Diffusion-weighted magnetic resonance spectroscopy enables cell-specific monitoring of astrocyte reactivity in vivo

Abstract

Reactive astrocytes exhibit hypertrophic morphology and altered metabolism. Deciphering astrocytic status would be of great importance to understand their role and dysregulation in pathologies, but most analytical methods remain highly invasive or destructive. The diffusion of brain metabolites, as non-invasively measured using diffusion-weighted magnetic resonance spectroscopy (DW-MRS) in vivo, depends on the structure of their micro-environment. Here we perform advanced DW-MRS in a mouse model of reactive astrocytes to determine how cellular compartments confining metabolite diffusion are changing. This reveals myo-inositol as a specific intraastrocytic marker whose diffusion closely reflects astrocytic morphology, enabling non-invasive detection of astrocyte hypertrophy (subsequently confirmed by confocal microscopy ex vivo). Furthermore, we measure massive variations of lactate diffusion properties, suggesting that intracellular lactate is predominantly astrocytic under control conditions, but predominantly neuronal in case of astrocyte reactivity. This indicates massive remodeling of lactate metabolism, as lactate compartmentation is tightly linked to the astrocyte-to-neuron lactate shuttle mechanism.

Revisiting double diffusion encoding MRS in the mouse brain at 11.7T: Which microstructural features are we sensitive to?

“…Consequently, a heavy fitting pipeline may be needed to analyze experimental data, for instance involving a dictionary approach based on Monte Carlo simulations (Ligneul et al, 2019;Palombo et al, 2016;Rensonnet et al, 2019). The sensitivity to a specific feature of the cell microstructure, i.e.…”

“…We then wondered if a realistic soma, rather than a branched fiber structure, was also able to better account for DDE behavior. Hence we set dsoma =10 µm, while imposing a very simple fiber structure, so we set Nbranch=1 and Lseg=450 µm, so that fibers can be considered "almost" infinite and simultaneously ensures that the soma occupies ~16% of the total cellular volume, which is a realistic volume fraction for both neurons and astrocytes (Chklovskii et al, 2002;Chvatal et al, 2007;Ligneul et al, 2019;Sherwood et al, 2004) (Figure 6C). A very strong drop of A is observed, while B is decreased as the same value as case 2.…”

“…Different diffusion-weighted MRS (DW-MRS) approaches have been proposed to quantify cell microstructure, such as high-b value experiments (mainly to probe fiber diameter but also presumably yielding some sensitivity to cell body diameter (Ligneul et al, 2019;Palombo et al, 2017;Palombo et al, 2018a, b)), or measurements of the apparent diffusion coefficient (ADC) up to very long diffusion times (td) to probe long-range cell structure (potentially enabling quantification of cell fiber branching and length (Ligneul et al, 2019;Palombo et al, 2016)). Such approaches were recently used to quantitatively estimate alterations of astrocytic morphology in a mouse model of reactive astrocytes, where the diffusion of Ins was significantly different compared to the control group, while the diffusion of other metabolites remained unchanged (Ligneul et al, 2019). In that work, modeling of Ins data resulted in higher values for fiber diameter and length in the case of activated astrocytes, which was confirmed by quantitative morphological analysis of astrocytes using confocal microscopy ex vivo, thus demonstrating the potential of the technique for microstructure quantification.…”

Questions and (some) answers on reactive astrocytes

“…(d) 3D rendering of the morphology of a normal (blue) and a hypertrophic ciliary neurotrophic factor (CNTF)-induced reactive astrocyte (brown) in the mouse striatum. Scale bar: 50 μm (Ligneul et al, 2019). (e) A nonreactive (up) and reactive (down) astrocyte in the mouse cortex, 4 days after cortical lesion.…”

“…Indeed, astrocytes may significantly contribute to imaging signals, such as those measured with blood oxygenation level-dependent functional MRI or [ 18 F]-fluorodeoxyglucose uptake by PET, and therefore their reactive state may impact these measurements (Carter et al, 2019;Mishra, 2017). Refined approaches that are more selective for reactive astrocytes are being developed (Carter et al, 2019;Ligneul et al, 2019;Rodriguez-Vieitez et al, 2016;Scholl et al, 2015). Such improvements will be facilitated by basic studies that establish the molecular and functional profile of reactive astrocytes (see Section 6) but also of their microglia or neuronal neighbors, allowing the identification of new and specific astrocyte targets.…”