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Journal of Pharmacology and Experimental Therapeutics

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OtherCELLULAR AND MOLECULAR PHARMACOLOGY

Activation of Guanosine 3′,5′-Cyclic Monophosphate (cGMP)-Dependent Protein Kinase in Rabbit Aorta by Nitroglycerin and Sodium Nitroprusside

Ashwinkumar I. Patel and Jack Diamond
Journal of Pharmacology and Experimental Therapeutics November 1997, 283 (2) 885-893;
Ashwinkumar I. Patel
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Jack Diamond
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Abstract

It is generally accepted that cGMP mediates the vascular relaxant effects of nitrovasodilators such as sodium nitroprusside (SNP) and nitroglycerin (NTG). It has been suggested that the relaxant effects of cGMP are mediated via activation of a specific, cGMP-dependent protein kinase (PKG). The objective of this study was to determine whether PKG can be activated by SNP and by NTG in intact strips of rabbit aorta and, if so, whether a good correlation exists between activation of PKG and relaxation of the arteries by the nitrovasodilators. PKG activity was measured by means of a recently described assay using a peptide substrate, BPDEtide, that exhibits good sensitivity and specificity for PKG compared with other protein kinases. Verification of the specificity of the assay for PKG was obtained using MonoQ chromatography to resolve soluble extracts of the rabbit aorta and subsequent immunoblotting to identify the kinase by means of a PKG-specific antibody. The role of PKG in vascular relaxation was investigated by simultaneously monitoring the effects of SNP and NTG on cGMP levels, PKG activity ratios and tension in isolated strips of rabbit aorta exposed to varying concentrations of the nitrovasodilators for varying times. The results indicate that PKG can be activated in a concentration- and time-dependent manner by both SNP and NTG in intact vascular preparations and that reasonably good correlations exist between PKG activation and relaxation in these experiments. Although a causal relationship between the two parameters has not been definitely established, these results are consistent with the proposed role for PKG as a mediator of the vascular relaxant effects of cGMP-elevating agents such as SNP and NTG.

Footnotes

  • Send reprint requests to: Dr. Jack Diamond, Division of Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences, University of British Columbia, 2146 East Mall, Vancouver, B.C., Canada. V6T 1Z3.

  • ↵1 This work was supported by a grant from the Heart and Stroke Foundation of British Columbia and Yukon.

  • Abbreviations:
    PKG
    cGMP-dependent protein kinase
    NTG
    nitroglycerin
    PE
    phenylephrine
    SNP
    sodium nitroprusside
    v.s.m.
    vascular smooth muscle
    ANF
    atrial natriuretic factor
    PKI
    protein kinase inhibitor
    PMSF
    phenylmethylsulfonyl fluoride
    PKA
    cAMP-dependent protein kinase
    SDS-PAGE
    sodium dodecyl sulfate polyacrylamide gel electrophoresis
    TTBS
    Tris-buffered saline with Tween 20
    • Received October 25, 1996.
    • Accepted July 28, 1997.
  • The American Society for Pharmacology and Experimental Therapeutics
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Journal of Pharmacology and Experimental Therapeutics
Vol. 283, Issue 2
1 Nov 1997
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OtherCELLULAR AND MOLECULAR PHARMACOLOGY

Activation of Guanosine 3′,5′-Cyclic Monophosphate (cGMP)-Dependent Protein Kinase in Rabbit Aorta by Nitroglycerin and Sodium Nitroprusside

Ashwinkumar I. Patel and Jack Diamond
Journal of Pharmacology and Experimental Therapeutics November 1, 1997, 283 (2) 885-893;

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OtherCELLULAR AND MOLECULAR PHARMACOLOGY

Activation of Guanosine 3′,5′-Cyclic Monophosphate (cGMP)-Dependent Protein Kinase in Rabbit Aorta by Nitroglycerin and Sodium Nitroprusside

Ashwinkumar I. Patel and Jack Diamond
Journal of Pharmacology and Experimental Therapeutics November 1, 1997, 283 (2) 885-893;
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