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Neuronal network activity controls microglial process surveillance in awake mice via norepinephrine signaling

Nature Neuroscience volume 22, pages 1771–1781 (2019)Cite this article

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Abstract

Microglia dynamically survey the brain parenchyma. Microglial processes interact with neuronal elements; however, what role neuronal network activity plays in regulating microglial dynamics is not entirely clear. Most studies of microglial dynamics use either slice preparations or in vivo imaging in anesthetized mice. Here we demonstrate that microglia in awake mice have a relatively reduced process area and surveillance territory and that reduced neuronal activity under general anesthesia increases microglial process velocity, extension and territory surveillance. Similarly, reductions in local neuronal activity through sensory deprivation or optogenetic inhibition increase microglial process surveillance. Using pharmacological and chemogenetic approaches, we demonstrate that reduced norepinephrine signaling is necessary for these increases in microglial process surveillance. These findings indicate that under basal physiological conditions, noradrenergic tone in awake mice suppresses microglial process surveillance. Our results emphasize the importance of awake imaging for studying microglia–neuron interactions and demonstrate how neuronal activity influences microglial process dynamics.

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Fig. 1: Microglial process surveillance is increased after general anesthesia.
Fig. 2: Whisker trimming increases microglial process surveillance in the barrel cortex of awake mice.
Fig. 3: Intracerebral application of muscimol increases microglial process surveillance.
Fig. 4: Optogenetic suppression of neuronal activity through VGAT-ChR2-mediated inhibition increases microglial process surveillance.
Fig. 5: NE signaling, but not P2Y12 or CX3CR1 signaling, contributes to microglial surveillance increases.
Fig. 6: NE administration before whisker trimming or optogenetic inhibition prevents the increase in microglial process surveillance.
Fig. 7: Blocking adrenergic receptors or lowering endogenous NE signaling increases microglial process surveillance.
Fig. 8: NE decreases microglial process surveillance in acute brain slices.

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Data availability

Source data for all graphs are available from the corresponding author upon request.

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Acknowledgements

This work is supported by the National Institutes of Health (R01NS088627, R21DE025689 and R01NS112144 to L.-J.W.) and by a postdoctoral fellowship from the Mayo Clinic Center for Multiple Sclerosis and Autoimmune Neurology (to T.C). The authors thank M. Mattson (National Institute on Aging) for critical reading of the paper and members of the Wu Lab at the Mayo Clinic for insightful discussions.

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Authors and Affiliations

  1. Department of Neurology, Mayo Clinic, Rochester, MN, USA

    Yong U. Liu, Yanlu Ying, Yujiao Li, Ukpong B. Eyo, Tingjun Chen, Jiaying Zheng, Anthony D. Umpierre, Jia Zhu, Dale B. Bosco & Long-Jun Wu

  2. Department of Neuroscience and Center for Brain Immunology and Glia, University of Virginia, Charlottesville, VA, USA

    Ukpong B. Eyo

  3. Department of Anesthesiology, Xijing Hospital, Fourth Military Medical University, Xi’an, China

    Hailong Dong

  4. Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA

    Long-Jun Wu

  5. Department of Immunology, Mayo Clinic, Rochester, MN, USA

    Long-Jun Wu

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Contributions

Y.U.L., H.D. and L.-J.W. designed the studies. Y.U.L., A.D.U. and L.-J.W. wrote and revised the manuscript. Y.U.L. performed the electrophysiology experiments. Y.U.L., Y.Y., Y.L., U.B.E., T.C., J. Zheng, A.D.U., J .Zhu and D.B.B. performed animal surgery, image collection and data analyses. Y.U.L. and J. Zheng performed the in situ hybridization and immunofluorescence staining experiments.

Corresponding author

Correspondence to Long-Jun Wu.

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Supplementary information

Supplementary Information

Supplementary Figs. 1–12.

Reporting Summary

Supplementary Video 1

Microglial process surveillance increases after anesthesia. Time-lapse imaging of microglia (50–64 µm in depth) in the somatosensory cortex before and after isoflurane anesthesia. Experiments were repeated three times independently with similar results.

Supplementary Video 2

Neuronal network activity in the somatosensory cortex immediately decreases after isoflurane anesthesia. Calcium activity in excitatory CamKII-positive soma (150 µm in depth, left) or dendrites (50 µm, right) before and after isoflurane induction. Experiments were repeated three times independently with similar results.

Supplementary Video 3

Neuronal network activity in the barrel cortex decreases after contralateral whisker trimming. Calcium activity in CamKII neuronal dendrites (50 µm in depth, right) of the contralateral barrel cortex before and after whisker trimming. Experiments were repeated three times independently with similar results.

Supplementary Video 4

Microglial process surveillance increases after contralateral whisker trimming. Time-lapse imaging of microglia (50–64 µm in depth) in the contralateral barrel cortex before and after whisker trimming. Experiments were repeated three times independently with similar results.

Supplementary Video 5

Intracerebral administration of muscimol (870 µM) reduces neuronal network activity. Calcium activity of CamKII neuronal somas (150 µm in depth) in the somatosensory cortex before and after muscimol administration. Experiments were repeated three times independently with similar results.

Supplementary Video 6

Microglial process surveillance increases after intracerebral administration of muscimol (870 µM). Time-lapse imaging of microglia (50–64 µm depth) in somatosensory cortex before and after muscimol administration. Experiments were repeated three times independently with similar results.

Supplementary Video 7

Microglial process surveillance increases after optogenetic stimulation of VGAT-positive inhibitory neurons. Time-lapse imaging of microglia (50–64 µm depth) in the somatosensory cortex before and after optogenetic stimulation (10 Hz, 1 ms pulse, 10 min of stimulation) of ChR2-expressing VGAT-positive interneurons. Experiments were repeated three times independently with similar results.

Supplementary Video 8

In vivo imaging dense noradrenergic neuronal projections from the locus coeruleus. Z-stack movie showing td-Tomato-labeled axons, which project from noradrenergic neurons in the LC. The video progresses from the pial surface down to a 100 µm in depth in the somatosensory cortex. Experiments were repeated three times independently with similar results.

Supplementary Video 9

Neuronal network activity in acute brain slices exposed to glutamate. Calcium activity in excitatory CamKII-positive soma in three cortical slices exposed to vehicle, 50 µM glutamate or 1 mM glutamate. Experiments were repeated three times independently with similar results.

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Liu, Y.U., Ying, Y., Li, Y. et al. Neuronal network activity controls microglial process surveillance in awake mice via norepinephrine signaling. Nat Neurosci 22, 1771–1781 (2019). https://doi.org/10.1038/s41593-019-0511-3

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