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Noninvasive 40-Hz light flicker to recruit microglia and reduce amyloid beta load


Microglia, the primary immune cells of the brain, play a key role in pathological and normal brain function. Growing efforts aim to reveal how these cells may be harnessed to treat both neurodegenerative diseases such as Alzheimer’s and developmental disorders such as schizophrenia and autism. We recently showed that using noninvasive exposure to 40-Hz white-light (4,000 K) flicker to drive 40-Hz neural activity transforms microglia into an engulfing state and reduces amyloid beta, a peptide thought to initiate neurotoxic events in Alzheimer’s disease (AD). This article describes how to construct an LED-based light-flicker apparatus, expose animals to 40-Hz flicker and control conditions, and perform downstream assays to study the effects of these stimuli. Light flicker is simple, faster to implement, and noninvasive, as compared with driving 40-Hz activity using optogenetics; however, it does not target specific cell types, as is achievable with optogenetics. This noninvasive approach to driving 40-Hz neural activity should enable further research into the interactions between neural activity, molecular pathology, and the brain’s immune system. Construction of the light-flicker system requires ~1 d and some electronics experience or available guidance. The flicker manipulation and assessment can be completed in a few days, depending on the experimental design.

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Fig. 1: Diagram of exposure timing and sample stimulus conditions.
Fig. 2: Alternative circuit option: soldered circuit.
Fig. 3: Schematic of the breadboard circuit.
Fig. 4: Flicker exposure setup.
Fig. 5: Coronal sections showing mouse visual cortex.
Fig. 6: Measurement of microglia cell body diameter and process length using ImageJ.
Fig. 7: Expected results.

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We are grateful to H. Iaccarino, S. Kale, and members of the Tsai, Singer, and Boyden laboratories for methods development, technical assistance, and comments on the paper. A.C.S. acknowledges the Lane family and the Packard Foundation; A.J.M. was supported by T32 GM007484 Integrative Neuronal Systems, MIT Champions of the Brain (Alder) Fellowship, and MIT Henry E. Singleton Fellowship; L.-H.T. acknowledges the JPB Foundation, Belfer Neurodegeneration Consortium, Halis Family Foundation, and NIH grant RF1 AG047661.

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Authors and Affiliations



A.C.S. developed the light-flicker methods; A.C.S., J.M.D., A.J.M., F.A., H.M., and C.A. performed the experiments; A.C.S., J.M.D., M.K.A., and J.T. constructed and tested the flicker circuitry; and A.C.S., J.M.D., A.J.M., F.A., and L.-H.T. wrote the manuscript.

Corresponding authors

Correspondence to Annabelle C. Singer or Li-Huei Tsai.

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Competing interests

L.-H.T. is the scientific founder and serves on the scientific advisory board of Cognito Therapeutics, and A.C.S. owns shares of Cognito Therapeutics. The remaining authors declare no competing interests.

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Related links

Key references using this protocol

1. Iaccarino, H. F. et al. Nature 540, 230–235 (2016)

2. Mathys, H. et al. Cell Rep. 21, 366–380 (2017)

3. Seo, J. et al. Cell 157, 486–498 (2014)

Supplementary information

Supplementary Data

Example code to control the Arduino.

Reporting Summary

Supplementary Video

Mouse exposed to light flicker.

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Singer, A.C., Martorell, A.J., Douglas, J.M. et al. Noninvasive 40-Hz light flicker to recruit microglia and reduce amyloid beta load. Nat Protoc 13, 1850–1868 (2018).

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