Figure legends
Figure 1. (A) Schematic of a heart with a window cut into the right atrium to visualize the location of the atrioventricular node (AVN). (B) Image of the AVN preparation used for constructing the model, the location of sections and the angle of the knife.
Figure 2. The 13th, 39th, and 79th groups of serial sections are shown. (A) Masson’s trichrome-stained section. (B) AChE staining: AChE expression was largely positive around the tricuspid ring and dispersed in distribution (arrow). (C) Cx43 (green) and HCN4 (red) double-staining showed reciprocal Cx43 and HCN4 expression. (D) Cx40 (green) and Cx45 (red) double-label staining showed abundant Cx45 expression in the penetrating bundle (PB) tissue and areas along the atrial side where Cx43 expression was absent. Cx40 was expressed only in the PB tissue. (E) Masson’s trichrome-stained section. (F) AChE staining: Rusty-colored tissues highlighted a linear connection with the atrioventricular node (AVN) from the atrial side. (G) Cx43 (green) and HCN4 (red) double-staining revealed a correlation with AChE staining. (H) Cx40 (green) and Cx45 (red) double-staining showed abundant Cx45 expression in areas without Cx43 expression. PB tissue showed a longitudinal dissociation, with the volume of Cx40 expression decreasing compared with that at the former level (elliptical area). Cx40 expression was not present on the atrial side. (I) Masson’s trichrome staining showed the inferior extension of the AVN located on the atrial side above the tricuspid annules. (J) AChE staining was limited to the INE area. (K) Cx43 expression was absent in the inferior extension where HCN4 expression was present. The inferior extension divided into the right extension and the left extension (arrow). (L) Cx45 expression was more abundant in the INE than in other atrial areas (a slight fold in the section). No Cx40 expression was observed in the inferior extension.
Figure B(a) shows the location of images shown in Figures C(a) and D(a), while Figure B(b) shows the location of images shown in Figures C(b) and D(b). Figure F(a) shows the location of images shown in Figures G(a) and 2H(a), while Figure F(b) shows the location of images shown in Figures G(b) and H(b).
Figure 3. Continuous sections at the level of start of the CN and the end of the RN from another sample. (A1-A3) Part of the node-like tissue was separated from the RN and penetrated into the fibrous rings between the atrium and ventricle (dotted box). (B1-B3) The following continuous sections showed that the nodal tissue continued in the fibrous rings. (C1-C3) The following continuous sections showed that the nodal tissue descended into the CN. (A3, B3, C3) HCN4 labeling further suggested that the penetrated tissue was node-like tissue (dotted box). (D-F) The region connecting the RN and CN was also near the start of the tendon of Todaro (black arrow). Two narrow connections between the RN and CN at different levels near the tendon of Todaro (dotted box). After the node-like tissue descended into the CN, fibrous tissue surrounding the other part of the CN seemed to have no direct connection with the atrium.
Figure 4. (Left) Eighty model sections. The distance between groups is 6 * 4 = 12 μm. (Right) Anatomical model views of the rat atrioventricular node (AVN). (A and B) Nodal tissue model viewed from the right and left. (C and D) Nodal (yellow) and penetrating bundle (red) tissue models viewed from the right and left. (E and F) Conduction system tissue and valve ring (blue) model viewed from the right and left.
Figure 5. (A) Nav1.5 expression was abundant in the atrial and ventricular muscles but weak in the PB and RN. Cav3.1 was mainly expressed in the PB and RN. (B) and (C) The CN and INE also weakly expressed Nav1.5 but positively expressed Cav3.1. (D) Analysis of the Nav1.5 immunofluorescence staining intensity showed that the RN, CN and INE all presented significantly low Nav1.5 expression (P <0.001). No significant differences in expression were observed among the RN, CN and INE (P =0.104), but a trend toward slightly higher Nav1.5 expression in the RN than in the other two regions was observed.
Figure 6. Hypothesis of the structure and functional relationship of the AVN. (A) During sinus rhythm, the action potential propagates preferentially along the RN due to the presence of fewer connective tissue barriers to the atrial tissue. (B) In the slow-fast form of AVNRT, anterograde conduction occurs through INE inputs, and retrograde conduction occurs through the RN. Multiple breakthroughs of atrial activation occur during A-V nodal reentry due to the dispersed distribution of the RN.