Shortly after cardiac Na channels activate and initiate the action potential, inactivation ensues within milliseconds, attenuating the peak Na current, I and allowing the cell membrane to repolarize. A very limited number of Na channels that do not inactivate carry a persistent I, or late I. While late I is only a small fraction of peak magnitude, it significantly prolongs ventricular action potential duration, which predisposes patients to arrhythmia. Here, we review our current understanding of inactivation mechanisms, their regulation, and how they have been modeled computationally. Based on this body of work, we conclude that inactivation and its connection to late I would be best modeled with a "feet-on-the-door" approach where multiple channel components participate in determining inactivation and late I. This model reflects experimental findings showing that perturbation of many channel locations can destabilize inactivation and cause pathological late I.
Several inherited arrhythmias, including Brugada syndrome and arrhythmogenic cardiomyopathy, primarily affect the right ventricle and can lead to sudden cardiac death. Among many differences, right and left ventricular cardiomyocytes derive from distinct progenitors, prompting us to investigate how embryonic programming may contribute to chamber-specific conduction and arrhythmia susceptibility. Here, we show that developmental perturbation of Wnt signaling leads to chamber-specific transcriptional regulation of genes important in cardiac conduction that persists into adulthood. Transcriptional profiling of right versus left ventricles in mice deficient in Wnt transcriptional activity reveals global chamber differences, including genes regulating cardiac electrophysiology such as Gja1 and Scn5a. In addition, the transcriptional repressor Hey2, a gene associated with Brugada syndrome, is a direct target of Wnt signaling in the right ventricle only. These transcriptional changes lead to perturbed right ventricular cardiac conduction and cellular excitability. Ex vivo and in vivo stimulation of the right ventricle is sufficient to induce ventricular tachycardia in Wnt transcriptionally inactive hearts, while left ventricular stimulation has no effect. These data show that embryonic perturbation of Wnt signaling in cardiomyocytes leads to right ventricular arrhythmia susceptibility in the adult heart through chamber-specific regulation of genes regulating cellular electrophysiology.
Histology and immunohistochemistry. Immunohistochemistry was performed on paraffin-embedded sections. Gross heart morphology and collagen content were examined using Masson's trichrome stain (American MasterTech Scientific). Wheat germ agglutinin staining was used to visualize cell membranes and enable quantification of CM cell size. Microelectrode recordings. Investigators were blinded to the sample group allocation during the experiment and analysis of experimental outcome. Mouse hearts were Langendorff perfused and were recorded while in sinus rhythm and when stimulated at 10 Hz (approximately 600 beats per minute). Using glass sharp microelectrodes, single LA CMs were sampled near the epicardial surface. To decrease noise from motion artifacts, blebbistatin (0.2 mg/mL) was used to arrest motion and allow for stable microelectrode recording without requiring the use of floating electrodes. Additional information. All methods related to mouse RT-qPCR (Supplemental Table 14) and RNA-sequencing and analysis are detailed in the Supplemental Methods. Expanded methods for human tissue acquisition, CMN isolation, histology and immunohistochemistry, and microelectrode recordings are also supplied in the Supplemental Methods. RA RNA-sequencing accession number. RA RNA-sequencing data discussed in this manuscript have been deposited in the National Center for Biotechnology Information's (NCBI) Gene Expression Omnibus (GEO) database and are accessible through GSE100244. LA RNA-sequencing accession number. LA RNA-sequencing data discussed in this manuscript have been deposited in NCBI's GEO and are accessible through GSE138253. Human RNA-sequencing accession number. Data have been deposited in NCBI's GEO and are accessible through GSE138252. Statistics. All data are expressed as mean ± SEM. Statistical analyses were performed after assessing for normal distribution using either paired or unpaired Student's 2-tailed t tests for comparison of 2 groups with a Welch's correction. Values of P < 0.05 were considered statistically significant. Study approval. Animal protocols were approved by the Animal Studies Committee at Washington University in St. Louis, and animals were handled in accordance with the NIH's Guide for the Care and Use of Laboratory Animals (National Academies Press, 2011). Protocols involving human tissue acquisition were approved by the Washington University in St. Louis IRB. Informed consent was obtained for all tissue before inclusion in this study. Methods described in this manuscript were performed in accordance with all human research guidelines. Author contributions SLR was responsible for conceptualization of the study. SLR, CEL, JJ, and QG contributed to experimental design. CEL, JJ, QG, TY, and SB conducted experiments, acquired data, and performed data analysis. SCH conducted histology staining. GL contributed to data analysis. CEL, JJ, QG, GL, TY, SCH, DMZ, UG, KT, and BDB contributed to human tissue acquisition. RDN and CPC performed mouse RNA-sequencing and statistical analysis. SL and BZ perfo...
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