Potential role of A 2A adenosine receptor in traumatic optic neuropathy Saif Ahmad a,b, ⁎, Nadeem Fatteh a , Nehal M. El-Sherbiny a,c , Mohammad Naime a,1 , Ahmed S. Ibrahim c , Ahmed M. El-Sherbini a , Sally A. El-Shafey a , Sohail Khan d , Sadanand Fulzele e , Joyce Gonzales f , Gregory I. Liou a, ⁎ a Department of Ophthalmology, Georgia Regents University (GRU), Augusta, GA, USA b Departmet of Biological Sciences, College of Science and Arts, King Abdulaziz University, Rabigh, Saudi Arabia c Department of Biochemistry, Faculty of Pharmacy, Mansoura University, Mansoura, Egypt d South Western Medical Center, Dallas, TX, USA e Department of Orthopedics, Georgia Regents University (GRU), Augusta, GA, USA f Vascular Biology Center, Georgia Regents University (GRU), Augusta, GA, USA abstract article info Article history: Received 30 June 2013 Received in revised form 12 September 2013 Accepted 14 September 2013 Keywords: Traumatic optic neuropathy Adenosine A 2A receptor Microglia Inflammatory cytokines MAPKinase Oxidative stress In traumatic optic neuropathy (TON), apoptosis of retinal ganglion cells is closely related to the local production of reactive oxygen species and inflammatory mediators from activated microglial cells. Adenosine receptor A 2A (A 2A AR) has been shown to possess anti-inflammatory properties that have not been studied in TON. In the present study, we examined the role of A 2A AR in retinal complications associated with TON. Initial studies in wild-type mice revealed that treatment with the A 2A AR agonist resulted in marked decreases in the TON- induced microglial activation, retinal cell death and releases of reactive oxygen species and pro-inflammatory cytokines TNF-α and IL-6. To further assess the role of A 2A AR in TON, we studied the effects of A 2A AR ablation on the TON-induced retinal abnormalities. A 2A AR-/- mice with TON showed a significantly higher mRNA level of TNF-α, Iba1-1 in retinal tissue, and ICAM-1 expression in retinal sections compared with wild-type mice with TON. To explore a potential mechanism by which A 2A AR-signaling regulates inflammation in TON, we performed additional studies using hypoxia- or LPS-treated microglial cells as an in vitro model for TON. Activation of A 2A AR attenuates hypoxia or LPS-induced TNF-α release and significantly repressed the inflammatory signaling, ERK in the activated microglia. Collectively, this work provides pharmacological and genetic evidence for A 2A AR signaling as a control point of cell death in TON and suggests that the retinal protective effect of A 2A AR is mediated by atten- uating the inflammatory response that occurs in microglia via interaction with MAPKinase pathway. © 2013 Elsevier B.V. All rights reserved. 1. Introduction Traumatic optic nerve injury is commonly seen in motor vehicle acci- dents, assaults, war and in the natural disaster. Traumatic optic nerve in- jury is usually the consequence of a severe blunt head trauma, often a frontal blow severe enough to cause loss of consciousness. Unfortunately, there are currently no proven treatments that can prevent the damage associated with an acute traumatic optic neuropathy (TON). Prognosis for the recovery of vision in TON is still poor, nevertheless, animal models for TON are often used, mostly because they are easy to perform and can be well standardized (Levkovitch-Verbin, 2004). Retinal ganglion cell (RGC) death is known to be a fundamental pathological process in trau- matic optic injury including TON. Several common mechanisms have been hypothesized to underlie apoptotic processes, including interrup- tion of trophic support, oxidative stress, and increased extracellular glu- tamate levels that result in excitotoxicity. These stimuli associated with the injured RGCs often activate retinal microglia, which release pro- inflammatory cytokines and cytotoxic molecules to further exacerbate the degenerative process (Kreutzberg, 1996). These findings suggest that pharmacological interventions that reduce inflammation may be effective neuroprotectants for TON. Under stress and ischemic conditions, the local tissue concentration of extracellular adenosine is increased due to its synthesis from the re- leased ATP. This nucleoside has been proposed to modulate a variety of physiological responses by stimulating specific adenosine receptors (AR), which are classified as A1, A2A, A2B, and A3 subtypes (Collis and Hourani, 1993). These receptors can be distinguished based on their affinities for adenosine agonists and antagonists. In addition, these receptors are classified based on their mechanism of signal Journal of Neuroimmunology 264 (2013) 54–64 Abbreviations: TON, traumatic optic neuropathy; TNF-α, tumor necrosis factor-α; ELISA, Enzyme-linked immunosorbent assay; ROS, reactive oxygen species; MAP kinase, Mitogen- activated protein kinase; ERK, extracellular signal-regulated kinase; AR, adenosine receptor; CGS21680, 2-p-[2-Carboxyethyl]phenethylamino-5′-N-ethylcarboxamidoadenosine; LPS, lipopolysaccharides; NECA, 5′-N-Ethylcarboxamidoadenosine; ZM241385, 4-(2-[7-Amino- 2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol. ⁎ Corresponding authors at: Department of Ophthalmology, Georgia Regents University, 1120 15th Street, Augusta, GA 30912, USA. Tel.: +1 706 721 4599; fax: +1 706 721 1158. E-mail addresses: sahmad@gru.edu, asaif77@yahoo.com (S. Ahmad), giliou@gru.edu (G.I. Liou). 1 Biochemistry Lab, Regional Research Institute of Unani Medicine, CCRUM, Srinagar, J&K-190006, India. 0165-5728/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jneuroim.2013.09.015 Contents lists available at ScienceDirect Journal of Neuroimmunology journal homepage: www.elsevier.com/locate/jneuroim