Advertisement

Primary Cultures for Testing Neuroprotective Drugs

  • Heike Oberpichler-Schwenk
  • Josef Krieglstein
Part of the Advances in Neuroprotection book series (volume 22)

Abstract

Cerebral ischemia causes neuronal death and impairment of brain function. Numerous experimental approaches have been used to elucidate the underlying pathophysiologic mechanisms and to develop pharmacologic means for the protection of neurons against damage caused by ischemia. For the development of neuroprotective drugs, neuronal cell cultures have become widely used tools. They enable researchers to investigate drug effects on neuronal cells without consideration of pharmacokinetics. Primary neuronal cultures as well as neuronal cell lines are employed for such studies. The present chapter reviews the use of primary neuronal cultures for the investigation of neuroprotective drugs. We shall give a short survey of commonly used primary culture systems and their characteristics. Then we shall discuss various possible ways to experimentally induce and measure neuronal damage in these cultures in analogy to ischemic damage in vivo. Several examples of neuroprotective drug effects demonstrated in vitro will be reviewed.

Keywords

NMDA Receptor Neuronal Culture Cerebellar Granule Cell Primary Neuronal Culture Glutamate Toxicity 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Abele AE, Scholz KP, Scholz WK, Miller RJ (1990): Excitotoxicity induced by enhanced excitatory neurotransmission in cultured hippocampal pyramidal neurons. Neuron 2: 413–419CrossRefGoogle Scholar
  2. Ahlemeyer B, Krieglstein J (1990): Testing drug effects against hypoxic damage of cultured neurons during long-term recovery. Life Sci 45: 835–842CrossRefGoogle Scholar
  3. Atkinson DE (1968): The energy charge of the adenylate pool as a regulatory parameter. Interaction with feed-back modifiers. Biochemistry 7: 4030–4034CrossRefGoogle Scholar
  4. Balâzs R, Jorgensen OS, Hack N (1988): N-Methyl-D-aspartate promotes the survival of cerebellar granule cells in culture. Neuroscience 27: 437–451CrossRefGoogle Scholar
  5. Banker GA, Cowan M (1977): Rat hippocampal neurons in dispersed cell culture. Brain Res 126: 397–425CrossRefGoogle Scholar
  6. Banker GA, Cowan WM (1979): Further observations on hippocampal neurons in dispersed cell culture. J Comp Neurol 187: 469–494CrossRefGoogle Scholar
  7. Benveniste H, Drejer J, Schousboe A, Diemer NH (1984): Elevation of the extracellular concentrations of glutamate and aspartate in rat hippocampus during transient cerebral ischemia monitored by intracerebral microdialysis. J Neurochem 43: 1369–1374CrossRefGoogle Scholar
  8. Bielicki L, Krieglstein J (1977): Solubilization of brain mitochondrial hexokinase by thiopental. Naunyn-Schmiedeberg’s Arch Pharmacol 298: 61–65CrossRefGoogle Scholar
  9. Braestrup C, Honoré T, Nielsen M, Petersen EN, Jensen LH (1984): Ligands of benzodiazepine receptors with positive and negative efficacy. Biochem Pharmacol 33: 859–862CrossRefGoogle Scholar
  10. Choi DW (1988): Glutamate neurotoxicity and diseases of the nervous system. Neuron 1: 623–634CrossRefGoogle Scholar
  11. Choi DW, Maulucci-Gedde MA, Kriegstein AR (1987): Glutamate neurotoxicity in cortical cell culture. J Neurosci 7: 357–368Google Scholar
  12. Choi DW, Koh JY, Peters S (1988): Pharmacology of glutamate neurotoxicity in cortical cell culture: Attenuation by NMDA antagonists. J Neurosci 8: 185–196Google Scholar
  13. Dichter MA (1978): Rat cortical neurons in cell culture: Culture methods, cell morphology, electrophysiology, and synapse formation. Brain Res 149: 279–293CrossRefGoogle Scholar
  14. Favaron M, Manev H, Alho H, Bertolino M, Ferret B, Guidotti A, Costa E (1988): Gangliosides prevent glutamate and kainate neurotoxicity in primary neuronal cultures of neonatal rat cerebellum and cortex. Proc Natl Acad Sci USA 85: 7351–7355CrossRefGoogle Scholar
  15. Finkbeiner S, Stevens CF (1988): Applications of quantitative measurements for assessing glutamate neurotoxicity. Proc Natl Acad Sci USA 85: 4071–4074CrossRefGoogle Scholar
  16. Fujikara H, Kato H, Nakano S, Kogure K (1989): A serotonin S2 antagonist, naftidrofuryl, exhibited a protective effect on ischemic neuronal damage in the gerbil. Brain Res 494: 387–390CrossRefGoogle Scholar
  17. Gähwiler BH (1981): Organotypic monolayer cultures of nervous tissue. JNeurosci Meth 4: 329–342CrossRefGoogle Scholar
  18. Giffard RG, Monyer H, Christine CW, Choi DW (1990): Acidosis reduces NMDA receptor activation, glutamate neurotoxicity, and oxygen-glucose deprivation neuronal injury in cortical cultures. Brain Res 506: 339–342CrossRefGoogle Scholar
  19. Gill R, Foster AC, Woodruff GN (1987): Systemic administration of MK-801 protects against ischemia-induced hippocampal neurodegeneration in the gerbil. J Neurosci 7: 3343–3349Google Scholar
  20. Goldberg MP, Weiss JH, Pham PC, Choi DW (1987): N-Methyl-D-aspartate receptors mediate hypoxic neuronal injury in cortical culture. J Pharmacol Exp Ther 243: 784–791Google Scholar
  21. Goldberg MP, Viseskul V, Choi DW (1988): Phencyclidine receptor ligands attenuate cortical injury after N-methyl-D-aspartate exposure or hypoxia. J Pharmacol Exp Ther 245: 1081–1087Google Scholar
  22. Goldberg MP, Giffard R, Choi DW (1989): Presynaptic approaches to reduction of hypoxic neuronal injury. In: Pharmacology of Cerebral Ischemia 1988, Krieglstein J, ed. Stuttgart: Wissenschaftliche Verlagsgesellschaft, pp 151–156Google Scholar
  23. Huettner JE, Baughman RW (1986): Primary culture of identified neurons from the visual cortex of postnatal rats. J Neurosci 6: 3044–3060Google Scholar
  24. Johnson JW, Ascher P (1987): Glycine potentiates the NMDA response in cultured mouse brain neurons. Nature 325: 529–531CrossRefGoogle Scholar
  25. Karkoutly C, Backhauss C, Nuglisch J, Krieglstein J (1990): The measurement of the infarcted area after middle cerebral artery occlusion in the mouse: A screening model. In: Pharmacology of Cerebral Ischemia 1990, Krieglstein J, Oberpichler H, eds. Stuttgart: Wissenschaftliche Verlagsgesellschaft, pp 63–69Google Scholar
  26. Kirino T (1982): Delayed neuronal death in the gerbil hippocampus following ischemia. Brain Res 239: 57–69CrossRefGoogle Scholar
  27. Koh JY, Choi DW (1987): Quantitative determination of glutamate mediated cortical neuronal injury in cell culture by lactate dehydrogenase efflux assay. J Neurosci Meth 20: 83–90CrossRefGoogle Scholar
  28. Krieglstein J, Mwasekaga S (1987): Effect of methohexital on the relationship between hexokinase distribution and energy metabolism in neuroblastoma cells. Drug Res 37: 291–295Google Scholar
  29. Krieglstein J, Brungs H, Peruche B (1988): Cultured neurons for testing cerebro-protective drug effects in vitro. J Pharmacol Meth 20: 39–46CrossRefGoogle Scholar
  30. Krieglstein J, Sauer D, Nuglisch J, Rossberg C, Beck T, Bielenberg GW, Mennel HD (1989): Naftidrofuryl protects neurons against ischemic damage. Eur Neurol 29: 224–228CrossRefGoogle Scholar
  31. Louis JC, Pettmann B, Courageot J, Rumigny JF, Mandel P, Sensenbrenner M (1981): Developmental changes in cultured neurons from chick embryo cerebral hemispheres. Exp Brain Res 42: 63–72CrossRefGoogle Scholar
  32. Lysko PG, Cox JA, Vigano MA, Henneberry RC (1989): Excitatory amino acid neurotoxicity at the N-methyl-D-aspartate receptor in cultured neurons: Pharmacological characterization. Brain Res 499: 258–266CrossRefGoogle Scholar
  33. Manev H, Favaron M, Vicini S, Guidotti A, Costa E (1990): Glutamate-induced neuronal death in primary cultures of cerebellar granule cells: Protection by synthetic derivatives of endogenous sphingolipids. J Pharmacol Exp Ther 252: 419–427Google Scholar
  34. Mattson MP (1990): Antigenic changes similar to those seen in neurofibrillary tangles are elicited by glutamate and Cat+ influx in cultured hippocampal neurons. Neuron 2: 105–117CrossRefGoogle Scholar
  35. Mattson MP, Kater SB (1989): Development and selective neurodegeneration in cell cultures from different hippocampal regions. Brain Res 490: 110–125CrossRefGoogle Scholar
  36. Mattson MP, Lee RE, Adams ME, Guthrie PB, Kater SB (1988): Interactions between entorhinal axons and target hippocampal neurons: A role for glutamate in the development of hippocampal circuitry. Neuron 1: 865–876CrossRefGoogle Scholar
  37. Mattson MP, Murrain M, Guthrie PB, Kater SB (1989): Fibroblast growth factor and glutamate: Opposing roles in the generation and degeneration of hippocampal neuroarchitecture. J Neurosci 9: 3728–3740Google Scholar
  38. Messer A (1977): The maintenance and identification of mouse cerebellar granule cells in monolayer culture. Brain Res 130: 1–12CrossRefGoogle Scholar
  39. Miller RJ, Abele AE, Glaum SR, Scholz KP, Scholz WK (1990): Pharmacological aspects of NMDA mediated neuronal death in vitro. In: Pharmacology of Cerebral Ischemia 1990, Krieglstein J, Oberpichler H, eds. Stuttgart: Wissenschaftliche Verlagsgesellschaft, pp 227–238Google Scholar
  40. Nakajima Y, Nakajima S, Leonard RJ, Yamaguchi K (1986): Acetylcholine raises excitability by inhibiting the fast transient potassium current in cultured hippocampal neurons. Proc Natl Acad Sci USA 83: 3022–3026CrossRefGoogle Scholar
  41. Novelli A, Reilly JA, Lysko PG, Henneberry RC (1988): Glutamate becomes neurotoxic via the N-methyl-D-aspartate receptor when intracellular energy levels are reduced. Brain Res 451: 205–212CrossRefGoogle Scholar
  42. Nowak L, Bregestovski P, Ascher P, Herbet A, Prochiantz A (1984): Magnesium gates glutamate-activated channels in mouse central neurons. Nature 307: 462–465CrossRefGoogle Scholar
  43. Oberpichler H, Brungs H, Krieglstein J (1990): Effects of delayed administration of methohexital and ketamine on posthypoxic cell damage of primary neuronal cultures. Pharmacology 40: 165–173CrossRefGoogle Scholar
  44. Olney J (1978): Neurotoxicity of excitatory amino acids. In: Kainic Acid as a Tool in Neurobiology, McGeer EG, Olney JW, McGeer PL, eds. New York: Raven PressGoogle Scholar
  45. Ozyurt E, Graham D, Woodruff GN, McCulloch J (1988): Protective effect of the glutamate antagonist, MK-801, in focal cerebral ischemia in the cat. J Cereb Blood Flow Metab 8: 138–143CrossRefGoogle Scholar
  46. Pauwels PJ, Opperdoes FR, Trouet A (1985): Effects of antimycin, glucose deprivation, and serum on cultures of neurons, astrocytes, and neuroblastoma cells. J Neurochem 44: 143–148CrossRefGoogle Scholar
  47. Peruche B, Ahlemeyer A, Brungs H, Krieglstein J (1990): Cultured neurons for testing antihypoxic drug effects. J Pharmacol Meth 23: 63–77CrossRefGoogle Scholar
  48. Peterson C, Cotman CW (1990): Decreased survival of hippocampal neurons in medium conditioned by fibroblasts from aged and Alzheimer donors. Brain Res 515: 39–44CrossRefGoogle Scholar
  49. Pettmann B, Louis JC, Sensenbrenner M (1979): Morphological and biochemical maturation of neurons cultured in the absence of glial cells. Nature 281: 378–380CrossRefGoogle Scholar
  50. Prehn JHM, Krieglstein J (1991): Primary neuronal cultures from chick embryo cerebral hemispheres: A model for studying trophic and toxic effects of excitatory amino acids. J Cereb Blood Flow Metabol 11 (Suppl 2): S317Google Scholar
  51. Prehn JHM, Peruche B, Karkoutly C, Rossberg C, Mennel HD, Krieglstein J (1990): Dihydrolipoic acid protects neurons against ischemic/hypoxic damage. In: Pharmacology of Cerebral Ischemia 1990, Krieglstein J, Oberpichler H, eds. Stuttgart: Wissenschaftliche Verlagsgesellschaft, pp 357–362Google Scholar
  52. Reynolds IJ, Miller RJ (1988): Tricyclic antidepressants block N-methyl-D-aspartate receptors: Similarities to the action of zinc. Br J Pharmacol 95: 95–102Google Scholar
  53. Rothman SM (1983): Synaptic activity mediates death on hypoxic neurons. Science 220: 536–537CrossRefGoogle Scholar
  54. Rothman S (1984): Synaptic release of excitatory amino acid neurotransmitter mediates anoxic neuronal death. J Neurosci 4: 1884–1891Google Scholar
  55. Rothman SM, Olney JW (1986): Glutamate and the pathophysiology of hypoxicischemic brain damage. Ann Neurol 19: 105–111CrossRefGoogle Scholar
  56. Rothman SM, Samaie M (1985): Physiology of excitatory synaptic transmission in cultures of dissociated rat hippocampus. J Neurophysiol 54: 701–713Google Scholar
  57. Rothman SM, Thurston JH, Hauhart RE (1987a): Delayed neurotoxicity of excitatory amino acids in vitro. Neuroscience 22: 471–480CrossRefGoogle Scholar
  58. Rothman SM, Thurston JH, Hauhart RE, Clark GD, Solomon JS (1987b): Ketamine protects hippocampal neurons from anoxia in vitro. Neuroscience 21: 673–678CrossRefGoogle Scholar
  59. Scott BS (1982): Adult neurons in cell culture: Electrophysiological characterization and use in neurobiological research. Progr Neurobiol 19: 187–211CrossRefGoogle Scholar
  60. Seif el Nasr M, Sauer D, Rossberg C, Mennel HD, Krieglstein J (1989): Effects of NMDA antagonists against neuronal damage after forebrain ischemia in the rat. In: Pharmacology of Cerebral Ischemia 1988, Krieglstein J, ed. Stuttgart: Wissenschaftliche Verlagsgesellschaft, pp 211–215Google Scholar
  61. Seif el Nasr M, Peruche B, Rossberg C, Mennel HD, Krieglstein J (1990): Neuroprotective effect of memantine demonstrated in vivo and in vitro. Eur J Pharmacol 185: 19–24CrossRefGoogle Scholar
  62. Sensenbrenner M, Maderspach K, Latzkovits L, Jaros GG (1978): Neuronal cells from chick embryo cerebral hemispheres cultivated on polylysine-coated surfaces. Dey Neurosci 1: 90–101CrossRefGoogle Scholar
  63. Sheardown MJ, Hansen AJ, Eskesen K, Suzdak P, Diemer NH, Honoré T (1990): Blockade of AMPA receptors in the CAI region of the hippocampus prevents ischaemia induced cell death. In: Pharmacology of Cerebral Ischemia 1990, Krieglstein J, Oberpichler H, eds. Stuttgart: Wissenschaftliche Verlagsgesellschaft, pp 245–253Google Scholar
  64. Siesjö BK, Lundgren J, Pahlmark K (1990): The role of free radicals in ischemic brain damage: A hypothesis. In: Pharmacology of Cerebral Ischemia 1990, Krieglstein J, Oberpichler H, eds. Stuttgart: Wissenschaftliche Verlagsgesellschaft, pp 319–323Google Scholar
  65. Söderbäck M, Hansson E, Tottmar O, Rönnbäck L (1989): Neurons in primary cultures from five defined rat brain regions: Cellular composition and morphological appearance. Cell Mol Biol 35: 1–16Google Scholar
  66. Trussell LO, Thio LL, Zorumski CF, Fischbach GD (1988): Rapid desensitization of glutamate receptors in vertebrate cultured neurons. Proc Natl Acad Sci USA 85: 4562–4566CrossRefGoogle Scholar
  67. Weiss J, Goldberg MP, Choi DW (1986a): Ketamine protects cultured neocortical neurons from hypoxic injury. Brain Res 380: 186–190CrossRefGoogle Scholar
  68. Weiss S, Pin JP, Sebben M, Kemp DE, Sladeczek F, Gabrion J, Bockaert J (1986b): Synaptogenesis of cultured striatal neurons in serum-free medium: A morphological and biochemical study. Proc Natl Acad Sci USA 83: 2238–2242CrossRefGoogle Scholar

Copyright information

© Birkhäuser Boston 1992

Authors and Affiliations

  • Heike Oberpichler-Schwenk
  • Josef Krieglstein

There are no affiliations available

Personalised recommendations