Ventricular Myocyte Shortening Assay

Unloaded sarcomere shortening was measured in freshly isolated ventricular myocytes, as described previously [1 (link),2 (link)]. Briefly, isolated myocytes were transferred into a recording chamber mounted on an Olympus X51 inverted microscope and superfused with normal Tyrode solution saturated with room air. Additions to the Tyrode solution are described in the text. The mitochondrial calcium uptake inhibitor, Ru-360, was obtained from EMD Biosciences; all other chemicals were purchased from Sigma. Typically, cells were field stimulated to contract at 1 Hz. When thapsigargin was applied to cells, the stimulation frequency was reduced to 0.5 Hz. Video images were acquired using a Myocam camera and IonWizard software (IonOptix, Inc.). All experiments were performed at room temperature.