Electric stimulation of the vagus nerve reduced mouse neuroinflammation induced by lipopolysaccharide - PubMed (original) (raw)

doi: 10.1186/s12950-016-0140-5. eCollection 2016.

M Bautista 1, A Florentino 1, G Díaz 1, G Acero 1, H Besedovsky 2, D Meneses 3, A Fleury 4, A Del Rey 2, G Gevorkian 1, G Fragoso 1, E Sciutto 1

Affiliations

Electric stimulation of the vagus nerve reduced mouse neuroinflammation induced by lipopolysaccharide

G Meneses et al. J Inflamm (Lond). 2016.

Abstract

Background: Neuroinflammation (NI) is a key feature in the pathogenesis and progression of infectious and non-infectious neuropathologies, and its amelioration usually improves the patient outcome. Peripheral inflammation may promote NI through microglia and astrocytes activation, an increased expression of inflammatory mediators and vascular permeability that may lead to neurodegeneration. Several anti-inflammatory strategies have been proposed to control peripheral inflammation. Among them, electrical stimulation of the vagus nerve (VNS) recently emerged as an alternative to effectively attenuate peripheral inflammation in a variety of pathological conditions with few side effects. Considering that NI underlies several neurologic pathologies we explored herein the possibility that electrically VNS can also exert anti-inflammatory effects in the brain.

Methods: NI was experimentally induced by intraperitoneal injection of bacterial lipopolysaccharide (LPS) in C57BL/6 male mice; VNS with constant voltage (5 Hz, 0.75 mA, 2 ms) was applied for 30 s, 48 or 72 h after lipopolysaccharide injection. Twenty four hours later, pro-inflammatory cytokines (IL-1β, IL-6, TNFα) levels were measured by ELISA in brain and spleen extracts and total brain cells were isolated and microglia and macrophage proliferation and activation was assessed by flow cytometry. The level of ionized calcium binding adaptor molecule (Iba-1) and glial fibrillary acidic protein (GFAP) were estimated in whole brain extracts and in histologic slides by Western blot and immunohistochemistry, respectively.

Results: VNS significantly reduced the central levels of pro-inflammatory cytokines and the percentage of microglia (CD11b/CD45low) and macrophages (CD11b/CD45high), 24 h after the electrical stimulus in LPS stimulated mice. A significantly reduced level of Iba-1 expression was also observed in whole brain extracts and in the hippocampus, suggesting a reduction in activated microglia.

Conclusions: VNS is a feasible therapeutic tool to attenuate the NI reaction. Considering that NI accompanies different neuropathologies VNS is a relevant alternative to modulate NI, of particular interest for chronic neurological diseases.

Keywords: Antiinflammatory; Lipopolysaccharide neuropathologies; Microglia; Neuroinflammation; Stimulation of vagus nerve.

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Figures

Fig. 1

Fig. 1

Procedure of implanted electrodes and location by CTscan. a Procedure of electrode insertion into the vagus nerve. 1. Photograph showing the trachea and the sternocleidomastoid muscle, carotid. 2. Sheath with carotid and vagus nerve. 3. Vagus nerve is shown isolated. 4. Electrodes applied to the vagus nerve. t = trachea, ECOM = sternocleidomastoid, c = carotid, VN = vagus nerve. b Representative view of the electrodes inserted in the vagus nerve by Computed Tomography Scan

Fig. 2

Fig. 2

Experimental design line. a LPS or ISS were injected at day 0. The inflammatory peripheral and central status was evaluated before and at the different times after injection. b Electrodes were implanted at day 0. Five days later, mice were randomly divided into five groups: sham, isotonic saline solution treated mice (ISS), Vagus Nerve Stimulation (VNS), LPS and LPS + VNS

Fig. 3

Fig. 3

a A representative dot plot of isolated brain cells from saline or LPS treated mice analyzed by flow cytometry. b Mean ± SD of the percentage of CD11b/CD45low and CD11b/CD45high before and 48, 72, and 96 h after LPS treatment and mean ± SD of the fluorescence intensities of CD11b (c). Different literals indicate significant differences between the different groups (p < 0.05) using the Kruskal-Wallis test (non-parametric ANOVA) plus the Dunn’s multiple comparisons test. Data are representative of four experiments

Fig. 4

Fig. 4

a Representative dot plot of isolated brain cells analyzed by flow citometry. b Mean ± SD of the percentage of CD11b/CD45low and CD11b/CD45high cells and the respective Mean ± SD of the fluorescence intensities of CD11b (c) of five mice per group. Data are representative of two different independent assays. Different literals indicate significant differences in the percent of CD11b/CD45low and CD11b/CD45high cells (P < 0.05) between the different groups using the Kruskal- Wallis test (non-parametric ANOVA) plus the Dunn’s multiple comparisons test

Fig. 5

Fig. 5

Analysis of brain Iba-1 expression in ISS, VNS, LPS, and LPS + VNS- treated mice. a Representative Western blot showing Iba-1 and β-actin. b Western blot analysis of Iba-1 in whole brain homogenates of SSI, VNS, LPS, and LPS + VNS-treated mice. Each column represents the level of Iba-1 expressed as mean ± SEM, normalized to β-actin in a same gel. Different literals indicate significant differences in the expression level of Iba-1 among the different groups using one-way Analysis of Variance plus the Tukey-Kramer multiple comparisons test. F (3,16) = 6.7, p < 0.004. c Representative mouse brain sections of the different groups stained with anti-Iba-1 antibody (to detect microglia). Bottom images represent a nine-fold magnification of the region outlined in the box in the corresponding upper image. In LPS-treated mice, higher numbers of microglia with morphological characteristics of activated cells were observed

Fig. 6

Fig. 6

Analysis of GFAP expression in the brain of ISS, VNS, LPS-treated and LPS + VNS-treated. a Representative Western blots showing GFAP and β-actin levels. b Western blot analysis of GFAP in whole brain homogenates of untreated, VNS, LPS-treated, and LPS + VNS-treated mice. Each column in the graph represents the level of GFAP expressed as Mean ± SEM, normalized to β-actin in a same gel. No significant differences between GFAP levels were observed using one-way Analysis of Variance plus the Tukey-Kramer multiple comparisons test. F (3,16) = 0.44, p = 0.72. c Representative mouse brain sections of the different groups of mice stained with anti-GFAP antibody (to detect astrocytes). Bottom images represent a nine-fold magnification of the region outlined in the box in the corresponding upper image

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References

    1. Cinel I, Opal SM. Molecular biology of inflammation and sepsis: a primer. Crit Care Med. 2009;37:291–304. doi: 10.1097/CCM.0b013e31819267fb. - DOI - PubMed
    1. Dubový P, Klusáková I, Hradilová Svíženská I. Inflammatory profiling of Schwann cells in contact with growing axons distal to nerve injury. Biomed Res Int. 2014;2014:691041. doi: 10.1155/2014/691041. - DOI - PMC - PubMed
    1. Lang BT, Wang J, Filous AR, Au NP, Ma CH, Shen Y. Pleiotropic molecules in axon regeneration and neuroinflammation. Exp Neurol. 2014;258:17–23. doi: 10.1016/j.expneurol.2014.04.031. - DOI - PubMed
    1. Singhal G, Jaehne EJ, Corrigan F, Toben C, Baune BT. Inflammasomes in neuroinflammation and changes in brain function: a focused review. Front Neurosci. 2014;8:315–328. doi: 10.3389/fnins.2014.00315. - DOI - PMC - PubMed
    1. Setiawan E, Wilson AA, Mizrahi R, Rusjan PM, Miler L, Rajkowska G, et al. Role of translocator protein density, a marker of neuroinflammation, in the brain during major depressive episodes. JAMA Psychiat. 2015;72:268–75. doi: 10.1001/jamapsychiatry.2014.2427. - DOI - PMC - PubMed

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