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Chronic Two-Photon Imaging of Neural Activity in the Anesthetized and Awake Behaving Rodent

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Optical Imaging of Neocortical Dynamics

Part of the book series: Neuromethods ((NM,volume 85))

Abstract

Chronic in vivo two-photon imaging of genetically encoded sensors has recently enabled the measurement of activity from the same individual neurons repeatedly in different imaging sessions over months, opening new possibilities to investigate the function and plasticity of neuronal activity in both anesthetized and awake animals. A successful chronic imaging experiment involves the combination of several key techniques, including expression of genetically encoded indicators, optical measurement with cellular resolution through a cranial window, and, in awake animals, behavioral paradigms adapted to the two-photon microscope. Here we cover methods and advances in chronic imaging of cortical activity in the awake, behaving rodent for the investigation of learning and performance of sensory and cognitive tasks.

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References

  1. Hofer SB, Mrsic-Flogel TD, Bonhoeffer T, Hübener M (2006) Prior experience enhances plasticity in adult visual cortex. Nat Neurosci 9:127–132

    Article  CAS  PubMed  Google Scholar 

  2. Kaneko M, Hanover JL, England PM, Stryker MP (2008) TrkB kinase is required for recovery, but not loss, of cortical responses following monocular deprivation. Nat Neurosci 11:497–504

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  3. Masino SA, Frostig RD (1996) Quantitative long-term imaging of the functional representation of a whisker in rat barrel cortex. Proc Natl Acad Sci USA 93:4942–4947

    Article  CAS  PubMed  Google Scholar 

  4. Polley DB, Chen-Bee CH, Frostig RD (1999) Two directions of plasticity in the sensory-deprived adult cortex. Neuron 24:623–637

    Article  CAS  PubMed  Google Scholar 

  5. Polley DB, Kvasnak E, Frostig RD (2004) Naturalistic experience transforms sensory maps in the adult cortex of caged animals. Nature 429:67–71

    Article  CAS  PubMed  Google Scholar 

  6. Slovin H, Arieli A, Hildesheim R, Grinvald A (2002) Long-term voltage-sensitive dye imaging reveals cortical dynamics in behaving monkeys. J Neurophysiol 88:3421–3438

    Article  PubMed  Google Scholar 

  7. Bozza T, McGann JP, Mombaerts P, Wachowiak M (2004) In vivo imaging of neuronal activity by targeted expression of a genetically encoded probe in the mouse. Neuron 42:9–21

    Article  CAS  PubMed  Google Scholar 

  8. Minderer M, Liu W, Sumanovski LT, Kügler S, Helmchen F, Margolis DJ (2012) Chronic imaging of cortical sensory map dynamics using a genetically encoded calcium indicator. J Physiol 590:99–107

    CAS  PubMed  Google Scholar 

  9. Dickey AS, Suminski A, Amit Y, Hatsopoulos NG (2009) Single-unit stability using chronically implanted multielectrode arrays. J Neurophysiol 102:1331–1339

    Article  PubMed  Google Scholar 

  10. Tolias AS, Ecker AS, Siapas AG, Hoenselaar A, Keliris GA, Logothetis NK (2007) Recording chronically from the same neurons in awake, behaving primates. J Neurophysiol 98:3780–3790

    Article  PubMed  Google Scholar 

  11. Buzsaki G (2004) Large-scale recording of neuronal ensembles. Nat Neurosci 7:446–451

    Article  CAS  PubMed  Google Scholar 

  12. Holtmaat A, Svoboda K (2009) Experience-dependent structural synaptic plasticity in the mammalian brain. Nat Rev Neurosci 10:647–658

    Article  CAS  PubMed  Google Scholar 

  13. Andermann ML, Kerlin AM, Reid RC (2010) Chronic cellular imaging of mouse visual cortex during operant behavior and passive viewing. Front Cell Neurosci 4:3

    PubMed Central  PubMed  Google Scholar 

  14. Huber D, Gutnisky DA, Peron S, O’Connor DH, Wiegert JS, Tian L, Oertner TG, Looger LL, Svoboda K (2012) Multiple dynamic representations in the motor cortex during sensorimotor learning. Nature 484:473–478

    Article  CAS  PubMed  Google Scholar 

  15. Mank M, Santos AF, Direnberger S, Mrsic-Flogel TD, Hofer SB, Stein V, Hendel T, Reiff DF, Levelt C, Borst A, Bonhoeffer T, Hübener M, Griesbeck O (2008) A genetically encoded calcium indicator for chronic in vivo two-photon imaging. Nat Methods 5:805–811

    Article  CAS  PubMed  Google Scholar 

  16. Margolis DJ, Lütcke H, Schulz K, Haiss F, Weber B, Kügler S, Hasan MT, Helmchen F (2012) Reorganization of cortical population activity imaged throughout long-term sensory deprivation. Nat Neurosci 15:1539–1546

    Article  CAS  PubMed  Google Scholar 

  17. Tian L, Hires SA, Mao T, Huber D, Chiappe ME, Chalasani SH, Petreanu L, Akerboom J, McKinney SA, Schreiter ER, Bargmann CI, Jayaraman V, Svoboda K, Looger LL (2009) Imaging neural activity in worms, flies and mice with improved GCaMP calcium indicators. Nat Methods 6:875–881

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  18. Akemann W, Mutoh H, Perron A, Rossier J, Knopfel T (2010) Imaging brain electric signals with genetically targeted voltage-sensitive fluorescent proteins. Nat Methods 7:643–649

    Article  CAS  PubMed  Google Scholar 

  19. Zhong H, Sia GM, Sato TR, Gray NW, Mao T, Khuchua Z, Huganir RL, Svoboda K (2009) Subcellular dynamics of type II PKA in neurons. Neuron 62:363–374

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  20. Garaschuk O, Milos RI, Grienberger C, Marandi N, Adelsberger H, Konnerth A (2006) Optical monitoring of brain function in vivo: from neurons to networks. Pflugers Arch 453:385–396

    Article  CAS  PubMed  Google Scholar 

  21. Göbel W, Helmchen F (2007) In vivo calcium imaging of neural network function. Physiology (Bethesda) 22:358–365

    Article  Google Scholar 

  22. Grewe BF, Helmchen F (2009) Optical probing of neuronal ensemble activity. Curr Opin Neurobiol 19:520–529

    Article  CAS  PubMed  Google Scholar 

  23. Chen TW, Wardill TJ, Sun Y, Pulver SR, Renninger SL, Baohan A, Schreiter ER, Kerr RA, Orger MB, Jayaraman V, Looger LL, Svoboda K, Kim DS (2013) Ultrasensitive fluorescent proteins for imaging neuronal activity, Nature 499, 295–300

    Article  CAS  Google Scholar 

  24. Hendel T, Mank M, Schnell B, Griesbeck O, Borst A, Reiff DF (2008) Fluorescence changes of genetic calcium indicators and OGB-1 correlated with neural activity and calcium in vivo and in vitro. J Neurosci 28:7399–7411

    Article  CAS  PubMed  Google Scholar 

  25. Pologruto TA, Yasuda R, Svoboda K (2004) Monitoring neural activity and [Ca2+] with genetically encoded Ca2+ indicators. J Neurosci 24:9572–9579

    Article  CAS  PubMed  Google Scholar 

  26. Dombeck DA, Khabbaz AN, Collman F, Adelman TL, Tank DW (2007) Imaging large-scale neural activity with cellular resolution in awake, mobile mice. Neuron 56:43–57

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  27. Mrsic-Flogel TD, Hofer SB, Ohki K, Reid RC, Bonhoeffer T, Hübener M (2007) Homeostatic regulation of eye-specific responses in visual cortex during ocular dominance plasticity. Neuron 54:961–972

    Article  CAS  PubMed  Google Scholar 

  28. Shevtsova Z, Malik JM, Michel U, Bahr M, Kügler S (2005) Promoters and serotypes: targeting of adeno-associated virus vectors for gene transfer in the rat central nervous system in vitro and in vivo. Exp Physiol 90:53–59

    Article  CAS  PubMed  Google Scholar 

  29. Nathanson JL, Yanagawa Y, Obata K, Callaway EM (2009) Preferential labeling of inhibitory and excitatory cortical neurons by endogenous tropism of adeno-associated virus and lentivirus vectors. Neuroscience 161:441–450

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  30. Dreosti E, Odermatt B, Dorostkar MM, Lagnado L (2009) A genetically encoded reporter of synaptic activity in vivo. Nat Methods 6:883–889

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  31. Harvey CD, Coen P, Tank DW (2012) Choice-specific sequences in parietal cortex during a virtual-navigation decision task. Nature 484:62–68

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  32. Lütcke H, Murayama M, Hahn T, Margolis DJ, Astori S, Zum Alten Borgloh SM, Gobel W, Yang Y, Tang W, Kügler S, Sprengel R, Nagai T, Miyawaki A, Larkum ME, Helmchen F, Hasan MT (2010) Optical recording of neuronal activity with a genetically-encoded calcium indicator in anesthetized and freely moving mice. Front Neural Circuits 4:9

    PubMed Central  PubMed  Google Scholar 

  33. O’Connor DH, Peron SP, Huber D, Svoboda K (2010) Neural activity in barrel cortex underlying vibrissa-based object localization in mice. Neuron 67:1048–1061

    Article  PubMed  Google Scholar 

  34. Petreanu L, Gutnisky DA, Huber D, Xu NL, O’Connor DH, Tian L, Looger L, Svoboda K (2012) Activity in motor-sensory projections reveals distributed coding in somatosensation. Nature 489:299–303

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  35. Wallace DJ, Meyer zum Alten Borgloh S, Astori S, Yang Y, Bausen M, Kügler S, Palmer AE, Tsien RY, Sprengel R, Kerr JN, Denk W, Hasan MT (2008) Single-spike detection in vitro and in vivo with a genetic Ca2+ sensor. Nat Methods 5:797–804

    Article  CAS  PubMed  Google Scholar 

  36. Xu NL, Harnett MT, Williams SR, Huber D, O’Connor DH, Svoboda K, Magee JC (2012) Nonlinear dendritic integration of sensory and motor input during an active sensing task. Nature 492(7428):247–251

    Article  CAS  PubMed  Google Scholar 

  37. Hübener M, Bonhoeffer T (2010) Searching for engrams. Neuron 67:363–371

    Article  PubMed  Google Scholar 

  38. Zariwala HA, Borghuis BG, Hoogland TM, Madisen L, Tian L, De Zeeuw CI, Zeng H, Looger LL, Svoboda K, Chen TW (2012) A Cre-dependent GCaMP3 reporter mouse for neuronal imaging in vivo. J Neurosci 32:3131–3141

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  39. Chen Q, Cichon J, Wang W, Qiu L, Lee SJ, Campbell NR, Destefino N, Goard MJ, Fu Z, Yasuda R, Looger LL, Arenkiel BR, Gan WB, Feng G (2012) Imaging neural activity using Thy1-GCaMP transgenic mice. Neuron 76:297–308

    Article  CAS  PubMed  Google Scholar 

  40. Wolfe J, Houweling AR, Brecht M (2010) Sparse and powerful cortical spikes. Curr Opin Neurobiol 20:306–312

    Article  CAS  PubMed  Google Scholar 

  41. Miyawaki A, Llopis J, Heim R, McCaffery JM, Adams JA, Ikura M, Tsien RY (1997) Fluorescent indicators for Ca2+ based on green fluorescent proteins and calmodulin. Nature 388:882–887

    Article  CAS  PubMed  Google Scholar 

  42. Yamada Y, Michikawa T, Hashimoto M, Horikawa K, Nagai T, Miyawaki A, Hausser M, Mikoshiba K (2011) Quantitative comparison of genetically encoded Ca indicators in cortical pyramidal cells and cerebellar Purkinje cells. Front Cell Neurosci 5:18

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  43. Ohkura, M., Sasaki, T., Sadakari, J., Gengyo-Ando, K., Kagawa-Nagamura, Y., Kobayashi, C., Ikegaya, Y., and Nakai, J. (2012) Genetically encoded green fluorescent Ca2+ indicators with improved detectability for neuronal Ca2+ signals, PLoS One 7, e51286

    Article  CAS  Google Scholar 

  44. Nagai T, Yamada S, Tominaga T, Ichikawa M, Miyawaki A (2004) Expanded dynamic range of fluorescent indicators for Ca2+ by circularly permuted yellow fluorescent proteins. Proc Natl Acad Sci USA 101:10554–10559

    Article  CAS  PubMed  Google Scholar 

  45. Akerboom J, Chen TW, Wardill TJ, Tian L, Marvin JS, Mutlu S, Calderon NC, Esposti F, Borghuis BG, Sun XR, Gordus A, Orger MB, Portugues R, Engert F, Macklin JJ, Filosa A, Aggarwal A, Kerr RA, Takagi R, Kracun S, Shigetomi E, Khakh BS, Baier H, Lagnado L, Wang SS, Bargmann CI, Kimmel BE, Jayaraman V, Svoboda K, Kim DS, Schreiter ER, Looger LL (2012) Optimization of a GCaMP calcium indicator for neural activity imaging. J Neurosci 32:13819–13840

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  46. Horikawa K, Yamada Y, Matsuda T, Kobayashi K, Hashimoto M, Matsu-ura T, Miyawaki A, Michikawa T, Mikoshiba K, Nagai T (2010) Spontaneous network activity visualized by ultrasensitive Ca2+ indicators, yellow Cameleon-Nano. Nat Methods 7:729–732

    Article  CAS  PubMed  Google Scholar 

  47. Mank M, Griesbeck O (2008) Genetically encoded calcium indicators. Chem Rev 108:1550–1564

    Article  CAS  PubMed  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  49. Holtmaat A, Bonhoeffer T, Chow DK, Chuckowree J, De Paola V, Hofer SB, Hubener M, Keck T, Knott G, Lee WC, Mostany R, Mrsic-Flogel TD, Nedivi E, Portera-Cailliau C, Svoboda K, Trachtenberg JT, Wilbrecht L (2009) Long-term, high-resolution imaging in the mouse neocortex through a chronic cranial window. Nat Protoc 4:1128–1144

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  50. Dombeck DA, Harvey CD, Tian L, Looger LL, Tank DW (2010) Functional imaging of hippocampal place cells at cellular resolution during virtual navigation. Nat Neurosci 13:1433–1440

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  51. Stettler DD, Yamahachi H, Li W, Denk W, Gilbert CD (2006) Axons and synaptic boutons are highly dynamic in adult visual cortex. Neuron 49:877–887

    Article  CAS  PubMed  Google Scholar 

  52. Yamahachi H, Marik SA, McManus JN, Denk W, Gilbert CD (2009) Rapid axonal sprouting and pruning accompany functional reorganization in primary visual cortex. Neuron 64:719–729

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  53. Drew PJ, Shih AY, Driscoll JD, Knutsen PM, Blinder P, Davalos D, Akassoglou K, Tsai PS, Kleinfeld D (2010) Chronic optical access through a polished and reinforced thinned skull. Nat Methods 7:981–984

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  54. Schwarz C, Hentschke H, Butovas S, Haiss F, Stuttgen MC, Gerdjikov TV, Bergner CG, Waiblinger C (2010) The head-fixed behaving rat—procedures and pitfalls. Somatosens Mot Res 27:131–148

    Article  PubMed Central  PubMed  Google Scholar 

  55. Yang G, Pan F, Parkhurst CN, Grutzendler J, Gan WB (2010) Thinned-skull cranial window technique for long-term imaging of the cortex in live mice. Nat Protoc 5:201–208

    Article  CAS  PubMed  Google Scholar 

  56. Xu HT, Pan F, Yang G, Gan WB (2007) Choice of cranial window type for in vivo imaging affects dendritic spine turnover in the cortex. Nat Neurosci 10:549–551

    Article  CAS  PubMed  Google Scholar 

  57. Lowenstein PR, Mandel RJ, Xiong WD, Kroeger K, Castro MG (2007) Immune responses to adenovirus and adeno-associated vectors used for gene therapy of brain diseases: the role of immunological synapses in understanding the cell biology of neuroimmune interactions. Curr Gene Ther 7:347–360

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  58. Reimsnider S, Manfredsson FP, Muzyczka N, Mandel RJ (2007) Time course of transgene expression after intrastriatal pseudotyped rAAV2/1, rAAV2/2, rAAV2/5, and rAAV2/8 transduction in the rat. Mol Ther 15:1504–1511

    Article  CAS  PubMed  Google Scholar 

  59. Helmchen F, Denk W (2005) Deep tissue two-photon microscopy. Nat Methods 2:932–940

    Article  CAS  PubMed  Google Scholar 

  60. Kerr JN, Denk W (2008) Imaging in vivo: watching the brain in action. Nat Rev Neurosci 9:195–205

    Article  CAS  PubMed  Google Scholar 

  61. Engelbrecht CJ, Johnston RS, Seibel EJ, Helmchen F (2008) Ultra-compact fiber-optic two-photon microscope for functional fluorescence imaging in vivo. Opt Express 16:5556–5564

    Article  CAS  PubMed  Google Scholar 

  62. Piyawattanametha W, Cocker ED, Burns LD, Barretto RP, Jung JC, Ra H, Solgaard O, Schnitzer MJ (2009) In vivo brain imaging using a portable 2.9 g two-photon microscope based on a microelectromechanical systems scanning mirror. Opt Lett 34:2309–2311

    Article  PubMed Central  PubMed  Google Scholar 

  63. Sawinski J, Wallace DJ, Greenberg DS, Grossmann S, Denk W, Kerr JN (2009) Visually evoked activity in cortical cells imaged in freely moving animals. Proc Natl Acad Sci USA 106:19557–19562

    Article  CAS  PubMed  Google Scholar 

  64. Euler T, Hausselt SE, Margolis DJ, Breuninger T, Castell X, Detwiler PB, Denk W (2009) Eyecup scope—optical recordings of light stimulus-evoked fluorescence signals in the retina. Pflugers Arch 457:1393–1414

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  65. Heider B, Nathanson JL, Isacoff EY, Callaway EM, Siegel RM (2010) Two-photon imaging of calcium in virally transfected striate cortical neurons of behaving monkey. PLoS One 5:e13829

    Article  PubMed Central  PubMed  Google Scholar 

  66. Mayrhofer JM, Skreb V, von der Behrens W, Musall S, Weber B, Haiss F (2013) Novel two-alternative forced choice paradigm for bilateral vibrotactile whisker frequency discrimination in head-fixed mice and rats. J Neurophysiol 109:273–284

    Article  PubMed  Google Scholar 

  67. Dombeck DA, Graziano MS, Tank DW (2009) Functional clustering of neurons in motor cortex determined by cellular resolution imaging in awake behaving mice. J Neurosci 29:13751–13760

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  68. Keller GB, Bonhoeffer T, Hübener M (2012) Sensorimotor mismatch signals in primary visual cortex of the behaving mouse. Neuron 74:809–815

    Article  CAS  PubMed  Google Scholar 

  69. Niell CM, Stryker MP (2010) Modulation of visual responses by behavioral state in mouse visual cortex. Neuron 65:472–479

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  70. Royer S, Zemelman BV, Losonczy A, Kim J, Chance F, Magee JC, Buzsaki G (2012) Control of timing, rate and bursts of hippocampal place cells by dendritic and somatic inhibition. Nat Neurosci 15:769–775

    Article  CAS  PubMed  Google Scholar 

  71. Nimmerjahn A, Mukamel EA, Schnitzer MJ (2009) Motor behavior activates Bergmann glial networks. Neuron 62:400–412

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  72. Harvey CD, Collman F, Dombeck DA, Tank DW (2009) Intracellular dynamics of hippocampal place cells during virtual navigation. Nature 461:941–946

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  73. Haiss F, Schwarz C (2005) Spatial segregation of different modes of movement control in the whisker representation of rat primary motor cortex. J Neurosci 25:1579–1587

    Article  CAS  PubMed  Google Scholar 

  74. Hentschke H, Haiss F, Schwarz C (2006) Central signals rapidly switch tactile processing in rat barrel cortex during whisker movements. Cereb Cortex 16:1142–1156

    Article  PubMed  Google Scholar 

  75. Gentet LJ, Avermann M, Matyas F, Staiger JF, Petersen CC (2010) Membrane potential dynamics of GABAergic neurons in the barrel cortex of behaving mice. Neuron 65:422–435

    Article  CAS  PubMed  Google Scholar 

  76. Greenberg DS, Kerr JN (2009) Automated correction of fast motion artifacts for two-photon imaging of awake animals. J Neurosci Methods 176:1–15

    Article  PubMed  Google Scholar 

  77. Grewe BF, Langer D, Kasper H, Kampa BM, Helmchen F (2010) High-speed in vivo calcium imaging reveals neuronal network activity with near-millisecond precision. Nat Methods 7:399–405

    Article  CAS  PubMed  Google Scholar 

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

    Article  PubMed  Google Scholar 

  79. Lütcke H, Margolis DJ, Helmchen F (2013) Steady or changing? Long-term monitoring of neuronal population activity. Trends Neurosci 36(7):375–384

    Article  CAS  PubMed  Google Scholar 

  80. Petreanu L, Mao T, Sternson SM, Svoboda K (2009) The subcellular organization of neocortical excitatory connections. Nature 457:1142–1145

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  81. Hasan MT, Friedrich RW, Euler T, Larkum ME, Giese G, Both M, Duebel J, Waters J, Bujard H, Griesbeck O, Tsien RY, Nagai T, Miyawaki A, Denk W (2004) Functional fluorescent Ca2+ indicator proteins in transgenic mice under TET control. PLoS Biol 2:e163

    Article  PubMed Central  PubMed  Google Scholar 

  82. Heim N, Garaschuk O, Friedrich MW, Mank M, Milos RI, Kovalchuk Y, Konnerth A, Griesbeck O (2007) Improved calcium imaging in transgenic mice expressing a troponin C-based biosensor. Nat Methods 4:127–129

    Article  CAS  PubMed  Google Scholar 

  83. Han X, Qian X, Bernstein JG, Zhou HH, Franzesi GT, Stern P, Bronson RT, Graybiel AM, Desimone R, Boyden ES (2009) Millisecond-timescale optical control of neural dynamics in the nonhuman primate brain. Neuron 62:191–198

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  84. Wilt BA, Burns LD, Wei Ho ET, Ghosh KK, Mukamel EA, Schnitzer MJ (2009) Advances in light microscopy for neuroscience. Annu Rev Neurosci 32:435–506

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  85. Scanziani M, Häusser M (2009) Electrophysiology in the age of light. Nature 461:930–939

    Article  CAS  PubMed  Google Scholar 

  86. Cheng A, Goncalves JT, Golshani P, Arisaka K, Portera-Cailliau C (2011) Simultaneous two-photon calcium imaging at different depths with spatiotemporal multiplexing. Nat Methods 8:139–142

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  87. Kawakami R, Sawada K, Sato A, Hibi T, Kozawa Y, Sato S, Yokoyama H, Nemoto T (2013) Visualizing hippocampal neurons with in vivo two-photon microscopy using a 1030 nm picosecond pulse laser. Sci Rep 3:1014

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  88. Mittmann W, Wallace DJ, Czubayko U, Herb JT, Schaefer AT, Looger LL, Denk W, Kerr JN (2011) Two-photon calcium imaging of evoked activity from L5 somatosensory neurons in vivo. Nat Neurosci 14:1089–1093

    Article  CAS  PubMed  Google Scholar 

  89. Horton NG, Wang K, Kobat D, Clark CG, Wise FW, Schaffer CB, Xu C (2013) In vivo three-photon microscopy of subcortical structures within an intact mouse brain. Nat Photonics 7:205–209

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Brain Research Institute, University of Zurich, Zurich, Switzerland

    David J. Margolis, Henry Lütcke & Fritjof Helmchen

  2. Institute of Pharmacology and Toxicology, University of Zurich, Zurich, Switzerland

    Bruno Weber & Florent Haiss

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  1. David J. Margolis
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  2. Henry Lütcke
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  3. Fritjof Helmchen
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  4. Bruno Weber
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  5. Florent Haiss
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  1. University of Zurich, Switzerland

    Bruno Weber

  2. University of Zurich, Switzerland

    Fritjof Helmchen

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Margolis, D.J., Lütcke, H., Helmchen, F., Weber, B., Haiss, F. (2014). Chronic Two-Photon Imaging of Neural Activity in the Anesthetized and Awake Behaving Rodent. In: Weber, B., Helmchen, F. (eds) Optical Imaging of Neocortical Dynamics. Neuromethods, vol 85. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-785-3_10

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  • DOI: https://doi.org/10.1007/978-1-62703-785-3_10

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