Low-Level Vagus Nerve Stimulation Reverses Obstructive Sleep Apnea-Related Atrial Fibrillation by Ameliorating Sympathetic Hyperactivity and Atrial Myocyte Injury

Figures

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  Figure 1. Flow chart of the study protocol. (A). Acute OSA protocol. (B). Changes in blood gases pre- and post-apnea between the OSA and OSA+LLVS groups. (C). *p<0.05 vs. the pre-apnea; OSA, obstructive sleep apnea; LLVS, low-level vago-sympathetic trunk stimulation; EP, electrophysiology; BP, blood pressure; ECG, electrocardiography; PaO2, partial pressures of oxygen; PaCO2, partial carbon dioxide tension; ELISA, enzyme-linked immunosorbent assay; H, hours; min, minute.

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  Figure 2. (A) Representative immunohistochemical staining of PGP 9.5, TH, and ChAT on the LA among the three groups. (B) Quantitative analysis of PGP 9.5, TH, and ChAT-positive nerve density in the LA of each group. (C) The concentration of NE among the three groups at baseline, the 3rd hour, and the 6th hour. (D) Representative silver staining in the LA among the three groups. (E) Quantitative analysis of silver staining-positive nerve density in the LA of each group. *p<0.05, **p<0.01 vs. the control group; #p<0.05, ##p<0.01 vs. the OSA+LLVS group; PGP 9.5, protein gene product; TH, tyrosine hydroxylase; ChAT, choline acetyltransferase; NE, norepinephrine; H, hours.

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  Figure 3. (A) Representative examples of SNRT in the OSA group at baseline, the 1st hour, the 2nd hour, the 3rd hour, the 4th hour, and the 6th hour. (B) Changes in SNRT between the OSA and OSA+LLVS groups at each hour. (C) Changes in ion channels via gene expression level among the three groups. (D) Representative immunohistochemical staining of Cx 40 and Cx 43 on the LA among the three groups. (E) Quantitative analysis of Cx 43 and Cx 40-positive cells in the LA of each group. *p<0.05, **p<0.01 vs. the control group; #p<0.05, ##p<0.01 vs. the OSA+LLVS group; SNRT, sinoatrial node recovery time; Cx 43, connexin 43; Cx40, connexin 40; H, hours.

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  Figure 4. (A) Neural activity recording during the process of apnea and re-ventilation, (a) representative examples of the neural recording at the first 40s of apnea, (b) representative examples of the neural recording at the second 40s of apnea, (c) representative examples of the neural recording at the third 40s of apnea, (d) representative examples of the neural recording at the first 40s of re-ventilation; (B) The dynamic changes in HR and BP before and after apnea in OSA; (C) Changes in BP pre and after apnea between the OSA and OSA+LLVS groups at baseline, the 2nd hour, the 4th hour, and the 6th hour. *p<0.05. BS, baseline; H, hours. HR, heart rate; BP, blood pressure.

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  Figure 5. (A) Representative examples of AF episodes in the OSA and OSA+LLVS groups at baseline, the 2nd hour, the 3rd hour, the 4th hour, and the 6th hour. (B) Changes in ERP, AF duration, AH interval, and HV interval at each hour among the three groups. *p<0.05, **p<0.01, ##p<0.05; ERP, effective refractory period; AF, atrial fibrillation; AH, Atrial-His; HV, His-Ventricle; H, hours.

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  Figure 6. (A) Echocardiography shows images of the LA among the three groups. (B) The LA gradually and significantly increased after OSA, and the LA decreased through LLVS treatment. (C) There was no statistically significant difference in RA among the three groups. (D) OSA-induced FS gradually shorten in the OSA group, and LLVS treatment significantly increased FS. (E) Changes in the mitochondrial and stellate ganglion (F) by transmission electron microscopy among the three groups. **p<0.05 vs. the control group; #p<0.05 vs. the OSA+LLVS group; LA, left atrium; RA, right atrium; FS, fractional shortening; BS, baseline; H, hours.

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  Figure 7. Changes in cardiomyocytes among the three groups. HE staining showed that cardiomyocytes in the LA (A) and RA (B) were arranged tightly and orderly across the three groups. (C) Representative TUNEL staining on the LA across the three groups. (D) Quantitative analysis of TUNEL-positive cells in the LA of each group. (E) Representative PAS staining in the LA among the three groups. (F) Quantitative analysis of the glycogen volume fraction in the LA of each group. (G) The concentrations of TNF-α and IL-6 among the three groups at baseline, the 3rd hour, and the 6th hour. *p<0.05, **p<0.01 vs. the control group; #p<0.05 vs. the OSA+LLVS group; HE, hematoxylin and eosin; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling; PAS, periodic acid-Schiff; TNF-α, tumor necrosis factor alpha; IL-6, interleukin 6; H, hour.

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  Figure 8. Graphic illustration of possible mechanism about acute OSA induced-atrial fibrillation.