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OBJECTIVE: Focal cortical dysplasias (FCDs) are a common cause of drug-resistant focal epilepsy but frequently remain undetected by conventional magnetic resonance imaging (MRI) assessment. The visual detection can be facilitated by morphometric analysis of T1-weighted images, for example, using the Morphometric Analysis Program (v2018; MAP18), which was introduced in 2005, independently validated for its clinical benefits, and successfully integrated in standard presurgical workflows of numerous epilepsy centers worldwide. Here we aimed to develop an artificial neural network (ANN) classifier for robust automated detection of FCDs based on these morphometric maps and probe its generalization performance in a large, independent data set. METHODS: In this retrospective study, we created a feed-forward ANN for FCD detection based on the morphometric output maps of MAP18. The ANN was trained and cross-validated on 113 patients (62 female, mean age ± SD =29.5 ± 13.6 years) with manually segmented FCDs and 362 healthy controls (161 female, mean age ± SD =30.2 ± 9.6 years) acquired on 13 different scanners. In addition, we validated the performance of the trained ANN on an independent, unseen data set of 60 FCD patients (28 female, mean age ± SD =30 ± 15.26 years) and 70 healthy controls (42 females, mean age ± SD = 40.0 ± 12.54 years). RESULTS: In the cross-validation, the ANN achieved a sensitivity of 87.4% at a specificity of 85.4% on the training data set. On the independent validation data set, our method still reached a sensitivity of 81.0% at a comparably high specificity of 84.3%. SIGNIFICANCE: Our method shows a robust automated detection of FCDs and performance generalizability, largely independent of scanning site or MR-sequence parameters. Taken together with the minimal input requirements of a standard T1 image, our approach constitutes a clinically viable and useful tool in the presurgical diagnostic routine for drug-resistant focal epilepsy. ispartof: EPILEPSIA vol:62 issue:4 pages:1005-1021 ispartof: location:United States status: published

Background: Evidence for the comparative teratogenic risk of antiepileptic drugs is insufficient, particularly in relation to the dosage used. Therefore, we aimed to compare the occurrence of major congenital malformations following prenatal exposure to the eight most commonly used antiepileptic drugs in monotherapy. Methods: We did a longitudinal, prospective cohort study based on the EURAP international registry. We included data from pregnancies in women who were exposed to antiepileptic drug monotherapy at conception, prospectively identified from 42 countries contributing to EURAP. Follow-up data were obtained after each trimester, at birth, and 1 year after birth. The primary objective was to compare the risk of major congenital malformations assessed at 1 year after birth in offspring exposed prenatally to one of eight commonly used antiepileptic drugs (carbamazepine, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, topiramate, and valproate) and, whenever a dose dependency was identified, to compare the risks at different dose ranges. Logistic regression was used to make direct comparisons between treatments after adjustment for potential confounders and prognostic factors. Findings: Between June 20, 1999, and May 20, 2016, 7555 prospective pregnancies met the eligibility criteria. Of those eligible, 7355 pregnancies were exposed to one of the eight antiepileptic drugs for which the prevalence of major congenital malformations was 142 (10·3%) of 1381 pregnancies for valproate, 19 (6·5%) of 294 for phenobarbital, eight (6·4%) of 125 for phenytoin, 107 (5·5%) of 1957 for carbamazepine, six (3·9%) of 152 for topiramate, ten (3·0%) of 333 for oxcarbazepine, 74 (2·9%) of 2514 for lamotrigine, and 17 (2·8%) of 599 for levetiracetam. The prevalence of major congenital malformations increased with the dose at time of conception for carbamazepine (p=0·0140), lamotrigine (p=0·0145), phenobarbital (p=0·0390), and valproate (p

Background Surgery is a widely accepted option for the treatment of pharmacoresistant epilepsies of extratemporal origin. Objective To analyze clinical and epileptological results and to provide prognostic factors influencing seizure outcome. Methods This retrospective single-center study comprises a consecutive series of 383 patients, most of whom had an identifiable lesion on MRI, who underwent resective surgery for extratemporal epilepsy. Data including diagnostic modalities, surgical treatment, histopathology, prognostic factors, and epileptological outcome were analyzed. Results Resective procedures were located as follows: frontal (n = 183), parietal (n = 44), occipital (n = 24), and insular (n = 24). In 108 cases resection included more than 1 lobe. Histopatholological evaluation revealed focal cortical dysplasias (n = 178), tumors (n = 110), cavernomas (n = 27), gliosis (n = 42), and nonspecific findings (n = 36). A distinct epileptogenic lesion was detected in 338 (88.7%) patients. After a mean follow-up of 54 mo, 227 (62.5%) patients remained free from disabling seizures (Engel class I), and 178 (49%) were completely seizure free (Engel class Ia). There was no perioperative mortality. Permanent morbidity was encountered in 46 cases (11.8%). The following predictors were significantly associated with excellent seizure outcome (Engel I): lesion visible on magnetic resonance imaging (MRI; P = .02), noneloquent location (P = .01), complete resection of the lesion (P = .001), absence of epileptic activity postoperatively (P = .001), circumscribed histological findings (P = .001), lower age at surgery (P = .008), and shorter duration of epilepsy (P = .02). Conclusion Surgical treatment of extratemporal epilepsy provides satisfying epileptological results with an acceptable morbidity. Best results can be achieved in younger patients with circumscribed MRI lesions, which can be resected completely.

Seizure manifestations may be difficult to describe in words alone. Thus, initially, 24 seizure images were developed to support communication and gain assistance during obtaining the patient's history. Before being used in clinical practice, these seizure images must be investigated for validity and reliability. We tested the images with untrained participants including patients with epilepsy, persons who had witnessed seizures, and participants who had neither had nor witnessed epileptic seizures. The participants filled in a questionnaire evaluating the images twice within 3 days. The participants were asked to choose one of the 2 written descriptions that best matched each seizure image. The validity was assessed using one-proportion z-test. The reliability was assessed by Gwet's AC1. The first analysis showed that the proportion of correctly identified seizure images was higher than 70%, except for 2 images representing dystonia and myoclonus. The dystonia image was modified, and the myoclonus image was removed. In the final evaluation, the seizure images were identified with an overall correctness ratio of 96%. The final AC1 of the seizure images was classified as very high. The final 23 seizure images are proved to be valid and have a high agreement that can be used in clinical practice.

SUMMARY Purpose: To provide an estimate of the frequency of dacrystic seizures in video-electroencephalography (EEG) long-term monitoring units of tertiary referral epilepsy centers and to describe the clinical presentation of dacrystic seizures in relationship to the underlying etiology. Methods: We screened clinical records and video-EEG reports for the diagnosis of dacrystic seizures of all patients admitted for video-EEG long-term monitoring at five epilepsy referral centers in the United States and Germany. Patients with a potential diagnosis of dacrystic seizures were identified, and their clinical charts and video-EEG recordings were reviewed. We included only patients with: (1) stereotyped lacrimation, sobbing, grimacing, yelling, or sad facial expression; (2) long-term video-EEG recordings (at least 12 h); and (3) at least one brain magnetic resonance imaging (MRI) study. Key Findings: Nine patients (four female) with dacrystic seizures were identified. Dacrystic seizures were identified in 0.06–0.53% of the patients admitted for long-term video-EEG monitoring depending on the specific center. Considering our study population as a whole, the frequency was 0.13%. The presence of dacrystic seizures without other accompanying clinical features was found in only one patient. Gelastic seizures accompanied dacrystic seizures in five cases, and a hypothalamic hamartoma was found in all of these five patients. The underlying etiology in the four patients with dacrystic seizures without gelastic seizures was left mesial temporal sclerosis (three

Ictal fear is a semiological feature which is commonly associated with mesial temporal lobe epilepsy. Here, we describe fear as a leading symptom in cryptogenic occipital lobe epilepsy. In a patient with negative MRI findings, intracranial EEG recordings documented a strict correlation between habitual ictal anxiety attacks and both spontaneous and stimulation-induced epileptic activity in a right occipital epileptogenic area with subsequent spreading to the symptomatogenic zone in the amygdala. Circumscribed occipital topectomy led to seizure freedom. Episodes of non-epileptic fear ceased shortly afterwards. This report provides insight into pathways of propagation of epileptic activity, illustrates different etiologies of pathologic fear and underlines the importance of ictal EEG recordings.

Ictal-related cardiac asystole is supposed to be a risk factor for sudden unexpected death in epilepsy (SUDEP). We retrospectively analyzed the occurrence of ictal asystole in 2003 epilepsy patients undergoing long-term video EEG/ECG monitoring from 1/1999 to 6/2010 at the Freiburg epilepsy centre. Seven patients had cardiac arrest with a duration of at least 3s; 6 ictal, one postictal. In all patients, the temporal lobe was involved in ictal activity based on neurophysiological investigations or morphological lesion. Whereas asystole was self-limited in six cases, one patient with insular seizure origin had to undergo cardiopulmonary resuscitation. Interestingly, also patients with a short history of epilepsy, low seizure frequency and under treatment in monotherapy showed episodes of asystole. In all cases, even with brief cardiac arrest, asystole was associated with subsequent EEG flattening.In conclusion, ictal asystole is a rare event even in a population undergoing major changes in antiepileptic medication. Temporal lobe epilepsy was associated with a risk for asystole; cardiac arrest also occurred in patients who, based on their history, might have not been considered at elevated risk for SUDEP.

SUMMARY Purpose: To gain new insights into the clinical presentation, causes, treatment and prognosis of epilepsia partialis continua (EPC), and to develop hypotheses to be tested in a prospective investigation. Methods: In this retrospective multicenter study, all cases were included that fulfilled these criteria: constantly repeated fragments of epileptic seizures, with preserved consciousness, lasting ‡1 h and representing locally restricted motor or sensory epileptic activity. Single episodes were included when they lasted for a minimum of 1 day. EPC with Rasmussen syndrome and acute stroke were excluded. Key Findings: Three time courses with two subtypes each were distinguished, that is, EPC as a solitary event (de novo or in preexistent epilepsy); chronic repetitive nonprogressive EPC (with frequent or rare episodes); and chronic persistent nonprogressive EPC (primarily or evolving out of an episodic course). These were unrelated to etiologies (morphologic lesions 34%, inflammatory 29%, systemic disorders 9%, idiopathic 5%, unknown 23%). Precipitation and inhibition of seizures is a frequent feature of EPC. Levetiracetam and topiramate have improved the possibilities for pharmacotherapy. Topiramate seems to be particularly effective with dysontogenetic etiologies. Significance: The existence of several clearly distinct courses of nonprogressive EPC is a new finding. These distinctions will be further investigated in a prospective study with precise protocols for electroencephalography (EEG), imaging, and other studies. This should better establish the relation of motor and somatosensory EPC; further clarify the relations, pathogenesis, and significance of the different types and their etiologies; and possibly identify more semiologic variants. It should also provide more precise knowledge about therapy and modification of ictogenesis by external stimuli.

Summary Purpose: Characterization of seizure semiology in patients with hypothalamic hamartoma (HH) based on video–electroencephalography (EEG) monitoring (VEM). Methods: We retrospectively analyzed seizure semiology of 31 patients (20 male, mean age 23.5 years) who underwent VEM at the University Hospitals Freiburg or Heidelberg, Germany. Inclusion criteria were magnetic resonance evidence of an HH, no prior surgical or radiosurgical treatment, and at least two video-documented seizures. A total of 263 seizures were included (mean number of seizures/patient 8.5, range 2–10). To analyze age-dependent changes in seizure semiology, patients were grouped into “children” (3–11 years, n = 5), “adolescents” (12–17 years, n = 4), and “adults” (≥18 years, n = 22). Results: According to patient history, gelastic seizures had occurred in all patients, in 74% as the initial seizure type at epilepsy onset. In VEM, epileptic laughter varied from facial grinning to intense contractions of the diaphragm and body shaking. Unilateral motor signs were seen ipsi- and contralaterally to the HH. Tonic seizures were frequent and did not depend on the state of vigilance. Children, in contrast to adults and adolescents, did not show secondarily generalized tonic–clonic seizures, the gelastic component was the dominating and initial semiologic element, and seizures were significantly shorter. Conclusion: Seizure semiology is highly variable and age dependent. This may reflect network modulations with different propagation of ictal activity and/or secondary epileptogenesis. Detailed knowledge about such changes may contribute to both earlier recognition of seizures during childhood and better assignment of seizure types to a hypothalamic origin.

To assess latencies to the first typical generalized spike-wave discharge (GSWD) and clinically manifest seizure during long-term video-EEG monitoring (VEM) in patients with idiopathic generalized epilepsy (IGE). This is a retrospective analysis of continuous long-term VEM of 39 patients (25 women; mean age, 28.7 years). Mean duration of VEM was 3 days (1-11 days). Latencies from start of VEM to the first appearance of GSWD and the first clinically manifest seizure were analyzed for IGE subsyndrome. Overall, mean latency from the beginning of VEM to the first typical GSWD was 853 minutes (range, 3-7,305 minutes). In 38.5% of the patients, the first typical GSWD occurred during the first hour of VEM and in 87.2% during the first day. Latencies were significantly shorter in juvenile absence epilepsy than in juvenile myoclonic epilepsy, epilepsy with generalized tonic-clonic seizures only, and IGE not further classified (P = 0.001). In 38.5% of the patients, clinically manifest seizures were recorded. Overall, mean latency to the first seizure was 1,984 minutes (range, 3-8,123 minutes). There were no significant differences in latencies to the first seizure between IGE syndromes. One day of VEM is sufficient for classification as IGE in the majority of patients. Patients with juvenile absence epilepsy had particularly low latencies to the appearance of the first typical GSWD. There is, however, a group of patients (12.8% in our sample) requiring a prolonged VEM period to achieve sufficient electroclinical evidence for syndromatic classification.

Einer 25-jahrigen Patientin wurde basierend auf dem magnetresonanztomographischen Nachweis eines links frontalen Kavernoms und anamestisch berichteten epileptischen Anfallen mit Kloni des rechten Armes ein epilepsiechirugischer Eingriff empfohlen. Nach Vorstellung im Epilepsiezentrum wies ein prolongiertes Video-EEG-Monitoring nach vollstandigem Absetzen von Valproat generalisierte Spike-wave-Komplexe (3/s) und bilaterale myoklonische Anfalle nach. Der bildgebende Nachweis einer Lasion kann zu einer falschen Interpretation der Anfallssemiologie verfuhren, die wiederum in eine falsche Klassifikation der Epilepsie sowie fehlerhafte und potenziell gefahrliche therapeutische Interventionen mundet. Der Nachweis eines kausalen Zusammenhanges zwischen Lasion und Epilepsie durch iktale Video-EEG-Aufzeichnungen vor einem epilepsiechirurgischen Eingriff ist unerlasslich.

ICln is an ion channel identified by expression cloning using a cDNA library from Madin-Darby canine kidney cells. In all organisms tested so far, only one transcript for the ICln protein could be identified. Here we show that two splice variants of the ICln ion channel can be found in Caenorhabditis elegans. Moreover, we show that these two splice variants of the ICln channel protein, which we termed IClnN1 and IClnN2, can be functionally reconstituted and tested in an artificial lipid bilayer. In these experiments, the IClnN1-induced currents showed no voltage-dependent inactivation, whereas the IClnN2-induced currents fully inactivated at positive potentials. The molecular entity responsible for the voltage-dependent inactivation of IClnN2 is a cluster of positively charged amino acids encoded by exon 2a, which is absent in IClnN1. Our experiments suggest a mechanism of channel inactivation that is similar to the “ball and chain” model proposed for the Shaker potassium channel,i.e. a cluster of positively charged amino acids hinders ion permeation through the channel by a molecular and voltage-dependent interaction at the inner vestibulum of the pore. This hypothesis is supported by the finding that synthetic peptides with the same amino acid sequence as the positive cluster can transform the IClnN1-induced current to the current observed after reconstitution of IClnN2. Furthermore, we show that the nematode ICln gene is embedded in an operon harboring two additional genes, which we termed Nx and Ny. Co-reconstitution of Nx and IClnN2 and functional analysis of the related currents revealed a functional interaction between the two proteins, as evidenced by the fact that the IClnN2-induced current in the presence of Nx was no longer voltage-sensitive. The experiments described indicate that the genome organization in nematodes allows an effective approach for the identification of functional partner proteins of ion channels.

To gain new insights into the clinical presentation, causes, treatment and prognosis of epilepsia partialis continua (EPC), and to develop hypotheses to be tested in a prospective investigation.In this retrospective multicenter study, all cases were included that fulfilled these criteria: constantly repeated fragments of epileptic seizures, with preserved consciousness, lasting ≥ 1 h and representing locally restricted motor or sensory epileptic activity. Single episodes were included when they lasted for a minimum of 1 day. EPC with Rasmussen syndrome and acute stroke were excluded.Three time courses with two subtypes each were distinguished, that is, EPC as a solitary event (de novo or in preexistent epilepsy); chronic repetitive nonprogressive EPC (with frequent or rare episodes); and chronic persistent nonprogressive EPC (primarily or evolving out of an episodic course). These were unrelated to etiologies (morphologic lesions 34%, inflammatory 29%, systemic disorders 9%, idiopathic 5%, unknown 23%). Precipitation and inhibition of seizures is a frequent feature of EPC. Levetiracetam and topiramate have improved the possibilities for pharmacotherapy. Topiramate seems to be particularly effective with dysontogenetic etiologies.The existence of several clearly distinct courses of nonprogressive EPC is a new finding. These distinctions will be further investigated in a prospective study with precise protocols for electroencephalography (EEG), imaging, and other studies. This should better establish the relation of motor and somatosensory EPC; further clarify the relations, pathogenesis, and significance of the different types and their etiologies; and possibly identify more semiologic variants. It should also provide more precise knowledge about therapy and modification of ictogenesis by external stimuli.

Characterization of seizure semiology in patients with hypothalamic hamartoma (HH) based on video-electroencephalography (EEG) monitoring (VEM).We retrospectively analyzed seizure semiology of 31 patients (20 male, mean age 23.5 years) who underwent VEM at the University Hospitals Freiburg or Heidelberg, Germany. Inclusion criteria were magnetic resonance evidence of an HH, no prior surgical or radiosurgical treatment, and at least two video-documented seizures. A total of 263 seizures were included (mean number of seizures/patient 8.5, range 2-10). To analyze age-dependent changes in seizure semiology, patients were grouped into "children" (3-11 years, n = 5), "adolescents" (12-17 years, n = 4), and "adults" (≥18 years, n = 22).According to patient history, gelastic seizures had occurred in all patients, in 74% as the initial seizure type at epilepsy onset. In VEM, epileptic laughter varied from facial grinning to intense contractions of the diaphragm and body shaking. Unilateral motor signs were seen ipsi- and contralaterally to the HH. Tonic seizures were frequent and did not depend on the state of vigilance. Children, in contrast to adults and adolescents, did not show secondarily generalized tonic-clonic seizures, the gelastic component was the dominating and initial semiologic element, and seizures were significantly shorter.Seizure semiology is highly variable and age dependent. This may reflect network modulations with different propagation of ictal activity and/or secondary epileptogenesis. Detailed knowledge about such changes may contribute to both earlier recognition of seizures during childhood and better assignment of seizure types to a hypothalamic origin.

Ictal laughter is the cardinal clinical sign of gelastic seizures in hypothalamic hamartomas and may also occur in extrahypothalamic epilepsies. Laughing consists of an affective and a motor component. It has been suggested that the affective component may result from an involvement of temporobasal structures, whereas the motor part is related to an involvement of the mesial frontal cortex. So far, evidence is based on a limited number of cases with spontaneously recorded seizures or in whom electrical stimulation of invasive intracranial EEG recordings has been performed. We report a patient who suffered from epigastric psychic auras, complex partial seizures with a gelastic component and secondarily generalized seizures. To evaluate a possible epileptogenic role of the hippocampus and dysplastic region in the right mid-temporal gyrus, intracranial monitoring with subdural electrodes over the temporobasal and temporolateral regions, as well as a deep brain electrode in the hippocampus, were performed. During the intial part of the seizure, consisting of an intense retrosternal ascending feeling with sexual connotation, rhythmic spikes in temporolateral contacts were detected. Concomitant with the development of smiling and laughter, a rhythmic activity over the temporobasal regions evolved. The patient became seizure-free following right temporal lobe resection. This case supports the assumption that ictal involvement of temporobasal structures is crucial for gelastic seizure components in patients with temporal lobe epilepsy. Progression to temporobasal regions was associated with the feeling of happiness whereas motor components of laughter occurred later. These findings are in accordance with the interpretation of surface recordings by Dericioglu and co-workers in a similar previous case. [Published with video sequences].

Epileptische Angstauren (AA) konnen typische Panikattacken imitieren und somit zu einem Verzug in der korrekten Therapie der Epilepsie fuhren. Die zentrale Bedeutung der Amygdala fur iktale AA sowie deren Rolle als fruhes Symptom bei mesiotemporalen Epilepsien wurde gezeigt. Wir berichten uber einen Patienten, bei welchem im Oberflachen- sowie im invasiven Video-EEG Panikattacken im Sinne epileptischer AA abgeleitet wurden, die von einem epileptogenen Focus rechts occipito-lateral ihren Ursprung nahmen. Der 47-jahrige Patient litt seit seinem 16. LJ an einer Epilepsie. Initiale grands maux ohne einleitende fokale Anfalle sind infolge der Verknupfung mit einem exzessiven Alkoholabusus moglicherweise als Alkoholanfalle einzustufen. Im Verlauf etablierte sich eine Semiologie mit leichteren und schwereren Angstgefuhlen einer Frequenz von 1–3/d, durch AA eingeleiteten komplex fokalen Anfallen und sekundar generalisiert tonisch-klonischen Anfallen nur in Phasen ohne adaquate Medikamentenspiegel. MR-tomographisch ergaben sich keine Hinweise auf eine epileptogene Lasion. Das FDG-PET zeigte einen Glucose-Hypometabolismus rechts temporo-basal, das interiktale und iktale SPECT eine Hypoperfusion in derselben Region, gut vereinbar mit der Anfallssemiologie und einer iktalen Involvierung der Amygdala. Gegen diese sich semiologisch und in der funktionellen Bildgebung aufdrangende Hypothese einer Epileptogenese temporal anterior sprach das Oberflachen-EEG mit einem Schwerpunkt der interiktalen und iktalen Aktivitat rechts temporal-posterior-occipital. Im invasiven Video-EEG-Monitoring mit subduralen Elektroden sowie einer Tiefenelektrode zu rechter Amygdala und Hippocampus zeigte sich ein primarer Anfallsursprung occipito-lateral mit rascher Propagation der epileptischen Aktivitat nach temporo-mesial. Mittels Elektrostimulation an occipitalen und an amygdalaren Kontakten liesen sich habituelle AA auslosen. Uber einen Zeitraum von 12 Monaten nach Topektomie rechts occipito-lateral besteht Anfallsfreiheit. Die Histologie zeigte eine FCD 1a. Aufschlussreich ist zum einen die fehlende raumliche Uberschneidung von mittels invasivem Monitoring und durch die postoperative Anfallsfreiheit gesicherter occipitaler Anfallsursprungszone und symptomatogener Zone temporal anterior, zum anderen die enge Korrelation zwischen Wahrnehmung des Patienten und Ausdehnung iktaler Aktivitat. Panikattacken konnen durch einen occipitalen epileptogenen Focus getriggert werden.

Complex partial status epilepticus is a non-convulsive state of persistent behavioural change from baseline functioning, with continuous or intermittent ictal epileptic EEG discharges.1–3 Although relatively common, it is often difficult to diagnose. Treatment with widely available antiepileptic drugs is of limited success and can have relevant side effects.1 4 Here we report the use of a newly released intravenous formula of levetiracetam in a patient who, despite intravenous loading with phenytoin and oral administration of lorazepam, developed complex partial status epilepticus following a series of generalised tonic–clonic seizures. Video EEG monitoring showed resolution of mental clouding and a reduction in ictal EEG patterns within 35 min of administration of levetiracetam. ### Patient history This 29-year-old female patient suffered from cryptogenic focal epilepsy since the age of 11 years. No typical risk factors for the development of epilepsy were present. In particular, there was no evidence of birth related injury, or neonatal or febrile convulsions. Her psychosocial, motor and cognitive development was normal until the onset of epilepsy and concomitant mental retardation. Seizures had been refractory to treatment with primidone, phenytoin, phenytouride, sultiam, topiramate, carbamazepine, oxcarbazepine, lamotrigine, zonisamide, pregabalin and valproate. Seizure types were complex partial seizures as well as tonic and generalised tonic–clonic seizures. Earlier oral treatment with valproic acid resulted …

ICln is an ion channel identified by expression cloning using a cDNA library from Madin-Darby canine kidney cells. In all organisms tested so far, only one transcript for the ICln protein could be identified. Here we show that two splice variants of the ICln ion channel can be found in Caenorhabditis elegans. Moreover, we show that these two splice variants of the ICln channel protein, which we termed IClnN1 and IClnN2, can be functionally reconstituted and tested in an artificial lipid bilayer. In these experiments, the IClnN1-induced currents showed no voltage-dependent inactivation, whereas the IClnN2-induced currents fully inactivated at positive potentials. The molecular entity responsible for the voltage-dependent inactivation of IClnN2 is a cluster of positively charged amino acids encoded by exon 2a, which is absent in IClnN1. Our experiments suggest a mechanism of channel inactivation that is similar to the "ball and chain" model proposed for the Shaker potassium channel, i.e. a cluster of positively charged amino acids hinders ion permeation through the channel by a molecular and voltage-dependent interaction at the inner vestibulum of the pore. This hypothesis is supported by the finding that synthetic peptides with the same amino acid sequence as the positive cluster can transform the IClnN1-induced current to the current observed after reconstitution of IClnN2. Furthermore, we show that the nematode ICln gene is embedded in an operon harboring two additional genes, which we termed Nx and Ny. Co-reconstitution of Nx and IClnN2 and functional analysis of the related currents revealed a functional interaction between the two proteins, as evidenced by the fact that the IClnN2-induced current in the presence of Nx was no longer voltage-sensitive. The experiments described indicate that the genome organization in nematodes allows an effective approach for the identification of functional partner proteins of ion channels.

The ICln protein is expressed ubiquitously in mammals. Experiments designed to knock down the ICln protein in NIH 3T3 fibroblasts as well as in epithelial cells led to the conclusion that this protein is crucially involved in volume regulation after cytoplasmic swelling. Reconstitution of the ICln protein in lipid bilayers revealed the ion channel nature of ICln. Here we describe a new human promoter sequence, composed of 89 nucleotides, which is responsible for a highly constitutive expression of the ICln protein. The promoter sequence lacks a TATA box, and the transcription can be effected at multiple sites. In addition to the starting sites, upstream sequence elements are mandatory for an efficient transcription of the ICln gene (CLNS1A). These new nucleotide elements were defined by site-directed mutagenesis.

Expression cloning revealed a chloride channel (ICln) that we found to be fundamental for the regulatory volume decrease in a variety of cells. The chromosomal localization of the human ICln-gene showed two loci, one at chromosome 11 in position q13.5–q14.1, termed CLNS1A, and a second one at chromosome 6 in position p12.1–q13, termed CLNS1B. In this study, we offer a detailed characterization of the CLNS1A gene and provide the exact position (6p12) and sequence data of CLNS1B, an intronless gene 91.3% homologous to the coding region of CLNS1A.