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Journal of Integrative Neuroscience  2019, Vol. 18 Issue (1): 51-56    DOI: 10.31083/j.jin.2019.01.114
Original Research Previous articles | Next articles
Acetylcholine suppresses microglial inflammatory response via $\alpha$7nAChR to protect hippocampal neurons
Lin Li1, #, Zhan Liu1, #, Yong-Ying Jiang1, Wei-Xing Shen1, Yu-Ping Peng1, *(), Yi-Hua Qiu1, *()
1Department of Physiology, School of Medicine, and Co-innovation Center of Neuroregeneration, Nantong University, 19 Qixiu Road, Nantong, Jiangsu Province 226001, China
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Abstract  

Neuroinflammation is principally linked to glial function and has been demonstrated to participate in the pathogenesis of Alzheimer's disease, a neurodegenerative disorder characterized by beta-amyloid ccumulation and neurotransmission disruption. Previous findings suggest acetylcholine exerts anti-inflammatory and neuroprotective properties in several neurodegenerative disorders. However, the underlying mechanisms remain elusive. Here evaluation of the influence of acetylcholine on neuroinflammation and neurodegeneration in Alzheimer's disease is reported and further neuroprotective mechanisms are investigated. Investigation of microglia in lipopolysaccharide-induced hippocampal neuronal toxicity employed $\alpha$7nAChR gene silencing and demonstrated that both the anti-inflammatory and neuroprotective effects of acetylcholine rely on $\alpha$7nAChR pathways. As expected, in neuron-microglia co-cultures lipopolysaccharide induced an increase in expression of pro-inflammatory factors, including inducible nitric oxide synthase, interleukin-1$\beta$, and tumor necrosis factor-$\alpha$, and decreased expression of neurotrophic factors such as insulin-like growth factor-1, and neuronal apoptosis. Acetylcholine protects against lipopolysaccharide-elicited neuronal injury by inhibiting the microglial inflammatory response and promoting microglial neurotrophic factor production via the action of $\alpha$7nAChR on microglia. These findings establish that ACh activates $\alpha$7nAChR in microglia, which in turn protects hippocampal neurons.

Key words:  Acetylcholine      $\alpha$7nAChR      hippocampal neuron      lipopolysaccharide      microglia     
Submitted:  06 December 2018      Accepted:  22 March 2019      Published:  30 March 2019     
Fund: 

  • 31371182, 31771293/National Natural Science Foundation of China
  • 15B25/Nantong University - Priority Academic Program Development (PAPD) of Jiangsu Higher Education Institutions

*Corresponding Author(s):  Yu-Ping Peng E-mail: yppeng@ntu.edu.cn; Yi-Hua Qiu E-mail: yhqiu@ntu.edu.cn   
# These authors contribute equally.

Cite this article: 

Lin Li, Zhan Liu, Yong-Ying Jiang, Wei-Xing Shen, Yu-Ping Peng, Yi-Hua Qiu. Acetylcholine suppresses microglial inflammatory response via $\alpha$7nAChR to protect hippocampal neurons. Journal of Integrative Neuroscience, 2019, 18(1): 51-56.

URL: 

https://jin.imrpress.com/EN/10.31083/j.jin.2019.01.114     OR     https://jin.imrpress.com/EN/Y2019/V18/I1/51

Figure 1.  ACh reduces LPS-induced upregulation of pro-inflammatory mediators in neuron-microglia co-cultures. Cultures treated with LPS (100 ng/mL) for two hours followed by ACh addition (10$^{-7}$ or 10$^{-9}$ mol/L) and a 24 hour incubation. Western blot assay is employed to detect the expression of pro-inflammatory and neurotrophic factors ($n$ = 4). ** $p$ < 0.01 versus control; $^{++}p$ < 0.01-versus LPS.

Figure 2.  Silencing of $\alpha$7nAChR gene in microglia abolishes the ability of ACh to inhibit LPS-induced inflammatory responses. Treatment: Microglia transfected with $\alpha$7nAChR-shRNA lentiviral vector co-cultured with hippocampal neurons, Control: Scr-shRNA lentiviral vector transfected microglia. Neuron-microglia co-cultures were treated with LPS (100 ng/mL) for two hours followed by ACh application, then incubated for 24 hours. (A) $\alpha$7nAChR expression levels in microglia exposed to various treatments. (B) Representative electrophoretic bands and statistical analysis of protein expression levels. (C) Concentration of TNF-a, IL-1 $\beta$, and IGF-1 in neuron-microglia co-culture supernatants. ** $p$ < 0.01 versus control; $^{++}p$ < 0.01 versus LPS; $^{{\#}}$ $p$ < 0.05, versus LPS + ACh treatment; $^{\& \&}p$ < 0.01 versus LPS + 10$^{-9}$ mol/L ACh, $n$= 4 or 6.

Figure 3.  Silencing of $\alpha$7nAChR gene in microglia abolishes the ability of ACh to inhibit LPS-induced neuronal apoptosis. Neuron-microglia co-cultures were treated as described in preceding figure legends. Cells grown on coverslips inside 24-well plates were subjected to NeuN immunostaining and TUNEL assay. (A) Representative photomicrographs are given. (B) Quantitative summary and statistical analysis. Histogram shows the relative proportion of TUNEL and NeuN double-positive cells in the NeuN-positive cell population ($n$ = 7). ** $p$ < 0.01 versus control; $^{++}p$ < 0.01 versus LPS; $^{{\#}{\#}}$ $p$ < 0.01 versus LPS + ACh treatment.

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