Skip to main content

Advertisement

SpringerLink
Log in

Glutamate-induced Toxicity in Hippocampal Slices Involves Apoptotic Features and p38MAPK Signaling

  • Original Paper
  • Published:
Neurochemical Research Aims and scope Submit manuscript

Abstract

Glutamate excitotoxicity may culminate with neuronal and glial cell death. Glutamate induces apoptosis in vivo and in cell cultures. However, glutamate-induced apoptosis and the signaling pathways related to glutamate-induced cell death in acute hippocampal slices remain elusive. Hippocampal slices exposed to 1 or 10 mM glutamate for 1 h and evaluated after 6 h, showed reduced cell viability, without altering membrane permeability. This action of glutamate was accompanied by cytochrome c release, caspase-3 activation and DNA fragmentation. Glutamate at low concentration (10 μM) induced caspase-3 activation and DNA fragmentation, but it did not cause cytochrome c release and, it did not alter the viability of slices. Glutamate-induced impairment of hippocampal cell viability was completely blocked by MK-801 (non-competitive antagonist of NMDA receptors) and GAMS (antagonist of KA/AMPA glutamate receptors). Regarding intracellular signaling pathways, glutamate-induced cell death was not altered by a MEK1 inhibitor, PD98059. However, the p38MAPK inhibitor, SB203580, prevented glutamate-induced cell damage. In the present study we have shown that glutamate induces apoptosis in hippocampal slices and it causes an impairment of cell viability that was dependent of ionotropic and metabotropic receptors activation and, may involve the activation of p38MAPK pathway.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Abbreviations

ERK 1/2:

Extracellular-regulated MAP kinase

Glu:

Glutamate

GAMS:

γ-d-glutamylamino-methylsulfonate

KA:

Kainic acid

MAPKs:

Mitogen-activated protein kinases

MK-801:

(+)-5-Methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate

NMDA:

N-methyl-d-aspartate

M-CPG:

(RS)-α-methyl-4-carboxyphenylglycine

LDH:

Lactate dehydrogenase

MTT:

3-(4,5-Dimethylthiazol-2-yl)-diphenyltetrazolium bromide

References

  1. Obrenovitch TP, Urenjak J (1997) Altered glutamatergic transmission in neurological disorders: from high extracellular glutamate to excessive synaptic efficacy. Prog Neurobiol 51:39–87

    Article  PubMed  CAS  Google Scholar 

  2. Maragakis NJ, Rothstein JD (2004) Glutamate transporters: animal models to neurological disease. Neurobiol Dis 15:461–473

    Article  PubMed  CAS  Google Scholar 

  3. Meldrum BS (2000) Glutamate as a neurotransmitter in the brain: review of physiology and pathology. J Nutr 130:1007S–1015S

    PubMed  CAS  Google Scholar 

  4. Mattson M (2000) Apoptosis in neurodegenerative disorders. Nat Rev Mol Cell Biol 1:120–129

    Article  PubMed  CAS  Google Scholar 

  5. Olney JW (1981) Kainic acid and other excitotoxins: a comparative analysis. Adv Biochem Psychopharmacol 27:375–384

    PubMed  CAS  Google Scholar 

  6. Chen TA, Yang F, Cole GM, Chan SO (2001) Inhibition of caspase-3-like activity reduces glutamate induced cell death in the adult retina. Brain Res 1:1–12

    Google Scholar 

  7. Matute C, Domercq C, Sanchez-Gomez MV (2006) Glutamate-mediated glial injury: mechanisms and clinical importance. Glia 15:212–24

    Article  Google Scholar 

  8. Ankarcrona M, Dypburkt JM, Bonfoco E, Zhyvotovsky B, Orrenius S, Lipton SA, Nicotera P (1995) Glutamate-induced neuronal death: a succession of necrosis or apoptosis depending on mitochondrial function. Neuron 15:961–973

    Article  PubMed  CAS  Google Scholar 

  9. Portera-Cailliau C, Price DL, Martin LJ (1997) Non-NMDA and NMDA receptor-mediated excitotoxic neuronal deaths in adult brain are morphologically distinct: further evidence for an apoptosis-necrosis continuum. J Comp Neurol 378:88–104

    Article  PubMed  CAS  Google Scholar 

  10. Cheung NS, Pascoe CJ, Giardina SF, John CA (1998) Micromolar l-glutamate induces extensive apoptosis in an apoptotic-necrotic continuum of insult-dependent, excitotoxic injury in cultured cortical neurones. Neuropharmacology 37:1419–1429

    Article  PubMed  CAS  Google Scholar 

  11. Sastry PS, Rao KS (2000) Apoptosis and the nervous system. J Neurochem 74:1–20

    Article  PubMed  CAS  Google Scholar 

  12. Olson M, Kornbluth S (2001) Mitochondria in apoptosis and human disease. Curr Mol Med 1:91–122

    Article  PubMed  CAS  Google Scholar 

  13. Hirashima Y, Kurimoto M, Nogami K, Endo S, Saitoh M, Ohtani O, Nagata T, Muraguchi A, Takaku A (1999) Correlation of glutamate-induced apoptosis with caspase activities in cultured rat cerebral neurons. Brain Res 849(1–2):109–118

    Article  PubMed  CAS  Google Scholar 

  14. Thomas CE, Mayle DA (2000) NMDA-sensitive neurons profoundly influence delayed staurosporine-induced apoptosis in rat mixed cortical neuronal cultures. Brain Res 889:163–173

    Article  Google Scholar 

  15. Yu X, Sun L, Luo X, Xu Z, An L (2003) Investigation of the neuronal death mode induced by glutamate treatment in serum-, antioxidant-free primary cultures cortical neurons. Brain Res Dev Brain Res 145:263–268

    Article  PubMed  CAS  Google Scholar 

  16. Leal RB, Gonçalves CA, Rodnight R (1997) Calcium-dependent phosphorylation acidic protein (GFAP) in the rat hippocampus: a comparison of de kinase/phosphatase balance in immature and mature slices using tryptic phosphopeptide mapping. Brain Res Dev Brain Res 104:1–10

    Article  PubMed  CAS  Google Scholar 

  17. Vanhoutte P, Barnier JV, Guibert N, Pagès C, Besson M, Hipskind RA, Caboche J (1999) Glutamate induces phosphorylation of ElK-1 and CREB, along with c-fos activation, via an extracellular signal-regulated kinase-dependent pathway in brain slices. Mol Cell Biol 19:136–146

    PubMed  CAS  Google Scholar 

  18. Banko JL, Hou L, Klann E (2004) NMDA receptor activation in PKA- and ERK-dependent MnK1 activation and increase in eIF4E phosphorylation in hippocampal area CA1. J Neurochem 91:462–470

    Article  PubMed  CAS  Google Scholar 

  19. Corvol JC, Valjent E, Toutant M, Enslet H, Irinopoulot T, Lev S, Hervé D, Girault JA (2005) Depolarization activates ERK and praline-rich tyrosine kinase 2 (PYK2) independently in different cellular compartments in hippocampal slices. J Biol Chem 1:660–668

    Google Scholar 

  20. Collingridge GL, Bliss TW (1995) Memories of NMDA receptors and LTP. Trends Neurosci 18:54–56

    Article  PubMed  CAS  Google Scholar 

  21. Gong CX, Lidsky T, Wegiel J, Grundke-Igbal I, Igbal K (2001) Metabolically active rat brain slices as a model to study the regulation of protein phosphorylation in mammalian brain. Brain Res Brain Res Protoc 6:134–140

    Article  PubMed  CAS  Google Scholar 

  22. Cordova FM, Rodrigues ALS, Giacomelli MBO, Oliveira CS, Posser T, Dunkley PR, Leal RB (2004) Lead stimulates ERK 1/2 and p38MAPK phosphorylation in the hippocampus of immature rats. Brain Res 998:65–72

    Article  PubMed  CAS  Google Scholar 

  23. Molz S, Oliveira IJL, Decker H, Souza DO, Tasca CI (2005) Neurotoxicity induced by glutamate in glucose-deprived rat hippocampal slices is prevented by GMP. Neurochem Res 30:83–89

    Article  PubMed  CAS  Google Scholar 

  24. Niswander JM, Dokas LA (2006) Phosphorylation of HSP27 and synthesis of 14-3-3 epsilon are parallel responses to hyperosmotic stress in the hippocampus. Brain Res 1116:19–30

    Article  PubMed  CAS  Google Scholar 

  25. Oliveira IJL, Molz S, Souza DO, Tasca CI (2002) Neuroprotective effect of GMP in hippocampal slices submitted to an in vitro model of ischemia. Cell Mol Neurobiol 22:335–344

    Article  PubMed  CAS  Google Scholar 

  26. Brongholi K, Sousa DG, Bainy AC, Dafré AL, Tasca CI (2006) Oxygen–glucose deprivation decreases glutathione levels and glutamate uptake in rat hippocampal slices. Brain Res 1083:211–218

    Article  PubMed  CAS  Google Scholar 

  27. Camacho A, Massieu L (2006) Role of glutamate transporters in the clearance and release of glutamate during ischemia and its relation to neuronal death. Arch Med Res 37:11–18

    Article  PubMed  CAS  Google Scholar 

  28. Chen Z, Gibson TB, Robinson F, Silvestro L, Pearson G, Xu B, Wright A, Vanderbilt C, Cobb M (2001) MAP kinases. Chem Rev 101:2449–2476

    Article  PubMed  CAS  Google Scholar 

  29. Chang L, Karin M (2001) Mammalian MAP kinase signaling cascades. Nature 410:37–40

    Article  PubMed  CAS  Google Scholar 

  30. Gallagher SM, Daly CA, Bear MF, Huber KM (2004) Extracellular signal-regulated protein kinases activation is required for metabotropic glutamate receptor-dependent long-term depression in hippocampal area CA1. J Neurosci 24:4864–4859

    Google Scholar 

  31. Sweatt JD (2004) Mitogen-activated protein kinases in synaptic plasticity and memory. Curr Opin Neurobiol 14:311–317

    Article  PubMed  CAS  Google Scholar 

  32. Jiang Q, Zhengling G, Zhang G, Guozhang J (2000) Diphosphorylation and involvement of extracellular signal-regulated kinases (ERK1/2) in glutamate-induced apoptotic-like death in cultured rat cortical neurons. Brain Res 857:71–77

    Article  PubMed  CAS  Google Scholar 

  33. Grant ER, Errico MA, Emanuel SL, Benjamin D, McMillian MK, Wadsworth SA, Zivin RA, Zhong Z (2001) Protection against glutamate toxicity through inhibition of the p44/p42 mitogen-activated protein kinase pathway in neuronally differentiated P19 cells. Biochem Pharmacol 62:283–296

    Article  PubMed  CAS  Google Scholar 

  34. Legos JJ, McLaughlin B, Skaper SD, Strijbos PJ, Parsons AA, Aizenman E, Herin GA, Barone FC, Erhardt JA (2002) The selective p38 inhibitor SB-239063 protects primary neurons from mild to moderate excitotoxic injury. Eur J Pharmacol 447:37–42

    Article  PubMed  CAS  Google Scholar 

  35. Park JE, Kim EJ, Kwon KJ, Jung YS, Moon CH, Lee SH, Baik EJ (2004) Neuroprotection by fructose-1,6-bisphosphate involves ROS alterations via p38 MAPK/ERK. Brain Res 1026:295–301

    Article  PubMed  CAS  Google Scholar 

  36. Munemasa Y, Ohtani-Kaneko R, Kitaoka Y, Kuribayashi K, Isenoumi K, Kogo J, Yamashita K, Kumai T, Kobayashi S, Hirata K, Ueno K (2005) Contribution of mitogen-activated protein kinases to NMDA-induced neurotoxicity in the rat retina. Brain Res 1044:227–240

    Article  PubMed  CAS  Google Scholar 

  37. Sugino T, Nozaki K, Tagaki Y, Hattori I, Hashimoto N, Moriguchi T, Nishida E (2000) Activation of mitogen-activated protein kinases after transient forebrain ischemia in gerbil and hippocampus. J Neurosci 20:4506–4514

    PubMed  CAS  Google Scholar 

  38. Namura S, Iihara K, Takami S, Nagata I, Kikuchi H, Matsushita K, Moskowitz MA, Bonventre JV, Alessandrini A (2001) Intravenous administration of MEK inhibitor UO126 affords brain protection against forebrain ischemia and focal cerebral ischemia. Proc Natl Acad Sci USA 98:11569–11574

    Article  PubMed  CAS  Google Scholar 

  39. Gähwiller BH, Capogna M, Debanne D, Mckinney RA, Thompson SM (1997) Organotypic slices cultures: a technique has come of age. Trends Neurosci 20:471–477

    Article  Google Scholar 

  40. Mosmann T (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity. J Immunol Methods 65:55–63

    Article  PubMed  CAS  Google Scholar 

  41. Leist M, Single B, Naumann H, Fava E, Simon B, Kühnle S, Nicotera P (1999) Nitric oxide inhibits execution of apoptosis at two distinct ATP-dependent and downstream of mitochondrial cytochrome c release. Biochem Biophys Res Commun 258:215–221

    Article  PubMed  CAS  Google Scholar 

  42. Leal R, Cordova FM, Lynn H, Dunkley PR (2002) Lead-stimulate p38MAPK-dependent Hsp27 phosphorylation. Toxicol Appl Pharmacol 178:44–51

    Article  PubMed  CAS  Google Scholar 

  43. Yang G-M, Paul SM (1997) Cultured cerebellar granule neurons as a model of neuronal apoptosis. In: Poirier J (ed) Neuromethods, vol 29. Humana Press Inc, Totowa, pp 47–66

    Google Scholar 

  44. Lowry OH, Rosembrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275

    PubMed  CAS  Google Scholar 

  45. Peterson GL (1977) A simplification of the protein assay method of Lowry et al. which is more generally applicable. Anal Biochem 83:346–356

    Article  PubMed  CAS  Google Scholar 

  46. Ientile R, Macaione V, Teletta M, Torre V, Macaione S (2001) Apoptosis and necrosis occurring in excitotoxic cell death in isolated chick embryo retina. J Neurochem 79:71–78

    Article  PubMed  CAS  Google Scholar 

  47. Zhang Y, Bhavnani BR (2005) Glutamate-induced apoptosis in primary cortical neurons is inhibited by equine estrogen via down-regulation of caspase-3 and prevention of mitochondrial cytochrome c release. BMC Neurosci 6:1–23

    Article  Google Scholar 

  48. Tenneti L, Lipton SA (2000) Involvement of activated caspase-3-like proteases in N-methyl-d-aspartate-induced apoptosis in cerebrocortical neurons. J Neurochem 74:134–142

    Article  PubMed  CAS  Google Scholar 

  49. Brewer GJ, Lim A, Capps NG, Torricelli JR (2005) Age-related calcium changes, oxyradical damage, caspase activation and nuclear condensation in hippocampal neurons in response to glutamate and beta-amyloid. Exp Gerontol 40:426–437

    Article  PubMed  CAS  Google Scholar 

  50. Ventura R, Harris KM (1999) Three-dimensional relationships between hippocampal synapses and astrocytes. J Neurosci 19:6897–6906

    PubMed  CAS  Google Scholar 

  51. Garrido R, Mattson M, Hennig B, Toborek M (2000) Nicotine protects against arachidonic-acid-induced caspase activation, cytochrome c release and apoptosis of cultured spinal cord neurons. J Neurochem 76:1395–1403

    Article  Google Scholar 

  52. Hirai K, Sugawara T, Chan PK, Basus VJ, James TL, Litt L (2002) Cytochrome c associated apoptosis during ATP recovery after hypoxia in neonatal rat cerebrocortical slices. J Neurochem 83:309–319

    Article  PubMed  CAS  Google Scholar 

  53. Cho S, Liu D, Fairman D, Li P, Jenkins L, McGonigle P, Wood A (2004) Spatiotemporal evidence of apoptosis-mediated ischemic injury in organotypic hippocampal slice cultures. Neurochem Int 45:117–127

    Article  PubMed  CAS  Google Scholar 

  54. Bonfoco E, Krainc D, Ankarcrona M, Nicotera P, Lipton SP (1995) Apoptosis and necrosis: two distinct events induced, respectively, by mild and intense insult with N-methyl-d-aspartate or nitric oxide in cortical cell cultures. Proc Natl Acad Sci USA 92:7162–7166

    Article  PubMed  CAS  Google Scholar 

  55. Du Y, Bales KR, Dodel RC, Hamilton-Byrd E, Horn JW, Czilli DL, Simmons LK, Ni B, Paul SM (1997) Activation of a caspase 3-related cysteine protease is required for glutamate-mediated apoptosis of cultured cerebellar granule neurons. Proc Natl Acad Sci USA 94(21):11657–11662

    Article  PubMed  CAS  Google Scholar 

  56. Baskys A, Fang L, Bayazitov I (2005) Activation of neuroprotective pathways by metabotropic group I glutamate receptors: a potential target for drug discovery? Ann NY Acad Sci 1053:55–73

    Article  PubMed  CAS  Google Scholar 

  57. Rivera-Cervantes MC, Torres JS, Faria-Velasco A, Armendariz-Borunda J, Beas-Zarate C (2004) NMDA and AMPA expression and cortical neuronal death are associated with p38 in glutamate-induced excitotoxicity in vivo. J Neurosci Res 1:678–687

    Article  Google Scholar 

  58. Torres JES, Chaparro-Huerta V, Rivera-Cervantes MC, Montez-González R, Flores Soto ME, Beas-Zárate C (2006) Neuronal cell death due to glutamate excitotoxicity is mediated by P38 activation in the rat cerebral cortex. Neurosci Lett 403:133–238

    Article  Google Scholar 

  59. Chen J, Errico SL, Freed WJ (2005) Reactive oxygen species and p38 phosphorylation regulate the protective effect of Δ9-tetrahydrocannabinol in the apoptotic response to NMDA. Neurosci Lett 389:99–103

    Article  PubMed  CAS  Google Scholar 

  60. Kawasaki H, Morooka T, Shimohama S, Kimura J, Hirama T, Gotoh Y, Nishida E (1997) Activation and involvement of p38 mitogen-activated protein kinase in glutamate-induced apoptosis in rat cerebellar granule cells. J Biol Chem 272:18518–18521

    Article  PubMed  CAS  Google Scholar 

  61. Alessandrini A, Namura S, Moskowitz MA, Bonventre JV (1999) MEK1 protein kinase inhibition protects against damage resulting from focal cerebral ischemia. Proc Natl Acad Sci USA 96:12866–12869

    Article  PubMed  CAS  Google Scholar 

  62. Jiang Q, Gu Z, Zhang G, Jing J (2000) N-Methyl-d-aspartate receptor activation results in regulation of extracellular signal-regulated kinases by protein kinases and phosphatases in glutamate-induced neuronal apoptotic-like death. Brain Res 887:285–292

    Article  PubMed  CAS  Google Scholar 

  63. Giardina SF, Beart PM (2002) Kainate receptor-mediated apoptosis in primary culture of cerebellar granule cells is attenuated by mitogen-activated protein and cyclin-dependent kinase inhibitors. Br J Pharmacol 135:1733–1742

    Article  PubMed  CAS  Google Scholar 

  64. Seo SR, Chong SA, Lee SI, Sung J, Ahn YS, Chung KC, Seo JT (2001) Zn2+-induced ERK activation mediated by reactive oxygen species causes cell death in differentiated PC12 cells. J Neurochem 78:600–610

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Departamento de Bioquímica, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina, Trindade, 88040-900, Florianopolis, SC, Brazil

    Simone Molz, Helena Decker, Tharine Dal-Cim, Carla Cremonez, Rodrigo B. Leal & Carla I. Tasca

  2. Curso de Farmácia, Universidade do Contestado, 89460-000, Canoinhas, SC, Brazil

    Simone Molz

  3. Setor de Patologia Veterinária, Universidade Federal do Tocantins, 132, Araguaina, TO, Brazil

    Fabiano M. Cordova

Authors
  1. Simone Molz
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Helena Decker
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tharine Dal-Cim
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Carla Cremonez
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Fabiano M. Cordova
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Rodrigo B. Leal
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Carla I. Tasca
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Carla I. Tasca.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Molz, S., Decker, H., Dal-Cim, T. et al. Glutamate-induced Toxicity in Hippocampal Slices Involves Apoptotic Features and p38MAPK Signaling. Neurochem Res 33, 27–36 (2008). https://doi.org/10.1007/s11064-007-9402-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11064-007-9402-1

Keywords

Search

Navigation