Immunological stimulation of single rat basophilic leukemia RBL-2H3 cells co-activates Ca(2+)-entry and K(+)-channels.
The relationship between type 1 Fc epsilon-receptor (Fc epsilon RI) mediated cell stimulation, Ca(2+)-signals and membrane currents was studied in rat mucosal mast cells, subline RBL-2H3 by combining patch-clamp, Fura-2, 45Ca(2+)-uptake and secretory response measurements. Cells were stimulated by Fc epsilon RI clustering either with IgE and antigen or by an IgE specific monoclonal antibody. Both stimuli induced a biphasic increase in the free intracellular Ca(2+)-concentration ([Ca2+]i). Fc epsilon RI clustering in Ca(2+)-free solution induces a transient increase in [Ca2+]i reflecting Ca2+ release from the Ins(1,4,5)P3 sensitive stores. Mn2+ applied to a nominally Ca(2+)-free solution, causes quenching of the Fura-2 emission during Fc epsilon RI clustering, indicating activation of a transmembrane pathway for the entry of extracellular calcium ions. Whole-cell current-voltage relationship of resting cells showed strong inward rectification. The inward current component at a potential of -100 mV is increased by 23 +/- 11% (n = 14) upon Fc epsilon RI clustering, whereas the outward component at +50 mV was enhanced by 45 +/- 6%. The Fc epsilon RI activated current was identified as due to K+ ions, because it reversed close to the K(+)-equilibrium potential, was blocked by Ba2+ or Cs+ containing or K(+)-free bath solutions. It was also inhibited by TEA and quinidine, while DIDS had no effect. Moreover, an inwardly rectifying K(+)-channel with a conductance of 28 pS was recorded in single channel measurements. The open probability of this channel increased by 39 +/- 16% (n = 8) upon Fc epsilon RI clustering. Superfusion of the cells with nominally K(+)-free solution also significantly inhibited both the Fc epsilon RI mediated 45Ca2+ uptake and the secretory response of the cells. We conclude that activation of K(+)-channels upon Fc epsilon RI clustering is functionally involved in the control and the maintenance of the secretory response of RBL-2H3 mast cells.
Gericke
M
M
KU Leuven, Laboratorium voor Fysiologie, Belgium.
Dar
O
O
Droogmans
G
G
Pecht
I
I
Nilius
B
B
D000818:Animals; D000941:Antigens; D017609:Barium Compounds / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002586:Cesium / Q000494:pharmacology; D002712:Chlorides / Q000494:pharmacology; D004140:Dinitrophenols / Q000276:immunology; D017136:Ion Transport; D015471:Leukemia, Basophilic, Acute / Q000276:immunology / Q000378:metabolism; D008564:Membrane Potentials; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000378:metabolism*; D051381:Rats; D017455:Receptors, IgE / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000276:immunology; D015398:Signal Transduction; D019789:Tetraethylammonium; D013757:Tetraethylammonium Compounds / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000941:Antigens; D017609:Barium Compounds; D002712:Chlorides; D004140:Dinitrophenols; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D017455:Receptors, IgE; D013757:Tetraethylammonium Compounds; C034638:dinitrophenyl-bovine serum albumin; C024986:barium chloride; D002586:Cesium; D012710:Serum Albumin, Bovine; D019789:Tetraethylammonium; C028019:cesium chloride; D002118:Calcium