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Research ArticleArticle

Activation of Protein Kinase A Increases Phospholipase D Activity and Inhibits Phospholipase D Activation by Acetylcholine in Tracheal Smooth Muscle

A. M. Mamoon, J. Smith, R. C. Baker and J. M. Farley
Journal of Pharmacology and Experimental Therapeutics December 1999, 291 (3) 1188-1195;
A. M. Mamoon
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J. Smith
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R. C. Baker
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J. M. Farley
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Abstract

Increased cAMP by stimulation of adenylyl cyclase with forskolin or by β-adrenoceptor activation with isoproterenol increased phospholipase D (PLD) activity in tracheal smooth muscle strips. PLD activity was measured by the accumulation of phosphatidylethanol. A linear increase in the concentration of phosphatidylethanol was observed over 20 min in muscle strips treated with either forskolin or isoproterenol. Cholinergic stimulation with acetylcholine (ACh), by contrast, caused a rapid increase in phosphatidylethanol followed by a slow decline in the concentration of phosphatidylethanol from 5 to 20 min in the continued presence of ACh. Concomitant treatment with ACh and either forskolin or isoproterenol eliminated the rapid increases in phosphatidylethanol associated with ACh treatment. The response to forskolin or isoproterenol was not influenced by ACh. Inhibition of protein kinase C with calphostin C or bisindolylmaleimide I had no effect on isoproterenol- or forskolin-stimulated PLD activity but inhibited ACh-activated PLD activity. Protein kinase A (PKA) inhibitors H-89 and KT5720 significantly decreased forskolin- and isoproterenol-mediated activation of PLD activity. PKA inhibition also eliminated inhibition of ACh-stimulated PLD activity by forskolin or isoproterenol. Activation of adenylyl cyclase by forskolin or by isoproterenol caused increased phosphorylation of phospholipase C-β2 isoform and reduced the formation of inositol phosphates after ACh stimulation of muscarinic receptors. These results suggest that increasing the concentration of cAMP activates PLD via activation of PKA and that the increased activity of PKA also inhibits cholinergic stimulation of PLD, in part at least by inhibiting the activation of phospholipase C by ACh.

Footnotes

  • Send reprint requests to: Jerry M. Farley, Ph.D., University of Mississippi Medical Center, Department of Pharmacology and Toxicology, 2500 North State St., Jackson, MS 39216-4505. E-mail:Jfarley{at}pharmacology.umsmed.edu

  • ↵1 This work was supported by the American Lung Association of Mississippi and National Institutes of Health Grants HL55544 and AA07157-10.

  • Abbreviations:
    ACh
    acetylcholine
    DAG
    diacylglycerol
    GFX
    bisindolylmaleimide I
    mAChR
    muscarinic acetylcholine receptor
    IP3
    inositol-1,4,5-trisphosphate
    PEth
    phosphatidylethanol
    PKA
    protein kinase A
    PKC
    protein kinase C
    PLC
    phospholipase C
    PLD
    phospholipase D
    PMA
    phorbol-12-myristate-13-acetate
    TLC
    thin-layer chromatography
    • Received January 27, 1999.
    • Accepted September 3, 1999.
  • The American Society for Pharmacology and Experimental Therapeutics
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Journal of Pharmacology and Experimental Therapeutics: 291 (3)
Journal of Pharmacology and Experimental Therapeutics
Vol. 291, Issue 3
1 Dec 1999
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Research ArticleArticle

Activation of Protein Kinase A Increases Phospholipase D Activity and Inhibits Phospholipase D Activation by Acetylcholine in Tracheal Smooth Muscle

A. M. Mamoon, J. Smith, R. C. Baker and J. M. Farley
Journal of Pharmacology and Experimental Therapeutics December 1, 1999, 291 (3) 1188-1195;

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Research ArticleArticle

Activation of Protein Kinase A Increases Phospholipase D Activity and Inhibits Phospholipase D Activation by Acetylcholine in Tracheal Smooth Muscle

A. M. Mamoon, J. Smith, R. C. Baker and J. M. Farley
Journal of Pharmacology and Experimental Therapeutics December 1, 1999, 291 (3) 1188-1195;
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