KCNA2 Research Articles

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 potentially increase risk of sudden unexpected death in epilepsy (SUDEP). Electroencephalography (EEG) and electrocardiography (ECG) are widely used clinical diagnostic tools to monitor for abnormal brain and cardiac rhythms in patients. Here, a technique to simultaneously record video, EEG, and ECG in mice to measure behavior, brain, and cardiac activities, respectively, is described. The technique described herein utilizes a tethered (i.e., wired) recording configuration in which the implanted electrode on the head of the mouse is hard-wired to the recording equipment. Compared to wireless telemetry recording systems, the tethered arrangement possesses several technical advantages such as a greater possible number of channels for recording EEG or other biopotentials; lower electrode costs; and greater frequency bandwidth (i.e., sampling rate) of recordings. The basics of this technique can also be easily modified to accommodate recording other biosignals, such as electromyography (EMG) or plethysmography for assessment of muscle and respiratory activity, respectively. In addition to describing how to perform the EEG-ECG recordings, we also detail methods to quantify the resulting data for seizures, EEG spectral power, cardiac function, and heart rate variability, which we demonstrate in an example experiment using a mouse with epilepsy due to Kcna1 gene deletion. Video-EEG-ECG monitoring in mouse models of epilepsy or other neurological disease provides a powerful tool to identify dysfunction at the level of the brain, heart, or brain-heart interactions.

Familial paroxysmal kinesigenic dyskinesia is associated with mutations in the KCNA1 gene.

Paroxysmal kinesigenic dyskinesia (PKD) is a heterogeneous movement disorder characterized by recurrent dyskinesia attacks triggered by sudden movement. PRRT2 has been identified as the first causative gene of PKD. However, it is only responsible for approximately half of affected individuals, indicating that other loci are most likely involved in the etiology of this disorder. To explore the underlying causative gene of PRRT2-negative PKD, we used a combination strategy including linkage analysis, whole-exome sequencing and copy number variations analysis to detect the genetic variants within a family with PKD. We identified a linkage locus on chromosome 12 (12p13.32-12p12.3) and detected a novel heterozygous mutation c.956 T>G (p.319 L>R) in the potassium voltage-gated channel subfamily A member 1, KCNA1. Whole-exome sequencing in another 58 Chinese patients with PKD who lacked mutations in PRRT2 revealed another novel mutation in the KCNA1 gene [c.765 C>A (p.255 N>K)] within another family. Biochemical analysis revealed that the L319R mutant accelerated protein degradation via the proteasome pathway and disrupted membrane expression of the Kv1.1 channel. Electrophysiological examinations in transfected HEK293 cells showed that both the L319R and N255K mutants resulted in reduced potassium currents and respective altered gating properties, with a dominant negative effect on the Kv1.1 wild-type channel. Our study suggests that these mutations in KCNA1 cause the Kv1.1 channel dysfunction, which leads to familial PKD. The current study further extended the genotypic spectrum of this disorder, indicating that Kv1.1 channel dysfunction maybe one of the underlying defects in PKD.

Respiratory dysfunction progresses with age in Kcna1-null mice, a model of sudden unexpected death in epilepsy.

Increased breathing rate, apnea, and respiratory failure are associated with sudden unexpected death in epilepsy (SUDEP). We recently demonstrated the progressive nature of epilepsy and mortality in Kcna1-/- mice, a model of temporal lobe epilepsy and SUDEP. Here we tested the hypothesis that respiratory dysfunction progresses with age in Kcna1-/- mice, thereby increasing risk of respiratory failure and sudden death (SD). Respiratory parameters were determined in conscious mice at baseline and following increasing doses of methacholine (MCh) using noninvasive airway mechanics (NAM) systems. Kcna1+/+ , Kcna1+/- , and Kcna1-/- littermates were assessed during 3 age ranges when up to ~30%, ~55%, and ~90% of Kcna1-/- mice have succumbed to SUDEP: postnatal day (P) 32-36, P40-46, and P48-56, respectively. Saturated arterial O2 (SaO2 ) was determined with pulse oximetry. Lung and brain tissues were isolated and Kcna1 gene and protein expression were evaluated by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and Western blot techniques. Airway smooth muscle responsiveness was assessed in isolated trachea exposed to MCh. Kcna1-/- mice experienced an increase in basal respiratory drive, chronic oxygen desaturation, frequent apnea-hypopnea (A-H), an atypical breathing sequence of A-H-tachypnea-A-H, increased tidal volume, and hyperventilation induced by MCh. The MCh-provoked hyperventilation was dramatically attenuated with age. Of interest, only Kcna1-/- mice developed seizures following exposure to MCh. Seizures were provoked by lower concentrations of MCh as Kcna1-/- mice approached SD. MCh-induced seizures experienced by a subset of younger Kcna1-/- mice triggered death. Respiratory parameters of these younger Kcna1-/- mice resembled older near-SD Kcna1-/- mice. Kcna1 gene and protein were not expressed in Kcna1+/+ and Kcna1+/- lungs, and MCh-mediated airway smooth muscle contractions exhibited similar half-maximal effective concentration( EC50 ) in isolated Kcna1+/+ and Kcna1-/- trachea. The Kcna1-/- model of SUDEP exhibits progressive respiratory dysfunction, which suggests a potential increased susceptibility for respiratory failure during severe seizures that may result in sudden death.

Familial paroxysmal kinesigenic dyskinesia is associated with mutations in the KCNA1 gene.

Paroxysmal kinesigenic dyskinesia (PKD) is a heterogeneous movement disorder characterized by recurrent dyskinesia attacks triggered by sudden movement. PRRT2 has been identified as the first causative gene of PKD. However, it is only responsible for approximately half of affected individuals, indicating that other loci are most likely involved in the etiology of this disorder. To explore the underlying causative gene of PRRT2-negative PKD, we used a combination strategy including linkage analysis, whole-exome sequencing and copy number variations analysis to detect the genetic variants within a family with PKD. We identified a linkage locus on chromosome 12 (12p13.32-12p12.3) and detected a novel heterozygous mutation c.956 T>G (p.319 L>R) in the potassium voltage-gated channel subfamily A member 1, KCNA1. Whole-exome sequencing in another 58 Chinese patients with PKD who lacked mutations in PRRT2 revealed another novel mutation in the KCNA1 gene [c.765 C>A (p.255 N>K)] within another family. Biochemical analysis revealed that the L319R mutant accelerated protein degradation via the proteasome pathway and disrupted membrane expression of the Kv1.1 channel. Electrophysiological examinations in transfected HEK293 cells showed that both the L319R and N255K mutants resulted in reduced potassium currents and respective altered gating properties, with a dominant negative effect on the Kv1.1 wild-type channel. Our study suggests that these mutations in KCNA1 cause the Kv1.1 channel dysfunction, which leads to familial PKD. The current study further extended the genotypic spectrum of this disorder, indicating that Kv1.1 channel dysfunction maybe one of the underlying defects in PKD.

Inherited neuromyotonia as a phenotypic variation associated with a new mutation in the KCNQ2 gene

Background: Neuromyotonia is a disorder of peripheral nerve hyperexcitability (PNH) characterized by muscle cramps, stiffness, myokymia, delayed muscle relaxation after contraction, and hyperhidrosis. Usually cause of neuromyotonia was acquired autoimmune mechanisms against voltage-gated potassium-channel and rare cause was genetic disorders of potassium channels. Mutations are reported mostly in the KCNA1 gene and only one inherited PNH-associated mutation has been described in KCNQ2 (p.Arg207Trp).

A channelopathy mutation in the voltage-sensor discloses contributions of a conserved phenylalanine to gating properties of Kv1.1 channels and ataxia

Channelopathy mutations prove informative on disease causing mechanisms and channel gating dynamics. We have identified a novel heterozygous mutation in the KCNA1 gene of a young proband displaying typical signs and symptoms of Episodic Ataxia type 1 (EA1). This mutation is in the S4 helix of the voltage-sensing domain and results in the substitution of the highly conserved phenylalanine 303 by valine (p.F303V). The contributions of F303 towards K+ channel voltage gating are unclear and here have been assessed biophysically and by performing structural analysis using rat Kv1.2 coordinates. We observed significant positive shifts of voltage-dependence, changes in the activation, deactivation and slow inactivation kinetics, reduced window currents, and decreased current amplitudes of both Kv1.1 and Kv1.1/1.2 channels. Structural analysis revealed altered interactions between F303V and L339 and I335 of the S5 helix of a neighboring subunit. The substitution of an aromatic phenylalanine with an aliphatic valine within the voltage-sensor destabilizes the open state of the channel. Thus, F303 fine-tunes the Kv1.1 gating properties and contributes to the interactions between the S4 segment and neighboring alpha helices. The resulting channel’s loss of function validates the clinical relevance of the mutation for EA1 pathogenesis.

Preliminary Results of Whole Exome Sequencing in a Cohort of Sicilian Children with Early-Onset Epileptic Encephalopaty

Objective: Early-Onset Epileptic Encephalopathies encompass a group of severe disorders characterized by refractory epilepsy and/or electroencephalographic abnormalities with progressive cerebral dysfunction. The modern technologies of molecular genetics are providing crucial diagnostic insights, however, this is only the tip of the iceberg and a great effort is still necessary to detect new genes, new mutations and define their causative role. We herein present the preliminary results in our cohort with EOEE included into extensive multicenter Whole Exome Sequencing study (SynaPs network). Methods: Twenty-four patients with electroclinical diagnosis of EOEE, no structural abnormalities at brain MRI scans and negative array-CGH were enrolled. Results: A de novo mutation on GRIN1 gene was detected in a 5-years-old girl with EOEE, severe hypotonia, pyramidal and extrapyramidal signs and Rett-like manual stereotypies. GRIN1 mutations impairs functioning of glutamate NMDA receptor. In an 8 years-old male patient with EOEE, hypotonia, ataxia and autism features, a de novo mutation in KCNA2 gene was found. It encodes for potassium channel Kv1.2 and has been associated to inherited episodic ataxia and epilepsy. In a 16 years-old boy with EOEE and subsequent Lennox-Gastaut ’s phenotype, WES analysis evidenced a damaging mutation in the contactin 2 (CNTN2) gene. A pathogenic role of CNTN2 has been hypothesized due to its interaction with Contactin-associated Protein-like 2 which is implicated in several neurological disorders. A SZT2 gene mutation was found in a 7 years-old male with EOEE, microcephaly and severe developmental delay. To date, two clinical reports described few children with SZT2 mutations and heterogeneous clinical phenotypes including epileptic encephalopathy, developmental delay and dysmorphisms. Conclusion: WES analysis may significantly enhance diagnostic chances in patients with EOEE. A synergic work between clinicians and geneticists is mandatory to assess genotype-phenotype correlation and provide proper genetic counseling. Disease- specific treatments could be attempted and taken into account in clinical practice in the future.

Pharmacogenetic and case-control study on potassium channel related gene variants and genetic generalized epilepsy.

Potassium channels are the targets of antiepileptic drugs (AEDs), which play important roles in the etiology of epilepsy. KCNA1 and KCNA2 encode mammalian Kv1.1 and Kv1.2 channels, which are essential roles in the initiation and shaping of action potentials. KCNV2 encodes Kv8.2, which is a regional overlap with Kv2 subunits as functional heterotetramers. In our study, we aim to investigate whether variants of KCNA1, KCNA2, and KCNV2 genes influence susceptibility to genetic generalized epilepsies (GGEs) and the efficacy of AEDs. Seven hundred sixty-seven subjects (284 healthy controls, 279 drug-responsive, and 204 drug-resistant GGE patients) were enrolled in our study. Eight variants of KCNA1, KCNA2, and KCNV2 were assessed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry method. Results showed that there were no statistically significant correlations between the 8 variants of KCNA1, KCNA2, and KCNV2 and the risk/drug resistance of GGEs. In conclusion, our study suggests that KCNA1, KCNA2, and KCNV2 variants may not be involved in the risk/drug resistance of GGEs. Further multicenter, multiethnic, and large sample size pharmacogenetic and case–control studies are warranted to confirm our negative results.

Clinical heterogeneity of Episodic Ataxia type 1 presentation in family with the c.677C>G KCNA1 gene heterozygosity

Clinical heterogeneity of Episodic Ataxia type 1 presentation in family with the c.677C>G KCNA1 gene heterozygosity

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

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.

Abstract PR317

Background & Objectives: Our previous study showed that results showed that MBD1 knockout mice showed dramatically higher pain thresholds in Mechanical, Thermal, and Cold test in acute pain and SNL-pain models compared to wild-type mice. However, it remains unknown how MBD1 to regulate genesis of acute pain and chronic pain. This observation led us to hypothesize that MBD1 might regulate some related factors of pain. So, we first investigated both the mRNA and protein level of pain-related opioid receptors and potassium channels in the dorsal root ganglia, spinal cord and cortex, and determined whether MBD1 targeted tissue-specific subtypes of opioid receptors and/or potassium channels. Second, we used MBD1-AAV5 overexpression viral to treat those specific tissue sites and observed pain behavior and related expression of pain related mRNA and proteins. Finally, we determined whether MBD1 co-localization with opioid receptors or potassium channels gene in a single DRG cell or not. Materials & Methods: All experiments were carried out with the approval of the Animal Care Committee at Rutgers New Jersey Medical School. For detected opioid receptors or potassium channels mRNA, we used RT-qPCR to detect Oprm1, Oprk1, Oprd1, Kcna1 and Kcna2 mRNA gene expression and used western blot to detected mu, kappa, delta, Kv1.1 and Kv1.2 protein expression on the level of dorsal root ganglia, the spinal cord and cortex. For in vivo experiments, wild-type mice were microinjected with GFP-AAV5 viral (1 µl) or MBD1-AAV5 viral (1 µl) in DRG. CPP test was carried out to test the spontaneous pain behavior. Overexpression of MBD1 in wild-type mice induced spontaneous pain behavior after 6 weeks. On week 8, DRG was collected. For in vitro experiments, cultured DRG neurons were exposed to GFP or MBD1-AAV5 viral and proteins were assessed by Western blot. Single cell RT-PCR was carried out to test whether MBD1 co-localization with opioid receptors or potassium channels gene in a single DRG cell or not. Results: Oprm1, Oprk1, and Kcna2 mRNA gene expressed at high levels in DRG, spinal cord and cortex, but not midbrain. Interestingly, only mu opioid receptor and Kv1.2 receptor protein significantly increased in DRG, other proteins and sites were not altered. These increases were blocked by MBD1 over-expression in DRG, and MBD1 over-expression can cause spontaneous pain behavior. Furthermore, MBD1 overexpression in cultured DRG neurons down-regulate mu opioid receptor and Kv1.2 receptor protein expression. Single cell RT-PCR assay revealed that MBD1 co-localization with mu opioid receptor and Kv1.2 receptor gene in a single DRG cell. Conclusion: Our findings indicate that Expression of mu opioid receptor and Kv1.2 controlled by MBD1 in the dorsal root ganglion. Disclosure of Interest: None declared

Dominant-negative mutation p.Arg324Thr in KCNA1 impairs Kv1.1 channel function in episodic ataxia.

Episodic ataxia type 1 is a rare autosomal dominant neurological disorder caused by mutations in the KCNA1 gene that encodes the α subunit of voltage-gated potassium channel Kv1.1. The functional consequences of identified mutations on channel function do not fully correlate with the clinical phenotype of patients. A clinical and genetic study was performed in a family with 5 patients with episodic ataxia type 1, with concurrent epilepsy in 1 of them. Protein expression, modeling, and electrophysiological analyses were performed to study Kv1.1 function. Whole-genome linkage and candidate gene analyses revealed the novel heterozygous mutation p.Arg324Thr in the KCNA1 gene. The encoded mutant Kv1.1 channel displays reduced currents and altered activation and inactivation. Taken together, we provide genetic and functional evidence that mutation p.Arg324Thr in the KCNA1 gene is pathogenic and results in episodic ataxia type 1 through a dominant-negative effect. © 2016 International Parkinson and Movement Disorder Society.

Editing of Kv1.1 RNA Transcripts is Altered by Human Mutations Respons‐ible for Episodic Ataxia Type‐1

Transcripts encoding the Kv1.1‐subtype of voltage‐gated potassium channel un‐der‐go an adenosine‐to‐inosine (A‐to‐I) RNA editing event resulting in the con‐ver‐sion of a genomically‐encoded isoleucine (AUU) to a valine codon (IUU) at amino acid position 400 (I400V). Previous studies have shown that this single amino acid change alters the recovery rate following channel inactivation and has been pro‐posed to mod‐u‐late the kinetics of neuronal repolarization due to its location within the predicted ion pore of the channel. Numerous nonsense and mis‐sense muta‐tions in the Kv1.1 gene (kcna1) have been associated with the hu‐man po‐tas‐sium channelopathy, Episodic Ataxia type‐1 (EA1), which is char‐ac‐ter‐ized by epi‐sodes of stress‐induced ataxia and myokymia, and can be associated with epi‐leptic seizures. Three human EA1 mu‐ta‐tions (V404I, I407M, and V408A) are within close prox‐imity to the I400V editing site, and are contained within the RNA duplex structure required for A‐to‐I con‐ver‐sion. We have employed both in vitro and in vivo systems to determine the impact that these EA1 mutations might have on Kv1.1 RNA editing, as well as determined the electro‐phys‐iological changes in chan‐nel function derived from tran‐scripts containing EA1 mutations with either an edited or non‐edited status. In vitro tran‐scribed RNA substrates containing EA1 mutations were incubated with nuclear extracts isolated from cells overexpres‐sing the ADAR2 editing enzyme. Each of the EA1 mu‐ta‐tions dif‐fer‐en‐tial‐ly de‐creased A‐to‐I conversion of Kv1.1 RNAs in relation to their over‐all prox‐im‐ity to the editing site. Furthermore, a mouse model of EA1, encoding the human V408A mutation at the endogenous kcna1 locus, also re‐vealed a significant decrease in Kv1.1 editing in all tissues examined. Analysis of EA1 mutant chan‐nels using a Xenopus oocyte expression system, in a non‐edited (I400) or edited (V400) background, revealed significant changes in the elec‐‐tro‐physiological char‐ac‐teristics of the channel. These observations suggest that altered Kv1.1 function in EA1 could result from both the missense mutation and alterations in editing ef‐fici‐ency. These results also are con‐sis‐tent with a gen‐et‐ically‐modified mouse mod‐el of Kv1.1 editing, where mice sole‐ly expressing the non‐edited Kv1.1 iso‐form exhibit stress‐in‐duced ataxia and an increase in induced‐sei‐zure suscepti‐bil‐i‐ty.Support or Funding InformationJoel G. Hardman Chair in Pharmacology (RBE)Multidisciplinary Training in Molecular Endocrinology (METP) NIH/NIDDK – T32 DK007563 (EFK)Ruth L. Kirschstein National Research Service Award NINDS F31 NS087911 (EFK)

Distinctive role of KV1.1 subunit in the biology and functions of low threshold K(+) channels with implications for neurological disease.

The diversity of pore-forming subunits of KV1 channels (KV1.1-KV1.8) affords their physiological versatility and predicts a range of functional impairments resulting from genetic aberrations. Curiously, identified so far human neurological conditions associated with dysfunctions of KV1 channels have been linked exclusively to mutations in the KCNA1 gene encoding for the KV1.1 subunit. The absence of phenotypes related to irregularities in other subunits, including the prevalent KV1.2 subunit of neurons is highly perplexing given that deletion of the corresponding kcna2 gene in mouse models precipitates symptoms reminiscent to those of KV1.1 knockouts. Herein, we critically evaluate the molecular and biophysical characteristics of the KV1.1 protein in comparison with others and discuss their role in the greater penetrance of KCNA1 mutations in humans leading to the neurological signs of episodic ataxia type 1 (EA1). Future research and interpretation of emerging data should afford new insights towards a better understanding of the role of KV1.1 in integrative mechanisms of neurons and synaptic functions under normal and disease conditions.

Clinical heterogeneity associated with KCNA1 mutations include cataplexy and nonataxic presentations

Mutations in the KCNA1 gene are known to cause episodic ataxia/myokymia syndrome type 1 (EA1). Here, we describe two families with unique presentations who were enrolled in an IRB-approved study, extensively phenotyped, and whole exome sequencing (WES) performed. Family 1 had a diagnosis of isolated cataplexy triggered by sudden physical exertion in multiple affected individuals with heterogeneous neurological findings. All enrolled affected members carried a KCNA1 c.941T>C (p.I314T) mutation. Family 2 had an 8-year-old patient with muscle spasms with rigidity for whom WES revealed a previously reported heterozygous missense mutation in KCNA1 c.677C>G (p.T226R), confirming the diagnosis of EA1 without ataxia. WES identified variants in KCNA1 that explain both phenotypes expanding the phenotypic spectrum of diseases associated with mutations of this gene. KCNA1 mutations should be considered in patients of all ages with episodic neurological phenotypes, even when ataxia is not present. This is an example of the power of genomic approaches to identify pathogenic mutations in unsuspected genes responsible for heterogeneous diseases.

New insights into the pathogenesis and therapeutics of episodic ataxia type 1.

Episodic ataxia type 1 (EA1) is a K+ channelopathy characterized by a broad spectrum of symptoms. Generally, patients may experience constant myokymia and dramatic episodes of spastic contractions of the skeletal muscles of the head, arms, and legs with loss of both motor coordination and balance. During attacks additional symptoms may be reported such as vertigo, blurred vision, diplopia, nausea, headache, diaphoresis, clumsiness, stiffening of the body, dysarthric speech, and difficulty in breathing. These episodes may be precipitated by anxiety, emotional stress, fatigue, startle response or sudden postural changes. Epilepsy is overrepresented in EA1. The disease is inherited in an autosomal dominant manner, and genetic analysis of several families has led to the discovery of a number of point mutations in the voltage-dependent K+ channel gene KCNA1 (Kv1.1), on chromosome 12p13. To date KCNA1 is the only gene known to be associated with EA1. Functional studies have shown that these mutations impair Kv1.1 channel function with variable effects on channel assembly, trafficking and biophysics. Despite the solid evidence obtained on the molecular mechanisms underlying EA1, how these cause dysfunctions within the central and peripheral nervous systems circuitries remains elusive. This review summarizes the main breakthrough findings in EA1, discusses the neurophysiological mechanisms underlying the disease, current therapies, future challenges and opens a window onto the role of Kv1.1 channels in central nervous system (CNS) and peripheral nervous system (PNS) functions.

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

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.

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

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.

Auditory deficits of Kcna1 deletion are similar to those of a monaural hearing impairment

Kv1.1 subunits of low voltage-activated (Kv) potassium channels are encoded by the Kcna1 gene and crucially determine the synaptic integration window to control the number and temporal precision of action potentials in the auditory brainstem of mammals and birds. Prior electrophysiological studies showed that auditory signaling is compromised in monaural as well as in binaural neurons of the auditory brainstem in Kv1.1 knockout mice (Kcna1−/−). Here we examine the behavioral effects of Kcna1 deletion on sensory tasks dependent on either binaural processing (detecting the movement of a sound source across the azimuth), monaural processing (detecting a gap in noise), as well as binaural summation of the acoustic startle reflex (ASR). Hearing thresholds measured by auditory brainstem responses (ABR) do not differ between genotypes, but our data show a much stronger performance of wild type mice (+/+) in each test during binaural hearing which was lost by temporarily inducing a unilateral hearing loss (through short term blocking of one ear) thus remarkably, leaving no significant difference between binaural and monaural hearing in Kcna1−/− mice. These data suggest that the behavioral effect of Kv1.1 deletion is primarily to impede binaural integration and thus to mimic monaural hearing.

Novel phenotype associated with a mutation in the KCNA1(Kv1.1) gene.

Episodic ataxia type 1 (EA1) is an autosomal dominant K+ channelopathy which manifests with short attacks of cerebellar ataxia and dysarthria, and may also show interictal myokymia. Episodes can be triggered by emotional or physical stress, startle response, sudden postural change or fever. Here we describe a 31-year-old man displaying markedly atypical symptoms, including long-lasting attacks of jerking muscle contractions associated with hyperthermia, severe migraine, and a relatively short-sleep phenotype. A single nucleotide change in KCNA1 (c.555C>G) was identified that changes a highly conserved residue (p.C185W) in the first transmembrane segment of the voltage-gated K+ channel Kv1.1. The patient is heterozygous and the mutation was inherited from his asymptomatic mother. Next generation sequencing revealed no variations in the CACNA1A, CACNB4, KCNC3, KCNJ10, PRRT2 or SCN8A genes of either the patient or mother, except for a benign variant in SLC1A3. Functional analysis of the p.C185W mutation in KCNA1 demonstrated a deleterious dominant-negative phenotype where the remaining current displayed slower activation kinetics, subtle changes in voltage-dependence and faster recovery from slow inactivation. Structural modeling also predicts the C185W mutation to be functionally deleterious. This description of novel clinical features, associated with a Kv1.1 mutation highlights a possibly unrecognized relationship between K+ channel dysfunction, hyperthermia and migraine in EA1, and suggests that thorough assessments for these symptoms should be carefully considered for all patients affected by EA1.