Nitric oxide prevents 6-hydroxydopamine-induced apoptosis in PC12 cells through cGMP-dependent PI3 kinase/Akt activation

FASEB J. 2003 Jun;17(9):1036-47. doi: 10.1096/fj.02-0738com.

Abstract

Nitric oxide (NO) functions not only as an important signaling molecule in the brain by producing cGMP, but also regulates neuronal cell apoptosis. The mechanism by which NO regulates apoptosis is unclear. In this study, we demonstrated that NO, produced either from the NO donor S-nitroso-N-acetyl-d,l-penicillamine (SNAP) or by transfection of neuronal NO synthase, suppressed 6-hydroxydopamine (6-OHDA)-induced apoptosis in PC12 cells by inhibiting mitochondrial cytochrome c release, caspase-3 and -9 activation, and DNA fragmentation. This protection was significantly reversed by the soluble guanylyl cyclase inhibitor 1H-(1,2,4)-oxadiazole[4,3-a]quinoxalon-1-one, indicating that cGMP is a key mediator in NO-mediated anti-apoptosis. Moreover, the membrane-permeable cGMP analog 8-Br-cGMP inhibited 6-OHDA-induced apoptosis. These anti-apoptotic effects of SNAP and 8-Br-cGMP were suppressed by cGMP-dependent protein kinase G (PKG) inhibitor KT5823, indicating that PKG is a downstream signal mediator in the suppression of apoptosis by NO and cGMP. Both SNAP and 8-Br-cGMP induced endogenous Akt activation and Bad phosphorylation, resulting in the inhibition of Bad translocation to mitochondria; these effects were inhibited by KT5823 and the phosphatidylinositol 3-kinase (PI3K) inhibitors LY294002 and Wortmannin. Our data suggest that the NO/cGMP pathway suppresses 6-OHDA-induced PC12 cell apoptosis by suppressing the mitochondrial apoptosis signal via PKG/PI3K/Akt-dependent Bad phosphorylation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anilides / metabolism
  • Animals
  • Apoptosis*
  • Caspase 3
  • Caspases / metabolism
  • Cell Differentiation
  • Cyclic GMP / physiology*
  • Cyclic GMP-Dependent Protein Kinases / metabolism
  • Cytochrome c Group / metabolism
  • Cytoprotection
  • Enzyme Activation
  • Neurons / cytology
  • Neurons / drug effects
  • Neurons / enzymology*
  • Nitric Oxide / physiology*
  • Nitric Oxide Donors / pharmacology
  • Nitric Oxide Synthase / genetics
  • Nitric Oxide Synthase Type I
  • Oligopeptides / metabolism
  • Oxidopamine / antagonists & inhibitors*
  • PC12 Cells
  • Penicillamine / analogs & derivatives*
  • Penicillamine / pharmacology
  • Phosphatidylinositol 3-Kinases / metabolism*
  • Protein-Serine-Threonine Kinases*
  • Proto-Oncogene Proteins / metabolism*
  • Proto-Oncogene Proteins c-akt
  • Rats
  • Transfection

Substances

  • Anilides
  • Cytochrome c Group
  • Nitric Oxide Donors
  • Oligopeptides
  • Proto-Oncogene Proteins
  • S-nitro-N-acetylpenicillamine
  • acetyl-aspartyl-glutamyl-valyl-aspartic acid p-nitroanilide
  • Nitric Oxide
  • Oxidopamine
  • Nitric Oxide Synthase
  • Nitric Oxide Synthase Type I
  • Nos1 protein, rat
  • Phosphatidylinositol 3-Kinases
  • Akt1 protein, rat
  • Protein-Serine-Threonine Kinases
  • Proto-Oncogene Proteins c-akt
  • Cyclic GMP-Dependent Protein Kinases
  • Casp3 protein, rat
  • Caspase 3
  • Caspases
  • Penicillamine
  • Cyclic GMP