Cellular and Molecular Neurobiology

, Volume 29, Issue 8, pp 1233–1244 | Cite as

Polysaccharides from Wolfberry Antagonizes Glutamate Excitotoxicity in Rat Cortical Neurons

  • Yuen-Shan Ho
  • Man-Shan Yu
  • Suet-Yi Yik
  • Kwok-Fai So
  • Wai-Hung Yuen
  • Raymond Chuen-Chung ChangEmail author
Original Paper


Glutamate excitotoxicity is involved in many neurodegenerative diseases including Alzheimer’s disease (AD). Attenuation of glutamate toxicity is one of the therapeutic strategies for AD. Wolfberry (Lycium barbarum) is a common ingredient in oriental cuisines. A number of studies suggest that wolfberry has anti-aging properties. In recent years, there is a trend of using dried Wolfberry as food supplement and health product in UK and North America. Previously, we have demonstrated that a fraction of polysaccharide from Wolfberry (LBA) provided remarkable neuroprotective effects against beta-amyloid peptide-induced cytotoxicity in primary cultures of rat cortical neurons. To investigate whether LBA can protect neurons from other pathological factors such as glutamate found in Alzheimer brain, we examined whether it can prevent neurotoxicity elicited by glutamate in primary cultured neurons. The glutamate-induced cell death as detected by lactate dehydrogenase assay and caspase-3-like activity assay was significantly reduced by LBA at concentrations ranging from 10 to 500 μg/ml. Protective effects of LBA were comparable to memantine, a non-competitive NMDA receptor antagonist. LBA provided neuroprotection even 1 h after exposure to glutamate. In addition to glutamate, LBA attenuated N-methyl-d-aspartate (NMDA)-induced neuronal damage. To further explore whether LBA might function as antioxidant, we used hydrogen peroxide (H2O2) as oxidative stress inducer in this study. LBA could not attenuate the toxicity of H2O2. Furthermore, LBA did not attenuate glutamate-induced oxidation by using NBT assay. Western blot analysis indicated that glutamate-induced phosphorylation of c-jun N-terminal kinase (JNK) was reduced by treatment with LBA. Taken together, LBA exerted significant neuroprotective effects on cultured cortical neurons exposed to glutamate.


Wolfberry Neuroprotection Glutamate Excitotoxicity c-Jun N-terminal kinase 



The authors would like to thank Professor J. N. Fang for his help in providing LBA, Miss Michelle Huie for critical reading of the manuscript. This work is supported by the HKU Alzheimer’s Disease Research Network, General Research Grant (7552/06 M) and NSFC/RGC Joint Research Scheme (N_HKU707/07M) from Research Grant Council, and HKU Seed Funding for Basic Research (200811159082) to RCCC. Also, the work is supported by Azalea (1972) Endowment Fund. WHY would like to thank for the support from the Department of Chemistry. YSH is supported by the Graduate School, MSY is supported by Postdoctoral Fellowship, The University of Hong Kong.


  1. Abib RT, Quincozes-Santos A, Nardin P, Wofchuk ST, Perry ML, Gonçalves CA, Gottfried C (2008) Epicatechin gallate increases glutamate uptake and S100B secretion in C6 cell lineage. Mol Cell Biochem 310:153–158. doi: 10.1007/s11010-007-9675-3 CrossRefPubMedGoogle Scholar
  2. Amagase H, Nance DM (2008) A randomized, double-blind, placebo-controlled, clinical study of the general effects of a standardized Lycium barbarum (Goji) juice, GoChi. J Altern Complement Med 14(4):403–412CrossRefPubMedGoogle Scholar
  3. Amodio R, Esposito E, De RC, Bellavia V, Amodio E, Carruba G (2006) Red wine extract prevents neuronal apoptosis in vitro and reduces mortality of transgenic mice. Ann NY Acad Sci 1089:88–97. doi: 10.1196/annals.1386.026 CrossRefPubMedGoogle Scholar
  4. Arthur PG, Matich GP, Pang WW, Yu DY, Bogoyevitch MA (2007) Necrotic death of neurons following an excitotoxic insult is prevented by a peptide inhibitor of c-jun N-terminal kinase. J Neurochem 102:65–76. doi: 10.1111/j.1471-4159.2007.04618.x CrossRefPubMedGoogle Scholar
  5. Arundine M, Tymianski M (2004) Molecular mechanisms of glutamate-dependent neurodegeneration in ischemia and traumatic brain injury. Cell Mol Life Sci 61:657–668. doi: 10.1007/s00018-003-3319-x CrossRefPubMedGoogle Scholar
  6. Baethmann A, Staub F, Kempski O, Plesnila N, Chang RCC, Schnezder GH, Eriskat J, Stoffel M, Ringel F (1996) Glutamate enhances brain damage from ischemia and trauma. In: Ito U (ed) Maturation phenomenon in cerebral ischemia II. Springer-Verlag, Berlin, pp 43–51Google Scholar
  7. Borsello T, Clarke PG, Hirt L, Vercelli A, Repici M, Schorderet DF, Bogousslavsky J, Bonny C (2003) A peptide inhibitor of c-Jun N-terminal kinase protects against excitotoxicity and cerebral ischemia. Nat Med 9:1180–1186. doi: 10.1038/nm911 CrossRefPubMedGoogle Scholar
  8. Chan HC, Chang RCC, Koon-Ching IA, Chiu K, Yuen WH, Zee SY, So KF (2007) Neuroprotective effects of Lycium barbarum Lynn on protecting retinal ganglion cells in an ocular hypertension model of glaucoma. Exp Neurol 203:269–273. doi: 10.1016/j.expneurol.2006.05.031 CrossRefPubMedGoogle Scholar
  9. Chang RCC, So KF (2008) Use of anti-aging herbal medicine, Lycium barbarum, against aging-associated diseases. What do we know so far? Cell Mol Neurobiol 28:643–652. doi: 10.1007/s10571-007-9181-x CrossRefPubMedGoogle Scholar
  10. Chang RCC, Suen KC, Ma CH, Elyaman W, Ng HK, Hugon J (2002) Involvement of double-stranded RNA-dependent protein kinase and phosphorylation of eukaryotic initiation factor-2alpha in neuronal degeneration. J Neurochem 83:1215–1225. doi: 10.1046/j.1471-4159.2002.01237.x CrossRefPubMedGoogle Scholar
  11. Chao J, Yu MS, Ho YS, Wang M, Chang RCC (2008) Dietary oxyresveratrol prevents parkinsonian mimetic 6-hydroxydopamine neurotoxiciy. Free Radic Biol Med 45:1019–1026. doi: 10.1016/j.freeradbiomed.2008.07.002 CrossRefPubMedGoogle Scholar
  12. Chen RW, Qin ZH, Ren M, Kanai H, Chalecka-Franaszek E, Leeds P, Chuang DM (2003) Regulation of c-Jun N-terminal kinase, p38 kinase and AP-1 DNA binding in cultured brain neurons: roles in glutamate excitotoxicity and lithium neuroprotection. J Neurochem 84:566–575. doi: 10.1046/j.1471-4159.2003.01548.x CrossRefPubMedGoogle Scholar
  13. Chi CW, Wang CN, Lin YL, Chen CF, Shiao YJ (2005) Tournefolic acid B methyl ester attenuates glutamate-induced toxicity by blockade of ROS accumulation and abrogating the activation of caspases and JNK in rat cortical neurons. J Neurochem 92:692–700. doi: 10.1111/j.1471-4159.2004.02912.x CrossRefPubMedGoogle Scholar
  14. Chicoine LM, Bahr BA (2007) Excitotoxic protection by polyanionic polysaccharide: evidence of a cell survival pathway involving AMPA receptor-MAPK interactions. J Neurosci Res 85:294–302. doi: 10.1002/jnr.21117 CrossRefPubMedGoogle Scholar
  15. Chicoine LM, Suppiramaniam V, Vaithianathan T, Gianutsos G, Bahr BA (2004) Sulfate- and size-dependent polysaccharide modulation of AMPA receptor properties. J Neurosci Res 75:408–416. doi: 10.1002/jnr.10871 CrossRefPubMedGoogle Scholar
  16. Choi DW (1988) Glutamate neurotoxicity and diseases of the nervous system. Neuron 1:623–634. doi: 10.1016/0896-6273(88)90162-6 CrossRefPubMedGoogle Scholar
  17. Choi DW, Maulucci-Gedde M, Kriegstein AR (1987) Glutamate neurotoxicity in cortical cell culture. J Neurosci 7:357–368PubMedGoogle Scholar
  18. Choi SH, Lee DY, Kim SU, Jin BK (2005) Thrombin-induced oxidative stress contributes to the death of hippocampal neurons in vivo: role of microglial NADPH oxidase. J Neurosci 25:4082–4090. doi: 10.1523/JNEUROSCI.4306-04.2005 CrossRefPubMedGoogle Scholar
  19. Coyle JT, Puttfarcken P (1993) Oxidative stress, glutamate, and neurodegenerative disorders. Science 262:689–695. doi: 10.1126/science.7901908 CrossRefPubMedGoogle Scholar
  20. Dicou E, Rangon CM, Guimiot F, Spedding M, Gressens P (2003) Positive allosteric modulators of AMPA receptors are neuroprotective against lesions induced by an NMDA agonist in neonatal mouse brain. Brain Res 970:221–225. doi: 10.1016/S0006-8993(03)02357-6 CrossRefPubMedGoogle Scholar
  21. Fang X, Yu MM, Yuen WH, Zee SY, Chang RCC (2005) Immune modulatory effects of Prunella vulgaris L. on monocytes/macrophages. Int J Mol Med 16:1109–1116PubMedGoogle Scholar
  22. Freudenthaler S, Pantev M (2008) Dose-response analysis to support dosage recommendations for memantine. Naunyn Schmiedebergs Arch Pharmacol 353(Suppl):R159Google Scholar
  23. Gardoni F, Di LM (2006) New targets for pharmacological intervention in the glutamatergic synapse. Eur J Pharmacol 545:2–10. doi: 10.1016/j.ejphar.2006.06.022 CrossRefPubMedGoogle Scholar
  24. Gilgun-Sherki Y, Rosenbaum Z, Melamed E, Offen D (2002) Antioxidant therapy in acute central nervous system injury: current state. Pharmacol Rev 54:271–284. doi: 10.1124/pr.54.2.271 CrossRefPubMedGoogle Scholar
  25. Gladstone DJ, Black SE, Hakim AM (2002) Toward wisdom from failure: lessons from neuroprotective stroke trials and new therapeutic directions. Stroke 33:2123–2136. doi: 10.1161/01.STR.0000025518.34157.51 CrossRefPubMedGoogle Scholar
  26. Golde TE (2006) Disease modifying therapy for AD? J Neurochem 99:689–707. doi: 10.1111/j.1471-4159.2006.04211.x CrossRefPubMedGoogle Scholar
  27. Ho YS, Yu MS, Lai CS, So KF, Yuen WH, Chang RCC (2007) Characterizing the neuroprotective effects of alkaline extract of Lycium barbarum on beta-amyloid peptide neurotoxicity. Brain Res 1158C:123–134. doi: 10.1016/j.brainres.2007.04.075 CrossRefGoogle Scholar
  28. Hyrc K, Handran SD, Rothman SM, Goldberg MP (1997) Ionized intracellular calcium concentration predicts excitotoxic neuronal death: observations with low-affinity fluorescent calcium indicators. J Neurosci 17:6669–6677PubMedGoogle Scholar
  29. Johnson JW, Kotermanski SE (2006) Mechanism of action of memantine. Curr Opin Pharmacol 6:61–67. doi: 10.1016/j.coph.2005.09.007 CrossRefPubMedGoogle Scholar
  30. Kogo J, Takeba Y, Kumai T, Kitaoka Y, Matsumoto N, Ueno S, Kobayashi S (2006) Involvement of TNF-alpha in glutamate-induced apoptosis in a differentiated neuronal cell line. Brain Res 1122:201–208. doi: 10.1016/j.brainres.2006.09.006 CrossRefPubMedGoogle Scholar
  31. Kornhuber J, Quack G (1995) Cerebrospinal fluid and serum concentrations of the N-methyl-d-aspartate (NMDA) receptor antagonist memantine in man. Neurosci Lett 195:137–139. doi: 10.1016/0304-3940(95)11785-U CrossRefPubMedGoogle Scholar
  32. Kornhuber J, Kennepohl EM, Bleich S, Wiltfang J, Kraus T, Reulbach U, Meineke I (2007) Memantine pharmacotherapy: a naturalistic study using a population pharmacokinetic approach. Clin Pharmacokinet 46:599–612. doi: 10.2165/00003088-200746070-00005 CrossRefPubMedGoogle Scholar
  33. Lai SW, Yu MS, Yuen WH, Chang RCC (2006) Novel neuroprotective effects of the aqueous extracts from Verbena officinalis Linn. Neuropharmacology 50:641–650. doi: 10.1016/j.neuropharm.2005.11.009 CrossRefPubMedGoogle Scholar
  34. Lai SW, Yu MS, Yuen WH, So KF, Zee SY, Chang RCC (2008) Antagonizing beta-amyloid peptide neurotoxicity of the anti-aging fungus Ganoderma lucidum. Brain Res 1190:215–224. doi: 10.1016/j.brainres.2007.10.103 CrossRefPubMedGoogle Scholar
  35. Lauri SE, Kaukinen S, Kinnunen T, Ylinen A, Imai S, Kaila K, Taira T, Rauvala H (1999) Reg1ulatory role and molecular interactions of a cell-surface heparan sulfate proteoglycan (N-syndecan) in hippocampal long-term potentiation. J Neurosci 19:1226–1235PubMedGoogle Scholar
  36. Leveugle B, Ding W, Laurence F, Dehouck MP, Scanameo A, Cecchelli R, Fillit H (1998) Heparin oligosaccharides that pass the blood-brain barrier inhibit beta-amyloid precursor protein secretion and heparin binding to beta-amyloid peptide. J Neurochem 70:736–744PubMedGoogle Scholar
  37. Li XM, Ma YL, Liu XJ (2006) Effect of the Lycium barbarum polysaccharides on age-related oxidative stress in aged mice. J Ethnopharmacol 111:504–511. doi: 10.1016/j.jep.2006.12.024 CrossRefPubMedGoogle Scholar
  38. Lipton SA (2005) The molecular basis of memantine action in Alzheimer’s disease and other neurologic disorders: low-affinity, uncompetitive antagonism. Curr Alzheimer Res 2:155–165. doi: 10.2174/1567205053585846 CrossRefPubMedGoogle Scholar
  39. Ma Q, Dudas B, Hejna M, Cornelli U, Lee JM, Lorens S, Mervis R, Hanin I, Fareed J (2002) The blood-brain barrier accessibility of a heparin-derived oligosaccharides C3. Thromb Res 105:447–453. doi: 10.1016/S0049-3848(02)00050-6 CrossRefPubMedGoogle Scholar
  40. McDonald DR, Brunden KR, Landreth GE (1997) Amyloid fibrils activate tyrosine kinase-dependent signaling and superoxide production in microglia. J Neurosci 17:2284–2294PubMedGoogle Scholar
  41. Miyamoto E (2006) Molecular mechanism of neuronal plasticity: induction and maintenance of long-term potentiation in the hippocampus. J Pharmacol Sci 100:433–442. doi: 10.1254/jphs.CPJ06007X CrossRefPubMedGoogle Scholar
  42. Parsons CG, Gilling KE, Jatzke C (2008) Memantine does not show intracellular block of the NMDA receptor channel. Eur J Pharmacol 587:99–103. doi: 10.1016/j.ejphar.2008.03.053 CrossRefPubMedGoogle Scholar
  43. Pietrzik C, Behl C (2005) Concepts for the treatment of Alzheimer’s disease: molecular mechanisms and clinical application. Int J Exp Pathol 86:173–185. doi: 10.1111/j.0959-9673.2005.00435.x CrossRefPubMedGoogle Scholar
  44. Portera-Cailliau C, Price DL, Martin LJ (1997) Excitotoxic neuronal death in the immature brain is an apoptosis-necrosis morphological continuum. J Comp Neurol 378:70–87PubMedGoogle Scholar
  45. Schubert D, Piasecki D (2001) Oxidative glutamate toxicity can be a component of the excitotoxicity cascade. J Neurosci 21:7455–7462PubMedGoogle Scholar
  46. Seveg MG (1934) Deproteinization and removal of capsular polysaccharides. Biochem Z 273:419–423Google Scholar
  47. Shigeri Y, Seal RP, Shimamoto K (2004) Molecular pharmacology of glutamate transporters, EAATs and VGLUTs. Brain Res Brain Res Rev 45:250–265. doi: 10.1016/j.brainresrev.2004.04.004 CrossRefPubMedGoogle Scholar
  48. Shih AY, Erb H, Sun X, Toda S, Kalivas PW, Murphy TH (2006) Cystine/glutamate exchange modulates glutathione supply for neuroprotection from oxidative stress and cell proliferation. J Neurosci 26:10514–10523. doi: 10.1523/JNEUROSCI.3178-06.2006 CrossRefPubMedGoogle Scholar
  49. Sinnarajah S, Suppiramaniam V, Kumar KP, Hall RA, Bahr BA, Vodyanoy V (1999) Heparin modulates the single channel kinetics of reconstituted AMPA receptors from rat brain. Synapse 31:203–209. doi: 10.1002/(SICI)1098-2396(19990301)31:3≤203::AID-SYN5≥3.0.CO;2-W CrossRefPubMedGoogle Scholar
  50. Sotogaku N, Tully SE, Gama CI, Higashi H, Tanaka M, Hsieh-Wilson LC, Nishi A (2007) Activation of phospholipase C pathways by a synthetic chondroitin sulfate-E tetrasaccharide promotes neurite outgrowth of dopaminergic neurons. J Neurochem 103:749–760. doi: 10.1111/j.1471-4159.2007.04849.x CrossRefPubMedGoogle Scholar
  51. Suen KC, Lin KF, Elyaman W, So KF, Chang RCC, Hugon J (2003) Reduction of calcium release from the endoplasmic reticulum could only provide partial neuroprotection against beta-amyloid peptide toxicity. J Neurochem 87:1413–1426PubMedCrossRefGoogle Scholar
  52. Suppiramaniam V, Vaithianathan T, Manivannan K, Dhanasekaran M, Parameshwaran K, Bahr BA (2006) Modulatory effects of dextran sulfate and fucoidan on binding and channel properties of AMPA receptors isolated from rat brain. Synapse 60:456–464. doi: 10.1002/syn.20319 CrossRefPubMedGoogle Scholar
  53. Tan S, Wood M, Maher P (1998) Oxidative stress induces a form of programmed cell death with characteristics of both apoptosis and necrosis in neuronal cells. J Neurochem 71:95–105PubMedGoogle Scholar
  54. Tan S, Schubert D, Maher P (2001) Oxytosis: a novel form of programmed cell death. Curr Top Med Chem 1:497–506. doi: 10.2174/1568026013394741 CrossRefPubMedGoogle Scholar
  55. Won SJ, Kim DY, Gwag BJ (2002) Cellular and molecular pathways of ischemic neuronal death. J Biochem Mol Biol 35:67–86PubMedGoogle Scholar
  56. Wu X, Zhu D, Jiang X, Okagaki P, Mearow K, Zhu G, McCall S, Banaudha K, Lipsky RH, Marini AM (2004) AMPA protects cultured neurons against glutamate excitotoxicity through a phosphatidylinositol 3-kinase-dependent activation in extracellular signal-regulated kinase to upregulate BDNF gene expression. J Neurochem 90:807–818. doi: 10.1111/j.1471-4159.2004.02526.x CrossRefPubMedGoogle Scholar
  57. Yazawa K, Kihara T, Shen H, Shimmyo Y, Niidome T, Sugimoto H (2006) Distinct mechanisms underlie distinct polyphenol-induced neuroprotection. FEBS Lett 580:6623–6628. doi: 10.1016/j.febslet.2006.11.011 CrossRefPubMedGoogle Scholar
  58. Yu MS, Lai SW, Lin KF, Fang JN, Yuen WH, Chang RCC (2004) Characterization of polysaccharides from the flowers of Nerium indicum and their neuroprotective effects. Int J Mol Med 14:917–924PubMedGoogle Scholar
  59. Yu MS, Leung SK, Lai SW, Che CM, Zee SY, So KF, Yuen WH, Chang RCC (2005) Neuroprotective effects of anti-aging oriental medicine Lycium barbarum against beta-amyloid peptide neurotoxicity. Exp Gerontol 40:716–727. doi: 10.1016/j.exger.2005.06.010 CrossRefPubMedGoogle Scholar
  60. Yu MS, Ho YS, So KF, Yuen WH, Chang RCC (2006) Cytoprotective effects of Lycium barbarum against reducing stress on endoplasmic reticulum. Int J Mol Med 17:1157–1161PubMedGoogle Scholar
  61. Yu MS, Wong AY, So KF, Fang JN, Yuen WH, Chang RCC (2007) New polysaccharide from Nerium indicum protects neurons via stress kinase signaling pathway. Brain Res 1153:221–230. doi: 10.1016/j.brainres.2007.03.074 CrossRefPubMedGoogle Scholar
  62. Zhang Y, Bhavnani BR (2006) Glutamate-induced apoptosis in neuronal cells is mediated via caspase-dependent and independent mechanisms involving calpain and caspase-3 proteases as well as apoptosis inducing factor (AIF) and this process is inhibited by equine estrogens. BMC Neurosci 7:49. doi: 10.1186/1471-2202-7-49 CrossRefPubMedGoogle Scholar
  63. Zhang Y, Lu X, Bhavnani BR (2003) Equine estrogens differentially inhibit DNA fragmentation induced by glutamate in neuronal cells by modulation of regulatory proteins involved in programmed cell death. BMC Neurosci 4:32. doi: 10.1186/1471-2202-4-32 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Yuen-Shan Ho
    • 1
    • 2
  • Man-Shan Yu
    • 1
  • Suet-Yi Yik
    • 1
  • Kwok-Fai So
    • 1
    • 2
    • 3
  • Wai-Hung Yuen
    • 4
  • Raymond Chuen-Chung Chang
    • 1
    • 2
    • 3
    Email author
  1. 1.Laboratory of Neurodegenerative Diseases, Department of Anatomy, LKS Faculty of MedicineThe University of Hong KongPokfulamHong Kong SAR, China
  2. 2.Research Centre of Heart, Brain, Hormone and Healthy Aging, LKS Faculty of MedicineThe University of Hong KongPokfulamHong Kong SAR, China
  3. 3.State Key Laboratory of Brain and Cognitive SciencesThe University of Hong KongPokfulamHong Kong SAR, China
  4. 4.Department of ChemistryThe University of Hong KongPokfulamHong Kong SAR, China

Personalised recommendations