Targeted Occlusion to Surface and Deep Vessels in Neocortex via Linear and Nonlinear Optical Absorption

  • David Kleinfeld
  • Beth Friedman
  • Patrick D. Lyden
  • Andy  Y. Shih
Part of the Springer Protocols Handbooks book series (SPH)

We discuss two complementary methods for the study of cerebral blood flow and brain function in response to the occlusion of individual, targeted blood vessels. These bear on the study of microstroke and vascular dysfunction in cortex. One method makes use of linear optical absorption by a photosensitizer, transiently circulated in the blood stream, to induce an occlusion in a surface or near-surface vessel. The second method makes use of nonlinear optical interactions, without the need to introduce an exogenous absorber, to induce an occlusion in a subsurface microvessel. A feature of both methods is that the dynamics of blood flow and functional aspects of the vas-culature and underlying neurons in the neighborhood of the occluded vessel may be monitored before, during, and after the occlusion. We present details of both methods and associated surgical procedures, along with exemplary data from published studies.


Ablation Clot Hypoxia Ischemia Microstroke Photosensitizer Plasma Rodent Two-photon 


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  1. 1.
    Iadecola C. Neurovascular regulation in the normal brain and in Alzheimer's disease. Nature Reviews of Neuroscience 2004;5:347–360CrossRefGoogle Scholar
  2. 2.
    Scremin OU. Cerebral vascular system. In: Paxinos G, ed. The Rat Nervous System. 2 ed. San Diego: Academic Press; 1995Google Scholar
  3. 3.
    Miyake K, Takeo S, Kaijihara H. Sustained decrease in brain regional blood flow after microsphere embolism in rats. Stroke 1993;24:415–420PubMedGoogle Scholar
  4. 4.
    Sugi T, Schuier FJ, Hossmann KA, Zulch KJ. The effect of mild microem-bolic injury on the energy metabolism of the cat brain. Journal of Neurology 1980;223(4):285–292PubMedCrossRefGoogle Scholar
  5. 5.
    Gerriets T, Li F, Silva MD, et al. The macrosphere model: evaluation of a new stroke model for permanent middle cerebral artery occlusion in rats. Journal of Neuroscience Methods 2003;122(2):201–211PubMedCrossRefGoogle Scholar
  6. 6.
    Zhang Z, Zhang RL, Jiang Q, Raman SB, Cantwell L, Chopp M. A new rat model of thrombotic focal cerebral ischemia. Journal of Cerebral Blood Flow and Metabolism 1997;17(2):123–135PubMedCrossRefGoogle Scholar
  7. 7.
    Fuxe K, Bjelke B, Andbjer B, Grahn H, Rimondini R, Agnati LF. Endothelin-1 induced lesions of the frontoparietal cortex of the rat. A possible model of focal cortical ischemia. Neuroreport 1997;8(11):2623–2629PubMedCrossRefGoogle Scholar
  8. 8.
    Macrae IM, Robinson MJ, Graham DI, Reid JL, McCulloch J. Endothelin-1-induced reductions in cerebral blood flow: dose dependency, time course, and neuropathological consequences. Journal of Cerebral Blood Flow and Metabolism 1993;13(2):276–284PubMedGoogle Scholar
  9. 9.
    Sharkey J, Ritchie IM, Kelly PA. Perivascular microapplication of endothelin-1: a new model of focal cerebral ischaemia in the rat. Journal of Cerebral Blood Flow and Metabolism 1993;13(5):865–871PubMedGoogle Scholar
  10. 10.
    Zhang S, Boyd J, Delaney KR, Murphy TH. Rapid reversible changes in dendritic spine structure in vivo gated by the degree of ischemia. Journal of Neuroscience 2005;25:5333–5228PubMedCrossRefGoogle Scholar
  11. 11.
    Wei L, Rovainen CM, Woolsey TA. Ministrokes in rat barrel cortex. Stroke 1995;26:1459–1462PubMedGoogle Scholar
  12. 12.
    Chen ST, Hsu C Y, Hogan EL, Maricq H, Balentine JD. A model of focal ischemic stroke in the rat: Reproducible extensive cortical infarction. Stroke 1986;17(4): 738–743PubMedGoogle Scholar
  13. 13.
    Buchan AM, Xue D, Slivka A. A new model of temporary focal neocortical ischemia in the rat. Stroke 1992;23(2):273–279PubMedGoogle Scholar
  14. 14.
    Tamura A, Asano T, Sano K. Corrrelation between rCBF and histological changes following temporary middle cerebral artery occlusion. Stroke 1980;11:487–493PubMedGoogle Scholar
  15. 15.
    Katsman D, Zheng J, Spinelli K, Carmichael ST. Tissue microenvironments within functional cortical subdivisions adjacent to focal stroke. Journal of Cerebral Blood Flow and Metabolism 2003;23(9):997–1009PubMedGoogle Scholar
  16. 16.
    Markgraf CG, Kraydieh S, Prado R, Watson BD, Dietrich WD, Ginsberg MD. Comparative histopathologic consequences of photothrombotic occlusion of the distal middle cerebral artery in Sprague-Dawley and Wistar rats. Stroke 1993;24: 286–293PubMedGoogle Scholar
  17. 17.
    Helmchen F, Denk W. Deep tissue two-photon microscopy. Nature Methods 2005;2:932–940PubMedCrossRefGoogle Scholar
  18. 18.
    Schaffer CB, Friedman B, Nishimura N, et al. Two-photon imaging of cortical surface microvessels reveals a robust redistribution in blood flow after vascular occlusion. Public Library of Science Biology 2006;4:258–270Google Scholar
  19. 19.
    Nishimura B, Schaffer CB, Friedman B, Lyden PD, Kleinfeld D. Penetrating arte-rioles are a bottleneck in the perfusion of neocortex. Proceedings of the National Academy of Sciences USA 2007;104:365–370CrossRefGoogle Scholar
  20. 20.
    Nishimura N, Schaffer CB, Friedman B, Tsai PS, Lyden PD, Kleinfeld D. Targeted insult to individual subsurface cortical blood vessels using ultrashort laser pulses: Three models of stroke. Nature Methods 2006;3:99–108PubMedCrossRefGoogle Scholar
  21. 21.
    Svoboda K, Denk W, Kleinfeld D, Tank DW. In vivo dendritic calcium dynamics in neocortical pyramidal neurons. Nature 1997;385:161–165PubMedCrossRefGoogle Scholar
  22. 22.
    Kleinfeld D, Mitra PP, Helmchen F, Denk W. Fluctuations and stimulus-induced changes in blood flow observed in individual capillaries in layers 2 through 4 of rat neocortex. Proceedings of the National Academy of Sciences USA 1998;95:15741–15746CrossRefGoogle Scholar
  23. 23.
    Kleinfeld D, Denk W. Two-photon imaging of neocortical microcirculation. In: Yuste R, Lanni F, Konnerth A, eds. Imaging Neurons: A Laboratory Manual. Cold Spring Harbor: Cold Spring Harbor Laboratory Press; 2000:23.1–23.15Google Scholar
  24. 24.
    Kleinfeld D, Denk W. Two-photon imaging of cortical microcirculation. In: Yuste R, Konnerth A, eds. Imaging in Neuroscience and Development. Cold Spring Harbor: Cold Spring Harbor Laboratory Press; 2005:701–705Google Scholar
  25. 25.
    Zhang S, Murphy TH. Imaging the impact of cortical microcirculation on synaptic structure and sensory-evoked hemodynamic responses in vivo. Public Library of Science Biology 2007;5:e119Google Scholar
  26. 26.
    Tsai PS, Nishimura N, Yoder EJ, Dolnick EM, White GA, Kleinfeld D. Principles, design, and construction of a two photon laser scanning microscope for in vitro and in vivo brain imaging. In: Frostig RD, ed. In Vivo Optical Imaging of Brain Function. Boca Raton: CRC Press; 2002:113–171Google Scholar
  27. 27.
    Majewska A, Yiu G, Yuste R. A custom-made two-photon microscope and decon-volution system. Pflugers Archives 2000;441:398–408CrossRefGoogle Scholar
  28. 28.
    Short CE. Principles and Practice of Veterinary Anesthesia. Baltimore: Williams and Willisms; 1987Google Scholar
  29. 29.
    Paxinos G, Watson C. The Rat Brain in Stereotaxic Coordinates. San Diego: Academic Press; 1986Google Scholar
  30. 30.
    Kleinfeld D, Delaney KR. Distributed representation of vibrissa movement in the upper layers of somatosensory cortex revealed with voltage sensitive dyes. Journal of Comparative Neurology 1996;375:89–108PubMedCrossRefGoogle Scholar
  31. 31.
    Dirnagl U, Villringer A, Einhaupl KM. In-vivo confocal scanning laser microscopy of the cerebral microcirculation. Journal of Microscopy 1992;165:147–157PubMedGoogle Scholar
  32. 32.
    Brozici M, van der Zwain A, Hillen B. Anatomy and functionality of leptomenin-geal anastomoses: A review. Stroke 2003;34:2750–2762PubMedCrossRefGoogle Scholar
  33. 33.
    Nishimura N, Rosidi NL, Mandell J, Iadecola C, Schaffer CB. Neighboring arte-rioles do not dilate to increase collateral flow after the occlusion of a cortical penetrating arteriole. In: Abstracts of the Society for Neuroscience Annual Meeting, Neuroscience 2007Google Scholar
  34. 34.
    Nimmerjahn A, Kirchhoff F, Helmchen F. Resting microglial cells are highly dynamic surveillants of brain parenchyma in vivo. Science 2005;308(5726): 1314–1318PubMedCrossRefGoogle Scholar
  35. 35.
    Clay GO, Schaffer CB, Kleinfeld D. Large two-photon absorptivity of hemoglobin in the infrared range of 780–880 nm. Journal of Chemical Physics 2007;126:025102PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science + Business Media, LLC 2009

Authors and Affiliations

  • David Kleinfeld
    • 1
  • Beth Friedman
    • 1
  • Patrick D. Lyden
    • 1
  • Andy  Y. Shih
    • 2
  1. 1.Department of Physics, Graduate Program in NeurosciencesUniversity of California at San DiegoLa JollaUSA
  2. 2.Neuroprotection Research Laboratory, Departments of Neurology and Radiology, Massachusetts General HospitalHarvard Medical SchoolCharlestownUSA

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