This study revealed that EAAT expression was increased

This study revealed that EAAT2 Wnt-C59 was increased 4–8 weeks after cauterization as our previous work (Park et al., 2009). RGC death could not be blocked despite activation of EAAT2, which is one of glutamate transporters in cone photoreceptors and some bipolar cells, in chronic ocular hypertension rats (Pow, 2001; Rauen, 2000; Rauen et al., 1996). Increased EAAT2 may suggest a compensatory reaction in the retina, activated by elevation of glutamate levels. Treatment with an α2 adrenergic agonist decreased glial activation, as evidenced by GFAP, and increased EAAT1 compared with eyes without treatment after cauterization. A reduction in GFAP expression with brimonidine treatment in chronic ocular hypertension models corresponded to previous experimental studies, although one study demonstrated that the GFAP expression remained elevated (Kim et al., 2007a; Vidal et al., 2010; WoldeMussie et al., 2001). The mechanism by which brimonidine application results in a reduction in GFAP immunoreactivity remains unclear. Decreased retinal stress caused by reduced IOP seems to lower GFAP immunoreactivity and activate EAAT1 in Müller cells. EAAT1 may be the main glutamate transporter in the mammalian retina and play important roles in glutatmate homeostasis (Izumi et al., 2002; Pow, 2001; Rauen, 2000; Rauen et al., 1996). Activation of EAAT1 may lead to increased clearance of synaptic glutamate. EAAT1 activation by an α2 agonist may contribute to the neuroprotective effects in RGCs by decreasing synaptic glutamate levels. The present study showed the first evidence that topical brimonidine application blocked upregulations of GluN1 subunit expression in a chronic ocular hypertension model. In previous studies using glaucoma models, elevated NMDA receptor expression has been demonstrated (Dong et al., 2013, Kim et al., 2007b), but NMDA receptor level following IOP reduction has not been determined. We speculate that IOP reduction after α2 agonist treatment itself may lower stress in retina (Kim et al., 2007a, Vidal et al., 2010). There is a possibility that stimulation of EAAT1 by relieving stress may decrease extracellular glutamate levels to induce down regulations of GluN1 subunit expression in the RGC, even though more studies are needed to confirm that (Bringmann et al., 2013; Sarthy et al., 2004; Vorwerk et al., 2000). The α2 agonist might decrease vitreal glutamate (Donello et al., 2001) or directly modulate GluN1 subunit expression (Dong et al., 2008, Dong et al., 2010, Lee et al., 2012). The application of α2 agonist will lead to the activation of K+ channels (Donello et al., 2001; Ma et al., 2004). Therefore, activated α2-adrenoceptors will hyperpolarize cells and block the glutamate release (Donello et al., 2001; Ma et al., 2004). In addition, Dong et al. showed that brimonidine application decreased NMDA-induced calcium signals in the isolated retina and in rat glaucoma model. In those studies, systemic brimonidine treatment did not affect IOP (Ahmed et al., 2001; Dong et al., 2008; Hernandez et al., 2008; WoldeMussie et al., 2001). Lee et al. also reported that brimonidine attenuated the upregulations of NMDA receptors in ischemic retinal injury model (Lee et al., 2012). In animal research, a 0.2 or 0.5% brimonidine results in similar vitreous concentration. (0.2%, 82nM; 0.5%, 97nM) (Acheampong et al., 2002). A newer formulation, brimonidine-purite 0.15% which was used in the current study, has been found to be equivalent to brimonidine 0.2% in lowering IOP and to penetrate into the vitreous at concentration above to activate α2-receptors (Kent et al., 2006). Given those findings, application topical brimonidine (0.15%) in the current study may be acceptable, although absence of the dose-ranging can be one of limitations in this study. The absence of the group treated with NMDA receptor blocker may be one of limitations in this study. We previously reported that memantine, uncompetitive NMDA channel blocker, reduced RGC loss with a decrease in GluN1 subunit expression in the eyes of rats with chronic ocular hypertension model (Kim et al., 2007b). In the current study, we focused on whether α2-adrenoceptor activation could modulate glutamate transporters and NMDA receptors in a chronic ocular hypertension model.