Molecular and Cellular Biochemistry

, Volume 419, Issue 1–2, pp 53–64 | Cite as

Salidroside protects cortical neurons against glutamate-induced cytotoxicity by inhibiting autophagy

  • Wei-Yong Yin
  • Qiang Ye
  • Huan-Jie Huang
  • Nian-Ge Xia
  • Yan-Yan Chen
  • Yi Zhang
  • Qiu-Min Qu


Recent evidence suggests that glutamate-induced cytotoxicity contributes to autophagic neuron death and is partially mediated by increased oxidative stress. Salidroside has been demonstrated to have neuroprotective effects in glutamate-induced neuronal damage. The precise mechanism of its regulatory role in neuronal autophagy is, however, poorly understood. This study aimed to probe the effects and mechanisms of salidroside in glutamate-induced autophagy activation in cultured rat cortical neurons. Cell viability assay, Western blotting, coimmunoprecipitation, and small interfering RNA were performed to analyze autophagy activities during glutamate-evoked oxidative injury. We found that salidroside protected neonatal neurons from glutamate-induced apoptotic cell death. Salidroside significantly attenuated the LC3-II/LC3-I ratio and expression of Beclin-1, but increased (SQSTM1)/p62 expression under glutamate exposure. Pretreatment with 3-methyladenine (3-MA), an autophagy inhibitor, decreased LC3-II/LC3-I ratio, attenuated glutamate-induced cell injury, and mimicked some of the protective effects of salidroside against glutamate-induced cell injury. Molecular analysis demonstrated that salidroside inhibited cortical neuron autophagy in response to glutamate exposure through p53 signaling by increasing the accumulation of cytoplasmic p53. Salidroside inhibited the glutamate-induced dissociation of the Bcl-2-Beclin-1 complex with minor affects on the PI3K/Akt/mTOR signaling pathways. These data demonstrate that the inhibition of autophagy could be responsible for the neuroprotective effects of salidroside on glutamate-induced neuronal injury.


Salidroside Cortical neurons Cytotoxicity Autophagy Neuronal injury 





The mammalian target of rapamycin


Dulbecco’s modified Eagle’s medium


Neurobasal medium


Small interfering RNA









This research was supported by the 863-Chinese National Science and Technology Project (2007AA02Z443).

Supplementary material

11010_2016_2749_MOESM1_ESM.docx (1.4 mb)
Supplementary material 1 (DOCX 1429 kb)


  1. 1.
    Mizushima N, Levine B, Cuervo AM, Klionsky DJ (2008) Autophagy fights disease through cellular self-digestion. Nature 451:1069–1075CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Steiger-Barraissoul S, Rami A (2009) Serum deprivation induced autophagy and predominantly an AIF-dependent apoptosis in hippocampal HT22 neurons. Apoptosis 14:1274–1288CrossRefPubMedGoogle Scholar
  3. 3.
    Carloni S, Buonocore G, Balduini W (2008) Protective role of autophagy in neonatal hypoxia-ischemia induced brain injury. Neurobiol Dis 32:329–339CrossRefPubMedGoogle Scholar
  4. 4.
    Shi R, Weng J, Zhao L, Li XM, Gao TM, Kong J (2012) Excessive autophagy contributes to neuron death in cerebral ischemia. CNS Neurosci Ther 18:250–260CrossRefPubMedGoogle Scholar
  5. 5.
    Koike M, Shibata M, Tadakoshi M, Gotoh K, Komatsu M, Waguri S, Kawahara N, Kuida K, Nagata S, Kominami E, Tanaka K, Uchiyama Y (2008) Inhibition of autophagy prevents hippocampal pyramidal neuron death after hypoxic-ischemic injury. Am J Pathol 172:454–469CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Luo T, Liu G, Ma H, Lu B, Xu H, Wang Y, Wu J, Ge P, Liang J (2014) Inhibition of autophagy via activation of PI3K/Akt pathway contributes to the protection of ginsenoside Rb1 against neuronal death caused by ischemic insults. Int J Mol Sci 15:15426–15442CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Qin H, Tan W, Zhang Z, Bao L, Shen H, Wang F, Xu F, Wang Z (2015) 15d-prostaglandin J2 protects cortical neurons against oxygen-glucose deprivation/reoxygenation injury: involvement of inhibiting autophagy through upregulation of Bcl-2. Cell Mol Neurobiol 35:303–312CrossRefPubMedGoogle Scholar
  8. 8.
    Rami A, Langhagen A, Steiger S (2008) Focal cerebral ischemia induces upregulation of Beclin 1 and autophagy-like cell death. Neurobiol Dis 29:132–141CrossRefPubMedGoogle Scholar
  9. 9.
    Wen YD, Sheng R, Zhang LS, Han R, Zhang X, Zhang XD, Han F, Fukunaga K, Qin ZH (2008) Neuronal injury in rat model of permanent focal cerebral ischemia is associated with activation of autophagic and lysosomal pathways. Autophagy 4:762–769CrossRefPubMedGoogle Scholar
  10. 10.
    Wang JY, Xia Q, Chu KT, Pan J, Sun LN, Zeng B, Zhu YJ, Wang Q, Wang K, Luo BY (2011) Severe global cerebral ischemia-induced programmed necrosis of hippocampal CA1 neurons in rat is prevented by 3-methyladenine: a widely used inhibitor of autophagy. J Neuropathol Exp Neurol 70:314–322CrossRefPubMedGoogle Scholar
  11. 11.
    Xu F, Gu JH, Qin ZH (2012) Neuronal autophagy in cerebral ischemia. Neurosci Bull 28:658–666CrossRefPubMedGoogle Scholar
  12. 12.
    Kim H, Choi J, Ryu J, Park SG, Cho S, Park BC, Lee DH (2009) Activation of autophagy during glutamate-induced HT22 cell death. Biochem Biophys Res Commun 388:339–344CrossRefPubMedGoogle Scholar
  13. 13.
    Chen Z, Lu T, Yue X, Wei N, Jiang Y, Chen M, Ni G, Liu X, Xu G (2010) Neuroprotective effect of ginsenoside Rb1 on glutamate-induced neurotoxicity: with emphasis on autophagy. Neurosci Lett 482:264–268CrossRefPubMedGoogle Scholar
  14. 14.
    Diaz Lanza AM, Abad Martinez MJ, Fernandez Matellano L, Recuero Carretero C, Villaescusa Castillo L, Silvan Sen AM, Bermejo Benito P (2001) Lignan and phenylpropanoid glycosides from Phillyrea latifolia and their in vitro anti-inflammatory activity. Planta Med 67:219–223CrossRefPubMedGoogle Scholar
  15. 15.
    Xiao L, Li H, Zhang J, Yang F, Huang A, Deng J, Liang M, Ma F, Hu M, Huang Z (2014) Salidroside protects Caenorhabditis elegans neurons from polyglutamine-mediated toxicity by reducing oxidative stress. Molecules 19:7757–7769CrossRefPubMedGoogle Scholar
  16. 16.
    Zhong X, Lin R, Li Z, Mao J, Chen L (2014) Effects of Salidroside on cobalt chloride-induced hypoxia damage and mTOR signaling repression in PC12 cells. Biol Pharm Bull 37:1199–1206CrossRefPubMedGoogle Scholar
  17. 17.
    Yu S, Liu M, Gu X, Ding F (2008) Neuroprotective effects of salidroside in the PC12 cell model exposed to hypoglycemia and serum limitation. Cell Mol Neurobiol 28:1067–1078CrossRefPubMedGoogle Scholar
  18. 18.
    Yang X, Xu W, Zhao W, Zhao Y, Yang Y, Ling Y (2013) Synthesis and neuroprotective effects of the fluorine substituted salidroside analogues in the PC12 cell model exposed to hypoglycemia and serum limitation. Chem Pharm Bull 61:1192–1196 (Tokyo) CrossRefPubMedGoogle Scholar
  19. 19.
    Chen X, Liu J, Gu X, Ding F (2008) Salidroside attenuates glutamate-induced apoptotic cell death in primary cultured hippocampal neurons of rats. Brain Res 1238:189–198CrossRefPubMedGoogle Scholar
  20. 20.
    Yang Z, Klionsky DJ (2010) Mammalian autophagy: core molecular machinery and signaling regulation. Curr Opin Cell Biol 22:124–131CrossRefPubMedGoogle Scholar
  21. 21.
    Cheng Y, Ren X, Hait WN, Yang JM (2013) Therapeutic targeting of autophagy in disease: biology and pharmacology. Pharmacol Rev 65:1162–1197CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Harris H, Rubinsztein DC (2012) Control of autophagy as a therapy for neurodegenerative disease. Nat Rev Neurol 8:108–117CrossRefGoogle Scholar
  23. 23.
    Ni H, Gong Y, Yan JZ, Zhang LL (2010) Autophagy inhibitor 3-methyladenine regulates the expression of LC3, Beclin-1 and ZnTs in rat cerebral cortex following recurrent neonatal seizures. World J Emerg Med 1:216–223PubMedPubMedCentralGoogle Scholar
  24. 24.
    Noursadeghi M, Tsang J, Haustein T, Miller RF, Chain BM, Katz DR (2008) Quantitative imaging assay for NF-kappaB nuclear translocation in primary human macrophages. J Immunol Methods 329:194–200CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Xie CM, Chan WY, Yu S, Zhao J, Cheng CH (2011) Bufalin induces autophagy-mediated cell death in human colon cancer cells through reactive oxygen species generation and JNK activation. Free Radic Biol Med 51:1365–1375CrossRefPubMedGoogle Scholar
  26. 26.
    Levine B, Yuan J (2005) Autophagy in cell death: an innocent convict? J Clin Invest 115:2679–2688CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Fulceri F, Ferrucci M, Lazzeri G, Paparelli S, Bartalucci A, Tamburini I, Paparelli A, Fornai F (2011) Autophagy activation in glutamate-induced motor neuron loss. Arch Ital Biol 149:101–111PubMedGoogle Scholar
  28. 28.
    Mizushima N, Yoshimori T (2007) How to interpret LC3 immunoblotting. Autophagy 3:542–545CrossRefPubMedGoogle Scholar
  29. 29.
    Komarov PG, Komarova EA, Kondratov RV, Christov-Tselkov K, Coon JS, Chernov MV, Gudkov AV (1999) A chemical inhibitor of p53 that protects mice from the side effects of cancer therapy. Science 285:1733–1737CrossRefPubMedGoogle Scholar
  30. 30.
    Saiki S, Sasazawa Y, Imamichi Y, Kawajiri S, Fujimaki T, Tanida I, Kobayashi H, Sato F, Sato S, Ishikawa K, Imoto M, Hattori N (2011) Caffeine induces apoptosis by enhancement of autophagy via PI3K/Akt/mTOR/p70S6K inhibition. Autophagy 7:176–187CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Kumari S, Mehta SL, Li PA (2012) Glutamate induces mitochondrial dynamic imbalance and autophagy activation: preventive effects of selenium. PLoS One 7:e39382CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Sattler R, Tymianski M (2001) Molecular mechanisms of glutamate receptor-mediated excitotoxic neuronal cell death. Mol Neurobiol 24:107–129CrossRefPubMedGoogle Scholar
  33. 33.
    Murphy TH, Miyamoto M, Sastre A, Schnaar RL, Coyle JT (1989) Glutamate toxicity in a neuronal cell line involves inhibition of cystine transport leading to oxidative stress. Neuron 2:1547–1558CrossRefPubMedGoogle Scholar
  34. 34.
    Kulbe JR, Mulcahy Levy JM, Coultrap SJ, Thorburn A, Bayer KU (2014) Excitotoxic glutamate insults block autophagic flux in hippocampal neurons. Brain Res 1542:12–19CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Vucicevic L, Misirkic-Marjanovic M, Paunovic V, Kravic-Stevovic T, Martinovic T, Ciric D, Maric N, Petricevic S, Harhaji-Trajkovic L, Bumbasirevic V, Trajkovic V (2014) Autophagy inhibition uncovers the neurotoxic action of the antipsychotic drug olanzapine. Autophagy 10:2362–2378CrossRefPubMedGoogle Scholar
  36. 36.
    Vousden KH, Lane DP (2007) p53 in health and disease. Nat Rev Mol Cell Biol 8:275–283CrossRefPubMedGoogle Scholar
  37. 37.
    Rao S, Tortola L, Perlot T, Wirnsberger G, Novatchkova M, Nitsch R, Sykacek P, Frank L, Schramek D, Komnenovic V, Sigl V, Aumayr K, Schmauss G, Fellner N, Handschuh S, Glosmann M, Pasierbek P, Schlederer M, Resch GP, Ma Y, Yang H, Popper H, Kenner L, Kroemer G, Penninger JM (2014) A dual role for autophagy in a murine model of lung cancer. Nat Commun 5:3056CrossRefPubMedGoogle Scholar
  38. 38.
    Tasdemir E, Chiara Maiuri M, Morselli E, Criollo A, D’Amelio M, Djavaheri-Mergny M, Cecconi F, Tavernarakis N, Kroemer G (2008) A dual role of p53 in the control of autophagy. Autophagy 4:810–814CrossRefPubMedGoogle Scholar
  39. 39.
    Morselli E, Tasdemir E, Maiuri MC, Galluzzi L, Kepp O, Criollo A, Vicencio JM, Soussi T, Kroemer G (2008) Mutant p53 protein localized in the cytoplasm inhibits autophagy. Cell Cycle 7:3056–3061CrossRefPubMedGoogle Scholar
  40. 40.
    Green DR, Kroemer G (2009) Cytoplasmic functions of the tumour suppressor p53. Nature 458:1127–1130CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Marino G, Lopez-Otin C (2004) Autophagy: molecular mechanisms, physiological functions and relevance in human pathology. Cell Mol Life Sci 61:1439–1454CrossRefPubMedGoogle Scholar
  42. 42.
    Crighton D, Wilkinson S, O’Prey J, Syed N, Smith P, Harrison PR, Gasco M, Garrone O, Crook T, Ryan KM (2006) DRAM, a p53-induced modulator of autophagy, is critical for apoptosis. Cell 126:121–134CrossRefPubMedGoogle Scholar
  43. 43.
    Maiuri MC, Criollo A, Kroemer G (2010) Crosstalk between apoptosis and autophagy within the Beclin 1 interactome. EMBO J 29:515–516CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Pattingre S, Tassa A, Qu X, Garuti R, Liang XH, Mizushima N, Packer M, Schneider MD, Levine B (2005) Bcl-2 antiapoptotic proteins inhibit Beclin 1-dependent autophagy. Cell 122:927–939CrossRefPubMedGoogle Scholar
  45. 45.
    Peltier J, O’Neill A, Schaffer DV (2007) PI3K/Akt and CREB regulate adult neural hippocampal progenitor proliferation and differentiation. Dev Neurobiol 67:1348–1361CrossRefPubMedGoogle Scholar
  46. 46.
    Morrison RS, Kinoshita Y, Johnson MD, Ghatan S, Ho JT, Garden G (2002) Neuronal survival and cell death signaling pathways. Adv Exp Med Biol 513:41–86CrossRefPubMedGoogle Scholar
  47. 47.
    Wu YT, Tan HL, Huang Q, Ong CN, Shen HM (2009) Activation of the PI3K-Akt-mTOR signaling pathway promotes necrotic cell death via suppression of autophagy. Autophagy 5:824–834CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Wei-Yong Yin
    • 1
  • Qiang Ye
    • 2
  • Huan-Jie Huang
    • 2
  • Nian-Ge Xia
    • 2
  • Yan-Yan Chen
    • 2
  • Yi Zhang
    • 3
  • Qiu-Min Qu
    • 1
  1. 1.Department of NeurologyThe First Affiliated Hospital of Xi’an Jiaotong UniversityXi’anChina
  2. 2.Department of NeurologyThe First Affiliated Hospital of Wenzhou Medical UniversityWenzhouChina
  3. 3.School of Pharmaceutical SciencesWenzhou Medical UniversityWenzhouChina

Personalised recommendations