INOSITOL PHOSPHATE FORMATION AND RELEASE OF INTRACELLULAR FREE CALCIUM BY BRADYKININ IN HACAT KERATINOCYTES

Phospholipase C-mediated release of inositol trisposphate, followed by an increase in free intracellular calcium, is an important signal tra nsduction pathway for several membrane receptors. In the present inves tigation, the coupling of various receptors to phospholipase C was stu died in the human keratinocyte line HaCaT. Inositol trisphosphate form ation was determined by anion-exchange chromatography, and the release of intracellular calcium was analysed with the fluorescence probe Fur a-2 AM. Activation of HaCaT keratinocytes with bradykinin resulted in a time- and dose-dependent release of inositol trisphosphate and intra cellular calcium, with an EC50 value of 50 nM for bradykinin-induced i nositol trisphosphate formation. The mediators and cytokines IL-1, IL- 4, IL-6, IL-8, EGF and TGFalpha, as well as bombesin, prolactin, carba chol, substance P and retinoic acid, did not activate this pathway. Th e inability of the mediators examined to activate phospholipase C may be due to lack of the respective cognate receptors or to the use of ot her signal transduction pathways.