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Expression of the cystine-glutamate exchanger (xc −) in retinal ganglion cells and regulation by nitric oxide and oxidative stress

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Abstract

The cystine-glutamate exchanger, system xc −, mediates the Na+-independent exchange of cystine into cells, coupled to the efflux of intracellular glutamate. System xc − plays a critical role in glutathione homeostasis. Early studies of brain suggested that system xc − was present primarily in astrocytes but not neurons. More recent work indicates that certain brain neurons have an active system xc −. In the retina, system xc − has been demonstrated in Müller and retinal pigment epithelial cells. We have recently suggested that two protein components of system xc −, xCT and 4F2hc, are present in ganglion cells of the intact retina. Here, we have used (1) molecular and immunohistochemical assays to determine whether system xc − is present in primary ganglion cells isolated from neonatal mouse retinas and (2) functional assays to determine whether its activity is regulated by oxidative stress in a retinal ganglion cell line (RGC–5). Primary mouse ganglion cells and RGC–5 cells express xCT and 4F2hc. RGC–5 cells take up [3H]glutamate in the absence of Na+, and this uptake is blocked by known substrates of system xc − (glutamate, cysteine, cystine, quisqualic acid). Treatment of RGC–5 cells with NO and reactive oxygen species donors leads to increased activity of system xc − associated with an increase in the maximal velocity of the transporter with no significant change in the substrate affinity. This is the first report of system xc − in primary retinal ganglion cells and RGC–5 cells. Oxidative stress upregulates this transport system in RGC–5 cells, and the process is associated with an increase in xCT mRNA and protein but no change in 4F2hc mRNA or protein.

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References

  • Allen JW, Shanker G, Aschner MJW (2001) Methylmercury inhibits the in vitro uptake of the glutathione precursor, cystine, in astrocytes, but not in neurons. Brain Res 894:131–140

    Article  PubMed  CAS  Google Scholar 

  • Barber AJ (2003) A new view of diabetic retinopathy: a neurodegenerative disease of the eye. Prog Neuropsychopharmacol Biol Psychiatry 27:283–290

    Article  PubMed  CAS  Google Scholar 

  • Barber AJ, Antonetti DA, Kern TS, Reiter CE, Soans RS, Krady JK, Levison SW, Gardner TW, Bronson SK (2005) The Ins2Akita mouse as a model of early retinal complications in diabetes. Invest Ophthalmol Vis Sci 46:2210–2218

    Article  PubMed  Google Scholar 

  • Barres BA, Silverstein BE, Corey DP, Chun LL (1988) Immunological morphological and electrophysiological variation among retinal ganglion cells purified by panning. Neuron 19:9791–9803

    Google Scholar 

  • Bonfoco E, Leist M, Zhivotovsky B, Orrenius S, Lipton SA, Nicotera P (1996) Cytoskeletal breakdown and apoptosis elicited by NO donors in cerebellar granule cells require NMDA receptor activation. J Neurochem 67:2484–2493

    Article  PubMed  CAS  Google Scholar 

  • Boyd ZS, Kriatchko A, Yang J, Agarwal N, Wax MB, Patil RV (2003) Interleukin–10 receptor signaling through STAT–3 regulates the apoptosis of retinal ganglion cells in response to stress. Invest Ophthalmol Vis Sci 44:5206–5211

    Article  PubMed  Google Scholar 

  • Bridges CC, Kekuda R, Wang H, Prasad PD, Mehta P, Huang W, Smith SB, Ganapathy V (2001) Structure, function, and regulation of human cystine/glutamate transporter in retinal pigment epithelial cells. Invest Ophthalmol Vis Sci 42:47–54

    PubMed  CAS  Google Scholar 

  • Bridges CC, Hu H, Miyauchi S, Siddaramappa UN, Ganapathy ME, Ignatowicz L, Maddox DM, Smith SB, Ganapathy V (2004) Induction of cystine–glutamate transporter xc – by human immunodeficiency virus type 1 transactivator protein tat in retinal pigment epithelium. Invest Ophthalmol Vis Sci 45:2906–2914

    Article  PubMed  Google Scholar 

  • Brzezinski JA, Brown NL, Tanikawa A, Bush RA, Sieving PA, Vitaterna MH, Takahashi JS, Glaser T (2005) Loss of circadian photoentrainment and abnormal retinal electrophysiology in Math5 mutant mice. Invest Ophthalmol Vis Sci 46:2540–2551

    Article  PubMed  Google Scholar 

  • Chase LA, Roon RJ, Wellman L, Beitz AJ, Koerner JF (2001) L–Quisqualic acid transport into hippocampal neurons by a cystine–sensitive carrier is required for the induction of quisqualate sensitization. Neuroscience 106:287–301

    Article  PubMed  CAS  Google Scholar 

  • Clementi E (1998) Role of nitric oxide and its intracellular signalling pathways in the control of Ca2+ homeostasis. Biochem Pharmacol 55:713–718

    Article  PubMed  CAS  Google Scholar 

  • Danbolt NC (2001) Glutamate uptake. Prog Neurobiol 65:1–105

    Article  PubMed  CAS  Google Scholar 

  • Du Y, Sarthy VP, Kern TS (2004) Interaction between NO and COX pathways in retinal cells exposed to elevated glucose and retina of diabetic rats. Am J Physiol Regul Integr Comp Physiol 287:R735–R741

    PubMed  CAS  Google Scholar 

  • Gardner TW, Antonetti DA, Barber AJ, LaNoue KF, Levison SW (2002) Diabetic retinopathy: more than meets the eye. Surv Ophthalmol 47 (Suppl 2):S253–S262

    Article  PubMed  Google Scholar 

  • Goldstein IM, Ostwald P, Roth S (1996) Nitric oxide: a review of its role in retinal function and disease. Vision Res 36:2979–2994

    Article  PubMed  CAS  Google Scholar 

  • Gukasyan HJ, Kannan R, Lee VH, Kim KJ (2003) Regulation of L–cystine transport and intracellular GSH level by a nitric oxide donor in primary cultured rabbit conjunctival epithelial cell layers. Invest Ophthalmol Vis Sci 44:1202–1210

    Article  PubMed  Google Scholar 

  • Hosoya K, Tomi M, Ohtsuki S, Hosoya K, Tomi M, Ohtsuki S, Takanaga H, Saeki S, Kanai Y, Endou H, Naito M, Tsuruo T, Terasaki T (2002) Enhancement of L–cystine transport activity and its relation to xCT gene induction at the blood–brain barrier by diethyl maleate treatment. J Pharmacol Exp Ther 302:225–231

    Article  PubMed  CAS  Google Scholar 

  • Kashii S, Mandai M, Kikuchi M, Honda Y, Tamura Y, Kaneda K, Akaike A (1996) Dual actions of nitric oxide in N–methyl–D–aspartate receptor–mediated neurotoxicity in cultured retinal neurons. Brain Res 711:93–101

    Article  PubMed  CAS  Google Scholar 

  • Kato S, Ishita S, Sugawara K, Mawatari K (1993) Cystine/glutamate antiporter expression in retinal Müller glial cells: implications for DL–alpha–aminoadipate toxicity. Neuroscience 57:473–482

    Article  PubMed  CAS  Google Scholar 

  • Kortuem K, Geiger LK, Levin LA (2000) Differential susceptibility of retinal ganglion cells to reactive oxygen species. Invest Ophthalmol Vis Sci 41:3176–3182

    PubMed  CAS  Google Scholar 

  • Krishnamoorthy RR, Agarwal P, Prasanna G, Vopat K, Lambert W, Sheedlo HJ, Pang IH, Shade D, Wordinger RJ, Yorio T, Clark AF, Agarwal N (2001) Characterization of a transformed rat retinal ganglion cell line. Brain Res Mol Brain Res 86:1–12

    Article  PubMed  CAS  Google Scholar 

  • Leist M, Volbracht C, Kuhnle S, Fava E, Ferrando–May E, Nicotera P (1997) Caspase–mediated apoptosis in neuronal excitotoxicity triggered by nitric oxide. Mol Med 3:750–764

    PubMed  CAS  Google Scholar 

  • Levin LA, Gordon LK (2002) Retinal ganglion cell disorders: types and treatments. Prog Retina Eye Res 21:465–484

    Article  CAS  Google Scholar 

  • Li Q, Puro DG (2002) Diabetes–induced dysfunction of the glutamate transporter in retinal Muller cells. Invest Ophthalmol Vis Sci 43:3109–3116

    PubMed  Google Scholar 

  • Martin PM, Roon P, Van Ells TK, Ganapathy V, Smith SB (2004) Death of retinal neurons in streptozotocin–induced diabetic mice. Invest Ophthalmol Vis Sci 45:3330–3336

    Article  PubMed  Google Scholar 

  • McBean GJ (2002) Cerebral cystine uptake: a tale of two transporters. Trends Pharmacol Sci 23:299–302

    Article  PubMed  CAS  Google Scholar 

  • Moreno MC, Campanelli J, Sande P, Sanez DA, Keller Sarmiento MI, Rosenstein RE (2004) Retinal oxidative stress induced by high intraocular pressure. Free Radic Biol Med 37:803–812

    Article  PubMed  CAS  Google Scholar 

  • Murphy TH, Schnaar RL, Coyle JT (1990) Immature cortical neurons are uniquely sensitive to glutamate toxicity by inhibition of cystine uptake. FASEB J4:1624–1633

    Google Scholar 

  • Palacin M, Estevez R, Bertran J, Zorzano A (1998) Molecular biology of mammalian plasma membrane amino acid transporters. Physiol Rev 78:969–1054

    PubMed  CAS  Google Scholar 

  • Pow DV (2001) Visualising the activity of the cystine–glutamate antiporter in glial cells using antibodies to aminoadipic acid, a selectively transported substrate. Glia 34:27–38

    Article  PubMed  CAS  Google Scholar 

  • Pow DV, Crook DK (1995) Immunocytochemical evidence for the presence of high levels of reduced glutathione in radial glial cells and horizontal cells in the rabbit retina. Neurosci Lett 193(1):25–28

    Article  PubMed  CAS  Google Scholar 

  • Sagara J, Miura K, Bannai S (1993a) Cystine uptake and glutathione level in fetal brain cells in primary culture and in suspension. J Neurochem 61:1667–1671

    Article  PubMed  CAS  Google Scholar 

  • Sagara J, Miura K, Bannai S (1993b) Maintenance of neuronal glutathione by glial cells. J Neurochem 61:1672–1676

    Article  PubMed  CAS  Google Scholar 

  • Sagara Y, Schubert D (1998) The activation of metabotropic glutamate receptors protects nerve cells from oxidative stress. J Neurosci 18:6662–6671

    PubMed  CAS  Google Scholar 

  • Sasaki H, Sato H, Kuriyama–Matsumura K, Sato K, Maebara K, Wang H, Tamba M, Itoh K, Yamamoto M, Bannai S (2002) Electrophile response element–mediated induction of the cystine/glutamate exchange transporter gene expression. J Biol Chem 277:44765–44771

    Article  PubMed  CAS  Google Scholar 

  • Sato H, Tamba M, Ishii T, Bannai S (1999) Cloning and expression of a plasma membrane cystine/glutamate exchange transporter composed of two distinct proteins. J Biol Chem 274:11455–11458

    Article  PubMed  CAS  Google Scholar 

  • Sato H, Kuriyama–Matsumura K, Hashimoto T, Sasaki H, Wang H, Ishii T, Mann GE, Bannai S (2001) Effect of oxygen on induction of the cystine transporter by bacterial lipopolysaccharide in mouse peritoneal macrophages. J Biol Chem 276:10407–10412

    Article  PubMed  CAS  Google Scholar 

  • Schmetterer L, Findl O, Fasching P, Ferber W, Strenn K, Breitendeder H, Adam H, Eichler HG, Wolzt M (1997) Nitric oxide and ocular blood flow in patients with IDDM. Diabetes 46:653–658

    Article  PubMed  CAS  Google Scholar 

  • Shanker G, Allen JW, Mutkus LA, Aschner M (2001) The uptake of cysteine in cultured primary astrocytes and neurons. Brain Res 902:156–163

    Article  PubMed  CAS  Google Scholar 

  • Sucher NJ, Lipton SA, Dreyer EB (1997) Molecular basis of glutamate toxicity in retinal ganglion cells. Vis Res 37:3483–3493

    Article  PubMed  CAS  Google Scholar 

  • Tomi M, Funaki T, Abukawa H, Katayama K, Kondo T, Ohtsuki S, Ueda M, Obinata M, Terasaki T, Hosoya K (2003) Expression and regulation of L–cystine transporter, system xc –, in the newly developed rat retinal Muller cell line (TR–MUL). Glia 43:208–217

    Article  PubMed  Google Scholar 

  • Tomi M, Hosoya K, Takanaga H, Ohtsuki S, Terasaki T (2002) Induction of xCT gene expression and L–cystine transport activity by diethyl maleate at the inner blood–retinal barrier. Invest Ophthalmol Vis Sci 43:774–779

    PubMed  Google Scholar 

  • Verrey F, Meier C, Rossier G, Kuhn LC (2000) Glycoprotein–associated amino acid exchangers: broadening the range of transport specificity. Eur J Physiol 440:503–512

    Article  CAS  Google Scholar 

  • Warr O, Takahashi M, Attwell D (1999) Modulation of extracellular glutamate concentration in rat brain slices by cystine–glutamate exchange. J Physiol (Lond) 514:783–793

    Article  CAS  Google Scholar 

  • Yilmaz G, Esser P, Kociek N, Aydin P, Heimann K (2000) Elevated vitreous nitric oxide levels in patients with proliferative diabetic retinopathy. Am J Ophthalmol 130:87–90

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

We are grateful to Dr. Neeraj Agarwal of the University of North Texas Health Science Center for the kind gift of the RGC–5 cell line, to our colleagues, Dr. Zheng Dong and Dr. Darren Browning, at the Medical College of Georgia for the generous use of their equipment, and to Dr. Lin Gan, University of Rochester, for helpful suggestions in using the protocol for culturing primary ganglion cells from mice.

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Authors and Affiliations

  1. Department of Cellular Biology and Anatomy, Medical College of Georgia, 1459 Laney-Walker Blvd., CB 2820, Augusta, GA 30912-2000, USA

    Y. Dun, B. Mysona, T. Van Ells, L. Amarnath, M. Shamsul Ola & S. B. Smith

  2. Department of Biochemistry and Molecular Biology, Medical College of Georgia, Augusta, USA

    V. Ganapathy

  3. Department of Cellular Biology/Anatomy and Ophthalmology, Medical College of Georgia, Augusta, USA

    S. B. Smith

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  1. Y. Dun
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  2. B. Mysona
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  3. T. Van Ells
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  5. M. Shamsul Ola
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  6. V. Ganapathy
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  7. S. B. Smith
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Correspondence to S. B. Smith.

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This work was supported by National Institutes of Health grants EY014560 and EY012830.

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Dun, Y., Mysona, B., Van Ells, T. et al. Expression of the cystine-glutamate exchanger (xc −) in retinal ganglion cells and regulation by nitric oxide and oxidative stress. Cell Tissue Res 324, 189–202 (2006). https://doi.org/10.1007/s00441-005-0116-x

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  • DOI: https://doi.org/10.1007/s00441-005-0116-x

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