The Lesion Analysis of Cholinergic Neurons in 5XFAD Mouse Model in the Three-Dimensional Level of Whole Brain

Abstract

Cholinergic system is very important for many higher brain functions, including learning and memory. Cholinergic neurons, especially those in the basal forebrain, are specifically susceptible in some neurodegenerative diseases, such as in Alzheimer’s disease (AD). Here, we studied the cholinergic system lesion effects of five familial AD mutations in 5XFAD mice, a transgenic mouse model of AD. Although the cholinergic system has been studied in this mouse model, the cholinergic deficits in AD mice have never been systematically mapped in a whole-brain three-dimensional (3D) reconstruction. Using the 3D reconstruction technology combined with immunohistochemistry (3D-IHC) and design-based stereology, we comprehensively compared the differences of the cholinergic neurons and fibers between the 5XFAD mice and C57BL/6 control mice at different age. Here, we found that the lesion of cholinergic fibers occurred earlier than the cholinergic neuron loss in 5XFAD mice. The cholinergic fiber lesions in the AD mice started sequentially in amygdala, cortex, hippocampus, and then basal forebrain. However, the basal forebrain was the first brain region observed with cholinergic neuron loss at the age of 9 months in 5XFAD mice, whereas such phenomenon first occurred at the age of 15 months in C57BL/6 control mice. Moreover, using 3D reconstruction to compare the lesion of cholinergic system of aged 5XFAD and C57BL/6 control mice, it is intuitive to notice the pathologic regions and severity of lesion. Therefore, the 3D-IHC provides detailed overview of the cholinergic neurons in the whole mouse brain, which will contribute to the study of the developing and pathologic mouse brain.

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Change history

  • 07 July 2017

    An erratum to this article has been published.

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Acknowledgements

We gratefully acknowledge Dr. Qing Tian, Dr. Linqiang Zhu, and Dr. Juan Chen for their helpful discussion and suggestions; Jiaming Liang for the image alignment code; and Shaohua Zhang for the assistance with equipment maintenance. This study was supported by the National Nature Science Foundation of China (Grant Nos. 81130079, 91232302, 81571078, and 3160047).

Author information

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Authors

Contributions

Y.L. conceived and designed the study. H.Y. and P.P. did the work of immunohistochemistry and three-dimensional reconstruction of cholinergic neurons. W.C., H.Z., H.L., Z.W., X.K., and J.W. conducted the counting work of cholinergic neurons and cholinergic lesions. T.Z. and K.P. conducted the statistical work. H.Y., L.P., Y.H., H.K., and Y.L. wrote the manuscript.

Corresponding authors

Correspondence to Yunyun Han or Youming Lu.

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The authors declare that they have no competing interests.

Additional information

The original version of this article was revised: The presentation of the name K. A. Henok needs to be updated to “Henok K.A.”. Also in authors contribution, the last sentence should be read as “H.Y., L.P., Y.H., H.K., and Y.L. wrote the manuscript.”

Electronic Supplementary Material

Supplementary Figure 1
figure8

The labeling of cholinergic fiber lesions in the aged 5XFAD mouse brain. (GIF 60 kb).

Supplementary Figure 2
figure9

The distribution diagram of cholinergic system in mouse brain. (GIF 37 kb).

Supplementary Figure 3
figure10

The cholinergic fiber lesions in the LEnt of aged C57BL/6 and 5XFAD mice (20 months old). Black arrow indicates the fiber lesion, and the green arrow .indicates the normal cholinergic fiber. (GIF 78 kb).

12035_2017_621_MOESM4_ESM.mov

The 3D movie of cholinergic neurons in the mouse brain. Firstly, there are all the ChAT-positive neurons in the hemisphere mouse brain. Then, these neurons in the different brain regions are labeled by different color balls. Last, all the cholinergic fiber lesions in the various brain regions are labeled by different color balls. (MOV 6910 kb).

High resolution image (TIFF 9676 kb).

High resolution image (TIFF 6507 kb).

High resolution image (TIFF 7985 kb).

Supplementary Video 1

The 3D movie of cholinergic neurons in the mouse brain. Firstly, there are all the ChAT-positive neurons in the hemisphere mouse brain. Then, these neurons in the different brain regions are labeled by different color balls. Last, all the cholinergic fiber lesions in the various brain regions are labeled by different color balls. (MOV 6910 kb).

ESM 1

The code, which was written based on the software of MATLAB R2015b, was used for the alignment of pictures of serial brain sections. (M 1 kb).

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Yan, H., Pang, P., Chen, W. et al. The Lesion Analysis of Cholinergic Neurons in 5XFAD Mouse Model in the Three-Dimensional Level of Whole Brain. Mol Neurobiol 55, 4115–4125 (2018). https://doi.org/10.1007/s12035-017-0621-4

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Keywords

  • Alzheimer’s disease
  • 5XFAD mouse model
  • Cholinergic neurons
  • Three dimension
  • Whole brain