Functional Neuroplasticity in the Nucleus Tractus Solitarius and Increased Risk of Sudden Death in Mice with Acquired Temporal Lobe Epilepsy

Sudden unexpected death in epilepsy (SUDEP) is the leading cause of death in individuals with refractory acquired epilepsy. Cardiorespiratory failure is the most likely cause in most cases, and central autonomic dysfunction has been implicated as a contributing factor to SUDEP. Neurons of the nucleus tractus solitarius (NTS) in the brainstem vagal complex receive and integrate vagally mediated information regarding cardiorespiratory and other autonomic functions, and GABAergic inhibitory NTS neurons play an essential role in modulating autonomic output. We assessed the activity of GABAergic NTS neurons as a function of epilepsy development in the pilocarpine-induced status epilepticus (SE) model of temporal lobe epilepsy (TLE). Compared with age-matched controls, mice that survived SE had significantly lower survival rates by 150 d post-SE. GABAergic NTS neurons from mice that survived SE displayed a glutamate-dependent increase in spontaneous action potential firing rate by 12 wks post-SE. Increased spontaneous EPSC frequency was also detected, but vagal afferent synaptic release properties were unaltered, suggesting that an increase in glutamate release from central neurons developed in the NTS after SE. Our results indicate that long-term changes in glutamate release and activity of GABAergic neurons emerge in the NTS in association with epileptogenesis. These changes might contribute to increased risk of cardiorespiratory dysfunction and sudden death in this model of TLE

Visualizing Mutation-Specific Differences in the Trafficking-Deficient Phenotype of Kv11.1 Proteins Linked to Long QT Syndrome Type 2

KCNH2 encodes the Kv11.1 α-subunit that underlies the rapidly activating delayed-rectifier K+ current in the heart. Loss-of-function KCNH2 mutations cause long QT syndrome type 2 (LQT2), and most LQT2-linked missense mutations inhibit the trafficking of Kv11.1 channel protein to the cell surface membrane. Several trafficking-deficient LQT2 mutations (e.g., G601S) generate Kv11.1 proteins that are sequestered in a microtubule-dependent quality control (QC) compartment in the transitional endoplasmic reticulum (ER). We tested the hypothesis that the QC mechanisms that regulate LQT2-linked Kv11.1 protein trafficking are mutation-specific. Confocal imaging analyses of HEK293 cells stably expressing the trafficking-deficient LQT2 mutation F805C showed that, unlike G601S-Kv11.1 protein, F805C-Kv11.1 protein was concentrated in several transitional ER subcompartments. The microtubule depolymerizing drug nocodazole differentially affected G601S- and F805C-Kv11.1 protein immunostaining. Nocodazole caused G601S-Kv11.1 protein to distribute into peripheral reticular structures, and it increased the diffuse immunostaining of F805C-Kv11.1 protein around the transitional ER subcompartments. Proteasome inhibition also affected the immunostaining of G601S- and F805C-Kv11.1 protein differently. Incubating cells in MG132 minimally impacted G601S-Kv11.1 immunostaining, but it dramatically increased the diffuse immunostaining of F805C-Kv11.1 protein in the transitional ER. Similar results were seen after incubating cells in the proteasome inhibitor lactacystin. Differences in the cellular distribution of G601S-Kv11.1 and F805C-Kv11.1 protein persisted in transfected human inducible pluripotent stem cell derived cardiomyocytes. These are the first data to visually demonstrate mutation-specific differences in the trafficking-deficient LQT2 phenotype, and this study has identified a novel way to categorize trafficking-deficient LQT2 mutations based on differences in intracellular retention

A Molecular Mechanism for Adrenergic-Induced Long QT Syndrome

ObjectivesThis study sought to explore molecular mechanisms underlying the adrenergic-induced QT prolongation associated with KCNQ1 mutations.BackgroundThe most frequent type of congenital long QT syndrome is LQT1, which is caused by mutations in the gene (KCNQ1) that encodes the alpha subunit of the slow component of delayed rectifier K+ current (IKs) channel. We identified 11 patients from 4 unrelated families that are heterozygous for KCNQ1-G269S. Most patients remained asymptomatic, and their resting corrected QT intervals ranged from normal to borderline but were prolonged significantly during exercise.MethodsWild-type (WT) KCNQ1 and/or KCNQ1-G269S (G269S) were expressed in mammalian cells with KCNE1. IKs-like currents were measured in control conditions or after isoproterenol or protein kinase A (PKA) stimulation using the patch-clamp technique. Additionally, experiments that incorporated the phosphomimetic KCNQ1 substitution, S27D, in WT or KCNQ1-G269S were also performed.ResultsThe coexpression of WT-KCNQ1 with varying amounts of G269S decreased IKs, shifted the current-voltage I-V relation of IKs to more positive potentials, and accelerated the IKs deactivation rates in a concentration-dependent manner. In addition, the coexpression of G269S and WT blunted the activation of IKs in response to isoproterenol or PKA stimulation. Lastly, a phosphomimetic substitution in G269S did not show an increased IKs.ConclusionsG269S modestly affected IKs in control conditions, but it almost completely blunted IKs responsiveness in conditions that simulate or mimic PKA phosphorylation of KCNQ1. This insensitivity to PKA stimulation may explain why patients with G269S mutation showed an excessive prolongation of QT intervals on exercise

Long QT Syndrome Type 2: Emerging Strategies for Correcting Class 2 \u3cem\u3eKCNH2 (hERG)\u3c/em\u3e Mutations and Identifying New Patients

Significant advances in our understanding of the molecular mechanisms that cause congenital long QT syndrome (LQTS) have been made. A wide variety of experimental approaches, including heterologous expression of mutant ion channel proteins and the use of inducible pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) from LQTS patients offer insights into etiology and new therapeutic strategies. This review briefly discusses the major molecular mechanisms underlying LQTS type 2 (LQT2), which is caused by loss-of-function (LOF) mutations in the KCNH2 gene (also known as the human ether-à-go-go-related gene or hERG). Almost half of suspected LQT2-causing mutations are missense mutations, and functional studies suggest that about 90% of these mutations disrupt the intracellular transport, or trafficking, of the KCNH2-encoded Kv11.1 channel protein to the cell surface membrane. In this review, we discuss emerging strategies that improve the trafficking and functional expression of trafficking-deficient LQT2 Kv11.1 channel proteins to the cell surface membrane and how new insights into the structure of the Kv11.1 channel protein will lead to computational approaches that identify which KCNH2 missense variants confer a high-risk for LQT2

Cardiomyocyte Deletion of \u3ci\u3eBmal1\u3c/i\u3e Exacerbates QT- and RR-Interval Prolongation in \u3ci\u3eScn5a\u3c/i\u3e\u3csup\u3e+/ΔKPQ\u3c/sup\u3e Mice

Circadian rhythms are generated by cell autonomous circadian clocks that perform a ubiquitous cellular time-keeping function and cell type-specific functions important for normal physiology. Studies show inducing the deletion of the core circadian clock transcription factor Bmal1 in adult mouse cardiomyocytes disrupts cardiac circadian clock function, cardiac ion channel expression, slows heart rate, and prolongs the QT-interval at slow heart rates. This study determined how inducing the deletion of Bmal1 in adult cardiomyocytes impacted the in vivo electrophysiological phenotype of a knock-in mouse model for the arrhythmogenic long QT syndrome (Scn5a+/ΔKPQ). Electrocardiographic telemetry showed inducing the deletion of Bmal1 in the cardiomyocytes of mice with or without the ΔKPQ-Scn5a mutation increased the QT-interval at RR-intervals that were ≥130 ms. Inducing the deletion of Bmal1 in the cardiomyocytes of mice with or without the ΔKPQ-Scn5a mutation also increased the day/night rhythm-adjusted mean in the RR-interval, but it did not change the period, phase or amplitude. Compared to mice without the ΔKPQ-Scn5a mutation, mice with the ΔKPQ-Scn5a mutation had reduced heart rate variability (HRV) during the peak of the day/night rhythm in the RR-interval. Inducing the deletion of Bmal1 in cardiomyocytes did not affect HRV in mice without the ΔKPQ-Scn5a mutation, but it did increase HRV in mice with the ΔKPQ-Scn5a mutation. The data demonstrate that deleting Bmal1 in cardiomyocytes exacerbates QT- and RR-interval prolongation in mice with the ΔKPQ-Scn5a mutation