Long-term high osmolality activates Na+-H+ exchange and protein kinase C in aortic smooth muscle cells

M. Soleimani, G. Singh, Jesus Dominguez, R. L. Howard

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

The effect of long-term exposure to hypertonic medium on Na+-H+ exchange activity was studied in cultured vascular smooth muscle (VSM) cells by using a combination of 22Na+ influx and pH measurement with the pH-sensitive dye BCECF. Incubation of VSM cells in high-osmolality medium (510 mOsm/L) for 48 hours significantly increased the acid-stimulated 22Na+ influx (control, 3.16±0.41 nmol/mg protein per minute; high osmolality, 6.40±0.66 nmol/mg protein pet minute; P<.01) and Na+-dependent pH(i) recovery (control, 0.29±0.06 pH/min; high osmolality, 0.65±0.13 pH/min; P<.03). Activation of Na+-H+ exchange was osmolality dependent and reached maximal stimulation at ≃700 mOsm/L. Na+-H+ exchanger stimulation was independent of serum in the culture media. Na+-H+ exchanger isoform (NHE-1) mRNA in VSM cells cultured in high-osmolality medium was un changed from that in VSM cells cultured in control medium, indicating an absence of transcriptional regulation by high osmolality. Long-term high osmolality significantly increased protein kinase C (PKC) activity in cultured VSM cells, as assessed by phosphorylation of a PKC-specific substrate (control, 20.9±2.1 pmol phosphorylation/mg protein per minute; high osmolality, 33.6±2.9 pmol phosphorylation/mg protein per minute; P<.01). Downregulation of PKC by preincubation of VSM cells with 0.1 μmol/L phorbol 12-myristate 13-acetate (PMA) prevented osmolality-induced stimulation of the Na+-H+ exchanger (control plus PMA, 0.27±0.05 pH/min; high osmolality plus PMA, 0.33±0.08 pH/min; P>.05). These results indicate that long-term exposure to hypertonic medium stimulates Na+-H+ exchange activity in cultured VSM cells and that this effect is independent of antiporter gent expression regulation. The results further demonstrate that the stimulatory effect of osmolality on Na+-H+ exchanger is mediated via posttranslational modification of the Na+-H+ exchanger by chronic PKC activation. The Na+-H+ exchanger may be involved in VSM cell volume regulation in long-term high osmolality.

Original languageEnglish
Pages (from-to)530-535
Number of pages6
JournalCirculation Research
Volume76
Issue number4
StatePublished - 1995

Fingerprint

Vascular Smooth Muscle
Osmolar Concentration
Protein Kinase C
Sodium-Hydrogen Antiporter
Smooth Muscle Myocytes
Antiporters
Pets
Post Translational Protein Processing
Cell Size
Proteins
Coloring Agents
Acids

Keywords

  • high osmolality
  • Na-H exchanger
  • vascular smooth muscle cells

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

Cite this

Long-term high osmolality activates Na+-H+ exchange and protein kinase C in aortic smooth muscle cells. / Soleimani, M.; Singh, G.; Dominguez, Jesus; Howard, R. L.

In: Circulation Research, Vol. 76, No. 4, 1995, p. 530-535.

Research output: Contribution to journalArticle

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abstract = "The effect of long-term exposure to hypertonic medium on Na+-H+ exchange activity was studied in cultured vascular smooth muscle (VSM) cells by using a combination of 22Na+ influx and pH measurement with the pH-sensitive dye BCECF. Incubation of VSM cells in high-osmolality medium (510 mOsm/L) for 48 hours significantly increased the acid-stimulated 22Na+ influx (control, 3.16±0.41 nmol/mg protein per minute; high osmolality, 6.40±0.66 nmol/mg protein pet minute; P<.01) and Na+-dependent pH(i) recovery (control, 0.29±0.06 pH/min; high osmolality, 0.65±0.13 pH/min; P<.03). Activation of Na+-H+ exchange was osmolality dependent and reached maximal stimulation at ≃700 mOsm/L. Na+-H+ exchanger stimulation was independent of serum in the culture media. Na+-H+ exchanger isoform (NHE-1) mRNA in VSM cells cultured in high-osmolality medium was un changed from that in VSM cells cultured in control medium, indicating an absence of transcriptional regulation by high osmolality. Long-term high osmolality significantly increased protein kinase C (PKC) activity in cultured VSM cells, as assessed by phosphorylation of a PKC-specific substrate (control, 20.9±2.1 pmol phosphorylation/mg protein per minute; high osmolality, 33.6±2.9 pmol phosphorylation/mg protein per minute; P<.01). Downregulation of PKC by preincubation of VSM cells with 0.1 μmol/L phorbol 12-myristate 13-acetate (PMA) prevented osmolality-induced stimulation of the Na+-H+ exchanger (control plus PMA, 0.27±0.05 pH/min; high osmolality plus PMA, 0.33±0.08 pH/min; P>.05). These results indicate that long-term exposure to hypertonic medium stimulates Na+-H+ exchange activity in cultured VSM cells and that this effect is independent of antiporter gent expression regulation. The results further demonstrate that the stimulatory effect of osmolality on Na+-H+ exchanger is mediated via posttranslational modification of the Na+-H+ exchanger by chronic PKC activation. The Na+-H+ exchanger may be involved in VSM cell volume regulation in long-term high osmolality.",
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AU - Howard, R. L.

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