Multi-Spectral Imaging of Blood Volume, Metabolism, Oximetry, and Light Scattering

  • Mingrui Zhao
  • Hongtao Ma
  • Samuel Harris
  • Theodore H. Schwartz
Part of the Neuromethods book series (NM, volume 88)


Advances in functional imaging techniques including fMRI, PET, and SPECT have improved our understanding of the relationship between brain activity and brain energy supply. Neurovascular and neurometabolic coupling are critical to supply the energy demands of brain tissue during both normal physiological function and pathological conditions. With the use of multi-spectral imaging techniques, one can simultaneously measure changes in cerebral blood volume, oxyhemoglobin, deoxyhemoglobin, light scattering, and local metabolism during epilepsy.

Key words

Neurovascular coupling Epilepsy Intrinsic optical imaging Autofluorescence Brain activity map 



This work was supported by the NINDS RO1 NS49482 (T.H.S), CURE Taking Flight Award (H.M.), the Clinical and Translational Science Center (CTSC) Grant UL1 RR 024996 Pilot Grant (M.Z), and the Cornell University Ithaca-WCMC seed grant (M.Z.). We thank Dr. Yevgeniy B. Sirotin for help with LED setup and AFI analysis.


  1. 1.
    Logothetis NK, Wandell BA (2004) Interpreting the BOLD signal. Annu Rev Physiol 66:735–769PubMedCrossRefGoogle Scholar
  2. 2.
    Raichle ME, Mintun MA (2006) Brain work and brain imaging. Annu Rev Neurosci 29:449–476. doi: 10.1146/annurev.neuro.29.051605.112819 PubMedCrossRefGoogle Scholar
  3. 3.
    Girouard H, Iadecola C (2006) Neurovascular coupling in the normal brain and in hypertension, stroke, and Alzheimer disease. J Appl Physiol 100(1):328–335. doi: 10.1152/japplphysiol.00966.2005 PubMedCrossRefGoogle Scholar
  4. 4.
    Frostig RD, Lieke EE, Ts’o DY et al (1990) Cortical functional architecture and local coupling between neuronal activity and the microcirculation revealed by in vivo high-resolution optical imaging of intrinsic signals. Proc Natl Acad Sci 87:6082–6086PubMedCentralPubMedCrossRefGoogle Scholar
  5. 5.
    Malonek D, Grinvald A (1996) Interactions between electrical activity and cortical microcirculation revealed by imaging spectroscopy: implications for functional brain mapping. Science 272:551–554PubMedCrossRefGoogle Scholar
  6. 6.
    Sheth SA, Nemoto M, Guiou M et al (2004) Columnar specificity of microvascular oxygenation and volume responses: implications for functional brain mapping. J Neurosci 24(3): 634–641PubMedCrossRefGoogle Scholar
  7. 7.
    Salzberg BM, Obaid AL, Gainer H (1985) Large and rapid changes in light scattering accompany secretion by nerve terminals in the mammalian neurohypophysis. J Gen Physiol 86(3):395–411PubMedCrossRefGoogle Scholar
  8. 8.
    Obaid AL, Flores R, Salzberg BM (1989) Calcium channels that are required for secretion from intact nerve terminals of vertebrates are sensitive to omega-conotoxin and relatively insensitive to dihydropyridines. Optical studies with and without voltage-sensitive dyes. J Gen Physiol 93(4):715–729PubMedCrossRefGoogle Scholar
  9. 9.
    MacVicar BA, Hochman D (1991) Imaging of synaptically evoked intrinsic optical signals in hippocampal slices. J Neurosci 11(5):1458–1469PubMedGoogle Scholar
  10. 10.
    Alivisatos AP, Chun M, Church George M et al (2012) The brain activity map project and the challenge of functional connectomics. Neuron 74(6):970–974PubMedCentralPubMedCrossRefGoogle Scholar
  11. 11.
    Hill DK, Keynes RD (1949) Opacity changes in stimulated nerve. J Physiol 108(3): 278–281PubMedCentralGoogle Scholar
  12. 12.
    Jobsis F (1977) Noninvasive, infrared monitoring of cerebral and myocardial oxygen sufficiency and circulatory parameters. Science 198(4323):1264–1267. doi: 10.1126/science.929199 PubMedCrossRefGoogle Scholar
  13. 13.
    Chance B, Cohen P, Jobsis F et al (1962) Intracellular oxidation-reduction states in vivo: the microfluorometry of pyridine nucleotide gives a continuous measurement of the oxidation state. Science 137(3529):499–508. doi: 10.1126/science.137.3529.499 PubMedCrossRefGoogle Scholar
  14. 14.
    Grinvald A (1985) Real-time optical mapping of neuronal activity: from single growth cones to the intact mammalian brain. Annu Rev Neurosci 8(1):263–305. doi: 10.1146/ PubMedCrossRefGoogle Scholar
  15. 15.
    Grinvald A, Lieke E, Frostig RD et al (1986) Functional architecture of cortex revealed by optical imaging of intrinsic signals. Nature 324(6095):361–364PubMedCrossRefGoogle Scholar
  16. 16.
    Bonhoeffer T, Grinvald A (1991) Iso-orientation domains in cat visual cortex are arranged in pinwheel-like patterns. Nature 353(6343):429–431PubMedCrossRefGoogle Scholar
  17. 17.
    Schwartz TH, Bonhoeffer T (2001) In vivo optical mapping of epileptic foci and surround inhibition in ferret cerebral cortex. Nat Med 7(9):1063–1067PubMedCrossRefGoogle Scholar
  18. 18.
    Schwartz TH, Chen LM, Friedman RM et al (2004) Intraoperative optical imaging of human face cortical topography: a case study. Neuroreport 15(9):1527–1531PubMedCrossRefGoogle Scholar
  19. 19.
    Zepeda A, Arias C, Sengpiel F (2004) Optical imaging of intrinsic signals: recent developments in the methodology and its applications. J Neurosci Methods 136(1):1–21PubMedCrossRefGoogle Scholar
  20. 20.
    Cohen LB (1973) Changes in neuron structure during action potential propagation and synaptic transmission. Physiol Rev 53(2):373–418PubMedGoogle Scholar
  21. 21.
    Sheth SA, Nemoto M, Guiou G et al (2004) Linear and nonlinear relationships between neuronal activity, oxygen metabolism, and hemodynamic response. Neuron 42:347–355PubMedCrossRefGoogle Scholar
  22. 22.
    Georgakoudi I, Quinn KP (2012) Optical imaging using endogenous contrast to assess metabolic state. Annu Rev Biomed Eng 14(1):351–367. doi: 10.1146/annurev-bioeng-071811-150108 PubMedCrossRefGoogle Scholar
  23. 23.
    Chance B, Ernster L, Garland PB et al (1967) Flavoproteins of the mitochondrial respiratory chain. Proc Natl Acad Sci U S A 57(5): 1498–1505PubMedCentralPubMedCrossRefGoogle Scholar
  24. 24.
    Reinert KC, Dunbar RL, Gao W et al (2004) Flavoprotein autofluorescence imaging of neuronal activation in the cerebellar cortex in vivo. J Neurophysiol 92(1):199–211. doi: 10.1152/jn.01275.2003, pii: 01275.2003PubMedCrossRefGoogle Scholar
  25. 25.
    Reinert KC, Gao W, Chen G et al (2007) Flavoprotein autofluorescence imaging in the cerebellar cortex in vivo. J Neurosci Res 85(15):3221–3232. doi: 10.1002/jnr.21348 PubMedCrossRefGoogle Scholar
  26. 26.
    Llano DA, Theyel BB, Mallik AK et al (2009) Rapid and sensitive mapping of long-range connections in vitro using flavoprotein autofluorescence imaging combined with laser photostimulation. J Neurophysiol 101(6):3325–3340. doi: 10.1152/jn.91291.2008, pii: 91291.2008PubMedCentralPubMedCrossRefGoogle Scholar
  27. 27.
    Sirotin YB, Das A (2010) Spatial relationship between flavoprotein fluorescence and the hemodynamic response in the primary visual cortex of alert macaque monkeys. Front Neuroenergetics 2:6. doi: 10.3389/fnene.2010.00006 PubMedCentralPubMedGoogle Scholar
  28. 28.
    Chance B, Sager R (1957) Oxygen and light induced oxidations of cytochrome, flavoprotein, and pyridine nucleotide in a chlamydomonas mutant. Plant Physiol 32(6):548–561PubMedCentralPubMedCrossRefGoogle Scholar
  29. 29.
    Chance B, Schoener B, Oshino R et al (1979) Oxidation-reduction ratio studies of mitochondria in freeze-trapped samples. NADH and flavoprotein fluorescence signals. J Biol Chem 254(11):4764–4771PubMedGoogle Scholar
  30. 30.
    Gao W, Chen G, Reinert KC et al (2006) Cerebellar cortical molecular layer inhibition is organized in parasagittal zones. J Neurosci 26(32):8377–8387. doi: 10.1523/JNEUROSCI.2434-06.2006, pii: 26/32/8377PubMedCrossRefGoogle Scholar
  31. 31.
    Theyel BB, Llano DA, Sherman SM (2010) The corticothalamocortical circuit drives higher-order cortex in the mouse. Nat Neurosci 13(1):84–88. doi: 10.1038/nn.2449, pii: nn.2449PubMedCentralPubMedCrossRefGoogle Scholar
  32. 32.
    Benson RC, Meyer RA, Zaruba ME et al (1979) Cellular autofluorescence—is it due to flavins? J Histochem Cytochem 27(1):44–48PubMedCrossRefGoogle Scholar
  33. 33.
    Lee IY, Chance B (1968) Activation of malate-linked reductions of NAD and flavoproteins in Ascaris muscle mitochondria by phosphate. Biochem Biophys Res Commun 32(3):547–553PubMedCrossRefGoogle Scholar
  34. 34.
    Hassinen I, Chance B (1968) Oxidation-reduction properties of the mitochondrial flavoprotein chain. Biochem Biophys Res Commun 31(6):895–900PubMedCrossRefGoogle Scholar
  35. 35.
    Chance B, Graham N, Mayer D (1971) A time sharing fluorometer for the readout of intracellular oxidation-reduction states of NADH and flavoprotein. Rev Sci Instrum 42(7):951–957PubMedCrossRefGoogle Scholar
  36. 36.
    Lee IY, Chance B (1977) Regulatory factors of acetaldehyde metabolism in isolated rat liver mitochondria. Adv Exp Med Biol 85A:203–224PubMedCrossRefGoogle Scholar
  37. 37.
    Barlow CH, Harden WR III, Harken AH et al (1979) Fluorescence mapping of mitochondrial redox changes in heart and brain. Crit Care Med 7(9):402–406PubMedCrossRefGoogle Scholar
  38. 38.
    Rehncrona S, Mela L, Chance B (1979) Cerebral energy state, mitochondrial function, and redox state measurements in transient ischemia. Fed Proc 38(11):2489–2492PubMedGoogle Scholar
  39. 39.
    Duchen MR, Biscoe TJ (1992) Mitochondrial function in type I cells isolated from rabbit arterial chemoreceptors. J Physiol 450:13–31PubMedCentralPubMedGoogle Scholar
  40. 40.
    Kosterin P, Kim GH, Muschol M et al (2005) Changes in FAD and NADH fluorescence in neurosecretory terminals are triggered by calcium entry and by ADP production. J Membr Biol 208(2):113–124. doi: 10.1007/s00232-005-0824-x PubMedCrossRefGoogle Scholar
  41. 41.
    Takano T, Tian G-F, Peng W et al (2007) Cortical spreading depression causes and coincides with tissue hypoxia. Nat Neurosci 10(6):754–762PubMedCrossRefGoogle Scholar
  42. 42.
    Zhao M, Nguyen J, Ma H et al (2011) Preictal and ictal neurovascular and metabolic coupling surrounding a seizure focus. J Neurosci 31(37):13292–13300. doi: 10.1523/jneurosci.2597-11.2011 PubMedCentralPubMedCrossRefGoogle Scholar
  43. 43.
    Prakash N, Biag JD, Sheth SA et al (2007) Temporal profiles and 2-dimensional oxy-, deoxy-, and total-hemoglobin somatosensory maps in rat versus mouse cortex. Neuroimage 37(suppl 1):S27–S36, Scholar
  44. 44.
    Kozberg MG, Chen BR, DeLeo SE et al (2013) Resolving the transition from negative to positive blood oxygen level-dependent responses in the developing brain. Proc Natl Acad Sci. doi: 10.1073/pnas.1212785110 PubMedCentralPubMedGoogle Scholar
  45. 45.
    Sun R, Bouchard MB, Hillman EM (2010) SPLASSH: open source software for camera-based high-speed, multispectral in-vivo optical image acquisition. Biomed Opt Express 1(2):385–397. doi: 10.1364/boe.1.000385 PubMedCentralPubMedCrossRefGoogle Scholar
  46. 46.
    Bouchard MB, Chen BR, Burgess SA et al (2009) Ultra-fast multispectral optical imaging of cortical oxygenation, blood flow, and intracellular calcium dynamics. Opt Express 17(18):15670–15678PubMedCentralPubMedCrossRefGoogle Scholar
  47. 47.
    Zhao M, Suh M, Ma H et al (2007) Focal increases in perfusion and decreases in hemoglobin oxygenation precede seizure onset in spontaneous human epilepsy. Epilepsia 48(11):2059–2067. doi: 10.1111/j.1528-1167.2007.01229.x, pii: EPI1229PubMedCrossRefGoogle Scholar
  48. 48.
    Ma H, Zhao M, Suh M et al (2009) Hemodynamic surrogates for excitatory membrane potential change during interictal epileptiform events in rat neocortex. J Neurophysiol 101(5):2550–2562. doi: 10.1152/jn.90694.2008, pii: 90694.2008PubMedCentralPubMedCrossRefGoogle Scholar
  49. 49.
    Ratzlaff EH, Grinvald A (1991) A tandem-lens epifluorescence macroscope: hundred-fold brightness advantage for wide-field imaging. J Neurosci Methods 36(2–3): 127–137PubMedCrossRefGoogle Scholar
  50. 50.
    Sato C, Nemoto M, Tamura M (2002) Reassessment of activity-related optical signals in somatosensory cortex by an algorithm with wavelength-dependent path length. Jpn J Physiol 52(3):301–312PubMedCrossRefGoogle Scholar
  51. 51.
    Zhao M, Ma H, Suh M et al (2009) Spatiotemporal dynamics of perfusion and oximetry during ictal discharges in the rat neocortex. J Neurosci 29(9):2814–2823. doi: 10.1523/JNEUROSCI.4667-08.2009, pii: 29/9/2814PubMedCentralPubMedCrossRefGoogle Scholar
  52. 52.
    Suh M, Bahar S, Mehta AD et al (2005) Temporal dependence in uncoupling of blood volume and oxygenation during interictal epileptiform events in rat neocortex. J Neurosci 25(1):68–77PubMedCrossRefGoogle Scholar
  53. 53.
    Arieli A, Shoham D, Hildesheim R et al (1995) Coherent spatiotemporal patterns of ongoing activity revealed by real-time optical imaging coupled with single-unit recording in the cat visual cortex. J Neurophysiol 73(5):2072–2093PubMedGoogle Scholar
  54. 54.
    Hohman B (2007) LED light source: major advance in fluorescence microscopy. Biomed Instrum Technol 41(6):461–464PubMedCrossRefGoogle Scholar
  55. 55.
    Albeanu DF, Soucy E, Sato TF et al (2008) LED arrays as cost effective and efficient light sources for widefield microscopy. PLoS One 3(5):e2146. doi: 10.1371/journal.pone.0002146 PubMedCentralPubMedCrossRefGoogle Scholar
  56. 56.
    Boison D (2005) Adenosine and epilepsy: from therapeutic rationale to new therapeutic strategies. Neuroscientist 11(1):25–36. doi: 10.1177/1073858404269112 PubMedCrossRefGoogle Scholar
  57. 57.
    Adamantidis AR, Zhang F, Aravanis AM et al (2007) Neural substrates of awakening probed with optogenetic control of hypocretin neurons. Nature 450(7168):420–424PubMedCrossRefGoogle Scholar
  58. 58.
    Boison D (2006) Adenosine kinase, epilepsy and stroke: mechanisms and therapies. Trends Pharmacol Sci 27(12):652–658PubMedCrossRefGoogle Scholar
  59. 59.
    Zhang F, Wang L-P, Brauner M et al (2007) Multimodal fast optical interrogation of neural circuitry. Nature 446(7136):633–639PubMedCrossRefGoogle Scholar
  60. 60.
    Zayat L, Noval MG, Campi J et al (2007) A new inorganic photolabile protecting group for highly efficient visible light GABA uncaging. Chembiochem 8(17):2035–2038. doi: 10.1002/cbic.200700354 PubMedCrossRefGoogle Scholar
  61. 61.
    Deisseroth K, Feng G, Majewska AK et al (2006) Next-generation optical technologies for illuminating genetically targeted brain circuits. J Neurosci 26(41):10380–10386. doi: 10.1523/jneurosci.3863-06.2006 PubMedCentralPubMedCrossRefGoogle Scholar
  62. 62.
    Grinvald A, Shoham D, Shmuel A et al (1999) In-vivo optical imaging of cortical architecture and dynamics. In: Windhorst U, Johansson H (eds) Modern techniques in neuroscience research. Springer, Berlin, pp 893–968CrossRefGoogle Scholar
  63. 63.
    Przybyszewski AW, Sato T, Fukuda M (2008) Optical filtering removes non-homogenous illumination artifacts in optical imaging. J Neurosci Methods 168(1):140–145, Scholar
  64. 64.
    Mayhew JEW, Askew S, Zheng Y et al (1996) Cerebral vasomotion: a 0.1-Hz oscillation in reflected light imaging of neural activity. Neuroimage 4(3):183–193, Scholar
  65. 65.
    Fekete T, Omer DB, Naaman S et al (2009) Removal of spatial biological artifacts in functional maps by local similarity minimization. J Neurosci Methods 178(1):31–39, Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Mingrui Zhao
    • 1
  • Hongtao Ma
    • 1
  • Samuel Harris
    • 2
    • 3
  • Theodore H. Schwartz
    • 4
  1. 1.Department of Neurological SurgeryBrain and Mind Research Institute, Weill Cornell Medical CollegeNew YorkUSA
  2. 2.Department of PsychologyThe University of SheffieldSheffieldUK
  3. 3.Department of Neurological SurgeryWeill Cornell Medical CollegeNew YorkUSA
  4. 4.Department of Neurological SurgeryBrain and Mind Research Institute, Weill Cornell Medical College, New York Presbyterian HospitalNew YorkUSA

Personalised recommendations