simultaneous-video-eeg-ecg-monitoring-to-identify-neurocardiac

Simultaneous Video-EEG-ECG Monitoring to Identify Neurocardiac Dysfunction in Mouse Models of Epilepsy

In epilepsy, seizures can evoke cardiac rhythm disturbances such as heart rate changes, conduction blocks, asystoles, and arrhythmias, which can ...

microelectrode-array-recording-sinoatrial-node-firing-rate-to

Microelectrode Array Recording of Sinoatrial Node Firing Rate to Identify Intrinsic Cardiac Pacemaking Defects in Mice

The sinoatrial node (SAN), located in the right atrium, contains the pacemaker cells of the heart, and dysfunction of this region can cause tachycardia or ...

louisiana state university health sciences center

Southern Methodist University

Sudden unexpected death in epilepsy (SUDEP) is the leading cause of epilepsy-related mortality, but how to predict which patients are at risk and how to prevent it remain uncertain. The underlying pathomechanisms of SUDEP are still largely unknown, but the general consensus is that seizures somehow disrupt normal cardiac or respiratory physiology leading to death. However, the proportion of SUDEP cases exhibiting cardiac or respiratory dysfunction as a critical factor in the terminal cascade of events remains unresolved. Although many general risk factors for SUDEP have been identified, the development of reliable patient-specific biomarkers for SUDEP is needed to provide more accurate risk prediction and personalized patient management strategies. Studies in animal models and patient groups have revealed at least nine different brain-heart genes that may contribute to a genetic susceptibility for SUDEP, making them potentially useful as genomic biomarkers. This review summarizes data on the relationship between these neurocardiac genes and SUDEP, discussing their brain-heart expression patterns and genotype-phenotype correlations in mouse models and people with epilepsy. These neurocardiac genes represent good first candidates for evaluation as genomic biomarkers of SUDEP in future studies. The development of validated reliable genomic biomarkers for SUDEP has the potential to transform the clinical treatment of epilepsy by pinpointing patients at risk of SUDEP and allowing optimized, genotype-guided therapeutic and prevention strategies.

Genomic biomarkers of SUDEP in brain and heart.

Mice lacking voltage-gated Kv1.1 channels as a result of deletion of the Kcna1 gene are an extensively utilized genetic model of human epilepsy and sudden unexpected death in epilepsy because of their frequent seizures and genotypic-phenotypic similarity to the human condition. Ictal behaviors, electrophysiological recordings, and gene expression studies suggest limbic circuits are critical for epilepsy in Kcna1-null mice, but the exact brain networks recruited by seizures remain unknown. In this study, Fos protein expression patterns were used to map limbic brain regions with increased neuronal activity at baseline and during spontaneous seizures in Kcna1-null mice by comparing seizing and non-seizing knockouts and wild-type controls. Basal Fos levels were unchanged in non-seizing knockout mice compared to wild types for all brain regions examined except the dentate gyrus granule cell layer which exhibited a significant decrease in Fos-positive cells. Following seizures, Kcna1-null brains exhibited significantly increased Fos labeling in the basolateral amygdala and the dentate hilus region, but not in other principal cell layers of the hippocampal formation. The selective Fos activation in the amygdala following seizures suggests that extra hippocampal limbic circuits may be critically involved with seizure generation or spread in Kcna1-null mice. Fos protein expression patterns were analyzed using immunohistochemistry to provide the first map of brain regions recruited by spontaneous seizures in mice lacking Kv1.1 channels, an extensively used genetic model of epilepsy. Seizures significantly increased Fos expression in the amygdala and hilus by about fourfold, suggesting an important contribution by extrahippocampal networks to epilepsy in this model.

Spontaneous seizures in Kcna1-null mice lacking voltage-gated Kv1.1 channels activate Fos expression in select limbic circuits.

Voltage-gated Kv1.1 channels encoded by the Kcna1 gene are traditionally regarded as being neural-specific with no known expression or intrinsic functional role in the heart. However, recent studies in mice reveal low-level Kv1.1 expression in heart and cardiac abnormalities associated with Kv1.1-deficiency suggesting that the channel may have a previously unrecognized cardiac role. Therefore, this study tests the hypothesis that Kv1.1 channels are associated with arrhythmogenesis and contribute to intrinsic cardiac function. In intra-atrial burst pacing experiments, Kcna1-null mice exhibited increased susceptibility to atrial fibrillation (AF). The atria of Kcna1-null mice showed minimal Kv1 family ion channel remodeling and fibrosis as measured by qRT-PCR and Masson's trichrome histology, respectively. Using RT-PCR, immunocytochemistry, and immunoblotting, KCNA1 mRNA and protein were detected in isolated mouse cardiomyocytes and human atria for the first time. Patients with chronic AF (cAF) showed no changes in KCNA1 mRNA levels relative to controls; however, they exhibited increases in atrial Kv1.1 protein levels, not seen in paroxysmal AF patients. Patch-clamp recordings of isolated human atrial myocytes revealed significant dendrotoxin-K (DTX-K)-sensitive outward current components that were significantly increased in cAF patients, reflecting a contribution by Kv1.1 channels. The concomitant increases in Kv1.1 protein and DTX-K-sensitive currents in atria of cAF patients suggest that the channel contributes to the pathological mechanisms of persistent AF. These findings provide evidence of an intrinsic cardiac role of Kv1.1 channels and indicate that they may contribute to atrial repolarization and AF susceptibility.

Expression and function of Kv1.1 potassium channels in human atria from patients with atrial fibrillation.

Implanted gradient index lenses have extended the reach of standard multiphoton microscopy from the upper layers of the mouse cortex to the lower cortical layers and even subcortical regions. These lenses have the clarity to visualize dynamic activities, such as calcium transients, with subcellular and millisecond resolution and the stability to facilitate repeated imaging over weeks and months. In addition, behavioral tests can be used to correlate performance with observed changes in network function and structure that occur over time. Yet, this raises the questions, does an implanted microlens have an effect on behavioral tests, and if so, what is the extent of the effect? To answer these questions, we compared the performance of three groups of mice in three common behavioral tests. A gradient index lens was implanted in the prefrontal cortex of experimental mice. We compared their performance with mice that had either a cranial window or a sham surgery. Three presurgical and five postsurgical sets of behavioral tests were performed over seven weeks. Behavioral tests included rotarod, foot fault, and Morris water maze. No significant differences were found between the three groups, suggesting that microlens implantation did not affect performance. The results for the current study clear the way for combining behavioral studies with gradient index lens imaging in the prefrontal cortex, and potentially other regions of the mouse brain, to study structural, functional, and behavioral relationships in the brain.

Gradient Index Microlens Implanted in Prefrontal Cortex of Mouse Does Not Affect Behavioral Test Performance over Time.

Fused in sarcoma (FUS) is an RNA-binding protein associated with the neurodegenerative diseases amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration. ALS manifests in patients as a progressive paralysis which leads to respiratory dysfunction and failure, the primary cause of death in ALS. We expressed human FUS in rats to determine if FUS would induce ALS relevant respiratory changes to serve as an early stage disease indicator. The FUS expression was initiated in adult rats by way of an intravenously administered adeno-associated virus vector serotype 9 (AAV9) providing an adult onset model.

Severe respiratory changes at end stage in a FUS-induced disease state in adult rats.

People with epilepsy have greatly increased probability of premature mortality due to sudden unexpected death in epilepsy (SUDEP). Identifying which patients are most at risk of SUDEP is hindered by a complex genetic etiology, incomplete understanding of the underlying pathophysiology and lack of prognostic biomarkers. Here we evaluated heterozygous Scn2a gene deletion (Scn2a+/-) as a protective genetic modifier in the Kcna1 knockout mouse (Kcna1-/-) model of SUDEP, while searching for biomarkers of SUDEP risk embedded in electroencephalography (EEG) and electrocardiography (ECG) recordings. The human epilepsy gene Kcna1 encodes voltage-gated Kv1.1 potassium channels that act to dampen neuronal excitability whereas Scn2a encodes voltage-gated Nav1.2 sodium channels important for action potential initiation and conduction. SUDEP-prone Kcna1-/- mice with partial genetic ablation of Nav1.2 channels (i.e. Scn2a+/-; Kcna1-/-) exhibited a two-fold increase in survival. Classical analysis of EEG and ECG recordings separately showed significantly decreased seizure durations in Scn2a+/-; Kcna1-/- mice compared with Kcna1-/- mice, without substantial modification of cardiac abnormalities. Novel analysis of the EEG and ECG together revealed a significant reduction in EEG-ECG association in Kcna1-/- mice compared with wild types, which was partially restored in Scn2a+/-; Kcna1-/- mice. The degree of EEG-ECG association was also proportional to the survival rate of mice across genotypes. These results show that Scn2a gene deletion acts as protective genetic modifier of SUDEP and suggest measures of brain-heart association as potential indices of SUDEP susceptibility.

Scn2a deletion improves survival and brain-heart dynamics in the Kcna1-null mouse model of sudden unexpected death in epilepsy (SUDEP).

Antiseizure drugs are the leading therapeutic choice for treatment of epilepsy, but their efficacy is limited by pharmacoresistance and the occurrence of unwanted side effects. Here, we examined the therapeutic efficacy of KCNQ channel activation by retigabine in preventing seizures and neurocardiac dysfunction in 2 potassium channelopathy mouse models of epilepsy with differing severity that have been associated with increased risk of sudden unexpected death in epilepsy (SUDEP): the Kcna1-/- model of severe epilepsy and the Kcnq1A340E/A340E model of mild epilepsy.

Pharmacogenetics of KCNQ channel activation in 2 potassium channelopathy mouse models of epilepsy.

Voltage-gated Kv1.1 potassium channel α-subunits are broadly expressed in the nervous system where they act as critical regulators of neuronal excitability. Mutations in the  gene, which encodes Kv1.1, are associated with the neurological diseases episodic ataxia and epilepsy. Studies in mouse models have shown that Kv1.1 is important for neural control of the heart and that  deletion leads to cardiac dysfunction that appears to be brain-driven. Traditionally,  was not believed to be expressed in the heart. However, recent studies have revealed that Kv1.1 subunits are not only present in cardiomyocytes, but that they also make an important heart-intrinsic functional contribution to outward K currents and action potential repolarization. This review recounts the winding history of discovery of  gene expression and neurocardiac function from fruit flies to mammals and from brain to heart and looks at some of the salient questions that remain to be answered regarding emerging cardiac roles of Kv1.1.

Kv1.1 channel subunits in the control of neurocardiac function.

Sudden unexpected death in epilepsy (SUDEP) is the leading cause of epilepsy-related mortality, but the precise cellular substrates involved remain elusive. Epilepsy-associated ion channel genes with co-expression in brain and heart have been proposed as SUDEP candidate genes since they provide a singular unifying link between seizures and lethal cardiac arrhythmias. Here, we generated a conditional knockout (cKO) mouse with neuron-specific deletion of Kcna1, a SUDEP-associated gene with brain-heart co-expression, to test whether seizure-evoked cardiac arrhythmias and SUDEP require the absence of Kv1.1 in both brain and heart or whether ablation in neurons is sufficient. To obtain cKO mice, we developed a floxed Kcna1 mouse which we crossed to mice with the Synapsin1-Cre transgene, which selectively deletes Kcna1 in most neurons. Molecular analyses confirmed neuron-specific Kcna1 deletion in cKO mice and corresponding loss of Kv1.1 except in cerebellum where Synapsin1-Cre is not highly expressed. Survival studies and electroencephalography, electrocardiography, and plethysmography recordings showed that cKO mice exhibit premature death, epilepsy, and cardiorespiratory dysregulation but to a lesser degree than global knockouts. Heart rate variability (HRV) was increased in cKO mice with peaks during daytime suggesting disturbed diurnal HRV patterns as a SUDEP biomarker. Residual Kv1.1 expression in cKO cerebellum suggests it may play an unexpected role in regulating ictal cardiorespiratory dysfunction and SUDEP risk. This work demonstrates the principle that channelopathies with brain-heart expression patterns can increase death risk by brain-driven mechanisms alone without a functionally compromised heart, reinforcing seizure control as a primary clinical strategy for SUDEP prevention.

Neuron-specific Kv1.1 deficiency is sufficient to cause epilepsy, premature death, and cardiorespiratory dysregulation.

Mutations in the  gene, which encodes voltage-gated Kv1.1 potassium channel α-subunits, cause a variety of human diseases, complicating simple genotype-phenotype correlations in patients.  mutations are primarily associated with a rare neurological movement disorder known as episodic ataxia type 1 (EA1). However, some patients have EA1 in combination with epilepsy, whereas others have epilepsy alone.  mutations can also cause hypomagnesemia and paroxysmal dyskinesia in rare cases. Why  variants are associated with such phenotypic heterogeneity in patients is not yet understood. In this review, literature databases (PubMed) and public genetic archives (dbSNP and ClinVar) were mined for known pathogenic or likely pathogenic mutations in  to examine whether patterns exist between mutation type and disease manifestation. Analyses of the 47 deleterious  mutations that were identified revealed that epilepsy or seizure-related variants tend to cluster in the S1/S2 transmembrane domains and in the pore region of Kv1.1, whereas EA1-associated variants occur along the whole length of the protein. In addition, insights from animal models of  channelopathy were considered, as well as the possible influence of genetic modifiers on disease expressivity and severity. Elucidation of the complex relationship between  variants and disease will enable better diagnostic risk assessment and more personalized therapeutic strategies for  channelopathy.

Clinical Spectrum of  Mutations: New Insights into Episodic Ataxia and Epilepsy Comorbidity.

Oxidative stress drives the pathogenesis of atrial fibrillation (AF), the most common arrhythmia. In the cardiovascular system, cystathionine γ-lyase (CSE) serves as the primary enzyme producing hydrogen sulfide (HS), a mammalian gasotransmitter that reduces oxidative stress. Using a case control study design in patients with and without AF and a mouse model of CSE knockout (CSE-KO), we evaluated the role of HS in the etiology of AF. Patients with AF (n = 51) had significantly reduced plasma acid labile sulfide levels compared to patients without AF (n = 65). In addition, patients with persistent AF (n = 25) showed lower plasma free sulfide levels compared to patients with paroxysmal AF (n = 26). Consistent with an important role for HS in AF, CSE-KO mice had decreased atrial sulfide levels, increased atrial superoxide levels, and enhanced propensity for induced persistent AF compared to wild type (WT) mice. Rescuing HS signaling in CSE-KO mice by Diallyl trisulfide (DATS) supplementation or reconstitution with endothelial cell specific CSE over-expression significantly reduced atrial superoxide, increased sulfide levels, and lowered AF inducibility. Lastly, low HS levels in CSE KO mice was associated with atrial electrical remodeling including longer effective refractory periods, slower conduction velocity, increased myocyte calcium sparks, and increased myocyte action potential duration that were reversed by DATS supplementation or endothelial CSE overexpression. Our findings demonstrate an important role of CSE and HS bioavailability in regulating electrical remodeling and susceptibility to AF.

Decreased bioavailability of hydrogen sulfide links vascular endothelium and atrial remodeling in atrial fibrillation.

Epilepsy-associated Kv1.1 voltage-gated potassium channel subunits encoded by the Kcna1 gene have traditionally been considered absent in heart, but recent studies reveal they are expressed in cardiomyocytes where they could regulate intrinsic cardiac electrophysiology. Although Kv1.1 now has a demonstrated functional role in atria, its role in the ventricles has never been investigated. In this work, electrophysiological, histological, and gene expression approaches were used to explore the consequences of Kv1.1 deficiency in the ventricles of Kcna1 knockout (KO) mice at the organ, cellular, and molecular levels to determine whether the absence of Kv1.1 leads to ventricular dysfunction that increases the risk of premature or sudden death. When subjected to intracardiac pacing, KO mice showed normal baseline susceptibility to inducible ventricular arrhythmias (VA) but resistance to VA under conditions of sympathetic challenge with isoproterenol. Echocardiography revealed cardiac contractile dysfunction manifesting as decreased ejection fraction and fractional shortening. In whole-cell patch-clamp recordings, KO ventricular cardiomyocytes exhibited action potential prolongation indicative of impaired repolarization. Imaging, histological, and transcript analyses showed no evidence of structural or channel gene expression remodeling, suggesting that the observed deficits are likely electrogenic due to Kv1.1 deficiency. Immunoblots of patient heart samples detected the presence of Kv1.1 at relatively high levels, implying that Kv1.1 contributes to human cardiac electrophysiology. Taken together, this work describes an important functional role for Kv1.1 in ventricles where its absence causes repolarization and contractility deficits but reduced susceptibility to arrhythmia under conditions of sympathetic drive.

Kv1.1 potassium channel subunit deficiency alters ventricular arrhythmia susceptibility, contractility, and repolarization.

Autism spectrum disorder (ASD) and epilepsy are highly comorbid, suggesting potential overlap in genetic etiology, pathophysiology, and neurodevelopmental abnormalities; however, the nature of this relationship remains unclear. This work investigated how two ion channel mutations, one associated with autism (Scn2a-null) and one with epilepsy (Kcna1-null), interact to modify genotype-phenotype relationships in the context of autism. Previous studies have shown that Scn2a ameliorates epilepsy in Kcna1 mice, improving survival, seizure characteristics, and brain-heart dynamics. Here, we tested the converse, whether Kcna1 deletion modifies ASD-like repetitive and social behaviors in Scn2a mice.

Kv1.1 deficiency alters repetitive and social behaviors in mice and rescues autistic-like behaviors due to Scn2a haploinsufficiency.

Cardiorespiratory collapse following a seizure is a suspected cause of sudden unexpected death in epilepsy (SUDEP), the leading cause of epilepsy-related mortality. In the commonly used Kcna1 gene knockout (Kcna1) mouse model of SUDEP, cardiorespiratory profiling reveals an array of aberrant breathing patterns that could contribute to risk of seizure-related mortality. However, the brain structures mediating these respiratory abnormalities remain unknown. We hypothesize that Kv1.1 deficiency in respiratory control centers of the brain contribute to respiratory dysfunction in Kcna1 mice leading to increased SUDEP risk. Thus, in this study, we first used immunohistochemistry to map expression of Kv1.1 protein in cardiorespiratory brain regions of wild-type Kcna1 (WT) mice. Next, GFAP and Iba1 immunostaining was used to test for the presence of astrogliosis and microgliosis, respectively, in the cardiorespiratory centers of Kcna1 mice, which could be indicative of seizure-related brain injury that could impair breathing. In WT mice, we detected Kv1.1 protein in all cardiorespiratory centers examined, including the basolateral amygdala, dorsal respiratory group, dorsal motor nucleus of vagus, nucleus ambiguus, ventral respiratory column, and pontine respiratory group, as well as chemosensory centers including the retrotrapezoid and median raphae nuclei. Extensive gliosis was observed in the same areas in Kcna1 mice suggesting that seizure-associated brain injury could contribute to respiratory abnormalities.

ArticleTotal : 2	Year
Publication title Cited by 21	2018
Publication title Cited by 2	2021

Article	Year
Genomic biomarkers of SUDEP in brain and heart. Epilepsy & behavior : E&B\| PubMed ID: 24139807	2014
Spontaneous seizures in Kcna1-null mice lacking voltage-gated Kv1.1 channels activate Fos expression in select limbic circuits. Journal of neurochemistry\| PubMed ID: 26112121	2015
Expression and function of Kv1.1 potassium channels in human atria from patients with atrial fibrillation. Basic research in cardiology\| PubMed ID: 26162324	2015
Gradient Index Microlens Implanted in Prefrontal Cortex of Mouse Does Not Affect Behavioral Test Performance over Time. PloS one\| PubMed ID: 26799938	2016
Severe respiratory changes at end stage in a FUS-induced disease state in adult rats. BMC neuroscience\| PubMed ID: 27793099	2016
Scn2a deletion improves survival and brain-heart dynamics in the Kcna1-null mouse model of sudden unexpected death in epilepsy (SUDEP). Human molecular genetics\| PubMed ID: 28334922	2017
Pharmacogenetics of KCNQ channel activation in 2 potassium channelopathy mouse models of epilepsy. Epilepsia\| PubMed ID: 29265344	2017
Kv1.1 channel subunits in the control of neurocardiac function. Channels (Austin, Tex.)\| PubMed ID: 31250689	2019
Neuron-specific Kv1.1 deficiency is sufficient to cause epilepsy, premature death, and cardiorespiratory dysregulation. Neurobiology of disease\| PubMed ID: 31978607	2020
Clinical Spectrum of Mutations: New Insights into Episodic Ataxia and Epilepsy Comorbidity. International journal of molecular sciences\| PubMed ID: 32316562	2020
Decreased bioavailability of hydrogen sulfide links vascular endothelium and atrial remodeling in atrial fibrillation. Redox biology\| PubMed ID: 33310503	2021
Kv1.1 potassium channel subunit deficiency alters ventricular arrhythmia susceptibility, contractility, and repolarization. Physiological reports\| PubMed ID: 33427415	2021
Kv1.1 deficiency alters repetitive and social behaviors in mice and rescues autistic-like behaviors due to Scn2a haploinsufficiency. Brain and behavior\| PubMed ID: 33484493	2021
Kv1.1 subunits localize to cardiorespiratory brain networks in mice where their absence induces astrogliosis and microgliosis. Molecular and cellular neurosciences\| PubMed ID: 33901631	2021

Edward Glasscock

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