Tara L. Klassen

Arrhythmia in Heart and Brain: KCNQ1 Mutations Link Epilepsy and Sudden Unexplained Death

Mice engineered to carry a human mutation that causes heart problems also have epilepsy, suggesting a cause of SUDEP, sudden unexplained death in epilepsy. Patients with epilepsy face an extra frightening burden. Occasionally, otherwise healthy individuals with this disease die unexpectedly for no apparent cause. The incidence of sudden death is ~10% for epilepsy patients—a risk far greater than that faced by a non-epileptic person. Sudden unexplained death in epilepsy (SUDEP) frequently follows a seizure, and patients who experience many seizures have a greater risk of SUDEP. But the causes of SUDEP remain a mystery. Now, Goldman et al. shed new light on how defective potassium channels contribute to this syndrome. Various causes of SUDEP have been proposed—some that produce irreversible cardiac dysfunction and some that produce respiratory distress. One suggested cause invokes the common dependence of the heart and brain on electrical activity for proper functioning. When ion channels—the membrane proteins that control electrical activity—go awry (by gene mutation or a drug), the brain becomes uncontrollably excited, producing a seizure, during which the regular beating of the heart is disrupted and can cease altogether. Both people and a mouse model display mutations in the KCNQ1 gene—which encodes a potassium channel in the heart—that give rise to heartbeat abnormalities and a higher risk for sometimes fatal arrhythmias. Goldman et al. studied KCNQ1-mutant mice and found that this same potassium channel that causes problems in the heart is also present in neurons in the brain and is in particularly high abundance in regions that are susceptible to epilepsy. A closer look at the brains of these mice disclosed that their electrical discharges display abnormalities characteristic of epilepsy and that these aberrations often occur at the same time as abnormal heartbeats. Continuous video surveillance of these mice revealed that many also experienced overt seizures. In one instance, a mouse with the mutant ion channel suffered increasingly frequent seizures accompanied by irregular abnormal cardiac activity and ultimately went into cardiac arrest, a mouse version of SUDEP. Taken together, these results reinforce hints in the literature that SUDEP may result from common excitability defects in the brain and heart. Cardiac abnormalities that resemble those in the mutant mice can be caused in humans by mutations in ~10 genes. The ability to screen epilepsy patients for these mutations would allow those who are at risk for cardiac-induced sudden death to take preventive measures. Sudden unexplained death is a catastrophic complication of human idiopathic epilepsy, causing up to 18% of patient deaths. A molecular mechanism and an identified therapy have remained elusive. Here, we find that epilepsy occurs in mouse lines bearing dominant human LQT1 mutations for the most common form of cardiac long QT syndrome, which causes syncopy and sudden death. KCNQ1 encodes the cardiac KvLQT1 delayed rectifier channel, which has not been previously found in the brain. We have shown that, in these mice, this channel is found in forebrain neuronal networks and brainstem nuclei, regions in which a defect in the ability of neurons to repolarize after an action potential, as would be caused by this mutation, can produce seizures and dysregulate autonomic control of the heart. That long QT syndrome mutations in KCNQ1 cause epilepsy reveals the dual arrhythmogenic potential of an ion channelopathy coexpressed in heart and brain and motivates a search for genetic diagnostic strategies to improve risk prediction and prevention of early mortality in persons with seizure disorders of unknown origin.

Effects of Task-Specific Locomotor and Strength Training in Adults Who Were Ambulatory After Stroke: Results of the STEPS Randomized Clinical Trial

Background and Purpose: A phase II, single-blinded, randomized clinical trial was conducted to determine the effects of combined task-specific and lower-extremity (LE) strength training to improve walking ability after stroke. Subjects: The participants were 80 adults who were ambulatory 4 months to 5 years after a unilateral stroke. Method: The exercise interventions consisted of body-weight–supported treadmill training (BWSTT), limb-loaded resistive leg cycling (CYCLE), LE muscle-specific progressive-resistive exercise (LE-EX), and upper-extremity ergometry (UE-EX). After baseline assessments, participants were randomly assigned to a combined exercise program that included an exercise pair. The exercise pairs were: BWSTT/UE-EX, CYCLE/UE-EX, BWSTT/CYCLE, and BWSTT/LE-EX. Exercise sessions were 4 times per week for 6 weeks (total of 24 sessions), with exercise type completed on alternate days. Outcomes were self-selected walking speed, fast walking speed, and 6-minute walk distance measured before and after intervention and at a 6-month follow-up. Results: The BWSTT/UE-EX group had significantly greater walking speed increases compared with the CYCLE/UE-EX group; both groups improved in distance walked. All BWSTT groups increased walking speed and distance whether BWSTT was combined with LE strength training or not. Discussion and Conclusion: After chronic stroke, task-specific training during treadmill walking with body-weight support is more effective in improving walking speed and maintaining these gains at 6 months than resisted leg cycling alone. Consistent with the overtraining literature, LE strength training alternated daily with BWSTT walking did not provide an added benefit to walking outcomes.

Kv1.1 Potassium Channel Deficiency Reveals Brain-Driven Cardiac Dysfunction as a Candidate Mechanism for Sudden Unexplained Death in Epilepsy

Mice lacking Kv1.1 Shaker-like potassium channels encoded by the Kcna1 gene exhibit severe seizures and die prematurely. The channel is widely expressed in brain but only minimally, if at all, in mouse myocardium. To test whether Kv1.1-potassium deficiency could underlie primary neurogenic cardiac dysfunction, we performed simultaneous video EEG–ECG recordings and found that Kcna1-null mice display potentially malignant interictal cardiac abnormalities, including a fivefold increase in atrioventricular (AV) conduction blocks, as well as bradycardia and premature ventricular contractions. During seizures the occurrence of AV conduction blocks increased, predisposing Kv1.1-deficient mice to sudden unexplained death in epilepsy (SUDEP), which we recorded fortuitously in one animal. To determine whether the interictal AV conduction blocks were of cardiac or neural origin, we examined their response to selective pharmacological blockade of the autonomic nervous system. Simultaneous administration of atropine and propranolol to block parasympathetic and sympathetic branches, respectively, eliminated conduction blocks. When administered separately, only atropine ameliorated AV conduction blocks, indicating that excessive parasympathetic tone contributes to the neurocardiac defect. We found no changes in Kv1.1-deficient cardiac structure, but extensive Kv1.1 expression in juxtaparanodes of the wild-type vagus nerve, the primary source of parasympathetic input to the heart, suggesting a novel site of action leading to Kv1.1-associated cardiac bradyarrhythmias. Together, our data suggest that Kv1.1 deficiency leads to impaired neural control of cardiac rhythmicity due in part to aberrant parasympathetic neurotransmission, making Kcna1 a strong candidate gene for human SUDEP.

Gait parameters associated with responsiveness to treadmill training with body-weight support after stroke: an exploratory study.

Background Task-specific training programs after stroke improve walking function, but it is not clear which biomechanical parameters of gait are most associated with improved walking speed. Objective The purpose of this study was to identify gait parameters associated with improved walking speed after a locomotor training program that included body-weight–supported treadmill training (BWSTT). Design A prospective, between-subjects design was used. Methods Fifteen people, ranging from approximately 9 months to 5 years after stroke, completed 1 of 3 different 6-week training regimens. These regimens consisted of 12 sessions of BWSTT alternated with 12 sessions of: lower-extremity resistive cycling; lower-extremity progressive, resistive strengthening; or a sham condition of arm ergometry. Gait analysis was conducted before and after the 6-week intervention program. Kinematics, kinetics, and electromyographic (EMG) activity were recorded from the hemiparetic lower extremity while participants walked at a self-selected pace. Changes in gait parameters were compared in participants who showed an increase in self-selected walking speed of greater than 0.08 m/s (high-response group) and in those with less improvement (low-response group). Results Compared with participants in the low-response group, those in the high-response group displayed greater increases in terminal stance hip extension angle and hip flexion power (product of net joint moment and angular velocity) after the intervention. The intensity of soleus muscle EMG activity during walking also was significantly higher in participants in the high-response group after the intervention. Limitations Only sagittal-plane parameters were assessed, and the sample size was small. Conclusions Task-specific locomotor training alternated with strength training resulted in kinematic, kinetic, and muscle activation adaptations that were strongly associated with improved walking speed. Changes in both hip and ankle biomechanics during late stance were associated with greater increases in gait speed.

High‐resolution molecular genomic autopsy reveals complex sudden unexpected death in epilepsy risk profile

Advanced variant detection in genes underlying risk of sudden unexpected death in epilepsy (SUDEP) can uncover extensive epistatic complexity and improve diagnostic accuracy of epilepsy‐related mortality. However, the sensitivity and clinical utility of diagnostic panels based solely on established cardiac arrhythmia genes in the molecular autopsy of SUDEP is unknown. We applied the established clinical diagnostic panels, followed by sequencing and a high density copy number variant (CNV) detection array of an additional 253 related ion channel subunit genes to analyze the overall genomic variation in a SUDEP of the 3‐year‐old proband with severe myoclonic epilepsy of infancy (SMEI). We uncovered complex combinations of single nucleotide polymorphisms and CNVs in genes expressed in both neurocardiac and respiratory control pathways, including SCN1A, KCNA1, RYR3, and HTR2C. Our findings demonstrate the importance of comprehensive high‐resolution variant analysis in the assessment of personally relevant SUDEP risk. In this case, the combination of de novo single nucleotide polymorphisms (SNPs) and CNVs in the SCN1A and KCNA1 genes, respectively, is suspected to be the principal risk factor for both epilepsy and premature death. However, consideration of the overall biologically relevant variant complexity with its extensive functional epistatic interactions reveals potential personal risk more accurately.

Defective fast inactivation recovery of Nav1.4 in congenital myasthenic syndrome

To describe the unique phenotype and genetic findings in a 57‐year‐old female with a rare form of congenital myasthenic syndrome (CMS) associated with longstanding muscle fatigability, and to investigate the underlying pathophysiology.

Novel brain expression of ClC-1 chloride channels and enrichment of CLCN1 variants in epilepsy

Objective: To explore the potential contribution of genetic variation in voltage-gated chloride channels to epilepsy, we analyzed CLCN family (CLCN1-7) gene variant profiles in individuals with complex idiopathic epilepsy syndromes and determined the expression of these channels in human and murine brain. Methods: We used parallel exomic sequencing of 237 ion channel subunit genes to screen individuals with a clinical diagnosis of idiopathic epilepsy and evaluate the distribution of missense variants in CLCN genes in cases and controls. We examined regional expression of CLCN1 in human and mouse brain using reverse transcriptase PCR, in situ hybridization, and Western immunoblotting. Results: We found that in 152 individuals with sporadic epilepsy of unknown origin, 96.7% had at least one missense variant in the CLCN genes compared with 28.2% of 139 controls. Nonsynonymous single nucleotide polymorphisms in the “skeletal” chloride channel gene CLCN1 and in CLCN2, a putative human epilepsy gene, were detected in threefold excess in cases relative to controls. Among these, we report a novel de novo CLCN1 truncation mutation in a patient with pharmacoresistant generalized seizures and a dystonic writers cramp without evidence of variants in other channel genes linked to epilepsy. Molecular localization revealed the unexpectedly widespread presence of CLCN1 mRNA transcripts and the ClC-1 subunit protein in human and murine brain, previously believed absent in neurons. Conclusions: Our findings support a possible comorbid contribution of the “skeletal” chloride channel ClC-1 to the regulation of brain excitability and the need for further elucidation of the roles of CLCN genes in neuronal network excitability disorders.

Combined task-specific training and strengthening effects on locomotor recovery post-stroke: a case study.

Background and Purpose Task-specific and strength training have demonstrated efficacy as therapeutic interventions post-stroke. The intent of this case study is to describe outcomes associated with a therapy program that combines task-specific and strength training in an individual post-stroke and to discuss some possible mechanisms and modulating factors that may affect post-stroke neurologic recovery and responsiveness to intervention. Case Description The participant was a 38-year-old female with right middle cerebral artery stroke, evaluated 15 months post-onset. She ambulated independently with an ankle-foot orthosis and straight cane. Her free and fast overground velocity was 0.50 m/s and 0.62 m/s, respectively. Body-weight supported treadmill training and a limb-loaded cycling exercise were alternated over 24 treatments sessions (4 times/wk for 6 wks). Measurements were taken pre-, post-treatment, and at a 6-mo follow-up. Instrumented gait and motion analysis with fine-wire EMG recording of LE muscle activity occurred pre- and post-treatment. Outcomes Post-treatment, walking speed increased 18% for free- (0.59 m/ s) and 14.4% for fast-velocity (0.71 m/s); 6-min walking distance increased 4% (184.4 m). At 6-mos, continued improvements in all walking outcomes were evident. Gait and motion analysis revealed that functional locomotor recovery was associated with increases in magnitude of paretic leg gluteus maximus and gluteus medius activation during gait. Motion analysis confirmed an increase of hip and knee extension motions throughout stance and swing. Discussion For the person in this case clinically meaningful changes in walking function were associated with a combined therapeutic program that included both task-specific and LE strength training. Possible mechanisms associated with response to therapy were related to improved motor unit activation associated with increased strength in key muscles used in gait.

Comparative Analytical Utility of DNA Derived from Alternative Human Specimens for Molecular Autopsy and Diagnostics

Genetic testing and research have increased the demand for high-quality DNA that has traditionally been obtained by venipuncture. However, venous blood collection may prove difficult in special populations and when large-scale specimen collection or exchange is prerequisite for international collaborative investigations. Guthrie/FTA card–based blood spots, buccal scrapes, and finger nail clippings are DNA-containing specimens that are uniquely accessible and thus attractive as alternative tissue sources (ATS). The literature details a variety of protocols for extraction of nucleic acids from a singular ATS type, but their utility has not been systematically analyzed in comparison with conventional sources such as venous blood. Additionally, the efficacy of each protocol is often equated with the overall nucleic acid yield but not with the analytical performance of the DNA during mutation detection. Together with a critical in-depth literature review of published extraction methods, we developed and evaluated an all-inclusive approach for serial, systematic, and direct comparison of DNA utility from multiple biological samples. Our results point to the often underappreciated value of these alternative tissue sources and highlight ways to maximize the ATS-derived DNA for optimal quantity, quality, and utility as a function of extraction method. Our comparative analysis clarifies the value of ATS in genomic analysis projects for population-based screening, diagnostics, molecular autopsy, medico-legal investigations, or multi-organ surveys of suspected mosaicisms.

Selective Loss of Presynaptic Potassium Channel Clusters at the Cerebellar Basket Cell Terminal Pinceau in Adam11 Mutants Reveals Their Role in Ephaptic Control of Purkinje Cell Firing.

A specialized axonal ending, the basket cell “pinceau,” encapsulates the Purkinje cell axon initial segment (AIS), exerting final inhibitory control over the integrated outflow of the cerebellar cortex. This nonconventional axo-axonic contact extends beyond the perisomatic chemical GABAergic synaptic boutons to the distal AIS, lacks both sodium channels and local exocytotic machinery, and yet contains a dense cluster of voltage-gated potassium channels whose functional contribution is unknown. Here, we show that ADAM11, a transmembrane noncatalytic disintegrin, is the first reported Kv1-interacting protein essential for localizing Kv1.1 and Kv1.2 subunit complexes to the distal terminal. Selective absence of these channels at the pinceau due to mutation of ADAM11 spares spontaneous GABA release from basket cells at the perisomatic synapse yet eliminates ultrarapid ephaptic inhibitory synchronization of Purkinje cell firing. Our findings identify a critical role for presynaptic K+ channels at the pinceau in ephaptic control over the speed and stability of spike rate coding at the Purkinje cell AIS in mice. SIGNIFICANCE STATEMENT This study identifies ADAM11 as the first essential molecule for the proper localization of potassium ion channels at presynaptic nerve terminals, where they modulate excitability and the release of neural transmitters. Genetic truncation of the transmembrane disintegrin and metalloproteinase protein ADAM11 resulted in the absence of Kv1 channels that are normally densely clustered at the terminals of basket cell axons in the cerebellar cortex. These specialized terminals are responsible for the release of the neurotransmitter GABA onto Purkinje cells and also display electrical signaling. In the ADAM11 mutant, GABAergic release was not altered, but the ultrarapid electrical signal was absent, demonstrating that the dense presynaptic cluster of Kv1 ion channels at these terminals mediate electrical transmission. Therefore, ADAM11 plays a critical role at this central synapse.