Imaging atrial arrhythmic intracellular calcium in intact heart

Abstract

Abnormalities in intracellular Ca(2+) signaling have been proposed to play an essential role in the pathophysiology of atrial arrhythmias. However, a direct observation of intracellular Ca(2+) in atrial myocytes during atrial arrhythmias is lacking. Here, we have developed an ex vivo model of simultaneous Ca(2+) imaging and electrocardiographic recording in cardiac atria. Using this system we were able to record atrial arrhythmic intracellular Ca(2+) activities. Our results indicate that atrial arrhythmias can be tightly linked to intracellular Ca(2+) waves and Ca(2+) alternans. Moreover, we applied this strategy to analyze Ca(2+) signals in the hearts of WT and knock-in mice harboring a 'leaky' type 2 ryanodine receptor (RyR2-R2474S). We showed that sarcoplasmic reticulum (SR) Ca(2+) leak increases the susceptibility to Ca(2+) alternans and Ca(2+) waves increasing the incidence of atrial arrhythmias. Reduction of SR Ca(2+) leak via RyR2 by acute treatment with S107 reduced both Ca(2+) alternans and Ca(2+) waves, and prevented atrial arrhythmias.

Keywords: Atrial arrhythmias; Ex vivo model; Intracellular Ca(2+) imaging.

Figure 1. Ex vivo model for simultaneous bipolar ECG recording and intracellular Ca²⁺ imaging in atrium

A. A close-up of the model system. A Langendorff-perfused heart was placed on the imaging chamber with the right atrium downwards on the glass bottom. 1, a pair of silver electrodes for ECG recording of ventricle; 2, a pair of platinum electrodes for pacing the left atrium; 3, perfusion tube; 4, a pair of 25-μm-diameter platinum electrodes for bipolar ECG recording of right atrium; 5, objective of the confocal microscope. B. representative X-Y scan and X-T scan of the atrium. C, D. Bipolar ECG recording and intracellular Ca²⁺ imaging of a right atrium of RyR2-R2474S^+/− mouse during burst atrial pacing induced atrial arrhythmic event. The upper, middle and lower traces show simultaneous atrial intracellular Ca²⁺ imaging, right atrial and ventricular bipolar ECG recording, respectively. The arrhythmic event has been split into initiation (C) and disappearance (D) part with some overlap labeled by the green lines above the linescan images. The red dotted line indicates the application of the 2-second burst pacing. E. Quantitative analysis of the relationship between arrhythmic atrial beating rate and the number of atrial myocytes exhibiting intracellular Ca²⁺ waves during atrial arrhythmias events. The arrhythmic events were divided into 2 groups according to the atrial beating rate ≤ or >1500 bpm.

Figure 2. Ca²⁺ alternans and Ca²⁺ waves in right atrial cardiomyocytes

A–D, representative images of 10 Hz pacing-induced Ca²⁺ alternans in right atria of WT (A), RyR2-R2474S^+/− (B) and Vehicle (C) or S107 treated (D) RyR2-R2474S^+/− mice. The right panel displays the Ca²⁺ transients in atrial myocytes. E, Quantification (right: ratio, left: percent) of Ca²⁺ alternans in the indicated groups. F, Ca²⁺ waves in atrial myocytes in the indicated groups. Hearts were perfused with 1 μmol/L isoproterenol for 30 minutes to induce Ca²⁺ waves. The number of hearts examined in each group is labeled below each column. *: p< 0.05, **p< 0.01.