Activation of cannabinoid receptor 2 attenuates Angiotensin II-induced atrial fibrillation via a potential NOX/CaMKII mechanism

Figures

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  FIGURE 1. Under Angiotensin II stimulation, the RAAS was activated, and blood pressure (BP) rose. Ang II (2,000 ng/kg/min) was subcutaneously infused into the mice for 21 days. In addition, solvent (0.1 ml), AM1241 (20 mg/kg), and AM630 (3 mg/kg) were administered intraperitoneally every day to the animals. Before the treatment, there were no significant differences in blood pressure (BP) among the five groups. After 21 days of different treatments, BP increased significantly in all groups except the control group (A). The content of Angiotensin II (B), aldosterone (ALD) (C), malondialdehyde (MDA) (D) and glutathione (GSH) (E), and the enzyme activity of superoxide dismutase (SOD) (F) were determined by serological examination. The data represent the mean ± SEM. *P < 0.05 vs. control group. &p < 0.05 vs. before treatment. #P < 0.05 vs. Ang II + AM1241 group.

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  FIGURE 2. Administration of AM1241 reduced the inducibility of atrial fibrillation (AF) in Angiotensin II-infused mice. (A) Simultaneous recordings of surface electrocardiogram (ECG) after 21-day infusion of Ang II (2,000 ng/kg/min). Solid underlines indicate the burst pacing period, and dashed underlines highlight the AF period. The above two ECGs recorded representative ECG waves of AF and SR after a burst of electrical stimuli. The bottom recorded ECG waves of AF followed by spontaneous conversion to SR. Bar = 300 ms. (B) Inducibility of AF. (C) Duration of AF. (D) Electrophysiological mapping images of spontaneous conduction in the LA. The bars from red to blue indicate the total time from the first to the last measurement within one heartbeat. (E) Mean conduction velocity. (F) Absolute inhomogeneity of left atrial electrical conduction. (G) Representative M-mode echocardiography of the LA. (H) Quantification of LAD. The data represent the mean ± SEM. *P < 0.05 vs. control group. #P < 0.05 vs. Ang II + AM1241 group.

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  FIGURE 3. Administration of AM1241 attenuated oxidative stress in the atrial tissue of Angiotensin II-infused mice by activating cannabinoid receptor 2 (CB2R). (A) Representative western blot image and relative expression of CB2R. (B) Representative picture of reactive oxygen species (ROS) (red) by dihydroethidium staining. Bar = 100 μm. (C) Quantitative analysis of reactive oxygen species (ROS) mean intensity. (D) Representative western blot image and relative expression of NOX2 and NOX4. (E) Representative western blot image and relative expression of ox-CaMKII and CaMKII. Lane 1: control group; Lane 2: Ang II group; Lane 3: Ang II + Veh group; Lane 4: Ang II + AM1241 group; Lane 5: Ang II + AM630 group. The data represent the mean ± SEM. *P < 0.05 vs. control group. #P < 0.05 vs. Ang II + AM1241 group. @P < 0.05 vs. Ang II group. $P < 0.05 vs. Ang II + Veh group.

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  FIGURE 4. Administration of AM1241 attenuated mitochondrial damage in Angiotensin II-infused mice. (A) Representative transmission electron microscopic images of atrial samples. Bar = 1 μm. (B) Fraction of swollen mitochondria. (C) Length ratio of the mitochondrial cristae membrane and outer membrane, a parameter of mitochondrial cristae density. For each analyzed mitochondrion, the total length of the cristae membrane was obtained by adding the lengths of all the cristae together and multiplying by 2. The ratio was determined by dividing the total length of the cristae membrane by the mitochondrion perimeter. (D) Representative western blot image and relative expression of p-Drp1 and Drp1. Lane 1: control group; Lane 2: Ang II group; Lane 3: Ang II + Veh group; Lane 4: Ang II + AM1241 group; Lane 5: Ang II + AM630 group. The data represent the mean ± SEM. *P < 0.05 vs. control group. #P < 0.05 vs. Ang II + AM1241 group.

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  FIGURE 5. Administration of AM1241 attenuated atrial fibrosis in Ang II-infused mice. (A) Representative images of Masson’s trichrome staining. Bar = 50 μm. (B) Relative myocardial fibrosis. (C) Representative images of H&E staining. Bar = 50 μm. (D) Representative western blot image and relative expression of TGF-β, MMP9, collagen I and collagen III. Lane 1: control group; Lane 2: Ang II group; Lane 3: Ang II + Veh group; Lane 4: Ang II + AM1241 group; Lane 5: Ang II + AM630 group. The data represent the mean ± SEM. *P < 0.05 vs. control group. #P < 0.05 vs. Ang II + AM1241 group.

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  FIGURE 6. Proposed model describing the role of cannabinoid receptor 2 (CB2R) activation in the treatment of atrial fibrillation (AF).