Proceedings of The Physiological Society

University of Oxford (2011) Proc Physiol Soc 23, PC259

Poster Communications

Telethonin regulates transverse (t)-tubule structure and Ca2+ induced Ca2+ release (CICR) in mouse ventricular myocytes.

M. Ibrahim1, S. Miocic-Lazarevic2, U. Siedlecka1, C. Rao1, A. Moshkov3, M. Yacoub1, J. Gorelik3, C. Terracciano1, R. Knoell2

1. Cell Electrophysiology, Imperial College London, London, United Kingdom. 2. Myocardial Genetics, Imperial College London, London, United Kingdom. 3. Functional Microscopy, Imperial College London, London, United Kingdom.

CICR is critical for contraction in cardiomyocytes. The t-tubule system promotes the proximity of the L-type Ca2+ channels (LTCC) and the Ryanodine receptors. T-tubule disruption is implicated early in the pathogenesis of heart failure (1). It is known that telethonin (T-Cap) is necessary for normal t-tubule structure in skeletal muscle, and that it can promote t-tubule formation in response to increases in load (2). We hypothesised that T-Cap was important for normal t-tubule structure and CICR in ventricular cardiomyocytes. We studied Ca2+ handling and t-tubule structure in ventricular cardiomyocytes from hearts lacking T-Cap (KO) at a young (<3 months) and advanced (>8 months) age compared to age-matched wild-type mice (WT). We used non-parametric t-tests. Using confocal microscopy in single cells stained with Fluo-4, we observed an increase in the variance of the time to peak of the Ca2+ transient in both young (WT 297 ± 131 ms (n=40) vs KO 393 ± 156 ms (n=48), p<0.05) and old KO mice (WT 308 ± 156 ms (n=48) vs. KO 430 ± 165 ms (n=43), p<0.05) suggesting a deterioration in the regulation of the Ca2+ transient. In old animals, there was a delayed time to peak of the Ca2+ transient (WT 35 ± 20 ms (n=48) vs KO 95 ± 18 ms (n=43), p<0.01) as well as impaired time to 90% decline (WT 375 ± 73 ms (n=43) vs. KO 460 ± 55 ms (n=20), p<0.05). To probe for local CICR, we recorded Ca2+ sparks and observed an increase in Ca2+ spark frequency in young (WT 0.346 ± 0.35 sp/s (n=32) vs KO 0.867 ± 0.5 sp/s (n=29), p<0.05) and old KO mice (WT 0.370 ±0.3 sp/s (n=15) vs. KO 0.843 ±0.8 sp/s (n=23) p<0.001). There was also a significantly reduced Ca2+ spark peak in older KO animals (WT 1.90 ± 0.3 F/F0 (n=45) vs KO 1.4 ± 0.8 F/F0 (n=43), p<0.05). In addition, we studied LTCC density using whole cell patch clamping and observed a significant reduction in peak LTCC current density in old KO mice (WT -4.75 ± 0.38 pA/pF (n=16) vs KO -3.37 ± 0.30 pA/pF (n=20), p<0.001). The action potential morphology was unchanged. To examine whether changes to the t-tubule structure may be involved in the impaired Ca2+ handling and reduced LTCC observed in cardiomyocytes lacking T-Cap, we used di-8-Anepps and confocal microscopy. We observed a reduced t-tubule density in old KO animals (36.8 ± 5 % (n=45) vs. 30.2 ± 4 % (n=44), p<0.05). We measured t-tubule regularity by comparing the peak of the power-frequency Fourier transform in di-8-Anneps images; we observed a less regular t-tubule structure in both young (WT 2.350 ± 1.09 (n=30) vs KO 1.570 ± 1.06 (n=32) and old (WT 2.10 ± 1.10 (n=39) vs KO 1.30 ± 1.0 (n=20), p<0.05) KO animals. T-tubule disarray may explain the changes to Ca2+ homeostasis observed, but the demonstration of a causal link requires further studies. T-Cap is essential for normal t-tubule structure and CICR in ventricular cardiomyocytes.

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