Live imaging of cell interactions in the normal and diseased brain

We are studying cell-cell interactions in the normal and diseased central nervous system. Many cell types interact sequentially in the diseased CNS and synergistically control the evolution of pathologies.  The dynamics of these interactions as well as their outcome are poorly known due to the lack of resolving non-invasive imaging methodologies.

We have set up two in vivo imaging modalities with different scales of spatial resolution, X-ray micro computed tomography (CT) and 2-photon (2P) microscopy, to visualize neural and immune cells as well as blood vessels in mouse models of central nervous system pathologies.  The mastering of 2P in vivo imaging combined to the use of mouse models whose different cell populations shine in different colors allows simultaneous multicolor imaging as well as unlimited number of examinations of the same field of view. This opens the way to quantitative and correlative analysis of cell distributions and interactions in a truly physiological environment. So far, we have described over time and space the interplay between neural cells and angiogenesis in the context of glioblastoma (GBM) and spinal cord injury (SCI).We showed that unexpectedly, GMB tumor growth was not directly related to blood supply and that the transient anti-angiogenic therapeutic effect, also observed in patients, was more likely due to an effect on the stroma. By contrast, in SCI, the axons trying to regenerate always regrowth in the vicinity of blood vessels. To better explore these issues, we are presently studying the dynamics of neuronal/vascular/inflammatory responses in these pathologies. Collectively, our data constitute a framework to investigate the involvement of sub-populations of interest in the diseases’ evolution by means of their specific deletion through the use of dedicated mouse models, or their pharmacological manipulations. For example, VEGF is up-regulated in both pathologies and  can act on several cellular targets in addition to its angiogenic action. By combining the use of transgenic multicolor fluorescent mice with VEGF gain and loss of function, we are clarifying VEGF-dependent events and their time window.  The acquired information should help optimizing VEGF treatment protocols.

In collaboration with physicists, we validated a high resolution computed tomography (micro-CT) system, based on a new generation of detectors called Hybrid Pixel Detectors (HPD) to observe the soft tissues of the mouse. Moreover, because hybrid pixel detector allows photons discrimination based on an energy threshold, this should allow to implement the contrast enhancing K-edge method.  In other word, we expect to implement a multicolor CT scanner to obtain complementary information on the same animals previously imaged with the 2P microscope at a local scale. Although offering far less resolution than 2P microscopy, CT scan presents the complementary advantage of exploring the whole body.

These technologies allow the collection of large amounts of data from a single animal hence allowing for reduction in the number of animals engaged in an experimental study and improving the significance of the results. 

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