Nanoscale Imaging of Dyadic Ion Channels in Hearts of Thyroidectomized Rats

Thyroid hormone (TH) deficiency results in adverse changes in cardiomyocyte shape causing increased wall stress and chamber dilatation, resulting in contractile dysfunction if left untreated. This study aimed to use single molecule imaging microscopy to quantify ion channel localization at the dyad structures in cardiomyocytes in response to thyroid status. Methods: Female Sprague-Dawley rats were surgically thyroidectomized (TX); controls were euthyroid (EU) rats (6 rats/group). After 4-5 months, echocardiographic and hemodynamic measurements were recorded, and left ventricle cardiomyocytes (CM) were enzymatically isolated for contractility and Ca 2+ recordings. T-tubule (TT) organization was measured by confocal imaging of ANEPPS-stained CM. Single molecule localization of ryanodine receptors (RyR2), L-type Ca2+ channels (LTCC) and Junctophilin-2 (Jph2) were obtained by 3D stochastic optical reconstruction microscopy (STORM). Results: Hemodynamic and echocardiographic results showed significant systolic and diastolic dysfunction in TX rats. Field stimulation (1 Hz, 4V) of isolated TX CM showed slower maximum contraction velocity (+Vmax), 2.047±0.30 vs 3.503±0.50 um/sec (p<0.001) and slower max relaxation velocity (-Vmax), 1.369±0.27 vs 2.44±0.73 um/sec (p<0.01). Ca 2+ transients in TX CM had longer time to peak Ca 2+ (0.076±0.01 vs 0.0516±0.003 sec, p<0.0001) and slower Ca 2+ reuptake velocity (0.184±0.005 vs 0.261±0.03 um/sec, p<0.0001). TT-integrity analysis indicated severe disorganization in TX CM (1.77±0.23 vs 5.02±0.53, p<0.001). STORM images showed fewer RyR2 clusters per cell area with reduced Jph2 associated with the clusters, and with more clusters completely devoid of Jph2 and LTCC. In summary, hearts of animals with TH-deficiency showed disrupted TT structures, slower Ca 2+ kinetics and severe contractile dysfunction. Reduced clusters of RyR2 and decreased co-localization of Jph2 and Ca 2+ channels at the dyadic structures likely result in altered contractility with thyroid status. This study was funded by NIH R15HL154068 and supported by the NYIT Imaging Center. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.