Abstract
Purpose: :
To determine the neuroprotective effect of 2-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 RGCs cultures were obtained from retinas of 6 to 8 day old Wistar rats, following a two-step immuno-panning procedure. After 72 hours of cultivation, the neuroprotective effect of brimonidine (0.01µM, 0.1µM and 1 µM) was investigated by culturing the RGCs in glutamate, oxidative and hypoxic stress for a further 72 hours, 24 hours and 12 hours respectively. Glutamate neurotoxicity was induced by adding glutamate (25µM), while for oxidative stress, culture medium with B27 supplement without anti-oxidants were substituted. Hypoxic stress was induced by cultivation in controlled-atmosphere incubator with oxygen levels of 5% normal partial pressure. The RGC viability in each condition normalized to that under normal condition was evaluated as live cell percentage based on a total of 7-8 experiments.
Results: :
The cell survival percentage of control cultures exposed to glutamate, oxidative and hypoxic stress were 58.2%, 59.3% and 53.2% respectively. Brimonidine (1 µM) significantly increased RGC survival in the presence of glutamate (80.6%), oxidative (79.8%) and hypoxic (77.4%) stress (P=0.0018, <0.0001 and <0.0001 respectively). In the presence of 2-adrenergic antagonist yohimbine (10 µM), RGC viability with brimonidine (1 µM) was not significantly different from controls.
Conclusions: :
Brimonidine increased survival of purified rat RGCs in the presence of glutamate neurotoxicity, oxidative and hypoxic stress. The neuroprotective effect of brimonidine is mediated via 2-adrenergic receptors at the RGC level.
Keywords: neuroprotection • retinal culture • apoptosis/cell death