Redox modulation of intracellular free calcium concentration in thyroid FRTL-5 cells: evidence for an enhanced extrusion of calcium.
Redox modulation is involved in the regulation of the intracellular free calcium concentration ([Ca2+]i) in several cell types. In thyroid cells, including thyroid FRTL-5 cells, changes in [Ca2+]i regulate important functions. In the present study we investigated the effects of the oxidizing compounds thimerosal and t-butyl hydroperoxide on [Ca2+]i in thyroid FRTL-5 cells. Thimerosal mobilized sequestered calcium, and evoked modest store-dependent calcium entry. Both compounds potently attenuated the increase in [Ca2+]i when store-operated calcium entry was evoked with thapsigargin. The entry of barium was not attenuated. Experiments performed with high extracellular pH, in sodium-free buffer and in the presence of vanadate suggested that thimerosal decreased [Ca2+]i by activating a calcium extrusion mechanism, probably a plasma membrane Ca2+-ATPase. All the observed effects were abrogated by the reducing agent beta-mercaptoethanol. The mechanism of action was apparently mediated via activation of protein kinase C, as thimerosal potently stimulated binding of [3H]phorbol 12, 13-dibutyrate, and was without effect on store-operated calcium entry in cells treated with staurosporine or in cells with down-regulated protein kinase C. Thimerosal did not depolarize the membrane potential, as evaluated using patch-clamp in the whole-cell mode. In immunoprecipitates obtained with an antibody against plasma membrane Ca2+-ATPase, we observed several phosphorylated bands in cells stimulated with thimerosal. In conclusion, we have shown that thimerosal attenuates an increase in [Ca2+]i, probably by activating a plasma membrane Ca2+-ATPase.
Törnquist
K
K
Department of Biology, Abo Akademi University, BioCity, Artillerigatan 6, 20520 Turku, Finland. kid.tornqvist@abo.fi
Vainio
P
P
Titievsky
A
A
Dugué
B
B
Tuominen
R
R
D000818:Animals; D001464:Barium / Q000378:metabolism / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007093:Imidazoles / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D008623:Mercaptoethanol / Q000494:pharmacology; D010084:Oxidation-Reduction / Q000187:drug effects; D015240:Phorbol 12,13-Dibutyrate / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D019311:Staurosporine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D013849:Thimerosal / Q000494:pharmacology; D013961:Thyroid Gland / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D014638:Vanadates / Q000494:pharmacology; D020122:tert-Butylhydroperoxide / Q000494:pharmacology
D007093:Imidazoles; D013849:Thimerosal; D001464:Barium; D015240:Phorbol 12,13-Dibutyrate; D014638:Vanadates; D008623:Mercaptoethanol; D019284:Thapsigargin; D020122:tert-Butylhydroperoxide; D011493:Protein Kinase C; D000252:Calcium-Transporting ATPases; D019311:Staurosporine; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium