Priming of microglia with IFN-γ slows neuronal gamma oscillations in situ.

Type II IFN (IFN-γ) is a proinflammatory T lymphocyte cytokine that serves in priming of microglia-resident CNS macrophages-during the complex microglial activation process under pathological conditions. Priming generally permits an exaggerated microglial response to a secondary inflammatory stimulus. The impact of primed microglia on physiological neuronal function in intact cortical tissue (in situ) is widely unknown, however. We explored the effects of chronic IFN-γ exposure on microglia in hippocampal slice cultures, i.e., postnatal parenchyma lacking leukocyte infiltration (adaptive immunity). We focused on fast neuronal network waves in the gamma-band (30-70 Hz). Such gamma oscillations are fundamental to higher brain functions, such as perception, attention, and memory, and are exquisitely sensitive to metabolic and oxidative stress. IFN-γ induced substantial morphological changes and cell population expansion in microglia as well as moderate up-regulation of activation markers, MHC-II, CD86, IL-6, and inducible nitric oxide synthase (iNOS), but not TNF-α. Cytoarchitecture and morphology of pyramidal neurons and parvalbumin-positive inhibitory interneurons were well-preserved. Notably, gamma oscillations showed a specific decline in frequency of up to 8 Hz, which was not mimicked by IFN-α or IL-17 exposure. The rhythm disturbance was caused by moderate microglial nitric oxide (NO) release demonstrated by pharmacological microglia depletion and iNOS inhibition. In conclusion, IFN-γ priming induces substantial proliferation and moderate activation of microglia that is capable of slowing neural information processing. This mechanism might contribute to cognitive impairment in chronic brain disease featuring elevated IFN-γ levels, blood-brain barrier leakage, and/or T cell infiltration, well before neurodegeneration occurs.

TLR2- and TLR3-activated microglia induce different levels of neuronal network dysfunction in a context-dependent manner.

Recognition of pathogen- or damage-associated molecular patterns (PAMPs, DAMPs) by innate Toll-like receptors (TLRs) is central to the activation of microglia (brain macrophages) in many CNS diseases. Notably, TLR-mediated microglial activation is complex and modulated by additional exogenous and endogenous immunological signals. The impact of different microglial reactive phenotypes on electrical activity and neurotransmission is widely unknown, however. We explored the effects of TLR ligands on microglia and neuronal network function in rat organotypic hippocampal slice cultures (in situ), i.e., postnatal cortical tissue lacking adaptive immunity. Single exposure of slice cultures to TLR2 or TLR3 ligands [PGN, poly(I:C)] for 2-3 days induced moderate microglial activation featuring IL-6 and TNF-α release and only mild alterations of fast neuronal gamma band oscillations (30-70 Hz) that are fundamental to higher cognitive functions, such as perception, memory and behavior. Paired exposure to TLR3/TLR2 or TLR3/TLR4 ligands (LPS) induced nitric oxide (NO) release, enhanced TNF-α release, and associated with advanced network dysfunction, including slowing to the beta frequency band (12-30 Hz) and neural bursts (hyperexcitability). Paired exposure to a TLR ligand and the leukocyte cytokine IFN-γ enhanced NO release and associated with severe network dysfunction, albeit sensitive parvalbumin- and somatostatin-positive inhibitory interneurons were preserved. Notably, the neuronal disturbance was prevented by either microglial depletion or pharmacological inhibition of oxidant-producing enzymes, inducible NO synthase (iNOS) and NADPH oxidase. In conclusion, TLR-activated microglia can induce different levels of neuronal network dysfunction, in which severe dysfunction is mainly caused by reactive oxygen and nitrogen species rather than proinflammatory cytokines. Our findings provide a mechanistic insight into microglial activation and functional neuronal network impairment, with relevance to neuroinflammation and neurodegeneration observed in, e.g., meningoencephalitis, multiple sclerosis and Alzheimer's disease.

GM-CSF induces noninflammatory proliferation of microglia and disturbs electrical neuronal network rhythms in situ.

BACKGROUND: The granulocyte-macrophage colony-stimulating factor (GM-CSF) (or CSF-2) is involved in myeloid cell growth and differentiation, and, possibly, a major mediator of inflammation in body tissues. The role of GM-CSF in the activation of microglia (CNS resident macrophages) and the consequent impacts on neuronal survival, excitability, and synaptic transmission are widely unknown, however. Here, we focused on electrical neuronal network rhythms in the gamma frequency band (30-70 Hz). Gamma oscillations are fundamental to higher brain functions, such as perception, attention, and memory, and they are exquisitely sensitive to metabolic and oxidative stress. METHODS: We explored the effects of chronic GM-CSF exposure (72 h) on microglia in male rat organotypic hippocampal slice cultures (in situ), i.e., postnatal cortex tissue lacking leukocyte invasion (adaptive immunity). We applied extracellular electrophysiological recordings of local field potential, immunohistochemistry, design-based stereology, biochemical analysis, and pharmacological ablation of microglia. RESULTS: GM-CSF triggered substantial proliferation of microglia (microgliosis). By contrast, the release of proinflammatory cytokines (IL-6, TNF-α) and nitric oxide, the hippocampal cytoarchitecture as well as the morphology of parvalbumin-positive inhibitory interneurons were unaffected. Notably, GM-CSF induced concentration-dependent, long-lasting disturbances of gamma oscillations, such as slowing (beta frequency band) and neural burst firing (hyperexcitability), which were not mimicked by the T lymphocyte cytokine IL-17. These disturbances were attenuated by depletion of the microglial cell population with liposome-encapsulated clodronate. In contrast to priming with the cytokine IFN-γ (type II interferon), GM-CSF did not cause inflammatory neurodegeneration when paired with the TLR4 ligand LPS. CONCLUSIONS: GM-CSF has a unique role in the activation of microglia, including the potential to induce neuronal network dysfunction. These immunomodulatory properties might contribute to cognitive impairment and/or epileptic seizure development in disease featuring elevated GM-CSF levels, blood-brain barrier leakage, and/or T cell infiltration.