Axial tubule junctions control rapid calcium signaling in atria
Sören Brandenburg, … , W. Jonathan Lederer, Stephan E. Lehnart
Sören Brandenburg, … , W. Jonathan Lederer, Stephan E. Lehnart
Published September 19, 2016
Citation Information: J Clin Invest. 2016;126(10):3999-4015. https://doi.org/10.1172/JCI88241.
View: Text | PDF
Research Article Cardiology Cell biology

Axial tubule junctions control rapid calcium signaling in atria

Abstract

The canonical atrial myocyte (AM) is characterized by sparse transverse tubule (TT) invaginations and slow intracellular Ca2+ propagation but exhibits rapid contractile activation that is susceptible to loss of function during hypertrophic remodeling. Here, we have identified a membrane structure and Ca2+-signaling complex that may enhance the speed of atrial contraction independently of phospholamban regulation. This axial couplon was observed in human and mouse atria and is composed of voluminous axial tubules (ATs) with extensive junctions to the sarcoplasmic reticulum (SR) that include ryanodine receptor 2 (RyR2) clusters. In mouse AM, AT structures triggered Ca2+ release from the SR approximately 2 times faster at the AM center than at the surface. Rapid Ca2+ release correlated with colocalization of highly phosphorylated RyR2 clusters at AT-SR junctions and earlier, more rapid shortening of central sarcomeres. In contrast, mice expressing phosphorylation-incompetent RyR2 displayed depressed AM sarcomere shortening and reduced in vivo atrial contractile function. Moreover, left atrial hypertrophy led to AT proliferation, with a marked increase in the highly phosphorylated RyR2-pS2808 cluster fraction, thereby maintaining cytosolic Ca2+ signaling despite decreases in RyR2 cluster density and RyR2 protein expression. AT couplon “super-hubs” thus underlie faster excitation-contraction coupling in health as well as hypertrophic compensatory adaptation and represent a structural and metabolic mechanism that may contribute to contractile dysfunction and arrhythmias.

Authors

Sören Brandenburg, Tobias Kohl, George S.B. Williams, Konstantin Gusev, Eva Wagner, Eva A. Rog-Zielinska, Elke Hebisch, Miroslav Dura, Michael Didié, Michael Gotthardt, Viacheslav O. Nikolaev, Gerd Hasenfuss, Peter Kohl, Christopher W. Ward, W. Jonathan Lederer, Stephan E. Lehnart

×

Figure 5

In situ regulation of atrial RyR2 cluster phosphorylation.

Options: View larger image (or click on image) Download as PowerPoint
In situ regulation of atrial RyR2 cluster phosphorylation. (A) Illustrat...
(A) Illustration showing cAMP-dependent responses of Epac1-camps, a ubiquitous cytosolic FRET sensor, during live-cell measurements. In AMs, signals from 4 transversally distributed FRET regions were sampled and signal averaged pairwise for corresponding central (2 + 3, red) versus surface (1 + 4, blue) regions. kOFF, cAMP dissociation constant; kON, cAMP association constant; CFP, cyan fluorescent protein; YFP, yellow fluorescent protein. (B) FRET ratio traces show the same spatiotemporal AM response to 0.1 μM ISO stimulation during increasing intracellular cAMP generation. Bar graph comparing the maximal rate of cAMP rise shows no significant difference between AM center and surface. NS, by Student’s t test. n = 12 AMs. (C) Confocal time series showing progressive RyR2-S2808 cluster phosphorylation following up to 60 seconds of β-adrenergic stimulation (1 μM ISO). RyR2-pS2808 cluster signals were averaged from 10 striations (see magnifications) through equal-sized central (red) and surface (blue) regions indicated by dashed-line yellow boxes. RyR2-pS2808 intensity plot shows progressive cluster phosphorylation, which was significantly faster for central AM regions after a 20-second ISO stimulation. Each time point represents 7 AM experiments. *P < 0.05, **P < 0.01, and ***P < 0.001, by ANOVA. (D and E) Maximal stimulation by combined ISO/RO (1 μM/10 μM) treatment converted lowphos into highphos RyR2 clusters. (D) Histogram shows bimodal pS2808-RyR2 cluster signal pattern under control conditions (black trace), resulting in a profound change of the lowphos RyR2 cluster signals by ISO/RO treatment: 1 major highphos peak, consistent with complete lowphos cluster conversion (red trace). **P < 0.01, by Mann-Whitney U test. Dara are representative of 13 control-treated and 18 ISO/RO-treated AMs. Scale bars: 10 μM, magnification ×4. (E) Images and histogram showing a distinct pS2814/RyR2 cluster phosphorylation shift after ISO/RO stimulation (note: control data are the same as in Figure 3F). *P < 0.05, by Mann-Whitney U test. Data are representative of 19 control-treated and 12 ISO/RO-treated AMs. Scale bars: 10 μM, magnification ×4.