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Simultaneous two-photon imaging of oxygen and blood flow in deep cerebral vessels

Nature Medicine volume 17, pages 893–898 (2011)Cite this article

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Abstract

Uncovering principles that regulate energy metabolism in the brain requires mapping of partial pressure of oxygen (PO2) and blood flow with high spatial and temporal resolution. Using two-photon phosphorescence lifetime microscopy (2PLM) and the oxygen probe PtP-C343, we show that PO2 can be accurately measured in the brain at depths up to 300 μm with micron-scale resolution. In addition, 2PLM allowed simultaneous measurements of blood flow and of PO2 in capillaries with less than one-second temporal resolution. Using this approach, we detected erythrocyte-associated transients (EATs) in oxygen in the rat olfactory bulb and showed the existence of diffusion-based arterio-venous shunts. Sensory stimulation evoked functional hyperemia, accompanied by an increase in PO2 in capillaries and by a biphasic PO2 response in the neuropil, consisting of an 'initial dip' and a rebound. 2PLM of PO2 opens new avenues for studies of brain metabolism and blood flow regulation.

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Figure 1: Measurements of PO2 in deep cerebral vessels using probe PtP-C343.
Figure 2: Temporal and spatial resolution of two-photon phosphorescence measurement.
Figure 3: Simultaneous measurements of RBC flow and PO2 using fluorescence and phosphorescence of PtP-C343.
Figure 4: Diffusional shunt of oxygen between arterial and venous compartments in the olfactory nerve layer.
Figure 5: Functional hyperemia, vascular and neuropil PO2 dynamics in response to odor stimulation.

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Acknowledgements

We thank V. De Sars and N. Chaari for technical assistance in the design of electronic circuits, G. Bouchery for her help in rat surgery, C. Pouzat for help with statistical analysis and L.E. Sinks for discussion of the phosphorescence microscopy data. Support was provided by INSERM, CNRS, the Région Ile de France (Sesame program), the Fondation Bettancourt Schueller the Leducq Foundation, the Human Frontier Science Program Organization, the European Commission FP6 (LSHM-CT-2007-037765), the Fondation pour la Recherche Médicale and the US National Institutes of Health (grant EB007279). Photophysical characterization of the probe was performed in the Ultrafast Optical Processes Laboratory at the University of Pennsylvania (US National Institutes of Health grant P41-RR001348).

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  1. Jérôme Lecoq

    Present address: Current address: James H. Clark Center for Biomedical Engineering and Sciences, Stanford University, Stanford, California, USA.,

  2. Jérôme Lecoq, Alexandre Parpaleix and Emmanuel Roussakis: These authors contributed equally to this work.

Authors and Affiliations

  1. Institut National de la Santé et de la Recherche Médicale (INSERM), U603, Paris, France

    Jérôme Lecoq, Alexandre Parpaleix, Mathieu Ducros, Yannick Goulam Houssen & Serge Charpak

  2. Centre National de la Recherche Scientifique (CNRS), UMR 8154, Paris, France

    Jérôme Lecoq, Alexandre Parpaleix, Mathieu Ducros, Yannick Goulam Houssen & Serge Charpak

  3. Laboratory of Neurophysiology and New Microscopies, Université Paris Descartes, Paris, France

    Jérôme Lecoq, Alexandre Parpaleix, Mathieu Ducros, Yannick Goulam Houssen & Serge Charpak

  4. Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, Pennsylvania, USA

    Emmanuel Roussakis & Sergei A Vinogradov

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Contributions

E.R. and S.A.V. designed and synthesized the oxygen probe. J.L. and M.D. designed and built the optical setup. J.L. and M.D. wrote the LabVIEW program controlling the system and analyzing the data. J.L., A.P., Y.G.H. and S.C. conducted the experiments and analyzed the data. J.L. and S.C. initiated the project. All authors edited the paper.

Corresponding authors

Correspondence to Sergei A Vinogradov or Serge Charpak.

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The authors declare no competing financial interests.

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Supplementary Figures 1–3 and Supplementary Methods (PDF 704 kb)

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Lecoq, J., Parpaleix, A., Roussakis, E. et al. Simultaneous two-photon imaging of oxygen and blood flow in deep cerebral vessels. Nat Med 17, 893–898 (2011). https://doi.org/10.1038/nm.2394

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