Tyrosine phosphorylation is involved in Ca2+ entry in human gingival fibroblasts

Abstract Bradykinin (1 μM) and histamine (100 μM) evoked an initial transient increase and a subsequent sustained increase in intracellular Ca 2+ concentration ([Ca 2+ ] i ) in fura-2-loaded human gingival fibroblasts, which may be attributed to Ca 2+ release from intracellular stores and Ca 2+ entry from extracellular sites, respectively. In fibroblasts pretreated with tyrosine kinase inhibitors such as herbimycin A (1 μM) and tyrphostin 47 (20 μM) , the sustained level of [Ca 2+ ] i induced by bradykinin and histamine increased, but not the initial peak level. In the absence of external Ca 2+ , bradykinin and histamine induced only the transient increase in [Ca 2+ ] i , but a subsequent addition of Ca 2+ to the medium resulted in a sustained increase in [Ca 2+ ] i caused by Ca 2+ entry. Thapsigargin, an inhibitor of Ca 2+ -ATPase in inositol 1,4,5-trisphosphate-sensitive Ca 2+ stores, mimicked the effect of bradykinin and histamine. In the fibroblasts pretreated with tyrosine kinase inhibitors, the bradykinin-, histamine- and thapsigargin-induced Ca 2+ entry was clearly enhanced, but not the transient [Ca 2+ ] i increase. Tyrosine phosphatase inhibitor benzylphosphonic acid (200 μM) had no effect on Ca 2+ entry or transient [Ca 2+ ] i increase. These results suggest that tyrosine phosphorylation is involved in Ca 2+ entry in human gingival fibroblasts.