Nanoscale Organization, Regulation, and Dynamic Reorganization of Cardiac Calcium Channels

Figures

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  Figure 1. Calcium channel organization/clustering in the Dyad. (A) Illustration depicting the cardiac dyad where CaV1.2 channels are delivered along microtubules to BIN1-hubs on the t-tubule membrane. There they concentrate and form clusters of variable sizes, opposite clusters of RyR2 on the junctional sarcoplasmic reticulum. The dyad is held together by JPH2 which anchors the jSR within 12–15 nm of the t-tubule sarcolemma. JPH2 interacts with both CaV1.2 and RyR2, affecting RyR2 gating, and potentially playing a role in CaV1.2 retention in the dyad. Not all CaV1.2 channels and RyR2 are found in these t-tubule:jSR junctional complexes, some are found in junctions at the sarcolemmal crest and others (not illustrated) are found at junctions along longitudinally oriented axial-tubules. (B) CaV1.2 channel clusters at the t-tubule (left) and crest (right) sarcolemma are seen in super-resolution Ground State Depletion (GSD) localization maps from fixed, immunostained mouse ventricular myocytes. Yellow boxes indicate zoomed-in regions (middle). Reproduced and modified with permission from (Del Villar et al., 2021). (C) RyR2 clusters are observed in 3D-renderings of 3D-STORM images of internal…

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  Figure 2. Phosphorylation-mediated expansion of CaV1.2 and RyR2. (A) Illustration of phosphorylation-dependent expansion of CaV1.2 and RyR2 in the dyad and its physiological consequences. (B) Super-resolution Ground State Depletion (GSD) localization maps from isolated cardiomyocytes immunostained to visualize CaV1.2 channel clusters at the t-tubules in untreated control cells (left) and cells treated with 100 nM ISO for 8 min at room temperature prior to fixation (right). CaV1.2 cluster expansion is evident. (C) Show the results of dynamic TIRF imaging performed on mouse ventricular myocytes transduced with AAV9-CaVβ2-paGFP which acts as a fluorescent CaV1.2 channel biosensor. The images show the GFP-fluorescence emission from channel located on the surface of a myocyte before and after addition of 100 nM ISO. Individual ROIs are indicated (i, ii, and iii) and show time courses of the changes in GFP-emission at each site where channel clusters are seen to undergo stimulated insertion in response to ISO. (D) Histogram summarizing the percentage increase in PM CaVβ2-paGFP stimulated by ISO treatment. Panels B, C, and D are reproduced, with permission from (Del Villar et al., 2021). (E)…

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  Figure 3. CaV1.2 and RyR2 reorganization and cluster fragmentation in Heart Failure. (A) Illustration of the current state of knowledge on CaV1.2 and RyR2 channel reorganization in heart failure. (B) dSTORM images of sham-operated control and post-infarction HF rat cardiomyocytes show RyR2 channel cluster fragmentation/dispersal. Yellow boxes indicate zoomed-in regions which are thresholded below to identify channel clusters. Panel B is reproduced and modified with permission from (Kolstad et al., 2018) under the terms of the Creative Commons Attribution 4.0 International Public License (CC-BY 4.0; https://creativecommons.org/licenses/by/4.0/) associated with that article. Illustrations were created in Biorender.