Digital Reconstruction of the Neuro-Glia-Vascular Architecture.

Astrocytes connect the vasculature to neurons mediating the supply of nutrients and biochemicals. They are involved in a growing number of physiological and pathophysiological processes that result from biophysical, physiological, and molecular interactions in this neuro-glia-vascular ensemble (NGV). The lack of a detailed cytoarchitecture severely restricts the understanding of how they support brain function. To address this problem, we used data from multiple sources to create a data-driven digital reconstruction of the NGV at micrometer anatomical resolution. We reconstructed 0.2 mm3 of the rat somatosensory cortex with 16 000 morphologically detailed neurons, 2500 protoplasmic astrocytes, and its microvasculature. The consistency of the reconstruction with a wide array of experimental measurements allows novel predictions of the NGV organization, allowing the anatomical reconstruction of overlapping astrocytic microdomains and the quantification of endfeet connecting each astrocyte to the vasculature, as well as the extent to which they cover the latter. Structural analysis showed that astrocytes optimize their positions to provide uniform vascular coverage for trophic support and signaling. However, this optimal organization rapidly declines as their density increases. The NGV digital reconstruction is a resource that will enable a better understanding of the anatomical principles and geometric constraints, which govern how astrocytes support brain function.

Simultaneous optical recording in multiple cells by digital holographic microscopy of chloride current associated to activation of the ligand-gated chloride channel GABA(A) receptor.

Chloride channels represent a group of targets for major clinical indications. However, molecular screening for chloride channel modulators has proven to be difficult and time-consuming as approaches essentially rely on the use of fluorescent dyes or invasive patch-clamp techniques which do not lend themselves to the screening of large sets of compounds. To address this problem, we have developed a non-invasive optical method, based on digital holographic microcopy (DHM), allowing monitoring of ion channel activity without using any electrode or fluorescent dye. To illustrate this approach, GABA(A) mediated chloride currents have been monitored with DHM. Practically, we show that DHM can non-invasively provide the quantitative determination of transmembrane chloride fluxes mediated by the activation of chloride channels associated with GABA(A) receptors. Indeed through an original algorithm, chloride currents elicited by application of appropriate agonists of the GABA(A) receptor can be derived from the quantitative phase signal recorded with DHM. Finally, chloride currents can be determined and pharmacologically characterized non-invasively simultaneously on a large cellular sampling by DHM.