Student Presenter(s): Sanketh Kumar
Faculty Mentor: Kaie Ojamaa
School/College: Arts and Sciences, Old Westbury

The contractile function of the heart is dependent on synchronous calcium release from intracellular storage compartments called sarcoplasmic reticulum (SR) through ion channels (Ryanodine Receptors, RyR). Super-resolution microscopy of single molecules of RyR, voltage-activated calcium channels (LTCC), and membrane tethering protein junctophilin-2 (Jph) was used to test our hypothesis that co-localization of these three proteins is disrupted in heart failure. Heart failure was produced by permanent ligation of a coronary artery (myocardial infarction, MI) in female rats. Cardiac myocytes were isolated from the hearts and labeled with fluorescent-tagged antibodies recognizing RyR, LTCC and Jph proteins. Images of these cardiac myocytes were captured using STORM (stochastic optical reconstruction microscopy) to identify clusters of RyR and LTCC, and co-localization of Jph proteins. The integrity of the T-tubule membranes was measured by staining the cardiomyocytes with a fluorescent dye. Echocardiography measured at 4 months post-MI verified heart failure with reduced ejection fraction. Staining showed disorganization and loss of integrity of the T-tubule network in failing cardiomyocytes. STORM imaging showed spatial re-organization of RyR and LTCC clusters with decreased Jph co-localizations. These adverse changes in ultrastructure in failing hearts support our hypothesis that approximation of these proteins at the T-tubule–SR junction is necessary for normal cardiac function.