Neuroinflammation driven by interferon-gamma (IFN-γ) and microglial activation has been linked to neurological disease. However, the effects of IFN-γ-activated microglia on hippocampal neurogenesis and behavior are unclear. In the present study, IFN-γ was administered to mice via intracerebroventricular injection. Mice received intraperitoneal injection of ruxolitinib to inhibit the JAK/STAT1 pathway or injection of minocycline to inhibit microglial activation. During a 7-day period, mice were assessed for depressive-like behaviors and cognitive impairment based on a series of behavioral analyses. Effects of the activated microglia on neural stem/ precursor cells (NSPCs) were examined, as was pro-inflammatory cytokine expression by activated microglia. We showed that IFN-γ-injected animals showed long-term adult hippocampal neurogenesis reduction, behavior despair, anhedonia, and cognitive impairment. Chronic activation with IFN-γ induces reactive phenotypes in microglia associated with morphological changes, population expansion, MHC II and CD68 up-regulation, and pro-inflammatory cytokine (IL-1β, TNF-α, IL-6) and nitric oxide (NO) release. Microglia isolated from the hippocampus of IFN-γ-injected mice suppressed NSPCs proliferation and stimulated apoptosis of immature neurons. Inhibiting of the JAK/STAT1 pathway in IFN-γ-injected animals to block microglial activation suppressed microglia-mediated neuroinflammation and neurogenic injury, and alleviated depressive-like behaviors and cognitive impairment. Collectively, these findings suggested that priming of microglia with IFN-γ impairs adult hippocampal neurogenesis and leads to depression-like behaviors and cognitive defects. Targeting microglia by modulating levels of IFN-γ the brain may be a therapeutic strategy for neurodegenerative diseases and psychiatric disorders.
The morphology of microglial cells is often closely related to their functions. The mechanisms that regulate microglial ramification are not well understood. Here we reveal the biological mechanisms by which astrocytes regulate microglial ramification. Morphological variation in mouse microglial cultures was measured in terms of cell area as well as branch number and length. Effects on microglial ramification were analyzed after microinjecting the toxin L-alpha-aminoadipic acid (L-AAA) in the mouse cortex or hippocampus to ablate astrocytes, and after culturing microglia on their own in an astrocyte-conditioned medium (ACM) or together with astrocytes in coculture. TGF-β expression was determined by Western blotting, immunohistochemistry, and ELISA. The TGF-β signaling pathway was blocked by the TGF-β antibody to assess the role of TGF-β on microglial ramification. The results showed that microglia had more and longer branches and smaller cell bodies in brain areas where astrocytes were abundant. In the mouse cortex and hippocampus, ablation of astrocytes by L-AAA decreased number and length of microglial branches and increased the size of cell bodies. Similar results were obtained with isolated microglia in culture. However, isolated microglia were able to maintain their multibranched structure for a long time when cultured on astrocyte monolayers. Ameboid microglia isolated from P0 to P3 mice showed increased ramification when cultured in ACM or on astrocyte monolayers. Microglia cultured on astrocyte monolayers showed more complex branching structures than those cultured in ACM. Blocking astrocyte-derived TGF-β decreased microglial ramification. Astrocytes induced the formation of protuberances on branches of microglia by forming glial fibers that increased traction. These experiments in mice suggest that astrocytes promote microglial ramification by forming glial fibers to create traction and by secreting soluble factors into the surroundings. For example, astrocyte-secreted TGFβ promotes microglia to generate primitive branches, whose ramification is refined by glial fibers.
Purpose: To investigate the effect of dexmedetomidine on streptozotocin (STZ)-induced diabetic neuropathy pain (DNP) in rats and elucidate its mechanism of action.Methods: The DNP rat model was established by injecting STZ (70 mg/kg) following dexmedetomidine treatment. Next BV-2 cells were stimulated using lipopolysaccharide (LPS, 200 ng/mL) and then administered 20 μM dexmedetomidine. Blood glucose levels, body weight, and paw withdrawal threshold (PWT) were measured once a week in DNP rats. Transfection was performed, and luciferasereporter assay was used to verify microRNA (miR)-337 binding to Rap1A mRNA. Reverse transcriptionpolymerase chain reaction (RT-PCR) was used to measure the levels of miR-618 and P2Y12 while the protein levels of P2Y12 and ionized calcium-binding adaptor molecule 1 (IBA-1) were determined by western blot analysis.Results: Dexmedetomidine treatment significantly increased PWT (p < 0.01) in DNP rats and decreased miR-618 expression (p < 0.01) but increased P2Y12 expression (p < 0.01) in the spinal cord of DNP rats. Luciferase reporter assay data showed that the presumed binding site of miR-618 is located in the 3′-untranslated regions of P2Y12. MiR-618 overexpression significantly reduced P2Y12levels (p < 0.01). Dexmedetomidine upregulated P2Y12 expression (p < 0.01) but decreased IBA-1 expression (p < 0.01).Conclusion: Dexmedetomidine application attenuates DNP by inhibiting microglial activation via the regulation of miR-618/P2Y12 pathway. This finding provides a potential therapeutic strategy for DNP management. Keywords: Dexmedetomidine, Diabetic neuropathy pain, Paw withdrawal threshold, Calcium-binding adaptor molecule 1, MiR-618, P2Y12
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