Antisense inhibition of Na+/Ca2+ exchange in primary cultured arterial myocytes

M. K. Slodzinski, M. Juhaszova, M. P. Blaustein

Research output: Contribution to journalArticlepeer-review

Abstract

The effects of chimeric phosphorothioated antisense oligodeoxynucleotides (AS-oligos) targeted against the Na+/Ca2+ exchanger (NCX) were tested in primary cultured rat mesenteric artery myocytes. In parallel cultures, myocytes proliferated and were morphologically normal in the presence of scrambled nonsense (NS-) or AS-oligos or no oligos (controls). NCX function was examined with digital imaging, using fura 2 to estimate the cytosolic free Ca2+ concentration ([Ca2+](cyt)). Resting [Ca2+](cyt) was higher (145 ± 4 nM; P < 0.05) in AS-oligo-treated cells than in controls (125 ± 5 nM) or NS-oligo-treated cells (131 ± 4 nM). Lowering external Na+, to promote Ca2+ entry via NCX, increased [Ca2+](cyt) transiently in controls and NS-oligo-treated cells but not in AS-oligo-treated cells. Raising the cytosolic free Na+ concentration with ouabain augmented the low-Na+- induced rise in [Ca2+](cyt) in controls and NS-oligo-treated cells, but AS- oligo-treated cells still did not respond. Nevertheless, serotonin (5-HT) increased [Ca2+](cyt) in all three groups. Thus AS-oligos selectively blocked NCX activity but not the 5-HT response. To determine the effect of NCX knockdown on the modulation of stored Ca2+, the 5-HT response was tested immediately after removal of external Ca2+. Ouabain augmented the 5- HT-induced rise in [Ca2+](cyt) in control and NS-oligo-treated cells but not AS-oligo-treated cells. This indicates that the NCX can modulate intracellular Ca2+ stores. We conclude that AS oligos are useful for investigating the physiological role of NCX in vascular smooth muscle.

Original languageEnglish (US)
Pages (from-to)C1340-C1345
JournalAmerican Journal of Physiology - Cell Physiology
Volume269
Issue number5 38-5
DOIs
StatePublished - 1995

Keywords

  • calcium
  • oligodeoxynucleotides
  • ouabain
  • sarcoplasmic reticulum
  • vascular smooth muscle

ASJC Scopus subject areas

  • Physiology
  • Cell Biology

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