Short-Chain Fatty Acids Improve Poststroke Recovery via Immunological Mechanisms.

Recovery after stroke is a multicellular process encompassing neurons, resident immune cells, and brain-invading cells. Stroke alters the gut microbiome, which in turn has considerable impact on stroke outcome. However, the mechanisms underlying gut-brain interaction and implications for long-term recovery are largely elusive. Here, we tested the hypothesis that short-chain fatty acids (SCFAs), key bioactive microbial metabolites, are the missing link along the gut-brain axis and might be able to modulate recovery after experimental stroke. SCFA supplementation in the drinking water of male mice significantly improved recovery of affected limb motor function. Using in vivo wide-field calcium imaging, we observed that SCFAs induced altered contralesional cortex connectivity. This was associated with SCFA-dependent changes in spine and synapse densities. RNA sequencing of the forebrain cortex indicated a potential involvement of microglial cells in contributing to the structural and functional remodeling. Further analyses confirmed a substantial impact of SCFAs on microglial activation, which depended on the recruitment of T cells to the infarcted brain. Our findings identified that microbiota-derived SCFAs modulate poststroke recovery via effects on systemic and brain resident immune cells.SIGNIFICANCE STATEMENT Previous studies have shown a bidirectional communication along the gut-brain axis after stroke. Stroke alters the gut microbiota composition, and in turn, microbiota dysbiosis has a substantial impact on stroke outcome by modulating the immune response. However, until now, the mediators derived from the gut microbiome affecting the gut-immune-brain axis and the molecular mechanisms involved in this process were unknown. Here, we demonstrate that short-chain fatty acids, fermentation products of the gut microbiome, are potent and proregenerative modulators of poststroke neuronal plasticity at various structural levels. We identified that this effect was mediated via circulating lymphocytes on microglial activation. These results identify short-chain fatty acids as a missing link along the gut-brain axis and as a potential therapeutic to improve recovery after stroke.

In vivo widefield calcium imaging of the mouse cortex for analysis of network connectivity in health and brain disease.

The organization of brain areas in functionally connected networks, their dynamic changes, and perturbations in disease states are subject of extensive investigations. Research on functional networks in humans predominantly uses functional magnetic resonance imaging (fMRI). However, adopting fMRI and other functional imaging methods to mice, the most widely used model to study brain physiology and disease, poses major technical challenges and faces important limitations. Hence, there is great demand for alternative imaging modalities for network characterization. Here, we present a refined protocol for in vivo widefield calcium imaging of both cerebral hemispheres in mice expressing a calcium sensor in excitatory neurons. We implemented a stringent protocol for minimizing anesthesia and excluding movement artifacts which both imposed problems in previous approaches. We further adopted a method for unbiased identification of functional cortical areas using independent component analysis (ICA) on resting-state imaging data. Biological relevance of identified components was confirmed using stimulus-dependent cortical activation. To explore this novel approach in a model of focal brain injury, we induced photothrombotic lesions of the motor cortex, determined changes in inter- and intrahemispheric connectivity at multiple time points up to 56 days post-stroke and correlated them with behavioral deficits. We observed a severe loss in interhemispheric connectivity after stroke, which was partially restored in the chronic phase and associated with corresponding behavioral motor deficits. Taken together, we present an improved widefield calcium imaging tool accounting for anesthesia and movement artifacts, adopting an advanced analysis pipeline based on human fMRI algorithms and with superior sensitivity to recovery mechanisms in mouse models compared to behavioral tests. This tool will enable new studies on interhemispheric connectivity in murine models with comparability to human imaging studies for a wide spectrum of neuroscience applications in health and disease.