Skip to main content
SpringerLink
Log in
Book cover

Isolated Central Nervous System Circuits pp 125–158Cite as

Physiological Properties of Human Fetal Cortex In Vitro

  • Protocol
  • First Online:

Part of the book series: Neuromethods ((NM,volume 73))

Abstract

Emergence of neuronal membrane excitability (e.g., voltage-gated ion conductances) and spontaneous electrical activity (e.g., spontaneous firing of action potentials) is an imperative for the normal development and maturation of brain circuits. Understanding the interplay between electrical activity and normal brain development may prove critical for prevention and therapy of devastating neurological and psychiatric diseases. Due to the limited availability of human fetal tissue, our current understanding of functional (physiological) maturation of the human cerebral cortex has been limited to in vivo and in vitro animal models. Although invaluable for the generation of basic insights into this process, animal models fall short in providing an accurate model of human cortical development. The discrepancy between animal and human models of brain development is largely due to evolutionary changes such as differences in the time course, number of cellular divisions, and cellular composition of cortical layers, making it difficult to ascertain the step by step physiological maturation in human. Here, we provide detailed methodology on how to handle human fetal tissue, generate acute brain slices, and perform whole-cell patch-clamp recordings of biophysical membrane properties in human cortical neurons in their natural surroundings, that is, preserved neighboring neurons, glia, and extracellular matrix.

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

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   179.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Zhang LI, Poo MM (2001) Electrical activity and development of neural circuits. Nat Neurosci 4:1207–1214

    Article  PubMed  CAS  Google Scholar 

  2. Spitzer NC (2006) Electrical activity in early neuronal development. Nature 444:707–712

    Article  PubMed  CAS  Google Scholar 

  3. Price DJ, Kennedy H, Dehay C, Zhou L, Mercier M, Jossin Y, Goffinet AM, Tissir F, Blakey D, Molnar Z (2006) The development of cortical connections. Eur J Neurosci 23:910–920

    Article  PubMed  Google Scholar 

  4. Sakmann B, Neher E (1984) Patch clamp techniques for studying ionic channels in excitable membranes. Annu Rev Physiol 46:455–472

    Article  PubMed  CAS  Google Scholar 

  5. Moody WJ, Bosma MM (2005) Ion channel development, spontaneous activity, and activity-dependent development in nerve and muscle cells. Physiol Rev 85:883–941

    Article  PubMed  CAS  Google Scholar 

  6. Moore AR, Filipovic R, Mo Z, Rasband MN, Zecevic N, Antic SD (2009) Electrical excitability of early neurons in the human cerebral cortex during the second trimester of gestation. Cereb Cortex 19:1795–1805

    Article  PubMed  Google Scholar 

  7. Luhmann HJ, Hanganu I, Kilb W (2003) Cellular physiology of the neonatal rat cerebral cortex. Brain Res Bull 60:345–353

    Article  PubMed  CAS  Google Scholar 

  8. Luhmann HJ, Reiprich RA, Hanganu I, Kilb W (2000) Cellular physiology of the neonatal rat cerebral cortex: intrinsic membrane properties, sodium and calcium currents. J Neurosci Res 62:574–584

    Article  PubMed  CAS  Google Scholar 

  9. Kerkovich DM, Sapp D, Weidenheim K, Brosnan CF, Pfeiffer SE, Yeh HH, Busciglio J (1999) Fetal human cortical neurons grown in culture: morphological differentiation, biochemical correlates and development of electrical activity. Int J Dev Neurosci 17:347–356

    Article  PubMed  CAS  Google Scholar 

  10. Khazipov R, Esclapez M, Caillard O, Bernard C, Khalilov I, Tyzio R, Hirsch J, Dzhala V, Berger B, Ben-Ari Y (2001) Early development of neuronal activity in the primate hippocampus in utero. J Neurosci 21:9770–9781

    PubMed  CAS  Google Scholar 

  11. Chiu FC, Rozental R, Bassallo C, Lyman WD, Spray DC (1994) Human fetal neurons in culture: intercellular communication and voltage- and ligand-gated responses. J Neurosci Res 38:687–697

    Article  PubMed  CAS  Google Scholar 

  12. Johnson MA, Weick JP, Pearce RA, Zhang SC (2007) Functional neural development from human embryonic stem cells: accelerated synaptic activity via astrocyte coculture. J Neurosci 27:3069–3077

    Article  PubMed  CAS  Google Scholar 

  13. Picken Bahrey HL, Moody WJ (2003) Early development of voltage-gated ion currents and firing properties in neurons of the mouse cerebral cortex. J Neurophysiol 89:1761–1773

    Article  PubMed  Google Scholar 

  14. Mo Z, Moore AR, Filipovic R, Ogawa Y, Kazuhiro I, Antic SD, Zecevic N (2007) Human cortical neurons originate from radial glia and neuron-restricted progenitors. J Neurosci 27:4132–4145

    Article  PubMed  CAS  Google Scholar 

  15. Molnar Z, Metin C, Stoykova A, Tarabykin V, Price DJ, Francis F, Meyer G, Dehay C, Kennedy H (2006) Comparative aspects of cerebral cortical development. Eur J Neurosci 23:921–934

    Article  PubMed  Google Scholar 

  16. Rakic P (1995) A small step for the cell, a giant leap for mankind: a hypothesis of neocortical expansion during evolution. Trends Neurosci 18:383–388

    Article  PubMed  CAS  Google Scholar 

  17. Jiang C, Haddad GG (1992) Differential responses of neocortical neurons to glucose and/or O2 deprivation in the human and rat. J Neurophysiol 68:2165–2173

    PubMed  CAS  Google Scholar 

  18. Beck H, Goussakov IV, Lie A, Helmstaedter C, Elger CE (2000) Synaptic plasticity in the human dentate gyrus. J Neurosci 20:7080–7086

    PubMed  CAS  Google Scholar 

  19. Broicher T, Speckmann EJ (2012) Living human brain slices: network analysis using voltage sensitive dyes. In Isolated Central Nervous System Circuits (Ed K Ballanyi), Neuromethods Series Vol. 73 (Ed W Walz). Springer Science+Business Media, LLC, New York, NY, pp 285–300

    Google Scholar 

  20. Vanhatalo S, Kaila K (2006) Development of neonatal EEG activity: from phenomenology to physiology. Semin Fetal Neonatal Med 11:471–478

    Article  PubMed  Google Scholar 

  21. Kostovic I, Judas M (2006) Prolonged coexistence of transient and permanent circuitry elements in the developing cerebral cortex of fetuses and preterm infants. Dev Med Child Neurol 48:388–393

    Article  PubMed  Google Scholar 

  22. Andre M, Lamblin MD, D’Allest AM, Curzi-Dascalova L, Moussalli-Salefranque F, Nguyen The Tich S, Vecchierini-Blineau MF, Wallois F, Walls-Esquivel E, Plouin P (2010) Electro­encephalography in premature and full-term infants. Developmental features and glossary. Neurophysiol Clin 40:59–124

    Article  PubMed  CAS  Google Scholar 

  23. Wilson CJ (2008) Up and down states. Scholarpedia 3, p 1410, revision #68992

    Google Scholar 

  24. Chauvette S, Volgushev M, Timofeev I (2010) Origin of active states in local neocortical networks during slow sleep oscillation. Cereb Cortex 20:2660–2674

    Article  PubMed  Google Scholar 

  25. Trapp S, Ballanyi K (2012) Autonomic nervous system in vitro: studying tonically active neurons controlling vagal outflow in rodent brainstem slices. In Isolated Central Nervous System Circuits (Ed K Ballanyi), Neuromethods Series Vol. 73 (Ed W Walz). Springer Science+Business Media, LLC, New York, NY, pp 1–59

    Google Scholar 

  26. Ruangkittisakul A, Panaitescu B, Secchia L, Bobocea N, Kantor C, Kuribayashi J, Iizuka M, Ballanyi K (2012) Anatomically ‘Calibrated’ Isolated Respiratory Networks from Newborn Rodents. In Isolated Central Nervous System Circuits (Ed K Ballanyi), Neuromethods Series Vol. 73 (Ed W Walz). Springer Science+Business Media, LLC, New York, NY, pp 61–124

    Google Scholar 

  27. Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch 391:85–100

    Article  PubMed  CAS  Google Scholar 

  28. Molleman A (2003) Patch clamping: an introductory guide to patch clamp electrophysiology. Wiley, West Sussex

    Google Scholar 

  29. Somjen GG (2002) Ion regulation in the brain: implications for pathophysiology. Neuro­scientist 8:254–267

    PubMed  CAS  Google Scholar 

  30. Sanchez-Vives MV (2012) Spontaneous rhythmic activity in the adult cerebral cortex in vitro. In Isolated Central Nervous System Circuits (Ed K Ballanyi), Neuromethods Series Vol. 73 (Ed W Walz). Springer Science+Business Media, LLC, New York, NY, pp 263–284

    Google Scholar 

  31. Kantor C, Panaitescu B, Kuribayashi J, Ruangkittisakul A, Jovanovic I, Leung V, Lee TF, MacTavish D, Jhamandas JH, Cheung PY, Ballanyi K (2012) Spontaneous neural network oscillations in hippocampus, cortex and locus coeruleus of newborn rat and piglet brain slices. In Isolated Central Nervous System Circuits (Ed K Ballanyi), Neuromethods Series Vol. 73 (Ed W Walz). Springer Science+Business Media, LLC, New York, NY, pp 315–356

    Google Scholar 

  32. Kostovic I, Rakic P (1990) Developmental history of the transient subplate zone in the visual and somatosensory cortex of the macaque monkey and human brain. J Comp Neurol 297:441–470

    Article  PubMed  CAS  Google Scholar 

  33. Allendoerfer KL, Shatz CJ (1994) The subplate, a transient neocortical structure: its role in the development of connections between thalamus and cortex. Annu Rev Neurosci 17:185–218

    Article  PubMed  CAS  Google Scholar 

  34. Kanold PO, Luhmann HJ (2010) The subplate and early cortical circuits. Annu Rev Neurosci 33:23–48

    Article  PubMed  CAS  Google Scholar 

  35. Moore AR, Zhou WL, Jakovcevski I, Zecevic N, Antic SD (2011) Spontaneous electrical activity in the human fetal cortex in vitro. J Neurosci 31:2391–2398

    Article  PubMed  CAS  Google Scholar 

  36. Frankenhaeuser B, Hodgkin AL (1957) The action of calcium on the electrical properties of squid axons. J Physiol 137:218–244

    PubMed  CAS  Google Scholar 

  37. Acker CD, Antic SD (2009) Quantitative assessment of the distributions of membrane conductances involved in action potential backpropagation along basal dendrites. J Neurophysiol 101:1524–1541

    Article  PubMed  Google Scholar 

  38. Zecevic N, Chen Y, Filipovic R (2005) Contributions of cortical subventricular zone to the development of the human cerebral cortex. J Comp Neurol 491:109–122

    Article  PubMed  Google Scholar 

  39. Poolos NP, Jones TD (2004) Patch-clamp recording from neuronal dendrites. Curr Protoc Neurosci Chapter 6: p Unit 6 19

    Google Scholar 

  40. Rakic S, Zecevic N (2003) Emerging complexity of layer I in human cerebral cortex. Cereb Cortex 13:1072–1083

    Article  PubMed  Google Scholar 

  41. Neher E, Sakmann B (1992) The patch clamp technique. Sci Am 266:44–51

    Article  PubMed  CAS  Google Scholar 

  42. Barbour B (2010) Electronics for electrophysiologists tutorial. http://www.biologie.ens.fr/perso/barbour/

  43. Neher E (1992) Correction for liquid junction potentials in patch clamp experiments. Methods Enzymol 207:123–131

    Article  PubMed  CAS  Google Scholar 

  44. Burgard EC, Hablitz JJ (1993) Developmental changes in NMDA and non-NMDA receptor-mediated synaptic potentials in rat neocortex. J Neurophysiol 69:230–240

    PubMed  CAS  Google Scholar 

  45. Kim HG, Fox K, Connors BW (1995) Properties of excitatory synaptic events in neurons of primary somatosensory cortex of neonatal rats. Cereb Cortex 5:148–157

    Article  PubMed  CAS  Google Scholar 

  46. Connors BW, Benardo LS, Prince DA (1983) Coupling between neurons of the developing rat neocortex. J Neurosci 3:773–782

    PubMed  CAS  Google Scholar 

  47. Loturco JJ, Blanton MG, Kriegstein AR (1991) Initial expression and endogenous activation of NMDA channels in early neocortical development. J Neurosci 11:792–799

    PubMed  CAS  Google Scholar 

  48. Mienville JM, Lange GD, Barker JL (1994) Reciprocal expression of cell-cell coupling and voltage-dependent Na current during embryogenesis of rat telencephalon. Brain Res Dev Brain Res 77:89–95

    Article  PubMed  CAS  Google Scholar 

  49. McCormick DA, Prince DA (1987) Post-natal development of electrophysiological properties of rat cerebral cortical pyramidal neurones. J Physiol 393:743–762

    PubMed  CAS  Google Scholar 

  50. Beckh S, Noda M, Lubbert H, Numa S (1989) Differential regulation of three sodium channel messenger RNAs in the rat central nervous system during development. EMBO J 8:3611–3616

    PubMed  CAS  Google Scholar 

  51. Goldin AL (2001) Resurgence of sodium channel research. Annu Rev Physiol 63:871–894

    Article  PubMed  CAS  Google Scholar 

  52. Boiko T, Rasband MN, Levinson SR, Caldwell JH, Mandel G, Trimmer JS, Matthews G (2001) Compact myelin dictates the differential targeting of two sodium channel isoforms in the same axon. Neuron 30:91–104

    Article  PubMed  CAS  Google Scholar 

  53. Hille B (2001) Ion channels of excitable membranes, 3rd edn. Sinauer, Sunderland

    Google Scholar 

  54. Rush AM, Dib-Hajj SD, Waxman SG (2005) Electrophysiological properties of two axonal sodium channels, Nav1.2 and Nav1.6, expressed in mouse spinal sensory neurones. J Physiol 564:803–815

    Article  PubMed  CAS  Google Scholar 

  55. Felts PA, Yokoyama S, Dib-Hajj S, Black JA, Waxman SG (1997) Sodium channel alpha-subunit mRNAs I, II, III, NaG, Na6 and hNE (PN1): different expression patterns in developing rat nervous system. Brain Res Mol Brain Res 45:71–82

    Article  PubMed  CAS  Google Scholar 

  56. Mechaly I, Scamps F, Chabbert C, Sans A, Valmier J (2005) Molecular diversity of voltage-gated sodium channel alpha subunits expressed in neuronal and non-neuronal excitable cells. Neuroscience 130:389–396

    Article  PubMed  CAS  Google Scholar 

  57. Meadows LS, Chen YH, Powell AJ, Clare JJ, Ragsdale DS (2002) Functional modulation of human brain Nav1.3 sodium channels, expressed in mammalian cells, by auxiliary beta 1, beta 2 and beta 3 subunits. Neuroscience 114:745–753

    Article  PubMed  CAS  Google Scholar 

  58. Pan F, Beam KG (1999) The absence of resurgent sodium current in mouse spinal neurons. Brain Res 849:162–168

    Article  PubMed  CAS  Google Scholar 

  59. Shibata R, Misonou H, Campomanes CR, Anderson AE, Schrader LA, Doliveira LC, Carroll KI, Sweatt JD, Rhodes KJ, Trimmer JS (2003) A fundamental role for KChIPs in determining the molecular properties and trafficking of Kv4.2 potassium channels. J Biol Chem 278:36445–36454

    Article  PubMed  CAS  Google Scholar 

  60. Mennerick S, Zorumski CF (2000) Neural activity and survival in the developing nervous system. Mol Neurobiol 22:41–54

    Article  PubMed  CAS  Google Scholar 

  61. Zito K, Svoboda K (2002) Activity-dependent synaptogenesis in the adult Mammalian cortex. Neuron 35:1015–1017

    Article  PubMed  CAS  Google Scholar 

  62. Zhang ZW (2004) Maturation of layer V pyramidal neurons in the rat prefrontal cortex: intrinsic properties and synaptic function. J Neurophysiol 91:1171–1182

    Article  PubMed  Google Scholar 

  63. Katz LC, Shatz CJ (1996) Synaptic activity and the construction of cortical circuits. Science 274:1133–1138

    Article  PubMed  CAS  Google Scholar 

  64. Rakic P, Komuro H (1995) The role of receptor/channel activity in neuronal cell migration. J Neurobiol 26:299–315

    Article  PubMed  CAS  Google Scholar 

  65. Arnold SE, Hyman BT, Van Hoesen GW, Damasio AR (1991) Some cytoarchitectural abnormalities of the entorhinal cortex in schizophrenia. Arch Gen Psychiatry 48:625–632

    Article  PubMed  CAS  Google Scholar 

  66. Akbarian S, Kim JJ, Potkin SG, Hetrick WP, Bunney WE Jr, Jones EG (1996) Maldistribution of interstitial neurons in prefrontal white matter of the brains of schizophrenic patients. Arch Gen Psychiatry 53:425–436

    Article  PubMed  CAS  Google Scholar 

  67. Kanold PO (2004) Transient microcircuits formed by subplate neurons and their role in functional development of thalamocortical connections. Neuroreport 15:2149–2153

    Article  PubMed  Google Scholar 

  68. Ruangkittisakul A, Secchia L, Bornes TD, Palathinkal DM, Ballanyi K (2007) Dependence on extracellular Ca2+/K+ antagonism of inspiratory centre rhythms in slices and en bloc preparations of newborn rat brainstem. J Physiol 584:489–508

    Article  PubMed  CAS  Google Scholar 

  69. Agopyan N, Avoli M (1988) Synaptic and non-synaptic mechanisms underlying low calcium bursts in the in vitro hippocampal slice. Exp Brain Res 73:533–540

    Article  PubMed  CAS  Google Scholar 

  70. Haas HL, Jefferys JG (1984) Low-calcium field burst discharges of CA1 pyramidal neurones in rat hippocampal slices. J Physiol 354:185–201

    PubMed  CAS  Google Scholar 

  71. Yaari Y, Konnerth A, Heinemann U (1983) Spontaneous epileptiform activity of CA1 hippocampal neurons in low extracellular calcium solutions. Exp Brain Res 51:153–156

    Article  PubMed  CAS  Google Scholar 

  72. Rasband MN, Trimmer JS (2001) Developmental clustering of ion channels at and near the node of Ranvier. Dev Biol 236:5–16

    Article  PubMed  CAS  Google Scholar 

  73. Berghs S, Aggujaro D, Dirkx R Jr, Maksimova E, Stabach P, Hermel JM, Zhang JP, Philbrick W, Slepnev V, Ort T, Solimena M (2000) betaIV spectrin, a new spectrin localized at axon initial segments and nodes of ranvier in the central and peripheral nervous system. J Cell Biol 151:985–1002

    Article  PubMed  CAS  Google Scholar 

  74. Stuart G, Schiller J, Sakmann B (1997) Action potential initiation and propagation in rat neocortical pyramidal neurons. J Physiol 505:617–632

    Article  PubMed  CAS  Google Scholar 

  75. Rasband MN, Peles E, Trimmer JS, Levinson SR, Lux SE, Shrager P (1999) Dependence of nodal sodium channel clustering on paranodal axoglial contact in the developing CNS. J Neurosci 19:7516–7528

    PubMed  CAS  Google Scholar 

  76. Inda MC, De Felipe J, Munoz A (2006) Voltage-gated ion channels in the axon initial segment of human cortical pyramidal cells and their relationship with chandelier cells. Proc Natl Acad Sci USA 103:2920–2925

    Article  PubMed  CAS  Google Scholar 

  77. Cummins TR, Jiang C, Haddad GG (1993) Human neocortical excitability is decreased during anoxia via sodium channel modulation. J Clin Invest 91:608–615

    Article  PubMed  CAS  Google Scholar 

  78. Zhang JH, Gibney GT, Xia Y (2001) Effect of prolonged hypoxia on Na+ channel mRNA subtypes in the developing rat cortex. Brain Res Mol Brain Res 91:154–158

    Article  PubMed  CAS  Google Scholar 

  79. Xia Y, Haddad GG (1999) Effect of prolonged O2 deprivation on Na+ channels: differential regulation in adult versus fetal rat brain. Neuroscience 94:1231–1243

    Article  PubMed  CAS  Google Scholar 

  80. Rakic S, Zecevic N (2000) Programmed cell death in the developing human telencephalon. Eur J Neurosci 12:2721–2734

    Article  PubMed  CAS  Google Scholar 

  81. Gavrieli Y, Sherman Y, Ben-Sasson SA (1992) Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol 119:493–501

    Article  PubMed  CAS  Google Scholar 

  82. Hodgkin AL, Huxley AF (1952) A quantitative description of membrane current and its application to conduction and excitation in nerve. J Physiol 117:500–544

    PubMed  CAS  Google Scholar 

  83. Engel D, Jonas P (2005) Presynaptic action potential amplification by voltage-gated Na+ channels in hippocampal mossy fiber boutons. Neuron 45:405–417

    Article  PubMed  CAS  Google Scholar 

  84. Bikson M, Hahn PJ, Fox JE, Jefferys JG (2003) Depolarization block of neurons during maintenance of electrographic seizures. J Neuro­physiol 90:2402–2408

    Article  PubMed  Google Scholar 

  85. Jones RS, Heinemann U (1987) Abolition of the orthodromically evoked IPSP of CA1 pyramidal cells before the EPSP during washout of calcium from hippocampal slices. Exp Brain Res 65:676–680

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Neuroscience, University of Connecticut Health Center, Farmington, CT, USA

    Anna R. Moore, Wen-Liang Zhou, Igor Jakovcevski, Nada Zecevic & Srdjan D. Antic

Authors
  1. Anna R. Moore
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Wen-Liang Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Igor Jakovcevski
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Nada Zecevic
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Srdjan D. Antic
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Srdjan D. Antic .

Editor information

Editors and Affiliations

  1. Faculty of Medicine & Dentistry, Department of Physiology and Centre for, University of Alberta, N/A 232D, Edmonton, T6G 2S2, Alberta, Canada

    Klaus Ballanyi

Rights and permissions

Reprints and permissions

Copyright information

© 2012 Springer Science+Business Media New York

About this protocol

Cite this protocol

Moore, A.R., Zhou, WL., Jakovcevski, I., Zecevic, N., Antic, S.D. (2012). Physiological Properties of Human Fetal Cortex In Vitro. In: Ballanyi, K. (eds) Isolated Central Nervous System Circuits. Neuromethods, vol 73. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-020-5_3

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-020-5_3

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-019-9

  • Online ISBN: 978-1-62703-020-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation