Disturbances in purinergic [Ca 2+] i signaling pathways in a transformed rat thyroid cell line

Abstract

It was previously shown that in rat thyroid PC-Cl3 cell line, a purinergic P2Y receptor increases the concentration of free cytosolic Ca 2+ ([Ca 2+] i) via phospholipase C activation. We here studied whether in a transformed cell line (PC-E1Araf) derived from parental PC-Cl3 cells, ATP is still able to transduce the [Ca 2+] i-based intracellular signal. We demonstrate the expression of mRNA for P2Y2 in both PC-Cl3 and PC-E1Araf cells; mRNAs for P2Y1, P2Y4, P2Y6 and P2Y11 were absent. In both cell lines activation of P2Y2 receptor provokes a transient increase in [Ca 2+] i followed by a lower sustained phase persisting for over 5 min in PC-Cl3 and only 1.5 min in PC-E1Araf cells. In both cell lines the [Ca 2+] i reached a plateau level significantly higher than the basal [Ca 2+] i level persisting for over 10 min. Removal of extracellular Ca 2+ reduced the initial transient response to ATP in PC-Cl3, but not in PC-E1Araf cells, and completely abolished the plateau phase in both cell lines. In the presence of extracellular Ca 2+ thapsigargin (TG) caused a rise in [Ca 2+] i significantly higher in PC-Cl3 than transformed PC-E1Araf cells, while in Ca 2+-free medium the effect of TG was similar in both cell lines. The capacitative Ca 2+-entry in PC-Cl3 resulted significantly higher than in PC-E1Araf cells. Further studies were performed in order to investigate whether the different effects of ATP on [Ca 2+] i was due to variation in divalent cation plasma membrane permeability. PC-E1Araf cells showed a much lower permeability to Ca 2+, Ba 2+, Sr 2+, Mn 2+, and Co 2+ that may be responsible for the differences in purinergic Ca 2+ signaling pathway with respect to parental PC-Cl3 cells.