Abstract 1244: Expression of the Cardiac Sodium Channel Nav1.5 is Required for the Differentiation of Cardiomyocyte Progenitor Cells in vivo
Background: While mutations in the cardiac sodium channel gene SCN5A are well-recognized in congenital arrhythmia syndromes, recent studies have implicated Nav1.5 dysfunction in dilated cardiomyopathy and cardiac fibrosis. Moreover, scn5a −/− mice have severe defects in ventricular development. The link between perturbed ion channel function and abnormalities of cardiac muscle, however, is not well understood. We used mice and zebrafish to study the expression and function of scn5a in the developing heart to identify mechanisms coupling excitability to morphogenesis.
Methods and Results: In mice, we detected scn5a transcripts at E8.0 in total embryonic RNA and observed expression in the atrial and ventricular myocardium at E9.5-E10.5 by in situ hybridization. Molecular cloning of the zebrafish ortholog to SCN5A demonstrated sequence conservation, and heterologous expression of zscn5a in CHO cells yielded typical INa. zscn5a transcripts were detected throughout early development (2 cells-D6) and in the embryonic heart (D2– 6). Knockdown of zscn5a using morpholino antisense oligos resulted in embryos with bradycardia, weak cardiac contractility, and impaired circulation. Morphant embryo hearts also failed to loop and displayed marked abnormalities of ventricular size and morphology. Although zscn5a morphant hearts have normal A-V patterning, high-resolution confocal imaging of mlc2:eGFP transgenic embryos revealed significantly reduced cardiomyocyte number in morphants vs. wild type embryos from 24s-D3 (e.g. D2: 170 vs. 230 cells, p<0.001). To determine the mechanism underlying this reduction, we examined the differentiation of cardiac progenitors and discovered that zscn5a knockdown significantly reduced expression of both early (Nkx2.5, Gata4) and late (VMHC, AMHC, CMLC2) markers of the cardiac lineage, without affecting the formation, viability, or proliferation of the anterior lateral mesoderm.
Conclusions: Voltage-gated sodium channel expression is required for the differentiation of cardiomyocyte progenitors and for normal embryonic heart size, morphology, and function in zebrafish. These observations support the concept that excitability is a critical modulator of gene expression during cardiac organogenesis.