Advertisement

Extending the Life Span of Acute Neuronal Tissue for Imaging and Electrophysiological Studies

  • Yossi BuskilaEmail author
  • Alba Bellot-Saez
  • Orsolya Kékesi
  • Morven Cameron
  • John Morley
Protocol
Part of the Neuromethods book series (NM, volume 152)

Abstract

Slice preparations of neuronal tissue are among the most commonly used experimental approaches in the field of neuroscience. They are employed for a variety of techniques addressing questions across the entire neuroscience spectrum, including immunohistochemical, anatomical, and electrophysiological methods to study the properties of individual, and networks of neurons. In the past decades, slice preparations have provided information that has allowed us to develop our understanding of the central nervous system. Unlike cultures, slice preparations leave the topography of neurons and glia intact and therefore retain a considerable degree of functionality that allows molecular, cellular, and network investigations. However, a major limitation of using acute brain slices is their life span which is limited to 6–8 h due to intrinsic and extrinsic factors. Recently, new technological and methodological modifications have proved efficient in extending the life span of acute neuronal tissue. In this chapter, we will review the mechanisms leading to tissue deterioration and describe in detail the steps required to achieve a significant enhancement in neuronal viability and longevity.

Key words

Brain slices Bacteria Ischemia Braincubator ATP Electrophysiology Incubation 

References

  1. 1.
    Warburg O, Wind F, Negelein E (1926) The metabolism of tumors in the body. J Gen Physiol 8:519–530CrossRefGoogle Scholar
  2. 2.
    Ashford CA, Dixon KC (1935) The effect of potassium on the glucolysis of brain tissue with reference to the Pasteur effect. Biochem J 29:157–168PubMedPubMedCentralGoogle Scholar
  3. 3.
    Dickens F, Greville G (1935) The metabolism of normal and tumour tissue: neutral salt effects. Biochem J 29:1468–1483PubMedPubMedCentralGoogle Scholar
  4. 4.
    Mcilwain H, Buchel L, Cheshire JD (1951) The inorganic phosphate and phosphocreatine of brain especially during metabolism in vitro. Biochem J 48:12–20PubMedPubMedCentralGoogle Scholar
  5. 5.
    Li C-L, Mcilwain H (1957) Maintenance of resting membrane potentials in slices of mammalian cerebral cortex and other tissues in vitro. J Physiol 139:178–190CrossRefGoogle Scholar
  6. 6.
    Hillman H, Mcilwain H (1961) Membrane potentials in mammalian cerebral tissues in vitro: dependence on ionic environment. J Physiol 157:263–278CrossRefGoogle Scholar
  7. 7.
    Yamamoto C, McIlwain H (1966) Electrical activities in thin sections from the mammalian brain maintained in chemically-defined media in vitro. J Neurochem 13:1333–1343CrossRefGoogle Scholar
  8. 8.
    Otsuka M, Konishi S (1974) Electrophysiology of the mammalian spinal curd in vitro. Nature 252:733–734.  https://doi.org/10.1038/252497a0CrossRefPubMedGoogle Scholar
  9. 9.
    Mitra P, Brownstone RM (2012) An in vitro spinal cord slice preparation for recording from lumbar motoneurons of the adult mouse. J Neurophysiol 107:728–741.  https://doi.org/10.1152/jn.00558.2011CrossRefPubMedGoogle Scholar
  10. 10.
    Kerkut G, Bagust J (1995) The isolated mammalian spinal cord. Prog Neurobiol 46:1–48CrossRefGoogle Scholar
  11. 11.
    Takahashi T (1978) Intracellular recording from visually identified motoneurons in rat spinal cord slices. Proc R Soc London 202:417–421CrossRefGoogle Scholar
  12. 12.
    Gibb A, Edwards F (1994) Patch clamp recording from cells in sliced tissues. Microelectrode Tech:255–274Google Scholar
  13. 13.
    Rekling JC, Funk GD, D a B et al (2000) Synaptic control of motoneuronal excitability. Physiol Rev 80:767–852.  https://doi.org/10.1001/jama.2014.15298.MetforminCrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Carp JS, Tennissen AM, Mongeluzi DL et al (2008) An in vitro protocol for recording from spinal motoneurons of adult rats. J Neurophysiol 100:474–481.  https://doi.org/10.1152/jn.90422.2008CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Carlin KP, Jiang Z, Brownstone RM (2000) Characterization of calcium currents in functionally mature mouse spinal motoneurons. Eur J Neurosci 12:1624–1634CrossRefGoogle Scholar
  16. 16.
    Bennett DJ, Li Y, Siu M (2001) Plateau potentials in sacrocaudal motoneurons of chronic spinal rats, recorded in vitro. J Neurophysiol 86:1955–1971CrossRefGoogle Scholar
  17. 17.
    Genovese T, Esposito E, Mazzon E et al (2009) Beneficial effects of ethyl pyruvate in a mouse model of spinal cord injury. Shock 32:217–227.  https://doi.org/10.1097/SHK.0b013e31818d4073CrossRefPubMedGoogle Scholar
  18. 18.
    Ottoson D, Svaetichin G (1953) The electrical activity of the retinal receptor layer. Acta Physiol Scand 29:31–39.  https://doi.org/10.1111/j.1748-1716.1953.tb00995.xCrossRefPubMedGoogle Scholar
  19. 19.
    Tomita T (1965) Electrophysiological study of the mechanisms subserving color coding in the fish retina. Cold Spring Harb Symp Quant Biol 30:559–566.  https://doi.org/10.1101/SQB.1965.030.01.054CrossRefPubMedGoogle Scholar
  20. 20.
    Buskila Y, Farkash S, Hershfinkel M, Amitai Y (2005) Rapid and reactive nitric oxide production by astrocytes in mouse neocortical slices. Glia 52:169–176CrossRefGoogle Scholar
  21. 21.
    Cameron MA, Al AA, Buskila Y et al (2017) Differential effect of brief electrical stimulation on voltage-gated potassium channels. J Neurophysiol.  https://doi.org/10.1152/jn.00915.2016CrossRefGoogle Scholar
  22. 22.
    Buskila Y, Crowe SE, Ellis-Davies GCR (2013) Synaptic deficits in layer 5 neurons precede overt structural decay in 5xFAD mice. Neuroscience 254:152–159CrossRefGoogle Scholar
  23. 23.
    Flynn JR, Conn VL, Boyle KA et al (2017) Anatomical and molecular properties of long descending propriospinal neurons in mice. Front Neuroanat 11:1–13.  https://doi.org/10.3389/fnana.2017.00005CrossRefGoogle Scholar
  24. 24.
    Agmon A, Connors B (1991) Thalamocortical responses of mouse somatosensory (barrel) cortexin vitro. Neuroscience 41:365–379CrossRefGoogle Scholar
  25. 25.
    Buskila Y, Morley JW, Tapson J, van Schaik A (2013) The adaptation of spike backpropagation delays in cortical neurons. Front Cell Neurosci 7:192.  https://doi.org/10.3389/fncel.2013.00192CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Kantevari S, Buskila Y, Ellis-Davies GCR (2012) Synthesis and characterization of cell-permeant 6-nitrodibenzofuranyl-caged IP3. Photochem Photobiol Sci 11:508–513CrossRefGoogle Scholar
  27. 27.
    Aghajanian GK, Rasmussen K (1989) Intracellular studies in the facial nucleus illustrating a simple new method for obtaining viable motoneurons in adult rat brain slices. Synapse 3:331–338.  https://doi.org/10.1002/syn.890030406CrossRefPubMedGoogle Scholar
  28. 28.
    Tanaka Y, Tanaka Y, Furuta T et al (2008) The effects of cutting solutions on the viability of GABAergic interneurons in cerebral cortical slices of adult mice. J Neurosci Methods 171:118–125.  https://doi.org/10.1016/j.jneumeth.2008.02.021CrossRefPubMedGoogle Scholar
  29. 29.
    Hong J, Zhang J, Xiao C, Kong J (2006) Patch-clamp studies in the CNS illustrate a simple new method for obtaining viable neurons in rat brain slices: glycerol replacement of NaCl protects CNS neurons. J Neurosci Methods 158:251–259.  https://doi.org/10.1016/j.jneumeth.2006.06.006CrossRefGoogle Scholar
  30. 30.
    Ting J, Daigle T, Chen Q, Feng G (2014) Acute brain slice methods for adult and aging animals: application of targeted patch clamp analysis and Optogenetics. Methods Mol Biol 1183:221–242.  https://doi.org/10.1007/978-1-4939-1096-0CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Brahma B, Forman RE, Stewart EE et al (2000) Ascorbate inhibits edema in brain slices. J Neurochem 74:1263–1270.  https://doi.org/10.1046/j.1471-4159.2000.741263.xCrossRefPubMedGoogle Scholar
  32. 32.
    Connors BW, Gutnick MJ, Prince DA (1982) Electrophysiological properties of neocortical neurons in vitro. J Neurophysiol 48:1302–1320CrossRefGoogle Scholar
  33. 33.
    Llinás R, Muhlethaler M (1988) An electrophysiological study of the in vitro, perfused brain stem-cerebellum of adult Guinea-pig. J Physiol 404:215–240CrossRefGoogle Scholar
  34. 34.
    Geiger JR, Bischofberger J, Vida I et al (2002) Patch-clamp recording in brain slices with improved slicer technology. Eur J Phys 443:491–501.  https://doi.org/10.1007/s00424-001-0735-3CrossRefGoogle Scholar
  35. 35.
    Buskila Y, Breen PP, Tapson J et al (2014) Extending the viability of acute brain slices. Sci Rep 4:4–10.  https://doi.org/10.1038/srep05309CrossRefGoogle Scholar
  36. 36.
    Aitken PG, Dudek FE, Eskessen K et al (1995) Making the best of brain slices: comparing preparative methods. J Neurosci Methods 59:151–156CrossRefGoogle Scholar
  37. 37.
    Cameron M, Kekesi O, Morley JW et al (2016) Calcium imaging of am dyes following prolonged incubation in acute neuronal tissue. PLoS One 11:e0155468.  https://doi.org/10.1371/journal.pone.0155468CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Breen PP, Buskila Y (2014) Braincubator: an incubation system to extend brain slice lifespan for use in neurophysiology. In: 2014 36th Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE, pp 4864–4867Google Scholar
  39. 39.
    Fujikawa DG (2015) The role of Excitotoxic programmed necrosis in acute brain injury. CSBJ 13:212–221.  https://doi.org/10.1016/j.csbj.2015.03.004CrossRefPubMedGoogle Scholar
  40. 40.
    Karnatovskaia LV, Wartenberg KE, Freeman WD (2014) Therapeutic hypothermia for neuroprotection: history, mechanisms, risks, and clinical applications. The Neurohospitalist 4:153–163.  https://doi.org/10.1177/1941874413519802CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Whittingham TS, Lust WD, Christakis DA, Passonneau JV (1984) Metabolic stability of hippocampal slice preparations during prolonged incubation. J Neurochem 43:689–696CrossRefGoogle Scholar
  42. 42.
    Feig S, Lipton P (1990) N-methyl-D-aspartate receptor activation and Ca2+ account for poor pyramidal cell structure in hippocampal slices. J Neurochem 55:473–483CrossRefGoogle Scholar
  43. 43.
    Davie JT, Kole MHP, Letzkus JJ et al (2006) Dendritic patch-clamp recording. Nat Protoc 1:1235–1247CrossRefGoogle Scholar
  44. 44.
    Rice ME (1999) Use of ascorbate in the preparation and maintenance of brain slices. Methods 18:144–149.  https://doi.org/10.1006/meth.1999.0767CrossRefPubMedGoogle Scholar
  45. 45.
    Lee J, Grabb MC, Zipfel GJ, Choi DW (2000) Tissue responses to ischemia brain tissue responses to ischemia. J Clin Invest 106:723–731CrossRefGoogle Scholar
  46. 46.
    Verkhratsky a KH (1996) Calcium signalling in glial cells. Trends Neurosci 19:346–352.  https://doi.org/10.1016/0166-2236(96)10048-5CrossRefPubMedGoogle Scholar
  47. 47.
    Won SJ, Kim DY, Gwag BJ (2002) Cellular and molecular pathways of ischemic neuronal death. J Biochem Mol Biol 35:67–86PubMedGoogle Scholar
  48. 48.
    Lee J, Grabb MC, Zipfel GJ, Choi DW (2000) Brain tissue responses to ischemia. J Clin Invest 106:723–731CrossRefGoogle Scholar
  49. 49.
    Silver I, Erecinska M (1998) Oxygen and ion concentrations in normoxic and hypoxic brain cells. In: Oxygen transport to tissue XX. Springer, Boston, MA, pp 7–16CrossRefGoogle Scholar
  50. 50.
    Moyer JR, Brown TH (2002) Patch-clamp techniques applied to brain slices. In: Patch-clamp anal. Humana Press, New Jersey, pp 135–193CrossRefGoogle Scholar
  51. 51.
    Buskila Y, Amitai Y (2010) Astrocytic iNOS-dependent enhancement of synaptic release in mouse neocortex. J Neurophysiol 103:1322–1328.  https://doi.org/10.1152/jn.00676.2009CrossRefPubMedGoogle Scholar
  52. 52.
    Aitken PG, Breese GR, Dudek FF et al (1995) Preparative methods for brain slices: a discussion. J Neurosci Methods 59:139–149CrossRefGoogle Scholar
  53. 53.
    Cohen P (1997) The search for physiological substrates of MAP and SAP kinases in mammalian cells. Trends Cell Biol 8924:353–361CrossRefGoogle Scholar
  54. 54.
    Espanol MT (1996) Adult rat brain-slice preparation for NMR studies of hypoxia. Anesthesiology 84:201–210CrossRefGoogle Scholar
  55. 55.
    Watson GB, Lopez OT, Charles VD, Lanthorn TH (1994) Assessment of long-term effects of transient anoxia on metabolic activity of rat hippocampal slices using triphenyltetrazolium chloride. J Neurosci Methods 53:203–208CrossRefGoogle Scholar
  56. 56.
    Ames A, Nesbett FB (1981) In vitro retina as an experimental model of the central nervous system. J Neurochem 37:867–877.  https://doi.org/10.1111/j.1471-4159.1981.tb04473.xCrossRefPubMedGoogle Scholar
  57. 57.
    Hudson B, Uphol WB, Devinny J, Vinograd J (1969) The use of an ethidium analogue in the dye-buoyant density procedure for the isolation of closed circular DNA: the variation of the superhelix density of mitochondrial DNA. Biochemistry 62:813–820Google Scholar
  58. 58.
    Latt SA (1973) Microfluorometric detection of deoxyribonucleic acid replication in human metaphase chromosomes. Proc Natl Acad Sci 70:3395–3399CrossRefGoogle Scholar
  59. 59.
    Williamson D, Fennell D (1975) The use of fluorescent DNA-binding agent for detecting and separating yeast mitochondrial DNA. Methods Cell Biol 12:335–352CrossRefGoogle Scholar
  60. 60.
    Monette R, Small DL, Mealing G, Morley P (1998) A fluorescence confocal assay to assess neuronal viability in brain slices. Brain Res Brain Res Protoc 2:99–108CrossRefGoogle Scholar
  61. 61.
    Khatri N, Man HY (2013) Synaptic activity and bioenergy homeostasis: implications in brain trauma and neurodegenerative diseases. Front Neurol 4:199.  https://doi.org/10.3389/fneur.2013.00199CrossRefPubMedPubMedCentralGoogle Scholar
  62. 62.
    Malarkey EB, Parpura V (2008) Mechanisms of glutamate release from astrocytes. Neurochem Int 52:142–154CrossRefGoogle Scholar
  63. 63.
    Reid KH, Edmonds HL, Schurr A et al (1988) Pitfalls in the use of brain slices. Prog Neurobiol 31:1–18CrossRefGoogle Scholar
  64. 64.
    Barone FC, Raymond ZF (1997) Brain cooling during transient focal ischemia provides complete neuroprotection. Neurosci Biobehav Rev 21:31–44CrossRefGoogle Scholar
  65. 65.
    Erecinska M, Thoresen M, Silver IA (2003) Effects of hypothermia on energy metabolism in mammalian central nervous system. J Cereb Blood Flow Metab 23:513–530.  https://doi.org/10.1097/01.WCB.0000066287.21705.21CrossRefPubMedGoogle Scholar
  66. 66.
    Attwell D, Laughlin SB (2001) An energy budget for signaling in the Grey matter of the brain. J Cereb Blood Flow Metab 21:1133–1145.  https://doi.org/10.1097/00004647-200110000-00001CrossRefPubMedGoogle Scholar
  67. 67.
    Waardes A, Van Thillart G, Van Den Erkelensy C et al (1990) Functional coupling of glycolysis and phosphocreatine utilization in anoxic fish muscle. J Biol Chem 265:914–923Google Scholar
  68. 68.
    Tisherman SA, Sterz F (2005) Therapeutic hypothermia. SpringerGoogle Scholar
  69. 69.
    Hicks SD, DeFranco DB, Callaway CW (2000) Hypothermia during reperfusion after asphyxial cardiac arrest improves functional recovery and selectively alters stress-induced protein expression. J Cereb Blood Flow Metab 20:520–530.  https://doi.org/10.1097/00004647-200003000-00011CrossRefPubMedGoogle Scholar
  70. 70.
    Canevari L, Console A, Tendi EA et al (1999) Effect of postischaemic hypothermia on the mitochondrial damage induced by ischaemia and reperfusion in the gerbil. Brain Res 817:241–245.  https://doi.org/10.1016/S0006-8993(98)01278-5CrossRefPubMedGoogle Scholar
  71. 71.
    Richerson GB, Messer C (1995) Effect of composition of experimental solutions on neuronal survival during rat brain slicing. Exp Neurol 131:133–143.  https://doi.org/10.1016/0014-4886(95)90015-2CrossRefPubMedGoogle Scholar
  72. 72.
    Yu DY, Cringle SJ (2001) Oxygen distribution and consumption within the retina in vascularised and avascular retinas and in animal models of retinal disease. Prog Retin Eye Res 20:175–208.  https://doi.org/10.1016/S1350-9462(00)00027-6CrossRefGoogle Scholar
  73. 73.
    Cameron MA, Suaning GJ, Lovell NH, Morley JW (2013) Electrical stimulation of inner retinal neurons in wild-type and retinally degenerate (rd/rd) mice. PLoS One 8:e68882.  https://doi.org/10.1371/journal.pone.0068882CrossRefGoogle Scholar
  74. 74.
    Medrano CJ, Fox DA (1995) Oxygen consumption in the rat outer and inner retina: light- and pharmacologically-induced inhibition. Exp Eye Res 61:273–284.  https://doi.org/10.1016/S0014-4835(05)80122-8CrossRefPubMedGoogle Scholar
  75. 75.
    Cameron M, Kékesi O, Morley JW et al (2016) Calcium imaging of AM dyes following prolonged incubation in acute neuronal tissue. PLoS One 11:e0155468.  https://doi.org/10.1371/journal.pone.0155468CrossRefPubMedPubMedCentralGoogle Scholar
  76. 76.
    Feigenspan A, Babai NZ (2017) Preparation of horizontal slices of adult mouse retina for electrophysiological studies. J Vis Exp:1–6.  https://doi.org/10.3791/55173
  77. 77.
    Kulkarni M, Schubert T, Baden T et al (2015) Imaging Ca2+ dynamics in cone photoreceptor axon terminals of the mouse retina. J Vis Exp:e52588.  https://doi.org/10.3791/52588
  78. 78.
    Buskila Y, Breen P, Wright J (2015) Device for storing a tissue sample WO2015021513Google Scholar
  79. 79.
    Cameron MA, Kekesi O, Morley JW et al (2017) Prolonged incubation of acute neuronal tissue for electrophysiology and. J Vis Exp 120:1–6.  https://doi.org/10.3791/55396CrossRefGoogle Scholar
  80. 80.
    Grøndahl TO, Langmoen IA (1993) Epileptogenic effect of antibiotic drugs. J Neurosurg 78:938–943.  https://doi.org/10.3171/jns.1993.78.6.0938CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Yossi Buskila
    • 1
    • 2
    Email author
  • Alba Bellot-Saez
    • 1
    • 2
  • Orsolya Kékesi
    • 1
    • 2
  • Morven Cameron
    • 2
  • John Morley
    • 2
  1. 1.The MARCS InstituteWestern Sydney UniversitySydneyAustralia
  2. 2.School of MedicineWestern Sydney UniversitySydneyAustralia

Personalised recommendations