Structural and functional remodelling of the atrioventricular node with ageing in rats: the role of Hyperpolarization-activated cyclic-nucleotide gated.

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Structural and functional remodelling of the atrioventricular node with ageing in rats: the role of Hyperpolarization-activated cyclic-nucleotide gated and Ryanodine 2 channels Yawer Saeed, PhD, MRCP, Ian Temple, PhD, MRCP, Zoltan Borbas, PhD, MRCP, Andrew Atkinson, BSc, Joseph Yanni, PhD, Michal Maczewski, MD, PhD, Urszula Mackiewicz, PhD, Mariam Aly, Sunil J.R.J. Logantha, PhD, Clifford Garratt, FRCP, Halina Dobrzynski, PhD Heart Rhythm DOI: 10.1016/j.hrthm.2017.12.027 Copyright © 2017 Terms and Conditions

Figure 1 Changes in the electrophysiological measurements in young (n=14) and old heart’s (n=6) AVN preparation with and without administration of the drugs. The data shows the changes with Cesium and Ryanodine. The AVNERP measurement was not possible in old hearts with ryanodine because of Wenkebach phenomenon with the S1-drive train. Heart Rhythm DOI: (10.1016/j.hrthm.2017.12.027) Copyright © 2017 Terms and Conditions

Figure 2 Masson’s trichome. Young and old proximal penetrating bundle (PPB). Top panel (A, B, C) show sections at PPB level at different magnifications in a young and bottom panel (E, F, G) shows the comparable images in an old heart. PPB is marked with a black dotted line. Myocytes are pink/purple and nuclei are black blue in high magnification images (C, G). Left atrium (LA), right atrium (RA), left ventricle (LV), right ventricle (RV). Bar is shown. Red colour seen in A is clotted blood. C and G showed disruption in cellular architecture seen with ageing. D: Number of nuclei measured in high magnification image in the regions of AVJ. H: Collagen signal estimation in young and old heart’s AVJ measured via picrosirius red and polarized microscopy. Compact node (CN). proximal penetrating bundle (PPB), distal penetrating/His bundle (DPB/His). Young hearts (n=6), Old hearts (n=6) Heart Rhythm DOI: (10.1016/j.hrthm.2017.12.027) Copyright © 2017 Terms and Conditions

Figure 3 Confocal microscope high magnification images of the AVJ with Cx40 (green) and Cav3 (red) immunolabelling. Top panel shows the AVJ components in a young and bottom panel in an old heart. Cx40 expression is also shown separately in B, D, F, H to clearly illustrate Cx40 expression and also its location within the myocardial cells. Bar=20μm. Young hearts (n=6), Old hearts (n=6) Heart Rhythm DOI: (10.1016/j.hrthm.2017.12.027) Copyright © 2017 Terms and Conditions

Figure 4 Changes in the expression of Cx40 (A), Cx43 (B), Nav1.5 (C), HCN4 (D) and cellular diameter (E) of the myocardial cells in the AVJ. Cellular diameter is measured in high magnification image in at least 20 myocardial cells in each heart. Mean±SEM are shown. * p <0.05. Young hearts (n=6), Old hearts (n=6) Heart Rhythm DOI: (10.1016/j.hrthm.2017.12.027) Copyright © 2017 Terms and Conditions

Figure 5 Top panel shows the AVJ regions in a young and bottom panel in an old heart. (A, C, E, G) Confocal microscope low magnification images of the AVJ with HCN4 (green) and Cx43 (red) immunolabelling. B, D, F, H showed corresponding high magnification images in the PPB and His. Bar is shown in each image. Young hearts (n=6), Old hearts (n=6) Heart Rhythm DOI: (10.1016/j.hrthm.2017.12.027) Copyright © 2017 Terms and Conditions

Figure 6 Confocal microscope high magnification images of the AVJ with RyR2 (red) immunolabelling, Top panel shows AVJ regions in a young and middle panel in an old heart. Bar=20μm. Bottom panel shows changes in RyR2 (G), SERCA 2a (H) and Cav1.3 (I) expression with ageing. Mean±SEM are shown. * p<0.05. Young hearts (n=6), Old hearts (n=6) Heart Rhythm DOI: (10.1016/j.hrthm.2017.12.027) Copyright © 2017 Terms and Conditions

Figure 7 Effect of reduced RyR2 expression with ageing on membrane depolarization. A: The young AVN myocyte. B: The old AVN myocyte. The reduced RyR2 expression with ageing results in decreased calcium sparks inhibiting the forward mode of sodium-calcium exchanger reducing inward INa-Ca. This will delay the diastolic membrane potential increase (towards positive membrane potential) thus inhibiting the activation of L-type calcium channel. Heart Rhythm DOI: (10.1016/j.hrthm.2017.12.027) Copyright © 2017 Terms and Conditions