Intracellular calcium signals in response to bradykinin in individual neuroblastoma cells

Abstract

The Ca indicator fura 2 was used to study the modulation of cytoplasmic Ca by bradykinin (Bk) in single N1E-115 murine neuroblastoma cells. Increases in cytoplasmic Ca in response to Bk were mediated by the B2 receptor subtype. Responses to high concentrations of Bk (1-100 nM) were homogeneous and characterized by a rapidly rising transient that decayed to baseline in the continued presence of agonist, with a half-time of 15 s. Responses to low concentrations of Bk (100-500 pM) were more heterogeneous, with longer latencies and often with oscillations. Pretreatment with thapsigargin for 20 min prevented the Ca response, showing that the Ca change results from intracellular Ca release. Removal of external Ca had little effect on the response to Bk, indicating that the agonist does not activate Ca influx. The extent of Ca release and refilling after Bk was tested with ionomycin. A saturating dose of Bk (20 nM) mobilizes > 90% of stored Ca within 30 s, and this is replaced slowly. Replacement of external Na by N-methyl-D-glucamine to block Na/Ca exchange affected the Ca response, causing decreases in latency and in the period of Ca oscillations and increases in overall duration and peak amplitude of the response.