Single-Molecule Localization of the Cardiac Voltage-Gated Sodium Channel Reveals Different Modes of Reorganization at Cardiomyocyte Membrane Domains

Abstract

Background: Mutations in the gene encoding the cardiac voltage-gated sodium channel Nav1.5 cause various cardiac arrhythmias. This variety may arise from different determinants of Nav1.5 expression between cardiomyocyte domains. At the lateral membrane and T-Tubules, Nav1.5 localization and function remain insufficiently characterized. Methods: We used novel single-molecule localization microscopy and computational modeling to define nanoscale features of Nav1.5 localization and distribution at the lateral membrane, the lateral membrane groove, and T-Tubules in cardiomyocytes from wild-Type (N=3), dystrophin-deficient (mdx; N=3) mice, and mice expressing C-Terminally truncated Nav1.5 (ΔSIV; N=3). We moreover assessed T-Tubules sodium current by recording whole-cell sodium currents in control (N=5) and detubulated (N=5) wild-Type cardiomyocytes. Results: We show that Nav1.5 organizes as distinct clusters in the groove and T-Tubules which density, distribution, and organization partially depend on SIV and dystrophin. We found that overall reduction in Nav1.5 expression in mdx and ΔSIV cells results in a nonuniform redistribution with Nav1.5 being specifically reduced at the groove of ΔSIV and increased in T-Tubules of mdx cardiomyocytes. A T-Tubules sodium current could, however, not be demonstrated. Conclusions: Nav1.5 mutations may site-specifically affect Nav1.5 localization and distribution at the lateral membrane and T-Tubules, depending on site-specific interacting proteins. Future research efforts should elucidate the functional consequences of this redistribution.