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• W2145301870 abstract "TRPM2 (transient receptor potential melastatin 2) is a Ca2+-permeable cation channel gated by ADPR (ADP-ribose) from the cytosolic side. To test whether endogenous concentrations of intracellular ADPR are sufficient for TRPM2 gating in neutrophil granulocytes, we devised an HPLC method to determine ADPR contents in HClO4 cell extracts. The reversed-phase ion-pair HPLC method with an Mg2+-containing isocratic eluent allows baseline resolution of one ADPR peak. Intracellular ADPR concentrations were approx. 5 μM in granulocytes and not significantly altered by stimulation with the chemoattractant peptide fMLP (N-formylmethionyl-leucylphenylalanine). We furthermore determined intracellular concentrations of cADPR (cyclic ADPR) with a cyclase assay involving enzymatic conversion of cADPR into NAD+ and fluorimetric determination of NAD+. Intracellular cADPR concentrations were approx. 0.2 μM and not altered by fMLP. In patch–clamp experiments, ADPR (0.1–100 μM) was dialysed into granulocytes to analyse its effects on whole-cell currents characteristic for TRPM2, in the presence of a low (&lt;10 nM) or a high (1 μM) intracellular Ca2+ concentration. TRPM2 currents were significantly larger at high than at low [Ca2+] (e.g. −225±27.1 versus −7±2.0 pA/pF at 5 μM ADPR), but no currents at all were observed in the absence of ADPR (ADPR concentration ≤0.3 μM). cADPR (0.1, 0.3 and 10 μM) was without effect even in the presence of subthreshold ADPR (0.1 μM). We conclude that ADPR enables an effective regulation of TRPM2 by cytosolic Ca2+. Thus ADPR and Ca2+ in concert behave as a messenger system for agonist-induced influx of Ca2+ through TRPM2 in granulocytes." @default.
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• W2145301870 date "2006-08-15" @default.
• W2145301870 modified "2023-10-15" @default.
• W2145301870 title "Endogenous ADP-ribose enables calcium-regulated cation currents through TRPM2 channels in neutrophil granulocytes" @default.
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• W2145301870 doi "https://doi.org/10.1042/bj20060183" @default.
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