Calcium signalling through nucleotide receptor P2Y2 in cultured human vascular endothelium.
Microfluorometric measurements in Fura-2-loaded single cultured human vascular endothelial cells were used to characterize the intracellular calcium [Ca2+]i responses triggered by extracellular application of adenosine 5'-triphosphate (ATP) and other nucleotides. Application of ATP or uridine 5'-triphosphate (UTP) gave rise to dose-dependent elevations of [Ca2+]i in all the cells tested. At saturating concentrations of agonist, the [Ca2+]i response was biphasic, with an early peak and a sustained plateau. Unlike peak responses, the sustained Ca2+ plateau was sensitive to removal of Ca2+ from the external medium. Mn2+ quenching revealed the presence of Ca2+ influx during the agonist-induced calcium plateau. The agonist-evoked calcium plateau was inhibited in a dose-dependent manner by the Cl-channel blocker NPPB, by the divalent cation Ni2+ and by the imidazole antimycotic econazole. Previously, these compounds have been shown to block store-operated Ca2+ entry. The two phases of the agonist-evoked [Ca2+]i response were blocked by the specific phospholipase C inhibitor U-73122 and by intracellular injection of low molecular weight heparin, suggesting the involvement of IP3-sensitive intracellular Ca2+ stores. The pharmacological profile of the response, using different nucleotides and analogues, ATP = UTP > ADP = UDP, and no responses to P2X1 and P2Y1 agonists, suggested the involvement of P2Y2 receptors. The expression of mRNA for the P2Y2 receptor was detected by RT-PCR analysis. These results indicate that P2Y2 receptors linked to intracellular Ca2+ mobilization are present in human vascular endothelial cells. The initial [Ca2+]i mobilization is followed by a phase of elevated [Ca2+]i influx.
Viana
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F
Katholieke Universiteit Leuven, Laboratorium voor Fysiologie, Belgium. Felix.Viana@umh.es
de Smedt
H
H
Droogmans
G
G
Nilius
B
B
D000255:Adenosine Triphosphate / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004963:Estrenes / Q000494:pharmacology; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008345:Manganese / Q000494:pharmacology; D009532:Nickel / Q000494:pharmacology; D009579:Nitrobenzoates / Q000494:pharmacology; D009711:Nucleotides / Q000494:pharmacology; D011760:Pyrrolidinones / Q000494:pharmacology; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000378:metabolism*; D058473:Receptors, Purinergic P2Y2; D010738:Type C Phospholipases / Q000187:drug effects / Q000378:metabolism; D014544:Uridine Triphosphate / Q000494:pharmacology
D004963:Estrenes; D009579:Nitrobenzoates; D009711:Nucleotides; C546385:P2RY2 protein, human; D011760:Pyrrolidinones; D018048:Receptors, Purinergic P2; D058473:Receptors, Purinergic P2Y2; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; D008345:Manganese; D009532:Nickel; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; D010738:Type C Phospholipases; D002118:Calcium; D014544:Uridine Triphosphate