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Calcium signalling through nucleotide receptor P2X1 in rat portal vein myocytes

J. Mironneau, F. Coussin, J. L. Morel, C. Barbot, L. H. Jeyakumar *, S. Fleischer * and C. Mironneau

1. ATP-mediated Ca²⁺ signalling was studied in freshly isolated rat portal vein myocytes by means of a laser confocal microscope and the patch-clamp technique.
2. In vascular myocytes held at -60 mV, ATP induced a large inward current that was supported mainly by activation of P2X1 receptors, although other P2X receptor subtypes (P2X3, P2X4 and P2X5) were revealed by reverse transcription-polymerase chain reaction.
3. Confocal Ca²⁺ measurements revealed that ATP-mediated Ca²⁺ responses started at initiation sites where spontaneous or triggered Ca²⁺ sparks were not detected, whereas membrane depolarizations triggered Ca²⁺ waves by repetitive activation of Ca²⁺ sparks from a single initiation site.
4. ATP-mediated Ca²⁺ responses depended on Ca²⁺ influx through non-selective cation channels that activated, in turn, Ca²⁺ release from the intracellular store via ryanodine receptors (RYRs). Using specific antibodies directed against the RYR subtypes, we show that ATP-mediated Ca²⁺ release requires, at least, RYR2, but not RYR3.
5. Our results suggest that, in vascular myocytes, Ca²⁺ influx through P2X1 receptors may trigger Ca²⁺-induced Ca²⁺ release at intracellular sites where RYRs are not clustered.

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