycin induced a rapid increase in Ca i 2+ and prostaglandin E 2 (PGE 2 ) formation in MC3T3‐E1 cells. Forskolin (10–30 μM) increased cyclic AMP accumulation but did not affect Ca i 2+ or PGE 2 formation. Depletion of extracellular Ca 2+ significantly reduced (but did not abolish) BK‐induced PGE 2 formation. The initial action of BK on Ca i 2+ is probably due to an inositol‐(1,4,5)‐trisphosphate‐mediated rapid release of Ca 2+ from intracellular stores in osteoblasts and is followed by an influx of extracellular Ca 2+ . The effect is due to B2 BK receptor occupancy and is not secondary to the prostaglandin synthesis. The BK‐induced breakdown of phosphatidylinositol‐(4,5)‐bisphosphate with a subsequent increase in Ca i 2+ may be involved in BK‐induced prostaglandin formation in osteoblasts.