Protein kinase C activator PMA reduces the Ca(2+) response to antigen stimulation of adherent RBL-2H3 mucosal mast cells by inhibiting depletion of intracellular Ca(2+) stores.
Activation of protein kinase C has been shown to reduce the Ca(2+) responses of a variety of cell types. In most cases, the reduction is due to inhibition of Ca(2+) influx, but acceleration of Ca(2+) efflux and inhibition of Ca(2+) store depletion by protein kinase C activation have also been described. For adherent RBL-2H3 mucosal mast cells, results from whole-cell patch clamp experiments suggest that protein kinase C activation reduces Ca(2+) influx, while experiments with intact, fura-2-loaded cells suggest that Ca(2+) influx is not affected. Here we present single-cell data from Ca(2+) imaging experiments with adherent RBL-2H3 cells, showing that antigen-stimulated Ca(2+) responses of phorbol 12-myristate 13-acetate (PMA)-treated cells are more transient than those of control cells. PMA also reduced the response to antigen in the absence of extracellular Ca(2+), indicating that depletion of intracellular Ca(2+) stores is inhibited. If PMA was added after stores had been depleted by thapsigargin, a small decrease in [Ca(2+)](i) was observed, consistent with a slight inhibition of Ca(2+) influx. However, the major effect of PMA on the antigen-stimulated Ca(2+) response is to inhibit depletion of intracellular Ca(2+) stores. We also show that inhibition of protein kinase C did not enhance the Ca(2+) response to antigen, suggesting that inhibition of the Ca(2+) response by activation of protein kinase C does not contribute to the physiological response to antigen.
Kuchtey
J
J
Department of Molecular Medicine, Cornell University, Ithaca, New York 14853, USA.
Fewtrell
C
C
D000941:Antigens / Q000276:immunology*; D002118:Calcium / Q000378:metabolism*; D002448:Cell Adhesion / Q000187:drug effects / Q000276:immunology; D002460:Cell Line; D004789:Enzyme Activation; D005456:Fluorescent Dyes; D016257:Fura-2; D016015:Logistic Models; D008407:Mast Cells / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D018408:Patch-Clamp Techniques; D011493:Protein Kinase C / Q000187:drug effects*; D013268:Stimulation, Chemical; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D019284:Thapsigargin / Q000494:pharmacology
D000941:Antigens; D005456:Fluorescent Dyes; D019284:Thapsigargin; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D016257:Fura-2