title: Blocking a vicious cycle nNOS/peroxynitrite/AMPK by S-nitrosoglutathione: implication for stroke therapy.
abstract: BACKGROUND
  Stroke immediately sets into motion sustained excitotoxicity and calcium dysregulation, causing aberrant activity in neuronal nitric oxide synthase (nNOS) and an imbalance in the levels of nitric oxide (NO). Drugs targeting nNOS-originated toxicity may therefore reduce stroke-induced damage. Recently, we observed that a redox-modulating agent of the NO metabolome, S-nitrosoglutathione (GSNO), confers neurovascular protection by reducing the levels of peroxynitrite, a product of aberrant NOS activity. We therefore investigated whether GSNO-mediated neuroprotection and improved neurological functions depend on blocking nNOS/peroxynitrite-associated injurious mechanisms using a rat model of cerebral ischemia reperfusion (IR).
  
  
  RESULTS
  IR increased the activity of nNOS, the levels of neuronal peroxynitrite and phosphorylation at Ser(1412) of nNOS. GSNO treatment of IR animals decreased IR-activated nNOS activity and neuronal peroxynitrite levels by reducing nNOS phosphorylation at Ser(1412). The Ser(1412) phosphorylation is associated with increased nNOS activity. Supporting the notion that nNOS activity and peroxynitrite are deleterious following IR, inhibition of nNOS by its inhibitor 7-nitroindazole or reducing peroxynitrite by its scavenger FeTPPS decreased IR injury. GSNO also decreased the activation of AMP Kinase (AMPK) and its upstream kinase LKB1, both of which were activated in IR brain. AMPK has been implicated in nNOS activation via Ser(1412) phosphorylation. To determine whether AMPK activation is deleterious in the acute phase of IR, we treated animals after IR with AICAR (an AMPK activator) and compound c (an AMPK inhibitor). While AICAR potentiated, compound c reduced the IR injury.
  
  
  CONCLUSIONS
  Taken together, these results indicate an injurious nNOS/peroxynitrite/AMPK cycle following stroke, and GSNO treatment of IR inhibits this vicious cycle, resulting in neuroprotection and improved neurological function. GSNO is a natural component of the human body, and its exogenous administration to humans is not associated with any known side effects. Currently, the FDA-approved thrombolytic therapy suffers from a lack of neuronal protective activity. Because GSNO provides neuroprotection by ameliorating stroke's initial and causative injuries, it is a candidate of translational value for stroke therapy.
authors.affiliation: Department of Pediatrics, Medical University of South Carolina, Charleston, SC, 29425, USA. khanm@musc.edu.
authors.forename: Mushfiquddin
authors.initials: M
authors.lastname: Khan
authors.affiliation: Department of Pediatrics, Medical University of South Carolina, Charleston, SC, 29425, USA. dhammu@musc.edu.
authors.forename: Tajinder S
authors.initials: TS
authors.lastname: Dhammu
authors.affiliation: Department of Pediatrics, Medical University of South Carolina, Charleston, SC, 29425, USA. fumiyo@health.nop.kagoshima-u.ac.jp.|School of Health Science, Kagoshima University, Kagoshima, Japan. fumiyo@health.nop.kagoshima-u.ac.jp.
authors.forename: Fumiyo
authors.initials: F
authors.lastname: Matsuda
authors.affiliation: Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, SC, USA. singha@musc.edu.|Ralph H. Johnson VA Medical Center, Charleston, SC, USA. singha@musc.edu.
authors.forename: Avtar K
authors.initials: AK
authors.lastname: Singh
authors.affiliation: Department of Pediatrics, Medical University of South Carolina, Charleston, SC, 29425, USA. singhi@musc.edu.
authors.forename: Inderjit
authors.initials: I
authors.lastname: Singh
mesh_terms: D000091162:AMP-Activated Protein Kinase Kinases; D000263:Adenylate Kinase / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D002545:Brain Ischemia / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D004195:Disease Models, Animal; D020244:Infarction, Middle Cerebral Artery; D008297:Male; D009474:Neurons / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D018696:Neuroprotective Agents / Q000494:pharmacology*; D052248:Nitric Oxide Synthase Type I / Q000037:antagonists & inhibitors / Q000378:metabolism*; D030421:Peroxynitrous Acid / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011897:Random Allocation; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D026422:S-Nitrosoglutathione / Q000494:pharmacology*; D020521:Stroke / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology
chemical_list: D018696:Neuroprotective Agents; D030421:Peroxynitrous Acid; D026422:S-Nitrosoglutathione; D052248:Nitric Oxide Synthase Type I; C496322:Nos1 protein, rat; D017346:Protein Serine-Threonine Kinases; C488755:Stk11 protein, rat; D000091162:AMP-Activated Protein Kinase Kinases; D000263:Adenylate Kinase