Glutamate-induced excitotoxicity is thought to play an important role in several
neurodegenerative diseases in the central nervous system (CNS). In this study, neuroprotection against
glutamate-induced excitotoxicity was analyzed using
acetylcholine (ACh),
nicotine and the α7 specific
nicotinic acetylcholine receptor (α7 nAChR) agonist, N-[(3R)-1-azabicyclo[2.2.2]oct-3-yl]-4-chlorobenzamide hydrochloride (PNU-282987), in cultured adult rat retinal neurons. Adult Long Evans rat retinas were dissociated and retinal ganglion cells (RGCs) were isolated from all other
retinal tissue using a two-step panning technique. Once isolated, RGCs were cultured under various pharmacological conditions to demonstrate excitotoxicity and neuroprotection against excitotoxicity. After 3 days, RGCs were immunostained with
antibodies against the
glycoprotein, Thy 1.1, counted and cell survival was assessed relative to control untreated conditions. 500 μM
glutamate induced excitotoxicity in large and small RGCs in an adult rat dissociated culture. After 3 days in culture with
glutamate, the cell survival of large RGCs decreased by an average of 48.16% while the cell survival of small RGCs decreased by an average of 42.03%. Using specific
glutamate receptor agonists and antagonists, we provide evidence that the excitotoxic response was mediated through α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic
acid (
AMPA)/
kainic acid (KA) and
N-methyl-d-aspartate (
NMDA)
glutamate receptors through an apoptotic mechanism. However, the excitotoxic effect of
glutamate on all RGCs was eliminated if cells were cultured for an hour with 10 μM ACh, 100 μM
nicotine or 100 nM of the α7 nAChR agonist,
PNU-282987, before the
glutamate insult. Inhibition studies using 10nM
methyllycaconitine (MLA) or α-bungarotoxin (α-Bgt) supported the hypothesis that neuroprotection against
glutamate-induced excitotoxicity on rat RGCs was mediated through α7 nAChRs. In immunocytochemical studies, double-labeled experiments using
antibodies against Thy 1.1 and α7 nAChR subunits demonstrated that both large and small RGCs contained α7 nAChR subunits. The data presented in this study support the hypothesis that ACh and
nicotinic acetylcholine receptor (nAChR) agonists provide neuroprotection against
glutamate-induced excitotoxicity in adult rat RGCs through activation of α7 nAChR subunits. These studies lay the groundwork required for analyzing the effect of specific α7 nAChR agonists using in vivo models of excitotoxicity. Understanding the type of
ACh receptors involved in neuroprotection in the rat retina could ultimately lead to therapeutic treatment for any
CNS disease that involves excitotoxicity.