Agonists for G-protein-coupled receptor 84 (GPR84) alter cellular morphology and motility but do not induce pro-inflammatory responses in microglia.

BACKGROUND: Several G-protein-coupled receptors (GPCRs) have been shown to be important signaling mediators between neurons and glia. In our previous screening for identification of nerve injury-associated GPCRs, G-protein-coupled receptor 84 (GPR84) mRNA showed the highest up-regulation by microglia after nerve injury. GPR84 is a pro-inflammatory receptor of macrophages in a neuropathic pain mouse model, yet its function in resident microglia in the central nervous system is poorly understood. METHODS: We used endogenous, natural, and surrogate agonists for GPR84 (capric acid, embelin, and 6-OAU, respectively) and examined their effect on mouse primary cultured microglia in vitro. RESULTS: 6-n-Octylaminouracil (6-OAU), embelin, and capric acid rapidly induced membrane ruffling and motility in cultured microglia obtained from C57BL/6 mice, although these agonists failed to promote microglial pro-inflammatory cytokine expression. Concomitantly, 6-OAU suppressed forskolin-induced increase of cAMP in cultured microglia. Pertussis toxin, an inhibitor of Gi-coupled signaling, completely suppressed 6-OAU-induced microglial membrane ruffling and motility. In contrast, no 6-OAU-induced microglial membrane ruffling and motility was observed in microglia from DBA/2 mice, a mouse strain that does not express functional GPR84 protein due to endogenous nonsense mutation of the GPR84 gene. CONCLUSIONS: GPR84 mediated signaling causes microglial motility and membrane ruffling but does not promote pro-inflammatory responses. As GPR84 is a known receptor for medium-chain fatty acids, those released from damaged brain cells may be involved in the enhancement of microglial motility through GPR84 after neuronal injury.

Time lapse in vivo microscopy reveals distinct dynamics of microglia-tumor environment interactions-a new role for the tumor perivascular space as highway for trafficking microglia.

Microglial cells are critical for glioma growth and progression. However, only little is known about intratumoral microglial behavior and the dynamic interaction with the tumor. Currently the scarce understanding of microglial appearance in malignant gliomas merely originates from histological studies and in vitro investigations. In order to understand the pattern of microglia activity, motility and migration we designed an intravital study in an orthotopic murine glioma model using CX3CR1-eGFP(GFP/wt) mice. We analysed the dynamics of intratumoral microglia accumulation and activity, as well as microglia/tumor blood vessel interaction by epi-illumination and 2-photon laser scanning microscopy. We further investigated cellular and tissue function, including the enzyme activity of intratumoral and microglial NADPH oxidase measured by in vivo fluorescence lifetime imaging. We identified three morphological phenotypes of tumor-associated microglia cells with entirely different motility patterns. We found that NADPH oxidase activation is highly divergent in these microglia subtypes leading to different production levels of reactive oxygen species (ROS). We observed that microglia motility is highest within the perivascular niche, suggesting relevance of microglia/tumor blood vessel interactions. In line, reduction of tumor blood vessels by antivascular therapy confirmed the relevance of the tumor vessel compartment on microglia biology in brain tumors. In summary, we provide new insights into in vivo microglial behavior, regarding both morphology and function, in malignant gliomas. GLIA 2016;64:1210-1226.

Enhanced microglial dynamics and paucity of tau seeding in the amyloid plaque microenvironment contributes to cognitive resilience in Alzheimer's disease.

Asymptomatic Alzheimer's disease (AsymAD) describes the status of subjects with preserved cognition but with identifiable Alzheimer's disease (AD) brain pathology (i.e. Aß-amyloid deposits, neuritic plaques, and neurofibrillary tangles) at autopsy. In this study, we investigated the postmortem brains of a cohort of AsymAD cases to gain insight into the underlying mechanisms of resilience to AD pathology and cognitive decline. Our results showed that AsymAD cases exhibit an enrichment of core plaques and decreased filamentous plaque accumulation, as well as an increase in microglia surrounding this last type. In AsymAD cases we found less pathological tau aggregation in dystrophic neurites compared to AD and tau seeding activity comparable to healthy control subjects. We used spatial transcriptomics to further characterize the plaque niche and found autophagy, endocytosis, and phagocytosis within the top upregulated pathways in the AsymAD plaque niche, but not in AD. Furthermore, we found ARP2, an actin-based motility protein crucial to initiate the formation of new actin filaments, increased within microglia in the proximity of amyloid plaques in AsymAD. Our findings support that the amyloid-plaque microenvironment in AsymAD cases is characterized by microglia with highly efficient actin-based cell motility mechanisms and decreased tau seeding compared to AD. These two mechanisms can potentially provide protection against the toxic cascade initiated by Aß that preserves brain health and slows down the progression of AD pathology.

Spatiotemporal analysis of impaired microglia process movement at sites of secondary neurodegeneration post-stroke.

It has recently been identified that after motor cortex stroke, the ability of microglia processes to respond to local damage cues is lost from the thalamus, a major site of secondary neurodegeneration (SND). In this study, we combine a photothrombotic stroke model in mice, acute slice and fluorescent imaging to analyse the loss of microglia process responsiveness. The peri-infarct territories and thalamic areas of SND were investigated at time-points 3, 7, 14, 28 and 56 days after stroke. We confirmed the highly specific nature of non-responsive microglia processes to sites of SND. Non-responsiveness was at no time observed at the peri-infarct but started in the thalamus seven days post-stroke and persisted for 56 days. Loss of directed process extension is not a reflection of general functional paralysis as phagocytic function continued to increase over time. Additionally, we identified that somal P2Y12 was present on non-responsive microglia in the first two weeks after stroke but not at later time points. Finally, both classical microglia activation and loss of process extension are highly correlated with neuronal damage. Our findings highlight the importance of microglia, specifically microglia dynamic functions, to the progression of SND post-stroke, and their potential relevance as modulators or therapeutic targets during stroke recovery.