cAMP and purinergic P2y receptors upregulate and enhance inducible NO synthase mRNA and protein in vivo

Am J Physiol. 1997 Nov;273(5):L967-79. doi: 10.1152/ajplung.1997.273.5.L967.


Adenosine 3',5'-cyclic monophosphate (cAMP) and purinergic P2y receptor agonists upregulate inducible nitric oxide (NO) synthase (iNOS) but inhibit Escherichia coli endotoxin lipopolysaccharide (LPS)- and cytokine-mediated upregulation of iNOS in cultured cells. We examined the effects of cAMP and P2y receptor agonists on the iNOS system in vivo. Intratracheal administration of dibutyryl-cAMP (DBcAMP, 0.1 and 1 mg/kg), a P2y receptor agonist [2-methylthioadenosine 5'-triphosphate (MeS-ATP), 5 mg/kg], or LPS (0.6 mg/kg) to rats 2 h before bronchoalveolar lavage (BAL) increased iNOS mRNA (competitor-equalized reverse transcription-polymerase chain reaction) and iNOS protein (Western blot) in rat alveolar macrophages compared with the effects of sterile phosphate-buffered saline (0.5 ml it). At equal levels of upregulation of iNOS mRNA, 1) LPS, but not DBcAMP or MeS-ATP, upregulated nuclear transcription factor-kappa B (NF-kappa B) and 2) iNOS protein and formation of NO were greater in alveolar macrophages from LPS- and MeS-ATP-treated rats than from DBcAMP-treated rats. Administration of DBcAMP or MeS-AMP 15 min before LPS did not inhibit LPS-induced alveolar macrophage-derived iNOS mRNA, iNOS protein, and NO. Diethyldithiocarbamate (DETC, 5 mg/kg it) inhibited LPS-induced iNOS mRNA but did not affect upregulation of iNOS mRNA produced by the other agonists. We conclude that an LPS-dependent and -independent pathway of iNOS mRNA induction exists in vivo. The former is activated by IPS and most cytokines, is associated with upregulation of NF-kappa B and inhibited by DETC, and elicits an inflammatory response. The latter, activated by DBcAMP and MeS-ATP, is not associated with upregulation of NF-kappa B, inhibition by DETC, or activation of inflammation. The two systems are additive in vivo rather than antagonistic. Speculatively, if the LPS-independent iNOS pathway exists in humans, the iNOS in tissues from patients taking drugs affecting cAMP or P2y receptors may be iatrogenic rather than pathogenetic in origin.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adenosine Triphosphate / analogs & derivatives*
  • Adenosine Triphosphate / pharmacology
  • Albuterol / pharmacology
  • Animals
  • Bronchoalveolar Lavage Fluid / chemistry
  • Bucladesine / pharmacology*
  • Chemotaxis, Leukocyte / drug effects
  • Cyclic AMP / metabolism*
  • Ditiocarb / pharmacology
  • Enzyme Induction
  • Isoproterenol / pharmacology
  • Lipopolysaccharides / pharmacology
  • Lung / drug effects
  • Lung / physiology*
  • Macrophages, Alveolar / drug effects
  • Macrophages, Alveolar / enzymology*
  • Macrophages, Alveolar / immunology
  • Male
  • NF-kappa B / biosynthesis
  • Nitric Oxide Synthase / biosynthesis*
  • Polymerase Chain Reaction
  • Purinergic P2 Receptor Agonists
  • RNA, Messenger / biosynthesis
  • Rats
  • Rats, Sprague-Dawley
  • Receptors, Purinergic P2 / physiology*
  • Thionucleotides / pharmacology*
  • Transcription, Genetic / drug effects
  • Transcription, Genetic / physiology*
  • Tumor Necrosis Factor-alpha / biosynthesis


  • Lipopolysaccharides
  • NF-kappa B
  • Purinergic P2 Receptor Agonists
  • RNA, Messenger
  • Receptors, Purinergic P2
  • Thionucleotides
  • Tumor Necrosis Factor-alpha
  • Bucladesine
  • Adenosine Triphosphate
  • Ditiocarb
  • Cyclic AMP
  • Nitric Oxide Synthase
  • Isoproterenol
  • Albuterol
  • 2-methylthio-ATP