Intracellular Ca2+ signals induced by ATP and thapsigargin in glioma C6 cells. Calcium pools sensitive to inositol 1,4,5-trisphosphate and thapsigargin.
In glioma C6 cells, extracellular ATP generates inositol 1,4,5-trisphosphate (InsP3), indicating the presence of purinergic receptors coupled to phosphoinositide turnover. To identify the effect of ATP (acting via InsP3) and thapsigargin (acting without InsP3 production as a specific inhibitor of the endoplasmic reticulum Ca(2+)-ATPase) on intracellular Ca2+ pools we used video imaging of Fura-2 loaded into single, intact glioma C6 cells. It has been shown that ATP and thapsigargin initiate Ca2+ response consistent with the capacitative model of Ca2+ influx. When the cells were stimulated by increasing concentrations of ATP (1, 10, 50 and 100 microM) the graded, quantal Ca2+ response was observed. In the absence of extracellular Ca2+ thapsigargin and ionomycin-releasable Ca2+ pools are overlapping, demonstrating that Ca2+ stores are located mainly in the endoplasmic reticulum. After maximal Ca2+ mobilization by ATP, thapsigargin causes further increase in cytosolic Ca2+ concentration, whereas emptying of thapsigargin-sensitive intracellular stores prevents any further Ca2+ release by ATP. Thus, the thapsigargin-sensitive intracellular pool of Ca2+ in glioma C6 cells seems to be larger than that sensitive to InsP3. Two hypothesis to explain this result are proposed. One postulates a presence of two different Ca2+ pools, sensitive and insensitive to InsP3 and both discharged by thapsigargin, and the other, the same intracellular pool of Ca2+ completely emptying by thapsigargin and only partially by InsP3. These results may contribute to understanding the mechanism of Ca2+ signalling mediated by ATP, the most potent intracellular Ca2+ mobilizing agonist in all types of glial cells.
Sabała
P
P
Department of Cellular Biochemistry, Nencki Institute of Experimental Biology, Warsaw, Poland.
Amler
E
E
Barańska
J
J
D000255:Adenosine Triphosphate / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D004353:Drug Evaluation, Preclinical; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005910:Glioma / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects*; D019284:Thapsigargin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; D002118:Calcium