Original article Slow Ca 2+ sparks de-synchronize Ca 2+ release in failing cardiomyocytes: Evidence for altered configuration of Ca 2+ release units? William E. Louch a, b, ⁎, Johan Hake c, d , Halvor K. Mørk a, b , Karina Hougen a, b , Biljana Skrbic a, b , Daniel Ursu e , Theis Tønnessen a, b, f , Ivar Sjaastad a, b , Ole M. Sejersted a, b a Institute for Experimental Medical Research, Oslo University Hospital Ullevål and University of Oslo, Kirkeveien 166, 4th floor Building 7, 0407 Oslo, Norway b KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0407 Oslo, Norway c Department of Bioengineering, University of California, San Diego, 9500 Gilman Drive, MC 0412, La Jolla, CA 92093-0412, USA d Simula Research Laboratory, P.O. Box 134, 1325 Lysaker, Norway e Department of Applied Physiology, University of Ulm, Albert-Einstein-Allee 11, 89081 Ulm, Germany f Department of Cardiothoracic Surgery, Oslo University Hospital Ullevål, Kirkeveien 166, 0407 Oslo, Norway abstract article info Article history: Received 1 October 2012 Received in revised form 14 December 2012 Accepted 17 January 2013 Available online 30 January 2013 Keywords: Heart failure Cardiomyocytes Ca 2+ sparks Ca 2+ transient In heart failure, cardiomyocytes exhibit slowing of the rising phase of the Ca 2+ transient which contributes to the impaired contractility observed in this condition. We investigated whether alterations in ryanodine re- ceptor function promote slowing of Ca 2+ release in a murine model of congestive heart failure (CHF). Myo- cardial infarction was induced by left coronary artery ligation. When chronic CHF had developed (10 weeks post-infarction), cardiomyocytes were isolated from viable regions of the septum. Septal myocytes from SHAM-operated mice served as controls. Ca 2+ transients rose markedly slower in CHF than SHAM myocytes with longer time to peak (CHF=152±12% of SHAM, P b 0.05). The rise time of Ca 2+ sparks was also in- creased in CHF (SHAM=9.6±0.6 ms, CHF=13.2±0.7 ms, P b 0.05), due to a sub-population of sparks (≈20%) with markedly slowed kinetics. Regions of the cell associated with these slow spontaneous sparks also exhibited slowed Ca 2+ release during the action potential. Thus, greater variability in spark kinetics in CHF promoted less uniform Ca 2+ release across the cell. Dyssynchronous Ca 2+ transients in CHF additionally resulted from T-tubule disorganization, as indicated by fast Fourier transforms, but slow sparks were not as- sociated with orphaned ryanodine receptors. Rather, mathematical modeling suggested that slow sparks could result from an altered composition of Ca 2+ release units, including a reduction in ryanodine receptor density and/or distribution of ryanodine receptors into sub-clusters. In conclusion, our findings indicate that slowed, dyssynchronous Ca 2+ transients in CHF result from alterations in Ca 2+ sparks, consistent with rearrangement of ryanodine receptors within Ca 2+ release units. This article is part of a Special Issue entitled “Calcium Signaling in Heart”. © 2013 Elsevier Ltd. All rights reserved. 1. Introduction In ventricular cardiomyocytes, contraction is triggered by a tran- sient increase in intracellular Ca 2+ . This Ca 2+ transient is generated by a process known as Ca 2+ -induced Ca 2+ release, as Ca 2+ influx through L-type Ca 2+ channels triggers Ca 2+ release from ryanodine receptors (RyRs) in the sarcoplasmic reticulum (SR) (for review see [1]). Ca 2+ release occurs at so-called “Ca 2+ release units” (CRUs), where clusters of RyRs are in close apposition to Ca 2+ channels in the T-tubules. Release of Ca 2+ from a single CRU can be visualized as a Ca 2+ spark [2]. During the action potential, elicited Ca 2+ sparks temporally and spatially summate to constitute the Ca 2+ transient [2]. Spontaneous Ca 2+ sparks can also occur in the absence of an L-type Ca 2+ current, and their characteristics can be used to investi- gate the properties of CRU function [3]. We and others have observed that T-tubules are lost and/or disor- ganized during heart failure [4–9]. While Ca 2+ channels are thus effectively removed from CRUs, RyRs remain distributed along the z-lines [7,8,10], meaning that a significant proportion of RyRs are “orphaned” [7]. Ca 2+ release is delayed at these sites, as it is depen- dent on Ca 2+ diffusion following release from intact CRUs [6–8]. The resulting de-synchronization and slowing of the overall Ca 2+ transient promotes slower and weaker contractions in failing cells [11,12]. Importantly, we previously reported that irregular gaps Journal of Molecular and Cellular Cardiology 58 (2013) 41–52 Abbreviations: CRU, Ca 2+ release unit; CHF, congestive heart failure; RyR, ryanodine receptor; FWHM, full width at half maximum; FDHM, full duration at half maximum. ⁎ Corresponding author at: Institute for Experimental Medical Research, Kirkeveien 166, 4.etg. Bygg 7, Oslo University Hospital Ullevål, 0407 Oslo, Norway. Tel.: +47 23 016831; fax: +47 23 016799. E-mail address: w.e.louch@medisin.uio.no (W.E. Louch). 0022-2828/$ – see front matter © 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.yjmcc.2013.01.014 Contents lists available at SciVerse ScienceDirect Journal of Molecular and Cellular Cardiology journal homepage: www.elsevier.com/locate/yjmcc