EGF raises cytosolic Ca2+ in A431 and Swiss 3T3 cells by a dual mechanism. Redistribution from intracellular stores and stimulated influx.
The changes in Ca2+ homeostasis and phosphoinositide hydrolysis induced by EGF were studied in human epidermoid carcinoma A431 cells both when attached to a substratum and after detachment and suspension. The cytosolic Ca2+ concentration was measured by the conventional fluorimetric technique, using the specific probe, quin2, as well as by a new microscopic technique in which single cells are investigated after loading with another probe, fura-2. EGF applied in the complete, Ca2+-containing medium caused a rapid rise in the cytosolic Ca2+ concentration, that remained elevated for several minutes. In Ca2+-free, EGTA-containing medium, part of this response persisted, as revealed by quin2 results in suspended cells and microscopic results with fura-2. The lack of Ca2+ rise seen in attached cells loaded with quin2 and treated with EGF in Ca2+-free medium was probably the result of a Ca2+ buffer artifact. Concomitantly to the Ca2+ signal, EGF induced phosphoinositide hydrolysis, with stimulated accumulation of inositol 1,3,4,trisphosphate and -1,3,4,5-tetrakisphosphate. These results, as well as additional microscopic fura-2 results in Swiss 3T3 fibroblasts, demonstrate that the Ca2+ signal elicited by EGF is due to two components: redistribution from an intracellular store (possibly mediated by generation of inositol trisphosphate) and stimulated influx across the plasmalemma. This latter process was not detected in 3T3 cells treated with either PDGF or bombesin (growth factors that cause much greater phosphoinositide hydrolysis and Ca2+ redistribution responses than EGF). It is therefore suggested that the Ca2+ influx effect of EGF is under the control of a separate, as yet unidentified mechanism.
D000634:Aminoquinolines; D000818:Animals; D001572:Benzofurans; D001693:Biological Transport, Active / Q000187:drug effects; D001839:Bombesin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005456:Fluorescent Dyes; D016257:Fura-2; D006706:Homeostasis; D006801:Humans; D007700:Kinetics; D051379:Mice; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011994:Recombinant Proteins / Q000494:pharmacology
D000634:Aminoquinolines; D001572:Benzofurans; D005456:Fluorescent Dyes; D010982:Platelet-Derived Growth Factor; D011994:Recombinant Proteins; D004815:Epidermal Growth Factor; C035739:Quin2; D001839:Bombesin; D002118:Calcium; D016257:Fura-2