The characterization and quantification of antigen-induced Ca2+ oscillations in a rat basophilic leukaemia cell line (RBL-2H3).
Using the ratiometric Ca2+ indicator, indo-1, the antigen-induced increase in intracellular Ca2+ concentration ([Ca2+]i) was measured in individual RBL-2H3 cells which had been passively sensitized with monoclonal antibody to the dintrophenyl (DNP) haptenic group. Antigenic stimulation using DNP-human serum albumin conjugate (DNP-HSA) induced concentration-dependent asynchronous Ca2+ oscillations, or irregular spikes. To achieve a quantitative comparison of the effects of different concentrations of antigen on changes in Ca2+[i, the area under the curve (AUC) of Ca2+ oscillations in each cell was calculated. The dose-response curve of the calculated AUC is consistent with the bell-shaped dose-response curve for antigen-induced mediator release, depolarization and 86Rb(+)-efflux. Ca2+ oscillations induced by antigenic stimulation were abolished by removal of external Ca2+ and the subsequent reintroduction of external Ca2+ caused their resumption. To investigate the role of Ca2+ oscillations in the secretory response, changes in [Ca2+]i induced by concanavalin A (Con-A), A23187, thapsigargin and NECA were also monitored. Con-A mimicked the response induced by antigen, whilst A23187 and thapsigargin induced a large transient non-oscillatory response. NECA, an adenosine receptor agonist, induced only a small transient rise in Ca2+[i without oscillatory behaviour. Since all these stimuli accept NECA-induced degranulation in these cells, it is suggested that, although Ca2+ oscillations are not essential for the initiation of secretion, they probably underlie the in-vivo physiological response of mast cells and basophils to an antigenic challenge. They also seem to enhance the efficacy of the Ca2+ signal.
Narenjkar
J
J
Department of Pharmacology, University College London, UK.
Marsh
S J
SJ
Assem
E S
ES
D019830:Adenosine-5'-(N-ethylcarboxamide) / Q000494:pharmacology; D000818:Animals; D000941:Antigens / Q000378:metabolism* / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D020013:Calcium Signaling; D016764:Cell Polarity / Q000187:drug effects; D003208:Concanavalin A / Q000494:pharmacology; D004140:Dinitrophenols / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D007476:Ionophores / Q000494:pharmacology; D007938:Leukemia / Q000188:drug therapy / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects; D011868:Radioisotopes / Q000378:metabolism; D051381:Rats; D017455:Receptors, IgE / Q000187:drug effects / Q000378:metabolism; D012413:Rubidium / Q000378:metabolism; D012709:Serum Albumin / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D001619:beta-N-Acetylhexosaminidases / Q000187:drug effects / Q000378:metabolism
D000941:Antigens; D004140:Dinitrophenols; D007476:Ionophores; D011868:Radioisotopes; D017455:Receptors, IgE; D012709:Serum Albumin; C058064:dinitrophenyl-human serum albumin conjugate; D003208:Concanavalin A; D019830:Adenosine-5'-(N-ethylcarboxamide); D000001:Calcimycin; D001619:beta-N-Acetylhexosaminidases; D012413:Rubidium