In previous studies, some channels, called the “sleeper channels,” were reported to contribute to the shaping of the action potential (AP) only under non-physiological conditions. These channels have been hypothesized to play a role in providing a protective mechanism to prevent damage from neuronal hyperexcitation. Here we applied two-electrode current clamp at the primary branch point (1°BP) and the presynaptic terminal simultaneously on crayfish axons. Cadmium had minimal effects on AP shaping, suggesting the absence of calcium-activated potassium channels. Application of 1 mM TEA had minimal impact on AP waveform. In the presence of 4-Aminopyridine (4-AP), the same tetraethylammonium (TEA) concentration significantly prolonged AP duration, resembling the behaviors of sleeper channels. The kinetics of the TEA-sensitive channel (Kv(TEA)) is similar to the Kv2 family of mammalian K+ channels. TEA depolarized the potential after an AP and increased the AP duration in a dose-dependent manner, indicating that these channels contributed to maintaining AP waveform majorly during the hyperpolarization. The terminals were more sensitive to the blockers, suggesting a probability of regulation on neurotransmitter release. However, the TEA-sensitive channels at the crayfish axon had a higher affinity to TEA than the Kv2 channels. Pharmacological profiles, spatial distinction and function of the Kv(TEA) in the crayfish axon require further study.

No description available.

Voltage-gated K+ (Kv) channels are known to cause serious disease upon their malfunction. Kv channels desensitised to voltage show inability to fully repolarise the membrane in excitable cells, which can make the membrane hyperexcited and in turn cause seizures such as in epilepsy, periodic ataxia or heart arrhythmia. Therefore, enhancers of Kv channels could serve as potential drugs. Some of these enhancers are polyunsaturated fatty acids and resin-acids which bind at the proteinlipid surface and affect the movement of the voltage sensor in the channel by a mechanism called the lipoelectric effect. To explore the lipoelectric modulation mechanism, we have performed an extensive computational study including docking and molecular dynamics simulations on resin-acid derivatives added to a model potassium channel called Shaker. Four derivatives, Wu32 and Wu50 that excite the channel and thus induce repolarisation of the membrane, as well as Wu18 and Wu27, who were found to be non-potent in previous experimental studies, have helped to point out a novel binding site in Shaker. The site is located between the pore and voltage-sensing domain of the channel and is in direct contact with the first gating charge arginine, R1, and the residue W454. We hypothesize that it is possible for resinacid derivatives to directly bind to the voltage-sensor when it is in an activated state, prolonging the time Shaker stays open. Further experimental studies on Shaker and human homologs are now needed to test our hypothesis. Therefore, we suggest recording the sensitivity of Shaker towards potent derivatives in combination with mutations of W454. If our findings of the novel binding site are correct, the suitability of Shaker as a model system for human Kv channel modulation by lipoelectric modulators can be questioned as W454 is replaced by small hydrophobic side chains in mammalian Shaker homologs.

No description available.

Pathogenic variants in KCNA2, encoding for the voltage-gated potassium channel Kv1.2, have been identified as the cause for an evolving spectrum of neurological disorders. Affected individuals show early-onset developmental and epileptic encephalopathy, intellectual disability, and movement disorders resulting from cerebellar dysfunction. In addition, individuals with a milder course of epilepsy, complicated hereditary spastic paraplegia, and episodic ataxia have been reported. By analyzing phenotypic, functional, and genetic data from published reports and novel cases, we refine and further delineate phenotypic as well as functional subgroups of KCNA2-associated disorders. Carriers of variants, leading to complex and mixed channel dysfunction that are associated with a gain- and loss-of-potassium conductance, more often show early developmental abnormalities and an earlier onset of epilepsy compared to individuals with variants resulting in loss- or gain-of- function. We describe seven additional individuals harboring three known and the novel KCNA2 variants p.(Pro407Ala) and p.(Tyr417Cys). The location of variants reported here highlights the importance of the proline(405)–valine(406)–proline(407) (PVP) motif in transmembrane domain S6 as a mutational hotspot. A novel case of self-limited infantile seizures suggests a continuous clinical spectrum of KCNA2-related disorders. Our study provides further insights into the clinical spectrum, genotype–phenotype correlation, variability, and predicted functional impact of KCNA2 variants.

Les ataxies épisodiques (EA) d’origine génétique sont un groupe de maladies possédant un phénotype et génotype hétérogènes, mais ont en commun la caractéristique d’un dysfonctionnement cérébelleux intermittent. Les EA de type 1 et 2 sont les plus largement reconnues des ataxies épisodiques autosomiques dominantes et sont causées par un dysfonctionnement des canaux ioniques voltage-dépendants dans les neurones. La présente étude se concentrera sur les mutations causant l'EA-1, retrouvées dans le senseur de voltage (VSD) de Kv1.1, un canal très proche de la famille des canaux Shaker. Nous avons caractérisé les propriétés électrophysiologiques de six mutations différentes à la position F244 et partiellement celles des mutations T284 A/M, R297 K/Q/A/H, I320T, L375F, L399I et S412 C/I dans la séquence du Shaker grâce à la technique du ‘’cut open voltage clamp’’ (COVC). Les mutations de la position F244 situées sur le S1 du canal Shaker sont caractérisées par un décalement des courbes QV et GV vers des potentiels dépolarisants et modifient le couplage fonctionnel entre le domaine VSD et le pore. Un courant de fuite est observé durant la phase d'activation des courants transitoires et peut être éliminé par l'application du 4-AP (4-aminopyridine) ou la réinsertion de l'inactivation de type N mais pas par le TEA (tétraéthylamonium). Dans le but de mieux comprendre les mécanismes moléculaires responsables de la stabilisation d’un état intermédiaire, nous avons étudié séparément la neutralisation des trois premières charges positives du S4 (R1Q, R2Q et R3Q). Il en est ressorti l’existence d’une interaction entre R2 et F244. Une seconde interface entre S1 et le pore proche de la surface extracellulaire agissant comme un second point d'ancrage et responsable des courants de fuite a été mis en lumière. Les résultats suggèrent une anomalie du fonctionnement du VSD empêchant la repolarisation normale de la membrane des cellules nerveuses affectées à la suite d'un potentiel d'action. / The genetic episodic ataxias form a group of disorders with heterogeneous phenotype and genotype, but share the common feature of intermittent cerebellar dysfunction. Episodic ataxia (EA) types 1 and 2 are most widely recognised amongst the autosomal dominant episodic ataxias and are caused by dysfunction of neuronal voltage-gated ion channels. The present study focuses on mutations causing EA-1 located in the voltage sensor domains (VSDs) of Kv1.1. A member of the Shaker channel family. Here, we have characterised the electrophysiological properties of six different mutations at the position of F244 and we also reported the partiality effects of these following mutations T284A/M, R297K/Q/A/H, I320T, L375F, L399I S412C/I on Shaker sequence using the cut open voltage clamp technique (COVC). We have shown that mutations of F244 in the S1 of the Shaker Kv channel positively shift the voltage dependence of the VSD movement and alter functional coupling between VSD and pore domain. The mutations causing immobilization of the VSD movement during activation and deactivation and responsible for creating a leak current during activation, are removed by the application of 4-AP (4-aminopyridine) or by reinsertion of N-type inactivation but not by TEA (tetraethylamonium). Insights into the molecular mechanisms responsible for the stabilization of the intermediate state have been investigated by separately neutralizing the first three charges (R1Q, R2Q and R3Q) in the S4 segment. The result suggests an interaction between R2 and F244 mutants. It was established that a second co-evolved interface exists between S1 and the pore helix near the extracellular surface and it acts as a second anchor point. It is also responsible for generation of leak currents. The results suggest a dysfunction of the VSD in which the affected nerve cells cannot efficiently repolarize following an action potential because of altered delayed rectifier function

Les ataxies épisodiques (EA) d’origine génétique sont un groupe de maladies possédant un phénotype et génotype hétérogènes, mais ont en commun la caractéristique d’un dysfonctionnement cérébelleux intermittent. Les EA de type 1 et 2 sont les plus largement reconnues des ataxies épisodiques autosomiques dominantes et sont causées par un dysfonctionnement des canaux ioniques voltage-dépendants dans les neurones. La présente étude se concentrera sur les mutations causant l'EA-1, retrouvées dans le senseur de voltage (VSD) de Kv1.1, un canal très proche de la famille des canaux Shaker. Nous avons caractérisé les propriétés électrophysiologiques de six mutations différentes à la position F244 et partiellement celles des mutations T284 A/M, R297 K/Q/A/H, I320T, L375F, L399I et S412 C/I dans la séquence du Shaker grâce à la technique du ‘’cut open voltage clamp’’ (COVC). Les mutations de la position F244 situées sur le S1 du canal Shaker sont caractérisées par un décalement des courbes QV et GV vers des potentiels dépolarisants et modifient le couplage fonctionnel entre le domaine VSD et le pore. Un courant de fuite est observé durant la phase d'activation des courants transitoires et peut être éliminé par l'application du 4-AP (4-aminopyridine) ou la réinsertion de l'inactivation de type N mais pas par le TEA (tétraéthylamonium). Dans le but de mieux comprendre les mécanismes moléculaires responsables de la stabilisation d’un état intermédiaire, nous avons étudié séparément la neutralisation des trois premières charges positives du S4 (R1Q, R2Q et R3Q). Il en est ressorti l’existence d’une interaction entre R2 et F244. Une seconde interface entre S1 et le pore proche de la surface extracellulaire agissant comme un second point d'ancrage et responsable des courants de fuite a été mis en lumière. Les résultats suggèrent une anomalie du fonctionnement du VSD empêchant la repolarisation normale de la membrane des cellules nerveuses affectées à la suite d'un potentiel d'action. / The genetic episodic ataxias form a group of disorders with heterogeneous phenotype and genotype, but share the common feature of intermittent cerebellar dysfunction. Episodic ataxia (EA) types 1 and 2 are most widely recognised amongst the autosomal dominant episodic ataxias and are caused by dysfunction of neuronal voltage-gated ion channels. The present study focuses on mutations causing EA-1 located in the voltage sensor domains (VSDs) of Kv1.1. A member of the Shaker channel family. Here, we have characterised the electrophysiological properties of six different mutations at the position of F244 and we also reported the partiality effects of these following mutations T284A/M, R297K/Q/A/H, I320T, L375F, L399I S412C/I on Shaker sequence using the cut open voltage clamp technique (COVC). We have shown that mutations of F244 in the S1 of the Shaker Kv channel positively shift the voltage dependence of the VSD movement and alter functional coupling between VSD and pore domain. The mutations causing immobilization of the VSD movement during activation and deactivation and responsible for creating a leak current during activation, are removed by the application of 4-AP (4-aminopyridine) or by reinsertion of N-type inactivation but not by TEA (tetraethylamonium). Insights into the molecular mechanisms responsible for the stabilization of the intermediate state have been investigated by separately neutralizing the first three charges (R1Q, R2Q and R3Q) in the S4 segment. The result suggests an interaction between R2 and F244 mutants. It was established that a second co-evolved interface exists between S1 and the pore helix near the extracellular surface and it acts as a second anchor point. It is also responsible for generation of leak currents. The results suggest a dysfunction of the VSD in which the affected nerve cells cannot efficiently repolarize following an action potential because of altered delayed rectifier function