Intracellular Ca2+ increase in Neuro-2A cells and rat astrocytes followingstimulation of bradykinin B-2 receptor

Murine neuroblastoma cell line Neuro-2A cells and rat brain astrocytes show ed a dose-dependent increase in intracellular Ca2+ in response to bradykini n, when assessed by a single cell image analyzing system. The Ca2+ increase in Neuro-2A cells by bradykinin was also examined by a suspension fluoresc ent assay using fura-2 loading. The Ca2+ increase in both cases was suppres sed by a bradykinin B-2 receptor antagonist, Hoe 140, but not by a B-1 rece ptor antagonist, des-Arg-Hoe 140, suggesting that the effect occurred via s pecific B-2 receptor activation. RT-PCR for bradykinin B-2 receptor mRNA sh owed that both Neuro-2A cells and the astrocytes expressed B-2 receptor mRN A. Binding of [H-3]bradykinin to Neuro-2A cells was assessed, and a specifi c binding constant of 0.75 nM was determined. Furthermore, the increase in [Ca2+](i) by bradykinin could be caused by a release of Ca2+ from storage s ites in the endoplasmic reticulum, since thapsigargin and U-73122 attenuate d the effect of bradykinin in Neuro-2A as well as in astrocytes. These resu lts indicate that both astrocytes and neuroblastoma Neuro-2A cells stimulat ed by bradykinin could express a bradykinin B-2 receptor-mediated intracell ular Ca2+ increase leading to signal transduction.