title: Effect of nucleotides on the cytosolic free calcium activity and inositol phosphate formation in human glomerular epithelial cells.
abstract: 1. Glomerular epithelial cells (GEC) were cultured from human kidneys and immunologically characterized. 2. The effect of extracellular nucleotides on the cytosolic free calcium activity [Ca2+]i was investigated with the fura-2 microfluorescence method. Extracellular UTP, UDP, UMP, ATP, adenosine 5'-O-(3-thio)-trisphosphate (ATP-gamma-S), inosine-triphosphate (ITP), guanyltriphosphate (GTP), 2-methylthio-ATP, AMP, alpha,beta-methylene-ATP and adenosine led to a rapid, transient, concentration-dependent increase of [Ca2+]i, followed by a plateau above the baseline level. 3. In a calcium-free extracellular solution, the rapid increase of [Ca2+]i was still present, whereas the plateau level was abolished. 4. ATP and UTP (ED50 both: 10(-5) M) stimulated inositol trisphosphate (InsP3) formation in GEC. 5. The order of potency for the purine nucleotides in stimulating InsP3 formation was ATP = ATP-gamma-S greater than ADP greater than 2-methylthio-ATP greater than AMP = a,beta methylene-ATP = adenosine. 6. The increase of InsP3 induced by ATP (10(-5) M) could be inhibited by the P2 receptor blocker suramin (greater than 10(-4) M). Reactive blue 2 exhibited a weak stimulating effect on the InsP3 formation and only a weak inhibitory effect at a concentration of 10(-3) M was observed. 7. Protein kinase C activation by preincubation of GEC with phorbol 12-myristate 13-acetate (PMA, 100 ng ml-1, 15 min) abolished the effect of ATP (10(-5) M) on InsP3 formation. Downregulation of protein kinase C by long term incubation (18 h) with PMA had no significant effect on the phosphoinositol turnover induced by ATP.8. The results indicate that an increase of [Ca2+]i and inositol phosphate breakdown can be mediated via activation of a P2 receptor in human GEC.
authors.affiliation: Medizinische Klinik, Nephrologische Abteilung, Freiburg, Germany.
authors.forename: H
authors.initials: H
authors.lastname: Pavenstädt
authors.forename: M
authors.initials: M
authors.lastname: Späth
authors.forename: G
authors.initials: G
authors.lastname: Schlunck
authors.forename: M
authors.initials: M
authors.lastname: Nauck
authors.forename: R
authors.initials: R
authors.lastname: Fischer
authors.forename: C
authors.initials: C
authors.lastname: Wanner
authors.forename: P
authors.initials: P
authors.lastname: Schollmeyer
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D015536:Down-Regulation; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D016257:Fura-2; D007293:Inosine Triphosphate / Q000494:pharmacology; D007295:Inositol Phosphates / Q000378:metabolism*; D007678:Kidney Glomerulus / Q000187:drug effects* / Q000378:metabolism; D009711:Nucleotides / Q000494:pharmacology*; D011493:Protein Kinase C / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013498:Suramin / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014530:Uridine Diphosphate / Q000494:pharmacology; D014544:Uridine Triphosphate / Q000494:pharmacology
chemical_list: D007295:Inositol Phosphates; D009711:Nucleotides; D007293:Inosine Triphosphate; D014530:Uridine Diphosphate; D013498:Suramin; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D016257:Fura-2; D014544:Uridine Triphosphate