2 DISTINCT CA2+ INFLUX PATHWAYS ACTIVATED BY THE BRADYKININ BETA(2) RECEPTOR

The hormone-induced depletion of cellular Ca stores provides a signal for the Ca2+ influx into electrically non-excitable cells; however, th e underlying molecular mechanisms remain elusive. Therefore, we analyz ed bradykinin-activated Ca2+ influx into human foreskin fibroblast cel ls, HF-15, by fura-2 and Ca-45 labeling to discriminate between Ca2+ i nflux into the fura-sensitive compartment and Ca uptake into fura-inse nsitive Ca stores. Bradykinin-activated Ca2+ influx into the fura-sens itive compartment was blocked by inhibitors of NO synthases. These inh ibitors also suppressed bradykinin-activated increases in cGMP, indica ting that the NO-dependent increase in cGMP is involved in the activat ion of the Ca2+ influx into the fura-sensitive compartment. The cGMP-d ependent kinase inhibitors KT5823 and Rp-8-(para-chlorophenylthio)-cGM P (Rp-8-pCPT-cGMPS) blocked bradykinin-activated Ca2+ influx into the fura-sensitive compartment, suggesting that a cGMP-dependent kinase st ep participates in the activation of this Ca2+ influx pathway. In addi tion to the NO/cGMP-mediated Ca2+ influx into the fura-sensitive compa rtment, bradykinin enhanced Ca-45 uptake into Ca stores that were not accessible to fura-2. This enhanced Ca-45 uptake was insensitive to bl ockers of the NO/cGMP pathway, indicating that the Ca-45 uptake pathwa y is distinct from the NO-dependent Ca2+ influx into the fura-sensitiv e compartment. Furthermore, bradykinin enhanced Ca-45 uptake into prol iferating but not into quiescent HF-15 fibroblasts. Hence, bradykinin stimulates two distinct Ca2+ influx pathways in HF-15 cells, (a) Ca2influx into the fura-sensitive compartment which is NO/cGMP-dependent and (b) Ca uptake into Ca stores which bypasses the cytoplasm, which i s NO insensitive and which is linked to cell proliferation.