The effects of extracellular ATP and other nucleotides on the cytosolic free Ca²⁺ concentration ([Ca²⁺]_i) have been studied in single primary human hepatocytes and in human Hep G2 and HuH-7 hepatoma cells. ATP, adenosine 5'-O-(3-thiotriphosphate) (ATPS), and UTP caused a concentration-dependent biphasic increase in [Ca²⁺]_i with an initial peak followed by a small sustained plateau in most cells. In some cells, however, repetitive Ca²⁺ transients were observed. The rank order of potency was ATP  UTP > ATPS, and complete cross-desensitization of the Ca²⁺ responses occurred between ATP and UTP. The initial transient peak in [Ca²⁺]_i was resistant to extracellular Ca²⁺ depletion, which demonstrates mobilization of internal Ca²⁺ by inositol 1,4,5-trisphosphate whose formation was enhanced by ATP and UTP. In contrast, the sustained plateau phase required influx of external Ca²⁺. Ca²⁺ influx occurs most likely through a capacitative Ca²⁺ entry mechanism, which was shown to exist in these cells by experiments performed with thapsigargin. On the molecular level, specific mRNA coding for the human P2Y₁, P2Y₂, P2Y₄, and P2Y₆ receptors could be detected by RT-PCR in Hep G2 and HuH-7 cells. However, ADP and UDP, which are agonists for P2Y₁ and P2Y₆ receptors, respectively, caused no changes in [Ca²⁺]_i, demonstrating that these receptors are not expressed at a functional level. Likewise, ,-methylene-ATP, ,-methylene-ATP, AMP, and adenosine were inactive in elevating [Ca²⁺]_i, suggesting that the ATP-induced increase in [Ca²⁺]_i was not caused by activation of P2X or P1 receptors. Thus, on the basis of the pharmacological profile of the nucleotide-induced Ca²⁺-responses, extracellular ATP and UTP increase [Ca²⁺]_i by activating P2Y₂ and possibly P2Y₄ receptors coupled to the Ca²⁺-phosphatidylinositol signaling cascade in human hepatocytes. This suggests that extracellular nucleotides from various sources may contribute to the regulation of human liver cell functions.

adenosine triphosphate; uridine triphosphate; nucleotide receptor; intracellular calcium; human liver

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