Effect of nucleotides on the cytosolic free calcium activity and inositol phosphate formation in human glomerular epithelial cells

Br J Pharmacol. 1992 Sep;107(1):189-95. doi: 10.1111/j.1476-5381.1992.tb14485.x.

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.

MeSH terms

  • Adenosine Triphosphate / pharmacology
  • Calcium / metabolism*
  • Cells, Cultured
  • Cytosol / drug effects
  • Cytosol / metabolism
  • Down-Regulation
  • Epithelium / drug effects
  • Epithelium / metabolism
  • Fura-2
  • Inosine Triphosphate / pharmacology
  • Inositol Phosphates / metabolism*
  • Kidney Glomerulus / drug effects*
  • Kidney Glomerulus / metabolism
  • Nucleotides / pharmacology*
  • Protein Kinase C / metabolism
  • Spectrometry, Fluorescence
  • Suramin / pharmacology
  • Tetradecanoylphorbol Acetate / pharmacology
  • Uridine Diphosphate / pharmacology
  • Uridine Triphosphate / pharmacology

Substances

  • Inositol Phosphates
  • Nucleotides
  • Inosine Triphosphate
  • Uridine Diphosphate
  • Suramin
  • Adenosine Triphosphate
  • Protein Kinase C
  • Tetradecanoylphorbol Acetate
  • Calcium
  • Fura-2
  • Uridine Triphosphate