title: Brimonidine is neuroprotective against glutamate-induced neurotoxicity, oxidative stress, and hypoxia in purified rat retinal ganglion cells.
abstract: OBJECTIVE
  To investigate the neuroprotective effect of alpha2-adrenergic agonist brimonidine in the presence of glutamate-induced neurotoxicity, oxidative stress, and hypoxia on in vitro cultures of purified rat retinal ganglion cells (RGCs).
  
  
  METHODS
  Purified RGC cultures were obtained from retinas of 6-8-day old Wistar rats, following a two-step immunopanning procedure. After 72 h of cultivation, the neuroprotective effect of brimonidine (0.01 microM, 0.1 microM, and 1 microM) was investigated by culturing the RGCs under glutamate, oxidative, and hypoxic stress for a further 72 h, 24 h, and 12 h, respectively. Glutamate neurotoxicity was induced by adding glutamate (25 microM), while oxidative stress was induced by substituting the culture medium with B27 supplement without antioxidants, and hypoxia was induced by cultivation in a controlled-atmosphere incubator with oxygen levels 5% of the normal partial pressure. The RGC viability under each stress condition normalized to that under normal condition was evaluated as live cell percentage based on a total of 7-8 full repeated experiments.
  
  
  RESULTS
  The cell survival percentages of cultures exposed to glutamate, oxidative, and hypoxic stress were 58.2%, 59.3%, and 53.2%, respectively. Brimonidine dose dependently increased RGC survival in the presence of glutamate (80.6% at 1 microM), oxidative (79.8% at 1 microM), and hypoxic (72.3 and 77.4% at 0.1 and 1 microM, respectively) stress. In the presence of alpha2-adrenergic antagonist yohimbine (10 microM), brimonidine (1 microM) showed no protective effects on RGC viability.
  
  
  CONCLUSIONS
  At a concentration of 0.1 microM or higher, brimonidine increased survival of purified rat RGCs in the presence of glutamate neurotoxicity, oxidative stress, and hypoxia. The neuroprotective effect of brimonidine is mediated via alpha2-adrenergic receptors at the RGC level.
authors.affiliation: Department of Ophthalmology, University of Tokyo, Graduate School of Medicine, Bunkyo-ku, Tokyo, Japan.
authors.forename: Kelvin Yoon Chiang
authors.initials: KY
authors.lastname: Lee
authors.forename: Mao
authors.initials: M
authors.lastname: Nakayama
authors.forename: Makoto
authors.initials: M
authors.lastname: Aihara
authors.forename: Yi-Ning
authors.initials: YN
authors.lastname: Chen
authors.forename: Makoto
authors.initials: M
authors.lastname: Araie
mesh_terms: D000818:Animals; D000068438:Brimonidine Tartrate; D015687:Cell Hypoxia / Q000187:drug effects; D002469:Cell Separation; D002470:Cell Survival / Q000187:drug effects; D018698:Glutamic Acid / Q000633:toxicity*; D018696:Neuroprotective Agents / Q000494:pharmacology*; D020258:Neurotoxicity Syndromes / Q000517:prevention & control*; D018384:Oxidative Stress / Q000187:drug effects*; D011810:Quinoxalines / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D012165:Retinal Ganglion Cells / Q000187:drug effects* / Q000473:pathology*
chemical_list: D018696:Neuroprotective Agents; D011810:Quinoxalines; D018698:Glutamic Acid; D000068438:Brimonidine Tartrate