Evidence for receptor‐mediated calcium entry and refilling of intracellular calcium stores in FRTL‐5 rat thyroid cells

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Abstract
The aim of the present study was to investigate the relationship between agonist‐induced changes in intracellular free Ca2+ ([Ca2+]j) and the refilling of intracellular Ca2+ stores in Fura 2–loaded thyroid FRTL‐5 cells. Stimulating the cells with ATP induced a dose‐dependent increase in ([Ca2+]j). The ATP‐induced increase in [Ca2+]j was dependent on both release of sequestered intracellular Ca2+ as well as influx of extracellular Ca2+. Addition of Ni2+ prior to ATP blunted the component of the ATP‐induced increase in [Ca2+]j dependent on influx of Ca2+. In cells stimulated with ATP in a Ca2+‐free buffer, readdition of Ca2+ induced a rapid increase in [Ca2+]j; this increase was inhibited by Ni2+. In addition, the ATP‐induced influx of 45Ca2+ was blocked by Ni2+. Stimulating the cells with noradrenaline (NA) also induced release of sequestered Ca2+ and an influx of extracellular Ca2+. When cells were stimulated first with NA, a subsequent addition of ATP induced a blunted increase in [Ca2+]j. If the action of NA was terminated by addition of prazosin, and ATP was then added, the increase in [Ca2+]j was restored to control levels. Addition of Ni2+ prior to prazosin inhibited the restoration of the ATP response. In the presence of extracellular Mn2+, ATP stimulated quenching of Fura 2 fluorescence. The quenching was probably due to influx of Mn2+, as it was blocked by Ni2+. The results thus suggested that stimulating release of sequestered Ca2+ in FRTL‐5 cells was followed by influx of extracellular Ca2+ and rapid refilling of intracellular Ca2+ stores.