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Juxtasomal Loose-Patch Recordings in Awake, Head-Fixed Rats to Study the Link Between Structure and Function of Individual Neurons

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Advanced Patch-Clamp Analysis for Neuroscientists

Part of the book series: Neuromethods ((NM,volume 113))

Abstract

The loose-patch juxtasomal recording method can be applied to characterize action potential spiking from single units in the extracellular configuration and includes the attractive option of labeling the neuron for post hoc identification and reconstruction. This ensures “observing without disturbing” (Schubert, J Physiol 581(Pt 1):5, 2007) since the juxtasomal loose-patch recording does not involve breaking into the neuron and modifying its intracellular environment until after all physiological parameters have been obtained. The fundamental difference with extracellular recordings is therefore that juxtasomal recordings generate a direct link between physiological properties and cellular morphology. The necessary step for juxtasomal labeling involves physical interaction between the recording patch pipette and somatic membrane to create a loose-seal patch-clamp recording (hence: juxtasomal) and electroporation for label dialysis (Joshi and Hawken, J Neurosci Methods 156(1–2):37–49, 2006; Pinault, J Neurosci Methods 65(2):113–136, 1996). Next, post hoc histology is performed to reveal cell-type identity and optionally to digitally reconstruct the recorded neuron. In this chapter, I will describe the basic experimental procedures to obtain juxtasomal recordings in primary somatosensory cortex of awake, head-fixed rats and illustrate the information content of these experiments.

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References

  1. Schubert D (2007) Observing without disturbing: how different cortical neuron classes represent tactile stimuli. J Physiol 581(Pt 1):5

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Joshi S, Hawken MJ (2006) Loose-patch-juxtacellular recording in vivo – a method for functional characterization and labeling of neurons in macaque V1. J Neurosci Methods 156(1–2):37–49

    Article  CAS  PubMed  Google Scholar 

  3. Pinault D (1996) A novel single-cell staining procedure performed in vivo under electrophysiological control: morpho-functional features of juxtacellularly labeled thalamic cells and other central neurons with biocytin or neurobiotin. J Neurosci Methods 65(2):113–136

    Article  CAS  PubMed  Google Scholar 

  4. DeFelipe J (2013) Cajal and the discovery of a new artistic world: the neuronal forest. Prog Brain Res 203:201–220

    Article  PubMed  Google Scholar 

  5. Sotelo C (2011) Camillo Golgi and Santiago Ramon y Cajal: the anatomical organization of the cortex of the cerebellum. Can the neuron doctrine still support our actual knowledge on the cerebellar structural arrangement? Brain Res Rev 66(1–2):16–34

    Article  CAS  PubMed  Google Scholar 

  6. Devor A et al (2013) The challenge of connecting the dots in the B.R.A.I.N. Neuron 80(2):270–274

    Article  CAS  PubMed  Google Scholar 

  7. Markram H (2006) The blue brain project. Nat Rev Neurosci 7(2):153–160

    Article  CAS  PubMed  Google Scholar 

  8. Markram H (2012) The human brain project. Sci Am 306(6):50–55

    Article  PubMed  Google Scholar 

  9. Markram H (2013) Seven challenges for neuroscience. Funct Neurol 28(3):145–151

    PubMed  PubMed Central  Google Scholar 

  10. Armstrong-James M, Fox K, Das-Gupta A (1992) Flow of excitation within rat barrel cortex on striking a single vibrissa. J Neurophysiol 68(4):1345–1358

    CAS  PubMed  Google Scholar 

  11. Mountcastle VB, Davies PW, Berman AL (1957) Response properties of neurons of cat’s somatic sensory cortex to peripheral stimuli. J Neurophysiol 20(4):374–407

    CAS  PubMed  Google Scholar 

  12. Simons DJ (1978) Response properties of vibrissa units in rat SI somatosensory neocortex. J Neurophysiol 41(3):798–820

    CAS  PubMed  Google Scholar 

  13. Gilbert CD, Wiesel TN (1979) Morphology and intracortical projections of functionally characterised neurones in the cat visual cortex. Nature 280(5718):120–125

    Article  CAS  PubMed  Google Scholar 

  14. Mitani A et al (1985) Morphology and laminar organization of electrophysiologically identified neurons in the primary auditory cortex in the cat. J Comp Neurol 235(4):430–447

    Article  CAS  PubMed  Google Scholar 

  15. Larkman A, Mason A (1990) Correlations between morphology and electrophysiology of pyramidal neurons in slices of rat visual cortex. I. Establishment of cell classes. J Neurosci 10(5):1407–1414

    CAS  PubMed  Google Scholar 

  16. Mason A, Larkman A (1990) Correlations between morphology and electrophysiology of pyramidal neurons in slices of rat visual cortex. II. Electrophysiology. J Neurosci 10(5):1415–1428

    CAS  PubMed  Google Scholar 

  17. Powell TP, Mountcastle VB (1959) Some aspects of the functional organization of the cortex of the postcentral gyrus of the monkey: a correlation of findings obtained in a single unit analysis with cytoarchitecture. Bull Johns Hopkins Hosp 105:133–162

    CAS  PubMed  Google Scholar 

  18. Hamill OP et al (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch 391(2):85–100

    Article  CAS  PubMed  Google Scholar 

  19. Horikawa K, Armstrong WE (1988) A versatile means of intracellular labeling: injection of biocytin and its detection with avidin conjugates. J Neurosci Methods 25(1):1–11

    Article  CAS  PubMed  Google Scholar 

  20. Marx M et al (2012) Improved biocytin labeling and neuronal 3D reconstruction. Nat Protoc 7(2):394–407

    Article  CAS  PubMed  Google Scholar 

  21. Markram H et al (1997) Physiology and anatomy of synaptic connections between thick tufted pyramidal neurones in the developing rat neocortex. J Physiol 500(Pt 2):409–440

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Feldmeyer D et al (1999) Reliable synaptic connections between pairs of excitatory layer 4 neurones within a single “barrel” of developing rat somatosensory cortex. J Physiol 521(Pt 1):169–190

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Brecht M, Sakmann B (2002) Whisker maps of neuronal subclasses of the rat ventral posterior medial thalamus, identified by whole-cell voltage recording and morphological reconstruction. J Physiol 538(Pt 2):495–515

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. de Kock CP et al (2007) Layer and cell type specific suprathreshold stimulus representation in primary somatosensory cortex. J Physiol 581(1):139–154

    Article  PubMed  PubMed Central  Google Scholar 

  25. Feldmeyer D et al (2012) Barrel cortex function. Prog Neurobiol 2013 Apr;103:3–27. doi: 10.1016/j.pneurobio.2012.11.002. Epub 2012 Nov 27. Review.

    Google Scholar 

  26. Mountcastle VB (1997) The columnar organization of the neocortex. Brain 120(Pt 4):701–722

    Article  PubMed  Google Scholar 

  27. Oberlaender M et al (2012) Cell type-specific three-dimensional structure of thalamocortical circuits in a column of rat vibrissal cortex. Cereb Cortex 22(10):2375–2391

    Article  PubMed  Google Scholar 

  28. Gentet LJ et al (2012) Unique functional properties of somatostatin-expressing GABAergic neurons in mouse barrel cortex. Nat Neurosci 15(4):607–612

    Article  CAS  PubMed  Google Scholar 

  29. Schubert D, Kotter R, Staiger JF (2007) Mapping functional connectivity in barrel-related columns reveals layer- and cell type-specific microcircuits. Brain Struct Funct 212(2):107–119

    Article  PubMed  Google Scholar 

  30. Brecht M, Roth A, Sakmann B (2003) Dynamic receptive fields of reconstructed pyramidal cells in layers 3 and 2 of rat somatosensory barrel cortex. J Physiol 553(Pt 1):243–265

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Brecht M, Sakmann B (2002) Dynamic representation of whisker deflection by synaptic potentials in spiny stellate and pyramidal cells in the barrels and septa of layer 4 rat somatosensory cortex. J Physiol 543(Pt 1):49–70

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Manns ID, Sakmann B, Brecht M (2004) Sub- and suprathreshold receptive field properties of pyramidal neurones in layers 5A and 5B of rat somatosensory barrel cortex. J Physiol 556(Pt 2):601–622

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Klausberger T et al (2003) Brain-state- and cell-type-specific firing of hippocampal interneurons in vivo. Nature 421(6925):844–848

    Article  CAS  PubMed  Google Scholar 

  34. Mileykovskiy BY, Kiyashchenko LI, Siegel JM (2005) Behavioral correlates of activity in identified hypocretin/orexin neurons. Neuron 46(5):787–798

    Article  CAS  PubMed  Google Scholar 

  35. Bevan MD et al (1998) Selective innervation of neostriatal interneurons by a subclass of neuron in the globus pallidus of the rat. J Neurosci 18(22):9438–9452

    CAS  PubMed  Google Scholar 

  36. Varga C, Golshani P, Soltesz I (2012) Frequency-invariant temporal ordering of interneuronal discharges during hippocampal oscillations in awake mice. Proc Natl Acad Sci U S A 109(40):E2726–E2734

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Voigt BC, Brecht M, Houweling AR (2008) Behavioral detectability of single-cell stimulation in the ventral posterior medial nucleus of the thalamus. J Neurosci 28(47):12362–12367

    Article  CAS  PubMed  Google Scholar 

  38. Aksay E et al (2000) Anatomy and discharge properties of pre-motor neurons in the goldfish medulla that have eye-position signals during fixations. J Neurophysiol 84(2):1035–1049

    CAS  PubMed  Google Scholar 

  39. Burgalossi A et al (2011) Microcircuits of functionally identified neurons in the rat medial entorhinal cortex. Neuron 70(4):773–786

    Article  CAS  PubMed  Google Scholar 

  40. de Kock CP, Sakmann B (2009) Spiking in primary somatosensory cortex during natural whisking in awake head-restrained rats is cell-type specific. Proc Natl Acad Sci U S A 106(38):16446–16450

    Article  PubMed  PubMed Central  Google Scholar 

  41. Jorntell H, Ekerot CF (2006) Properties of somatosensory synaptic integration in cerebellar granule cells in vivo. J Neurosci 26(45):11786–11797

    Article  PubMed  Google Scholar 

  42. Boudewijns ZS et al (2013) Layer-specific high-frequency action potential spiking in the prefrontal cortex of awake rats. Front Cell Neurosci 7:99

    Article  PubMed  PubMed Central  Google Scholar 

  43. Helmstaedter M et al (2007) Reconstruction of an average cortical column in silico. Brain Res Rev 55(2):193–203

    Article  CAS  PubMed  Google Scholar 

  44. Ray S et al (2014) Grid-layout and theta-modulation of layer 2 pyramidal neurons in medial entorhinal cortex. Science 343(6173):891–896

    Article  CAS  PubMed  Google Scholar 

  45. Burgalossi A, Brecht M (2014) Cellular, columnar and modular organization of spatial representations in medial entorhinal cortex. Curr Opin Neurobiol 24(1):47–54

    Article  CAS  PubMed  Google Scholar 

  46. Doron G et al (2014) Spiking irregularity and frequency modulate the behavioral report of single-neuron stimulation. Neuron 81(3):653–663

    Article  CAS  PubMed  Google Scholar 

  47. Houweling AR, Brecht M (2008) Behavioural report of single neuron stimulation in somatosensory cortex. Nature 451(7174):65–68

    Article  CAS  PubMed  Google Scholar 

  48. Narayanan RT et al (2014) Juxtasomal biocytin labeling to study the structure-function relationship of individual cortical neurons. J Vis Exp 84:e51359

    Google Scholar 

  49. O’Connor DH et al (2010) Neural activity in barrel cortex underlying vibrissa-based object localization in mice. Neuron 67(6):1048–1061

    Article  PubMed  Google Scholar 

  50. Bagdasarian K et al (2013) Pre-neuronal morphological processing of object location by individual whiskers. Nat Neurosci 16(5):622–631

    Article  CAS  PubMed  Google Scholar 

  51. Carvell GE, Simons DJ (1990) Biometric analyses of vibrissal tactile discrimination in the rat. J Neurosci 10(8):2638–2648

    CAS  PubMed  Google Scholar 

  52. Barth AL, Poulet JF (2012) Experimental evidence for sparse firing in the neocortex. Trends Neurosci 35(6):345–355

    Article  CAS  PubMed  Google Scholar 

  53. Curtis JC, Kleinfeld D (2009) Phase-to-rate transformations encode touch in cortical neurons of a scanning sensorimotor system. Nat Neurosci 12(4):492–501

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. de Kock CP, Sakmann B (2008) High frequency action potential bursts (>or = 100 Hz) in L2/3 and L5B thick tufted neurons in anaesthetized and awake rat primary somatosensory cortex. J Physiol 586(14):3353–3364

    Article  PubMed  PubMed Central  Google Scholar 

  55. Fee MS, Mitra PP, Kleinfeld D (1996) Variability of extracellular spike waveforms of cortical neurons. J Neurophysiol 76(6):3823–3833

    CAS  PubMed  Google Scholar 

  56. Gao P, Bermejo R, Zeigler HP (2001) Whisker deafferentation and rodent whisking patterns: behavioral evidence for a central pattern generator. J Neurosci 21(14):5374–5380

    CAS  PubMed  Google Scholar 

  57. Berg RW, Kleinfeld D (2003) Rhythmic whisking by rat: retraction as well as protraction of the vibrissae is under active muscular control. J Neurophysiol 89(1):104–117

    Article  PubMed  Google Scholar 

  58. Hill DN et al (2008) Biomechanics of the vibrissa motor plant in rat: rhythmic whisking consists of triphasic neuromuscular activity. J Neurosci 28(13):3438–3455

    Article  CAS  PubMed  Google Scholar 

  59. Knutsen PM, Derdikman D, Ahissar E (2005) Tracking whisker and head movements in unrestrained behaving rodents. J Neurophysiol 93(4):2294–2301

    Article  PubMed  Google Scholar 

  60. Wong-Riley M (1979) Changes in the visual system of monocularly sutured or enucleated cats demonstrable with cytochrome oxidase histochemistry. Brain Res 171(1):11–28

    Article  CAS  PubMed  Google Scholar 

  61. Sakmann B, Neher E (1995) Single-channel recording. Plenum Press, New York

    Book  Google Scholar 

  62. Bock DD et al (2011) Network anatomy and in vivo physiology of visual cortical neurons. Nature 471(7337):177–182

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Briggman KL, Helmstaedter M, Denk W (2011) Wiring specificity in the direction-selectivity circuit of the retina. Nature 471(7337):183–188

    Article  CAS  PubMed  Google Scholar 

  64. Margrie TW, Brecht M, Sakmann B (2002) In vivo, low-resistance, whole-cell recordings from neurons in the anaesthetized and awake mammalian brain. Pflugers Arch 444(4):491–498

    Article  CAS  PubMed  Google Scholar 

  65. Oberlaender M, Ramirez A, Bruno RM (2012) Sensory experience restructures thalamocortical axons during adulthood. Neuron 74(4):648–655

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Bruno RM et al (2009) Sensory experience alters specific branches of individual corticocortical axons during development. J Neurosci 29(10):3172–3181

    Article  CAS  PubMed  Google Scholar 

  67. Boudewijns ZS et al (2011) Semi-automated three-dimensional reconstructions of individual neurons reveal cell type-specific circuits in cortex. Commun Integr Biol 4(4):486–488

    Article  PubMed  PubMed Central  Google Scholar 

  68. Oberlaender M et al (2011) Three-dimensional axon morphologies of individual layer 5 neurons indicate cell type-specific intracortical pathways for whisker motion and touch. Proc Natl Acad Sci U S A 108(10):4188–4193

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  69. Helmstaedter M (2013) Cellular-resolution connectomics: challenges of dense neural circuit reconstruction. Nat Methods 10(6):501–507

    Article  CAS  PubMed  Google Scholar 

  70. Sarid L et al (2007) Modeling a layer 4-to-layer 2/3 module of a single column in rat neocortex: interweaving in vitro and in vivo experimental observations. Proc Natl Acad Sci U S A 104(41):16353–16358

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. Petreanu L et al (2009) The subcellular organization of neocortical excitatory connections. Nature 457(7233):1142–1145

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  72. Xu NL et al (2012) Nonlinear dendritic integration of sensory and motor input during an active sensing task. Nature 492(7428):247–251

    Article  CAS  PubMed  Google Scholar 

  73. O’Connor DH, Huber D, Svoboda K (2009) Reverse engineering the mouse brain. Nature 461(7266):923–929

    Article  PubMed  Google Scholar 

  74. Kleinfeld D, Deschenes M (2011) Neuronal basis for object location in the vibrissa scanning sensorimotor system. Neuron 72(3):455–468

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

I was introduced to the juxtasomal loose-patch technique by Randy Bruno (Columbia University, NY, USA) under the supervision of Prof. Dr. Bert Sakmann and thank both Randy and Prof. Sakmann for their continuous support, enthusiasm, and fruitful collaborations. Additionally, I'd like to thank Anton Pieneman for excellent technical support.

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Authors and Affiliations

  1. Department of Integrative Neurophysiology, Center for Neurogenomics and Cognitive Research, VU University Amsterdam, Neuroscience Campus Amsterdam, De Boelelaan 1085, 1081 HV, Amsterdam, The Netherlands

    Christiaan P. J. de Kock

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  1. Christiaan P. J. de Kock
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Editors and Affiliations

  1. The Leslie and Susan Gonda Multidisciplinary Brain Research Center, The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel

    Alon Korngreen

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de Kock, C.P.J. (2016). Juxtasomal Loose-Patch Recordings in Awake, Head-Fixed Rats to Study the Link Between Structure and Function of Individual Neurons. In: Korngreen, A. (eds) Advanced Patch-Clamp Analysis for Neuroscientists. Neuromethods, vol 113. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3411-9_2

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  • DOI: https://doi.org/10.1007/978-1-4939-3411-9_2

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3409-6

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