Disturbances in purinergic [Ca2+]i signaling pathways in a transformed rat thyroid cell line

It was previously shown that in rat thyroid PC-C13 cell line, a purinergic P2Y receptor increases the concentration of free cytosolic Ca²⁺ ([Ca²⁺]_i) via phospholipase C activation. We here studied whether in a transformed cell line (PC-E1Araf) derived from parental PC-C13 cells, ATP is still able to transduce the [Ca²⁺]_i-based intracellular signal. We demonstrate the expression of mRNA for P2Y2 in both PC-C13 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 [Ca²⁺]_i followed by a lower sustained phase persisting for over 5 min in PC-C13 and only 1.5 min in PC-E1Araf cells. In both cell lines the [Ca²⁺]_i reached a plateau level significantly higher than the basal [Ca²⁺]_i level persisting for over 10 min. Removal of extracellular Ca²⁺ reduced the initial transient response to ATP in PC-C13, but not in PC-E1Araf cells, and completely abolished the plateau phase in both cell lines. In the presence of extracellular Ca²⁺ thapsigargin (TG) caused a rise in [Ca²⁺]_i significantly higher in PC-C13 than transformed PC-E1Araf cells, while in Ca²⁺-free medium the effect of TG was similar in both cell lines. The capacitative Ca²⁺-entry in PC-C13 resulted significantly higher than in PC-E1Araf cells. Further studies were performed in order to investigate whether the different effects of ATP on [Ca²⁺]_i was due to variation in divalent cation plasma membrane permeability. PC-E1Araf cells showed a much lower permeability to Ca²⁺, Ba²⁺, Sr²⁺, Mn²⁺, and Co²⁺ that may be responsible for the differences in purinergic Ca²⁺ signaling pathway with respect to parental PC-C13 cells.