Depolarization Shifts Research Articles

The Role of Butyrate in Paclitaxel and Morphine Induced Peripheral Hypersensitivity

Recent evidence implicates the gut microbiome in peripheral hypersensitivity in chronic opioid use models, and in chemotherapeutic‐induced hypersensitivity in mice. An important constituent of the commensal microbiome are bacterial species that ferment fibers through complex enzymatic pathways and produce short chain fatty acids, acetate, propionate, and butyrate. Butyrate is known to improve the integrity of the epithelium and enhance antimicrobial peptides thus preventing colonization by pathogenic bacteria, as well as having an established role in reducing inflammation through the inhibition of histone deacetylase. Based on this, we hypothesized that sodium butyrate can relieve peripheral hypersensitivity as well as reduce primary nociceptor hyperexcitability in experimental models of peripheral hypersensitivity. We used both paclitaxel (PAC) and morphine (MOR) animal models, which were organized into the following treatment groups: PAC (8mg/kg i.p q.d, total 4 injections) ± Sodium Butyrate (10mg/kg i.p b.i.d), MOR (escalating from 20mg/kg i.p b.i.d on day 1 to 80mg/kg i.p b.i.d on day 4) ± Sodium Butyrate (10mg/kg i.p b.i.d). PAC animals were assessed for peripheral cold hypersensitivity using an acetone evaporation assay. MOR‐treated animals were assayed using a hot‐plate test for opioid‐induced hyperalgesia. Primary nociceptors from L4‐S1 dorsal root ganglia were collected from these mice for whole‐cell patch clamp electrophysiological recordings to assess for enhanced excitability. PAC‐treated mice developed significant cold hypersensitivity 7 days post PAC, when compared with vehicle (average time engaging stimulated paw 2.0s vs 8.6s respectively). Butyrate ameliorated this paclitaxel‐induced hypersensitivity (average time reduced to 2.5s). Similarly, MOR induced thermal hypersensitivity (Hot‐plate latency of 14.8s vs 28.16s saline), which was also reversed by sodium butyrate (Hot‐plate latency of 24.84s). Whole‐cell patch clamp recordings revealed that PAC and MOR‐treated neurons fired a greater number of regenerating action potentials compared to respective controls (PAC: 4 vs 2; MOR: 3.8 vs 2.1), indicating enhanced excitability. In‐vivo butyrate treatment attenuated this enhanced excitability. Hysteresis plots of hyperexcitable cells revealed a depolarizing shift in the membrane potential of regenerating action potentials. Sodium butyrate treatment recovered this shift in membrane potential within the repetitive action potentials. Therefore, similar to the in‐vivo peripheral hypersensitivity, butyrate reversed enhanced excitability of PAC and MOR‐treated DRG neurons. These findings demonstrate that the short chain fatty acid, butyrate, a major metabolite of the gut microbiome plays an important role in preventing the development of drug‐induced peripheral hypersensitivity.

Kinetic Alterations in Resurgent Sodium Currents of Mutant Nav1.4 Channel in Two Patients Affected by Paramyotonia Congenita

Simple SummaryParamyotonia congenita (PMC) is a rare muscle disorder that causes myotonia and weakness of facial and limb muscles. The electromyography in PMC shows continual spontaneous, high-frequency spike potentials in skeletal muscles. Genetic mutations in the Nav1.4 channel that cause hyperexcitability of muscle fibers are responsible for PMC. However, the genotype–phenotype relationship is highly diversified, and the molecular pathology remains unclear. Here, we investigated the electrophysiology in the Nav1.4 channel with mutations, p.V781I and p.A1737T, which were found in two Taiwanese patients. We identified the distinct changes in gating mechanisms altered by mutations which may underlie the clinical phenotype.Paramyotonia congenita (PMC) is a rare skeletal muscle disorder characterized by muscle stiffness upon repetitive exercise and cold exposure. PMC was reported to be caused by dominant mutations in the SCN4A gene encoding the α subunit of the Nav1.4 channel. Recently, we identified two missense mutations of the SCN4A gene, p.V781I and p.A1737T, in two PMC families. To evaluate the changes in electrophysiological properties caused by the mutations, both mutant and wild-type (WT) SCN4A genes were expressed in CHO-K1 and HEK-293T cells. Then, whole-cell patch-clamp recording was employed to study the altered gating of mutant channels. The activation curve of transient current showed a hyperpolarizing shift in both mutant Nav1.4 channels as compared to the WT channel, whereas there was a depolarizing shift in the fast inactivation curve. These changes confer to an increase in window current in the mutant channels. Further investigations demonstrated that the mutated channel proteins generate significantly larger resurgent currents as compared to the WT channel and take longer to attain the peak of resurgent current than the WT channel. In conclusion, the current study demonstrates that p.V781I and p.A1737T mutations in the Nav1.4 channel increase both the sustained and the resurgent Na+ current, leading to membrane hyperexcitability with a lower firing threshold, which may influence the clinical phenotype.

Pathological Networks Involving Dysmorphic Neurons in Type II Focal Cortical Dysplasia.

Focal cortical dysplasia (FCD) is one of the most common causes of drug-resistant epilepsy. Dysmorphic neurons are the major histopathological feature of type II FCD, but their role in seizure genesis in FCD is unclear. Here we performed whole-cell patch-clamp recording and morphological reconstruction of cortical principal neurons in postsurgical brain tissue from drug-resistant epilepsy patients. Quantitative analyses revealed distinct morphological and electrophysiological characteristics of the upper layer dysmorphic neurons in type II FCD, including an enlarged soma, aberrant dendritic arbors, increased current injection for rheobase action potential firing, and reduced action potential firing frequency. Intriguingly, the upper layer dysmorphic neurons received decreased glutamatergic and increased GABAergic synaptic inputs that were coupled with upregulation of the Na+-K+-Cl- cotransporter. In addition, we found a depolarizing shift of the GABA reversal potential in the CamKII-cre::PTENflox/flox mouse model of drug-resistant epilepsy, suggesting that enhanced GABAergic inputs might depolarize dysmorphic neurons. Thus, imbalance of synaptic excitation and inhibition of dysmorphic neurons may contribute to seizure genesis in type II FCD.

The Effect of Valproate Sodium on the Electrical Activity of Helix aspersa F1 Neurons in a Pentylenetetrazole-Induced Epileptic Model Using an Intracellular Recording System

Background: Epilepsy is a chronic central nervous system disorder with a high prevalence in modern society. Despite using common anticonvulsant drugs, its control is not adequately achieved. Animal models for seizures play the leading role in advancing our understanding of the cellular mechanisms of epilepsy. The present study was an attempt to elucidate the electrophysiological mechanism of the effect of sodium valproate on the cellular model of epilepsy. Understanding the cellular mechanisms of this drug may help clarify the pharmacological screening of other drugs. Methods: The intracellular recording was made from F1 cells of garden Helix aspersa in the presence of Ringer solution. Following the extracellular application of valproate sodium at a concentration of 10 mM after and before the use of (25mM) epileptogenic agent (pentylenetetrazol (PTZ)), we evaluated its effect on paroxysmal depolarization shift (PDS) and electrophysiological characteristics. Results: These results showed that valproate sodium could reduce neuronal excitability. It could significantly hyperpolarize rest action potential by decreasing the frequency of firing rate and increasing the amplitude of afterhyperpolarization (AHP) and can prevent depolarization of rest action potential by PTZ. Conclusions: The results suggested that valproate sodium could reduce the PTZ-induced hyperexcitation by hyperpolarization of resting membrane potential (RMP), a reduction in AHP amplitude, and firing the frequency.

Possible Interactions of Extracellular Loop IVP2-S6 With Voltage-Sensing Domain III in Cardiac Sodium Channel.

Motion transmission from voltage sensors to inactivation gates is an important problem in the general physiology of ion channels. In a cryo-EM structure of channel hNav1.5, residues N1736 and R1739 in the extracellular loop IVP2-S6 approach glutamates E1225 and E1295, respectively, in the voltage-sensing domain III (VSD-III). ClinVar-reported variants E1230K, E1295K, and R1739W/Q and other variants in loops IVP2-S6, IIIS1-S2, and IIIS3-S4 are associated with cardiac arrhythmias, highlighting the interface between IVP2-S6 and VSD-III as a hot spot of disease mutations. Atomic mechanisms of the channel dysfunction caused by these mutations are unknown. Here, we generated mutants E1295R, R1739E, E1295R/R1739E, and N1736R, expressed them in HEK-293T cells, and explored biophysical properties. Mutation E1295R reduced steady-state fast inactivation and enhanced steady-state slow inactivation. In contrast, mutation R1739E slightly enhanced fast inactivation and attenuated slow inactivation. Characteristics of the double mutant E1295R/R1739E were rather similar to those of the wild-type channel. Mutation N1736R attenuated slow inactivation. Molecular modeling predicted salt bridging of R1739E with the outermost lysine in the activated voltage-sensing helix IIIS4. In contrast, the loss-of-function substitution E1295R repelled R1739, thus destabilizing the activated VSD-III in agreement with our data that E1295R caused a depolarizing shift of the G-V curve. In silico deactivation of VSD-III with constraint-maintained salt bridge E1295-R1739 resulted in the following changes: 1) contacts between IIIS4 and IVS5 were switched; 2) contacts of the linker-helix IIIS4-S5 with IVS5, IVS6, and fast inactivation tripeptide IFM were modified; 3) contacts of the IFM tripeptide with helices IVS5 and IVS6 were altered; 4) mobile loop IVP2-S6 shifted helix IVP2 that contributes to the slow inactivation gate and helix IVS6 that contributes to the fast inactivation gate. The likelihood of salt bridge E1295-R1739 in deactivated VSD-III is supported by Poisson–Boltzmann calculations and state-dependent energetics of loop IVP2-S6. Taken together, our results suggest that loop IVP2-S6 is involved in motion transmission from VSD-III to the inactivation gates.

Role of L-Type Voltage-Gated Calcium Channels in Epileptiform Activity of Neurons.

Epileptic discharges manifest in individual neurons as abnormal membrane potential fluctuations called paroxysmal depolarization shift (PDS). PDSs can combine into clusters that are accompanied by synchronous oscillations of the intracellular Ca2+ concentration ([Ca2+]i) in neurons. Here, we investigate the contribution of L-type voltage-gated calcium channels (VGCC) to epileptiform activity induced in cultured hippocampal neurons by GABA(A)R antagonist, bicuculline. Using KCl-induced depolarization, we determined the optimal effective doses of the blockers. Dihydropyridines (nifedipine and isradipine) at concentrations ≤ 10 μM demonstrate greater selectivity than the blockers from other groups (phenylalkylamines and benzothiazepines). However, high doses of dihydropyridines evoke an irreversible increase in [Ca2+]i in neurons and astrocytes. In turn, verapamil and diltiazem selectively block L-type VGCC in the range of 1–10 μM, whereas high doses of these drugs block other types of VGCC. We show that L-type VGCC blockade decreases the half-width and amplitude of bicuculline-induced [Ca2+]i oscillations. We also observe a decrease in the number of PDSs in a cluster and cluster duration. However, the pattern of individual PDSs and the frequency of the cluster occurrence change insignificantly. Thus, our results demonstrate that L-type VGCC contributes to maintaining the required [Ca2+]i level during oscillations, which appears to determine the number of PDSs in the cluster.

P-EP028. Low-dose perampanel improves refractory cortical myoclonus by the dispersed and suppressed paroxysmal depolarization shifts in the sensorimotor cortex

Introduction. Although low-dose perampanel (PER) is reportedly effective on cortical myoclonus in a limited number of patients, its underlying mechanisms remain unclear. We aimed to extract and elaborate the clinical effects of PER on refractory cortical myoclonus for dose, etiology and also somatosensory-evoked potential (SEP) findings. Method. We examined 18 epilepsy patients with seizure and cortical myoclonus (10 males; mean age, 48.4 ± 16.2 years; Unverricht-Lundborg disease = 7, benign adult familial myoclonus epilepsy = 6, dentatorubral-pallidoluysian atrophy = 2, Gaucher disease = 1, and Lance-Adams syndrome = 2). Based on data accumulated before and after PER treatment, activities of daily life [ADL]), short latency, cortical components of SEP, and PER blood concentration were analyzed. Results. PER (mean dose: 3.2 ± 2.1 mg/day) significantly improved seizure frequency, myoclonus and ADL scores. It also significantly decreased the amplitude of and prolonged correlate with clinical scores among all patients. In contrast, the degree of P25 prolongation ms) significantly correlated with improved ADL score (p = 0.019) and blood PER concentration (p = 0.011). The degree of N33 prolongation (32.1 ± 4.0 to 33.7 ± 3.4 ms) was positively correlated with ADL score and blood PER concentration (p = 0.025 and p = 0.025, respectively). Conclusion. Low-dose PER markedly improved myoclonus and ADL in patients with refractory cortical myoclonus. SEP, especially P25 latency, is a potentially good biomarker to assess the objective effects of PER on intractable cortical myoclonus. ( https://doi.org/10.1016/j.clinph.2019.07.006 ).

On the Origin of Paroxysmal Depolarization Shifts: The Contribution of Cav1.x Channels as the Common Denominator of a Polymorphous Neuronal Discharge Pattern

Since their discovery in the 1960s, the term paroxysmal depolarization shift (PDS) has been applied to a wide variety of reinforced neuronal discharge patterns. Occurrence of PDS as cellular correlates of electrographic spikes during latent phases of insult-induced rodent epilepsy models and their resemblance to giant depolarizing potentials (GDPs) nourished the idea that PDS may be involved in epileptogenesis. Both GDPs and – in analogy – PDS may lead to progressive changes of neuronal properties by generation of pulsatile intracellular Ca2+ elevations. Herein, a key element is the gating of L-type voltage gated Ca2+ channels (LTCCs, Cav1.x family), which may convey Ca2+ signals to the nucleus. Accordingly, the present study investigates various insult-associated neuronal challenges for their propensities to trigger PDS in a LTCC-dependent manner. Our data demonstrate that diverse disturbances of neuronal function are variably suited to induce PDS-like events, and the contribution of LTCCs is essential to evoke PDS in rat hippocampal neurons that closely resemble GDPs. These PDS appear to be initiated in the dendritic sub-compartment. Their morphology critically depends on the position of recording electrodes and on their rate of occurrence. These results provide novel insight into induction mechanisms, origin, variability, and co-existence of PDS with other discharge patterns and thereby pave the way for future investigations regarding the role of PDS in epileptogenesis.

Elucidating the Structural Constituents of Fibroblast Growth Factor 14 that Confer its Modulatory Effects on the Voltage-Gated Sodium Channel 1.6

Disruption of protein:protein interactions (PPIs) between voltage-gated Na+ (Nav) channels and their auxiliary proteins is implicated in the etiology of channelopathies. Despite restoration of these perturbed PPIs serving as a novel therapeutic approach, such efforts to develop PPI-based pharmacotherapies are hindered by lack of insights into the structural constituents of regulatory proteins that confer their modulatory effects on Nav channel kinetics. Focusing on the PPI between Nav1.6 and its auxiliary protein fibroblast growth factor 14 (FGF14), we previously identified clusters of amino acids on the β12 sheet and β8-β9 loop of FGF14 that are at its PPI interface with the C-terminal domain (CTD) of Nav1.6 and are predicted to disproportionately contribute to the Gibbs energy of their protein:protein binding. To develop chemical probes, we derived peptidomimetics from the PLEV and EYYV motifs of the β12 sheet and β8-β9 loop, respectively. Crucially, a FGF14PLEV derivative caused a depolarizing shift in the V1/2 of Nav1.6 channel activation and steady-state inactivation, but did not affect long-term inactivation. Conversely, a FGF14EYYV derivate caused an increase in the long-term inactivation of Nav1.6 channels, but did not affect the V1/2 of activation or steady-state inactivation, suggesting that the β12 sheet and β8-β9 loop constituents of FGF14 regulate divergent kinetic properties of Nav1.6 channels. Based upon molecular docking of these two derivatives, it is hypothesized that their divergent regulation of Nav1.6 channel kinetics is conferred by differential interactions with the EF hand like (EFL) and IQ domains of the CTD. Exploration of this hypothesis will enable an increasingly refined understanding of Nav channel regulation and provide insights into developing therapeutics targeted against PPI interfaces between Nav channels and their auxiliary proteins.

Molecular mechanisms of action determine inhibition of paroxysmal depolarizing shifts by NMDA receptor antagonists in rat cortical neurons

N-methyl-d-aspartate glutamate receptors (NMDARs) are involved in numerous central nervous system (CNS) processes, including epileptiform activity. We used a picrotoxin-induced epileptiform activity model to compare the action of different types of NMDAR antagonists in rat brain slices. Paroxysmal depolarizing shifts (PDS) were evoked by external stimulation in the medial prefrontal cortex (mPFC) slices and recorded in pyramidal cells (PC) and in fast-spiking interneurons (FSI). The NMDAR antagonists APV and memantine reduced the duration of PDS. However, the competitive antagonist APV caused similar effects on the PC and FSI, while the open-channel blocker memantine had a much stronger effect on the PDS in the FSI than in the PC. This difference cannot be explained by a corresponding difference in NMDAR sensitivity to memantine because the drug inhibited the excitatory postsynaptic current (EPSC) similarly in both cell types. Importantly, the PDS were significantly longer in the FSI than in the PC. The degree of PDS inhibition by memantine correlated with individual PDS durations in each cell type. Computer modeling of a synaptic network in the mPFC suggests that the different effects of memantine on the PDS in the PC and FSI can be explained by use dependence of its action. An open-channel blocking mechanism and competition with Mg2+ ions for the binding site result in pronounced inhibition of the long PDS, whereas the short PDS are weakly sensitive. Our results show that peculiarities of kinetics and the mechanism of action largely determine the effects of NMDAR antagonists on physiological and/or pathological processes.

GABA receptor inhibition and severe hypoxia induce a paroxysmal depolarization shift in goldfish neurons.

Mammalian neurons undergo rapid excitotoxic cell death when deprived of oxygen; however, the common goldfish (Carassius auratus) has the unique ability of surviving in oxygen-free waters, under anoxia. This organism utilizes γ-amino butyric acid (GABA) signaling to suppress excitatory glutamatergic activity during anoxic periods. Although GABAA receptor antagonists are not deleterious to the cellular survival, coinhibition of GABAA and GABAB receptors is detrimental by abolishing anoxia-induced neuroprotective mechanisms. Here we show that blocking the anoxic GABAergic neurotransmission induces seizure-like activity (SLA) analogous to a paroxysmal depolarization shift (PDS), with hyperpolarization of action potential (AP) threshold and elevation of threshold currents. The observed PDS was attributed to an increase in excitatory postsynaptic currents (EPSCs) that are normally attenuated with decreasing oxygen levels. Furthermore, for the first time, we show that in addition to PDS, some neurons undergo depolarization block and do not generate AP despite a suprathreshold membrane potential. In conclusion, our results indicate that with severe hypoxia and absence of GABA receptor activity, telencephalic neurons of C. auratus manifest a paroxysmal depolarization shift, a key feature of epileptic discharge.NEW & NOTEWORTHY This work shows that the combination of anoxia and inhibition of GABA receptors induces seizure-like activities in goldfish telencephalic pyramidal and stellate neurons. Importantly, to prevent seizure-like activity, an intact GABA-mediated inhibitory pathway is required.

Non-polar ZnCdO/ZnO step-barrier quantum wells designed for THz emission

Symmetric and asymmetric quantum wells of ZnCdO structures grown on ZnO in non-polar orientations are investigated for their potential in THz emission due to the intersubband transitions between the subbands in the conduction band. The Schrödinger and Poisson equations are solved self-consistently for the square and step-barrier quantum wells of the Zn1−xCdxO layers for x up to 0.25. The results are represented as functions of the Cd concentration in the well and step-barrier layers and their dimensions. The role of the many-body effects on the intersubband transitions is discussed. It is found that emission wavelengths between 4.5 and 8.55 THz in the step-barrier quantum wells are possible in the case of an optical pumping at a wavelength in the range of 8.27–15.7 μm. The depolarization and excitonic shifts are found to contribute to the bare intersubband transition energies as much as their 15% at maximum.

A Novel Kv7.3 Variant in the Voltage-Sensing S4 Segment in a Family With Benign Neonatal Epilepsy: Functional Characterization and in vitro Rescue by β-Hydroxybutyrate.

Pathogenic variants in KCNQ2 and KCNQ3, paralogous genes encoding Kv7.2 and Kv7.3 voltage-gated K+ channel subunits, are responsible for early−onset developmental/epileptic disorders characterized by heterogeneous clinical phenotypes ranging from benign familial neonatal epilepsy (BFNE) to early−onset developmental and epileptic encephalopathy (DEE). KCNQ2 variants account for the majority of pedigrees with BFNE and KCNQ3 variants are responsible for a much smaller subgroup, but the reasons for this imbalance remain unclear. Analysis of additional pedigrees is needed to further clarify the nature of this genetic heterogeneity and to improve prediction of pathogenicity for novel variants. We identified a BFNE family with two siblings and a parent affected. Exome sequencing on samples from both parents and siblings revealed a novel KCNQ3 variant (c.719T>G; p.M240R), segregating in the three affected individuals. The M240 residue is conserved among human Kv7.2-5 and lies between the two arginines (R5 and R6) closest to the intracellular side of the voltage-sensing S4 transmembrane segment. Whole cell patch-clamp recordings in Chinese hamster ovary (CHO) cells revealed that homomeric Kv7.3 M240R channels were not functional, whereas heteromeric channels incorporating Kv7.3 M240R mutant subunits with Kv7.2 and Kv7.3 displayed a depolarizing shift of about 10 mV in activation gating. Molecular modeling results suggested that the M240R substitution preferentially stabilized the resting state and possibly destabilized the activated state of the Kv7.3 subunits, a result consistent with functional data. Exposure to β-hydroxybutyrate (BHB), a ketone body generated during the ketogenic diet (KD), reversed channel dysfunction induced by the M240R variant. In conclusion, we describe the first missense loss-of-function (LoF) pathogenic variant within the S4 segment of Kv7.3 identified in patients with BFNE. Studied under conditions mimicking heterozygosity, the M240R variant mainly affects the voltage sensitivity, in contrast to previously analyzed BFNE Kv7.3 variants that reduce current density. Our pharmacological results provide a rationale for the use of KD in patients carrying LoF variants in Kv7.2 or Kv7.3 subunits.

Excitons bound by photon exchange

In contrast to interband excitons in undoped quantum wells, doped quantum wells do not display sharp resonances due to excitonic bound states. In these systems the effective Coulomb interaction between electrons and holes typically only leads to a depolarization shift of the single-electron intersubband transitions. Non-perturbative light-matter interaction in solid-state devices has been investigated as a pathway to tune optoelectronic properties of materials. A recent theoretical work [Cortese et al., Optica 6, 354 (2019)] predicted that, when the doped quantum wells are embedded in a photonic cavity, emission-reabsorption processes of cavity photons can generate an effective attractive interaction which binds electrons and holes together, leading to the creation of an intraband bound exciton. Spectroscopically, this bound state manifests itself as a novel discrete resonance which appears below the ionisation threshold only when the coupling between light and matter is increased above a critical value. Here we report the first experimental observation of such a bound state using doped GaAs/AlGaAs quantum wells embedded in metal-metal resonators whose confinement is high enough to permit operation in strong coupling. Our result provides the first evidence of bound states of charged particles kept together not by Coulomb interaction, but by the exchange of transverse photons. Light-matter coupling can thus be used as a novel tool in quantum material engineering, tuning electronic properties of semiconductor heterostructures beyond those permitted by mere crystal structures, with direct applications to mid-infrared optoelectronics.

Isoform-specific regulation of HCN4 channels by a family of endoplasmic reticulum proteins

Ion channels in excitable cells function in macromolecular complexes in which auxiliary proteins modulate the biophysical properties of the pore-forming subunits. Hyperpolarization-activated, cyclic nucleotide-sensitive HCN4 channels are critical determinants of membrane excitability in cells throughout the body, including thalamocortical neurons and cardiac pacemaker cells. We previously showed that the properties of HCN4 channels differ dramatically in different cell types, possibly due to the endogenous expression of auxiliary proteins. Here, we report the discovery of a family of endoplasmic reticulum (ER) transmembrane proteins that associate with and modulate HCN4. Lymphoid-restricted membrane protein (LRMP, Jaw1) and inositol trisphosphate receptor-associated guanylate kinase substrate (IRAG, Mrvi1, and Jaw1L) are homologous proteins with small ER luminal domains and large cytoplasmic domains. Despite their homology, LRMP and IRAG have distinct effects on HCN4. LRMP is a loss-of-function modulator that inhibits the canonical depolarizing shift in the voltage dependence of HCN4 in response to the binding of cAMP. In contrast, IRAG causes a gain of HCN4 function by depolarizing the basal voltage dependence in the absence of cAMP. The mechanisms of action of LRMP and IRAG are independent of trafficking and cAMP binding, and they are specific to the HCN4 isoform. We also found that IRAG is highly expressed in the mouse sinoatrial node where computer modeling predicts that its presence increases HCN4 current. Our results suggest important roles for LRMP and IRAG in the regulation of cellular excitability, as tools for advancing mechanistic understanding of HCN4 channel function, and as possible scaffolds for coordination of signaling pathways.

Electrocardiological effects of ranolazine and lidocaine on normal and diabetic rat atrium.

Cellular changes occurring in diabetic cardiomyopathy include disturbances of calcium and sodium homeostasis. Voltage-gated sodium channels are responsible for the initiation of cardiac action potentials, and the excitability would create relevance. The effect of ranolazine as a sodium channel blocker on atrium electromechanical parameters is investigated and compared with lidocaine in streptozocin-treated diabetic rats. After an 8-week induction of diabetes type I, the effect of cumulative concentrations of ranolazine and lidocaine on the electrophysiology of isolated atrium was studied. Ranolazine's effects were evaluated on cardiac sodium current in normal- and high-glucose medium, with whole-cell patch-clamp technique. Ranolazine at therapeutic concentrations had no significant statistical effect on refractory period in normal and diabetic isolated heart. Ranolazine (10μM) caused a hyperpolarizing shift of V1/2 for steady-state inactivation in normal media, while it significantly elicited a depolarizing shift in high-glucose media (p < 0.05). It is concluded that in the isolated rat atrium preparation, ranolazine and lidocaine have no beneficial on diabetic cardiomyopathy. Although refractoriness and contractility were not much different in normal and diabetic atria, there was a definite effect of ranolazine and lidocaine on sodium current in varying concentrations. This may have significance in future therapeutics.

Carvacrol inhibits the neuronal voltage-gated sodium channels Nav1.2, Nav1.6, Nav1.3, Nav1.7, and Nav1.8 expressed in Xenopus oocytes with different potencies

Carvacrol is the predominant monoterpene in essential oils from many aromatic plants. Several animal studies showing analgesic effects of carvacrol indicate potential of carvacrol as a new medication for patients with refractory pain. Voltage-gated sodium channels (Nav) are thought to have crucial roles in the development of inflammatory and neuropathic pain, but there is limited information about whether the analgesic mechanism of carvacrol involves Nav. We used whole-cell, two-electrode, voltage-clamp techniques to examine the effects of carvacrol on sodium currents in Xenopus oocytes expressing α subunits of Nav1.2, Nav1.3, Nav1.6, Nav1.7, and Nav1.8. Carvacrol dose-dependently suppressed sodium currents at a holding potential that induced half-maximal current. The half-maximal inhibitory concentration values for Nav1.2, Nav1.3, Nav1.6, Nav1.7, and Nav1.8 were 233, 526, 215, 367, and 824 μmol/L, respectively, indicating that carvacrol had more potent inhibitory effects towards Nav1.2 and Nav1.6 than Nav1.3, Nav1.7, and Nav1.8. Gating analysis showed a depolarizing shift of the activation curve and a hyperpolarizing shift of the inactivation curve in all five α subunits following carvacrol treatment. Furthermore, carvacrol exhibits a use-dependent block for all five α Nav subunits. These findings provide a better understanding of the mechanisms associated with the analgesic effect of carvacrol.

Terahertz intersubband absorption of GaN/AlGaN step quantum wells grown by MOVPE on Si(111) and Si(110) substrates

We demonstrate terahertz intersubband absorptions in nitride step quantum wells (SQWs) grown by metal organic vapor phase epitaxy simultaneously on two different substrate orientations [Si(111) and Si(110)]. The structure of the SQWs consists of a 3 nm thick Al0.1Ga0.9N barrier, a 3 nm thick GaN well, and an Al0.05Ga0.95N step barrier with various thicknesses. This structure design has been optimized to approach a flatband potential in the wells to allow for an intersubband absorption in the terahertz frequency range and to maximize the optical dipole moment. Structural characterizations prove the high quality of the samples. Intersubband absorptions at frequencies of 5.6 THz (λ ≈ 54 μm), 7 THz (43 μm), and 8.9 THz (34 μm) are observed at 77 K on both substrate orientations. The observed absorption frequencies are in excellent agreement with calculations accounting for the depolarization shift induced by the electron concentration in the wells.

Reconsidering the neuropathogenic role of paroxysmal depolarization shifts in epilepsy, epileptogenesis and beyond

Reconsidering the neuropathogenic role of paroxysmal depolarization shifts in epilepsy, epileptogenesis and beyond

Involvement of Ca2+ Channels in Abnormal Excitability of Hippocampal CA3 Pyramidal Cells in Noda Epileptic Rats

Noda epileptic rat (NER) is a mutant rat, which spontaneously exhibits a tonic-clonic convulsion from 14 weeks of age. An intracellular recording study was performed to elucidate the abnormal excitability of NER hippocampal CA3 neurons. The recorded neurons were classified into two groups, group A and B neurons, according to the responses to a single stimulation of mossy fibers. In group A neurons, a stimulus elicited a long-lasting depolarization shift accompanying repetitive firings followed by after-hyperpolarization. In group B neurons, the same stimulus elicited a single spike without a long-lasting depolarization shift. Bath application of 1 mM Cd2+, a nonselective Ca2+ channel blocker, completely inhibited the abnormal excitation in group A neurons. We further examined the character of Ca2+ spikes in NER CA3 neurons. Ca2+ spikes were completely blocked by 10 μM Cd2+ in group A neurons, but not in either group B or control neurons, suggesting that Ca2+ channels in NER group A neurons have the hypersensitivity to Cd2+. Analysis using subtype specific blockers of Ca2+ channel raised the possible involvement of T-type Ca2+ channels. These results suggest that Ca2+ channel dysfunction is involved in the abnormal excitability of CA3 pyramidal neurons and pathogenesis of epilepsy in NER.