All Title Author
Keywords Abstract

PLOS ONE  2013 

Impairment of Neuronal Glutamate Uptake and Modulation of the Glutamate Transporter GLT-1 Induced by Retinal Ischemia

DOI: 10.1371/journal.pone.0069250

Full-Text   Cite this paper   Add to My Lib


Excitotoxicity has been implicated in the retinal neuronal loss in several ocular pathologies including glaucoma. Dysfunction of Excitatory Amino Acid Transporters is often a key component of the cascade leading to excitotoxic cell death. In the retina, glutamate transport is mainly operated by the glial glutamate transporter GLAST and the neuronal transporter GLT-1. In this study we evaluated the expression of GLAST and GLT-1 in a rat model of acute glaucoma based on the transient increase of intraocular pressure (IOP) and characterized by high glutamate levels during the reperfusion that follows the ischemic event associated with raised IOP. No changes were reported in GLAST expression while, at neuronal level, a reduction of glutamate uptake and of transporter reversal-mediated glutamate release was observed in isolated retinal synaptosomes. This was accompanied by modulation of GLT-1 expression leading to the reduction of the canonical 65 kDa form and upregulation of a GLT-1-related 38 kDa protein. These results support a role for neuronal transporters in glutamate accumulation observed in the retina following an ischemic event and suggest the presence of a GLT-1 neuronal new alternative splice variant, induced in response to the detrimental stimulus.


[1]  Pow DV (2001) Amino acids and their transporters in the retina. Neurochem Int 38: 463–484.
[2]  Lam DM (1997) Neurotransmitters in the vertebrate retina. Invest Ophthalmol Vis Sci 38: 553–556.
[3]  Danbolt NC (2001) Glutamate uptake. Prog Neurobiol 65: 1–105.
[4]  Bringmann A, Pannicke T, Biedermann B, Francke M, Iandiev I, et al. (2009) Role of retinal glial cells in neurotransmitter uptake and metabolism. Neurochem Int 54: 143–160.
[5]  Rauen T, Wiessner M (2000) Fine tuning of glutamate uptake and degradation in glial cells: common transcriptional regulation of GLAST1 and GS. Neurochem Int 37: 179–189.
[6]  Nicholls D, Attwell D (1990) The release and uptake of excitatory amino acids. Trends Pharmacol Sci 11: 462–468.
[7]  Siliprandi R, Canella R, Carmignoto G, Schiavo N, Zanellato A, et al. (1992) N-methyl-D-aspartate-induced neurotoxicity in the adult rat retina. Vis Neurosci 8: 567–573.
[8]  Rothstein JD, Dykes-Hoberg M, Pardo CA, Bristol LA, Jin L, et al. (1996) Knockout of glutamate transporters reveals a major role for astroglial transport in excitotoxicity and clearance of glutamate. Neuron 16: 675–686.
[9]  Liu CH, Jiao H, Guo ZH, Peng Y, Wang WZ (2013) Up-regulated GLT-1 resists glutamate toxicity and attenuates glutamate-induced calcium loading in cultured neurocytes. Basic Clin Pharmacol Toxicol 112: 19–24.
[10]  Vorwerk CK, Naskar R, Schuettauf F, Quinto K, Zurakowski D, et al. (2000) Depression of retinal glutamate transporter function leads to elevated intravitreal glutamate levels and ganglion cell death. Invest Ophthalmol Vis Sci 41: 3615–3621.
[11]  Adachi K, Kashii S, Masai H, Ueda M, Morizane C, et al. (1998) Mechanism of the pathogenesis of glutamate neurotoxicity in retinal ischemia. Graefes Arch Clin Exp Ophthalmol 236: 766–774.
[12]  Louzada-Junior P, Dias JJ, Santos WF, Lachat JJ, Bradford HF, et al. (1992) Glutamate release in experimental ischaemia of the retina: an approach using microdialysis. J Neurochem 59: 358–363.
[13]  Nucci C, Tartaglione R, Rombola L, Morrone LA, Fazzi E, et al. (2005) Neurochemical evidence to implicate elevated glutamate in the mechanisms of high intraocular pressure (IOP)-induced retinal ganglion cell death in rat. Neurotoxicology 26: 935–941.
[14]  Russo R, Rotiroti D, Tassorelli C, Nucci C, Bagetta G, et al. (2009) Identification of novel pharmacological targets to minimize excitotoxic retinal damage. Int Rev Neurobiol 85: 407–423.
[15]  Qu J, Wang D, Grosskreutz CL (2010) Mechanisms of retinal ganglion cell injury and defense in glaucoma. Exp Eye Res 91: 48–53.
[16]  Kowluru RA, Zhong Q (2011) Beyond AREDS: is there a place for antioxidant therapy in the prevention/treatment of eye disease? Invest Ophthalmol Vis Sci 52: 8665–8671.
[17]  Li SY, Fu ZJ, Lo AC (2012) Hypoxia-induced oxidative stress in ischemic retinopathy. Oxid Med Cell Longev 2012: 426769.
[18]  Russo R, Cavaliere F, Berliocchi L, Nucci C, Gliozzi M, et al. (2008) Modulation of pro-survival and death-associated pathways under retinal ischemia/reperfusion: effects of NMDA receptor blockade. J Neurochem 107: 1347–1357.
[19]  Sucher NJ, Lipton SA, Dreyer EB (1997) Molecular basis of glutamate toxicity in retinal ganglion cells. Vision Res 37: 3483–3493.
[20]  Hare WA, WoldeMussie E, Weinreb RN, Ton H, Ruiz G, et al. (2004) Efficacy and safety of memantine treatment for reduction of changes associated with experimental glaucoma in monkey, II: Structural measures. Invest Ophthalmol Vis Sci 45: 2640–2651.
[21]  WoldeMussie E, Yoles E, Schwartz M, Ruiz G, Wheeler LA (2002) Neuroprotective effect of memantine in different retinal injury models in rats. J Glaucoma 11: 474–480.
[22]  Beart PM, O'Shea RD (2007) Transporters for L-glutamate: an update on their molecular pharmacology and pathological involvement. Br J Pharmacol 150: 5–17.
[23]  Gillessen T, Budd SL, Lipton SA (2002) Excitatory amino acid neurotoxicity. Adv Exp Med Biol 513: 3–40.
[24]  Martin KR, Levkovitch-Verbin H, Valenta D, Baumrind L, Pease ME, et al. (2002) Retinal glutamate transporter changes in experimental glaucoma and after optic nerve transection in the rat. Invest Ophthalmol Vis Sci 43: 2236–2243.
[25]  Naskar R, Vorwerk CK, Dreyer EB (2000) Concurrent downregulation of a glutamate transporter and receptor in glaucoma. Invest Ophthalmol Vis Sci 41: 1940–1944.
[26]  Schuettauf F, Thaler S, Bolz S, Fries J, Kalbacher H, et al. (2007) Alterations of amino acids and glutamate transport in the DBA/2J mouse retina; possible clues to degeneration. Graefes Arch Clin Exp Ophthalmol 245: 1157–1168.
[27]  Russo R, Cavaliere F, Watanabe C, Nucci C, Bagetta G, et al. (2008) 17Beta-estradiol prevents retinal ganglion cell loss induced by acute rise of intraocular pressure in rat. Prog Brain Res 173: 583–590.
[28]  Milanese M, Zappettini S, Onofri F, Musazzi L, Tardito D, et al. (2011) Abnormal exocytotic release of glutamate in a mouse model of amyotrophic lateral sclerosis. J Neurochem 116: 1028–1042.
[29]  Dunkley PR, Heath JW, Harrison SM, Jarvie PE, Glenfield PJ, et al. (1988) A rapid Percoll gradient procedure for isolation of synaptosomes directly from an S1 fraction: homogeneity and morphology of subcellular fractions. Brain Res 441: 59–71.
[30]  Raiteri M, Bonanno G, Marchi M, Maura G (1984) Is there a functional linkage between neurotransmitter uptake mechanisms and presynaptic receptors? J Pharmacol Exp Ther 231: 671–677.
[31]  Iuvone PM, Brown AD, Haque R, Weller J, Zawilska JB, et al. (2002) Retinal melatonin production: role of proteasomal proteolysis in circadian and photic control of arylalkylamine N-acetyltransferase. Invest Ophthalmol Vis Sci 43: 564–572.
[32]  Fernandez DC, Bordone MP, Chianelli MS, Rosenstein RE (2009) Retinal neuroprotection against ischemia-reperfusion damage induced by postconditioning. Invest Ophthalmol Vis Sci 50: 3922–3930.
[33]  Barnett NL, Pow DV, Bull ND (2001) Differential perturbation of neuronal and glial glutamate transport systems in retinal ischaemia. Neurochem Int 39: 291–299.
[34]  Rauen T, Rothstein JD, Wassle H (1996) Differential expression of three glutamate transporter subtypes in the rat retina. Cell Tissue Res 286: 325–336.
[35]  Pow DV, Barnett NL (1999) Changing patterns of spatial buffering of glutamate in developing rat retinae are mediated by the Muller cell glutamate transporter GLAST. Cell Tissue Res 297: 57–66.
[36]  Rauen T (2000) Diversity of glutamate transporter expression and function in the mammalian retina. Amino Acids 19: 53–62.
[37]  Rauen T, Kanner BI (1994) Localization of the glutamate transporter GLT-1 in rat and macaque monkey retinae. Neurosci Lett 169: 137–140.
[38]  Chidlow G, Osborne NN (2003) Rat retinal ganglion cell loss caused by kainate, NMDA and ischemia correlates with a reduction in mRNA and protein of Thy-1 and neurofilament light. Brain Res 963: 298–306.
[39]  Dreyer EB, Zurakowski D, Schumer RA, Podos SM, Lipton SA (1996) Elevated glutamate levels in the vitreous body of humans and monkeys with glaucoma. Arch Ophthalmol 114: 299–305.
[40]  Harada T, Harada C, Watanabe M, Inoue Y, Sakagawa T, et al. (1998) Functions of the two glutamate transporters GLAST and GLT-1 in the retina. Proc Natl Acad Sci U S A 95: 4663–4666.
[41]  Barnett NL, Pow DV (2000) Antisense knockdown of GLAST, a glial glutamate transporter, compromises retinal function. Invest Ophthalmol Vis Sci 41: 585–591.
[42]  Sullivan RK, Woldemussie E, Macnab L, Ruiz G, Pow DV (2006) Evoked expression of the glutamate transporter GLT-1c in retinal ganglion cells in human glaucoma and in a rat model. Invest Ophthalmol Vis Sci 47: 3853–3859.
[43]  Park CK, Cha J, Park SC, Lee PY, Kim JH, et al. (2009) Differential expression of two glutamate transporters, GLAST and GLT-1, in an experimental rat model of glaucoma. Exp Brain Res 197: 101–109.
[44]  Napper GA, Pianta MJ, Kalloniatis M (1999) Reduced glutamate uptake by retinal glial cells under ischemic/hypoxic conditions. Vis Neurosci 16: 149–158.
[45]  Quigley HA (1999) Neuronal death in glaucoma. Prog Retin Eye Res 18: 39–57.
[46]  Rao VL, Bowen KK, Dempsey RJ (2001) Transient focal cerebral ischemia down-regulates glutamate transporters GLT-1 and EAAC1 expression in rat brain. Neurochem Res 26: 497–502.
[47]  Yeh TH, Hwang HM, Chen JJ, Wu T, Li AH, et al. (2005) Glutamate transporter function of rat hippocampal astrocytes is impaired following the global ischemia. Neurobiol Dis 18: 476–483.
[48]  Pow DV, Naidoo T, Lingwood BE, Healy GN, Williams SM, et al. (2004) Loss of glial glutamate transporters and induction of neuronal expression of GLT-1B in the hypoxic neonatal pig brain. Brain Res Dev Brain Res 153: 1–11.
[49]  Furuya T, Pan Z, Kashiwagi K (2012) Role of retinal glial cell glutamate transporters in retinal ganglion cell survival following stimulation of NMDA receptor. Curr Eye Res 37: 170–178.
[50]  Osborne NN, Casson RJ, Wood JP, Chidlow G, Graham M, et al. (2004) Retinal ischemia: mechanisms of damage and potential therapeutic strategies. Prog Retin Eye Res 23: 91–147.
[51]  Russo R, Berliocchi L, Adornetto A, Varano GP, Cavaliere F, et al. (2011) Calpain-mediated cleavage of Beclin-1 and autophagy deregulation following retinal ischemic injury in vivo. Cell Death Dis 2: e144.
[52]  Lee A, Anderson AR, Stevens MG, Pow DV (2011) Exon 4-skipping GLT-1: a new form of an abundantly expressed glutamate transporter. Neurosci Lett 504: 228–231.
[53]  Walton HS, Pow DV, Innes DJ, Dodd PR (2005) Proteomic analysis of excitatory amino acid transporter 2 isoforms in Alzheimer's disease. J Neurochem P.371.
[54]  Munch C, Ebstein M, Seefried U, Zhu B, Stamm S, et al. (2002) Alternative splicing of the 5′-sequences of the mouse EAAT2 glutamate transporter and expression in a transgenic model for amyotrophic lateral sclerosis. J Neurochem 82: 594–603.
[55]  Utsunomiya-Tate N, Endou H, Kanai Y (1997) Tissue specific variants of glutamate transporter GLT-1. FEBS Lett 416: 312–316.
[56]  Honig LS, Chambliss DD, Bigio EH, Carroll SL, Elliott JL (2000) Glutamate transporter EAAT2 splice variants occur not only in ALS, but also in AD and controls. Neurology 55: 1082–1088.
[57]  Meyer T, Munch C, Knappenberger B, Liebau S, Volkel H, et al. (1998) Alternative splicing of the glutamate transporter EAAT2 (GLT-1). Neurosci Lett 241: 68–70.
[58]  Scott HA, Gebhardt FM, Mitrovic AD, Vandenberg RJ, Dodd PR (2011) Glutamate transporter variants reduce glutamate uptake in Alzheimer's disease. Neurobiol Aging 32: 553 e551–511.
[59]  Maragakis NJ, Dykes-Hoberg M, Rothstein JD (2004) Altered expression of the glutamate transporter EAAT2b in neurological disease. Ann Neurol 55: 469–477.
[60]  Rauen T, Wiessner M, Sullivan R, Lee A, Pow DV (2004) A new GLT1 splice variant: cloning and immunolocalization of GLT1c in the mammalian retina and brain. Neurochem Int 45: 1095–1106.
[61]  Chen W, Aoki C, Mahadomrongkul V, Gruber CE, Wang GJ, et al. (2002) Expression of a variant form of the glutamate transporter GLT1 in neuronal cultures and in neurons and astrocytes in the rat brain. J Neurosci 22: 2142–2152.
[62]  Schmitt A, Asan E, Lesch KP, Kugler P (2002) A splice variant of glutamate transporter GLT1/EAAT2 expressed in neurons: cloning and localization in rat nervous system. Neuroscience 109: 45–61.


comments powered by Disqus