Diffusion in the Transverse-Axial Tubule System of Cardiac Myocytes

Excitation-contraction (E-C) coupling in cardiac ventricular myocytes is critically dependent on the structure of transverse-axial (t-) tubules, which are invaginations of the surface sarcolemma. Many currents have been shown to be preferentially located at the t-tubular membrane, including the L-Type calcium current, Na-Ca exchange, tetrodotoxin-sensitive Na and steady-state K currents (Orchard, Pasek, and Brette. 2009. Exp. Physiol. 94: 509-519). It has been suggested that membrane folding within the t-system may introduce a slow diffusion zone, which may have implications for ion balance for these currents during the E-C coupling cycle (Hong et al. 2014. Nat. Med. 20: 624-632.). To investigate diffusion in the t-tubule system, we examined t-tubule structure in rabbit and mouse ventricular myocytes as two common experimental models, using electron tomography. Cells were also superfused with solutions containing solutes of varying molecular weight to examine their penetration into the t-system. Fluorescence Recovery After Photobleaching (FRAP) was used to examine transport within the t-tubules of quiescent myocytes. We found that despite marked differences in t-tubule diameter and complexity in the two species, there were relatively small differences in solute penetration below ∼70kDa, and in the time-course of fluorescence recovery during FRAP. These data suggest that gross differences in t-tubule structure do not have marked effects on the rate of diffusion within t-tubules. This work was supported by the BHF and MRC