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Voltage-gated proton channel (Hv) has activity of proton transport following electrochemical gradient of proton. Hv is expressed in neutrophils and macrophages of which functions are physiologically temperature-sensitive. Hv is also expressed in human sperm cells and regulates their locomotion. H+ transport through Hv is both regulated by membrane potential and pH difference across biological membrane. It is also reported that properties of Hv such as proton conductance and gating are highly temperature-dependent. Hv consists of the N-terminal cytoplasmic domain, the voltage sensor domain (VSD), and the C-terminal coiled-coil domain, and H+ permeates through VSD voltage-dependently. The functional unit of Hv is a dimer via the interaction between C-terminal coiled-coils assembly domain. We have reported that the coiled-coil domain of Hv has the nature of dissociation around our bodily temperature and mutational change of the coiled-coil affected temperature-sensitive gating, especially its temperature threshold. The temperature-sensitive gating is assessed from two separate points: temperature threshold and temperature dependence. In this chapter, I describe physiological roles and molecular structure mechanisms of Hv by mainly focusing on thermosensitive properties.

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The tomato leafminer, Tuta absoluta (Meyrick), one of the most economically destructive pests of tomato, causes severe yields losses of tomato production globally. Rapid evolution of insecticide resistance requires the development of alternative control strategy for this pest. RNA interference (RNAi) represents a promising, innovative control strategy against key agricultural insect pests, which has recently been licensed for Colorado Potato Beetle control. Here two essential genes, voltage-gated sodium channel (Nav) and NADPH-cytochrome P450 reductase (CPR) were evaluated as targets for RNAi using an ex vivo tomato leaf delivery system. Developmental stage-dependent expression profiles showed TaNav was most abundant in adult stages, whereas TaCPR was highly expressed in larval and adult stages. T. absoluta larvae feeding on tomato leaflets treated with dsRNA targeting TaNav and TaCPR showed significant knockdown of gene expression, leading to reduction in adult emergence. Additionally, tomato leaves treated with dsRNA targeting these two genes were significantly less damaged by larval feeding and mining. Furthermore, bioassay with LC30 doses of λ-cyholthin showed that silencing TaNav and TaCPR increased T. absoluta mortality about 32.2 and 17.4%, respectively, thus indicating that RNAi targeting TaNav and TaCPR could increase the susceptibility to λ-cyholthin in T. absoluta. This study demonstrates the potential of using RNAi targeting key genes, like TaNav and TaCPR, as an alternative technology for the control of this most destructive tomato pests in the future.

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Key Clinical Message: Gliosarcoma, a rare cerebral neoplasm, has not been linked to hippocampal changes in cats. We report a case of complex partial seizures with orofacial involvement, revealing gliosarcoma concurrent with bilateral hippocampal sclerosis. Abstract: A 16-year-old neutered female domestic shorthair cat presented with acute inappetence, ataxia, disorientation, and vacant staring. Brain MRI revealed an ill-defined, round, intra-axial mass in the right piriform lobe, showing hyperintensity on T2W, T2-FLAIR, and T2*W, and hypointensity on T1W images. The lesion exhibited mass effect and contrast enhancement in its center. Bilateral hyperintensity on T2-FLAIR images and contrast enhancement were observed in the hippocampus. Brain histologic and immunohistochemical analysis revealed cerebral gliosarcoma with concurrent hippocampal sclerosis. Feline LGI1-antibody testing on the serum and/or CSF was not performed due to insufficient biomaterial. Although retrospective testing on brain tissue was considered, it ultimately proved unfeasible, preventing us from ruling out antibody-associated limbic encephalitis. In conclusion, cerebral gliosarcoma should be included in feline intracranial tumor differentials, warranting brain MRI and feline LGI1-antibody testing in cats showing complex partial seizures with orofacial involvement. In our case, the prognosis remained poor due to the presence of a high-grade glioma.

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Mutations in voltage-gated sodium (Nav) channels, which are essential for generating and propagating action potentials, can lead to serious neurological disorders, such as epilepsy. However, disease-causing Nav channel mutations do not always result in severe symptoms, suggesting that the disease conditions are significantly affected by other genetic factors and various environmental exposures, collectively known as the "exposome". Notably, recent research emphasizes the pivotal role of commensal bacteria in neural development and function. Although these bacteria typically benefit the nervous system under normal conditions, their impact during pathological states remains largely unknown. Here, we investigated the influence of commensal microbes on seizure-like phenotypes exhibited by paraShu-a gain-of-function mutant of the Drosophila Nav channel gene, paralytic. Remarkably, the elimination of endogenous bacteria considerably ameliorated neurological impairments in paraShu. Consistently, reintroducing bacteria, specifically from the Lactobacillus or Acetobacter genera, heightened the phenotypic severity in the bacteria-deprived mutants. These findings posit that particular native bacteria contribute to the severity of seizure-like phenotypes in paraShu. We further uncovered that treating paraShu with antibiotics boosted Nrf2 signaling in the gut, and that global Nrf2 activation mirrored the effects of removing bacteria from paraShu. This raises the possibility that the removal of commensal bacteria suppresses the seizure-like manifestations through augmented antioxidant responses. Since bacterial removal during development was critical for suppression of adult paraShu phenotypes, our research sets the stage for subsequent studies, aiming to elucidate the interplay between commensal bacteria and the developing nervous system in conditions predisposed to the hyperexcitable nervous system.

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Osteoarthritis (OA) represents a significant pain challenge globally, as current treatments are limited and come with substantial and adverse side effects. Voltage-gated calcium channels have proved to be pharmacologically effective targets, with multiple FDA-approved CaV2.2 modulators available for the treatment of pain. Although effective, drugs targeting CaV2.2 are complicated by the same obstacles facing other pain therapeutics- invasive routes of administration, narrow therapeutic windows, side effects, and addiction potential. We have identified a key regulator of CaV2.2 channels, collapsing response mediator protein 2 (CRMP2), that allows us to indirectly regulate CaV2.2 expression and function. We previously developed a peptidomimetic modulator of CRMP2, CBD3063, that effectively reverses neuropathic and inflammatory pain without negative side effects by reducing membrane expression of CaV2.2. The potent analgesic properties of CBD3063 combined with the lack of negative side effects prompted us to assess the efficacy of CBD3063 in a rodent model of OA pain. Here, we demonstrate the intraperitoneal administration of CBD3063 alleviates both evoked and non-evoked behavioral hallmarks of OA pain. Further, we reveal that CBD3063 reduces OA-induced increased neural activity in the parabrachial nucleus, a key supraspinal site modulating the pain experience. Together, these studies suggest CBD3063 is an effective analgesic for OA pain. PERSPECTIVE: Despite the high prevalence of osteoarthritis pain worldwide, current treatment options remain limited. We demonstrate that CBD3063-mediated disruption of the CaV2.2-CRMP2 interaction alleviates pain in a preclinical joint pain model, providing a promising basis for the development of new osteoarthritis pain treatments.

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This study identified genetic mutations linked to resistance to pyrethroid insecticides in the plant pest Lygus pratensis. The voltage-gated sodium channel (VGSC) gene was cloned, revealing two mutations (Met918Thr and Leu1014Phe) in laboratory strains and field populations from Inner Mongolia, resulting in variable pyrethroid resistance. A 3D model of LpVGSC was created using homology modeling, and pyrethroid binding patterns were analyzed via molecular docking. Molecular dynamics simulations confirmed structural stability changes and binding stability of pyrethroids to VGSC sites. Mutation frequencies of homozygous and heterozygous genotypes did not exceed 40 and 20%, respectively. Toxicity tests showed high resistance to λ-cyhalothrin (LC50:401.31 ng/cm2). The kdr (L1014F) and superkdr (M918T) mutations weakened interaction forces, reducing pyrethroid binding. M918T and L1014F mutations are predicted to reduce Type I pyrethroid affinity, suggesting Type II pyrethroids may be more effective against resistant strains. These findings aid in resistance management and insecticide design.

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Variants in KCNMA1, encoding the voltage- and calcium-activated K+ (BK) channel, are associated with human neurological disease. The effects of gain-of-function (GOF) and loss-of-function (LOF) variants have been predominantly studied on BK channel currents evoked under steady-state voltage and Ca2+ conditions. However, in their physiological context, BK channels exist in partnership with voltage-gated Ca2+ channels and respond to dynamic changes in intracellular Ca2+ (Ca2+i). In this study, an L-type voltage-gated Ca2+ channel present in the brain, CaV1.2, was co-expressed with wild type and mutant BK channels containing GOF (D434G, N999S) and LOF (H444Q, D965V) patient-associated variants in HEK-293T cells. Whole-cell BK currents were recorded under CaV1.2 activation using buffering conditions that restrict Ca2+i to nano- or micro-domains. Both conditions permitted wild type BK current activation in response to CaV1.2 Ca2+ influx, but differences in behavior between wild type and mutant BK channels were reduced compared to prior studies in clamped Ca2+i. Only the N999S mutation produced an increase in BK current in both micro- and nano-domains using square voltage commands and was also detectable in BK current evoked by a neuronal action potential within a microdomain. These data corroborate the GOF effect of N999S on BK channel activity under dynamic voltage and Ca2+ stimuli, consistent with its pathogenicity in neurological disease. However, the patient-associated mutations D434G, H444Q, and D965V did not exhibit significant effects on BK current under CaV1.2-mediated Ca2+ influx, in contrast with prior steady-state protocols. These results demonstrate a differential potential for KCNMA1 variant pathogenicity compared under diverse voltage and Ca2+ conditions.

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Voltage-gated sodium channels (NaVs) selectively permit diffusion of sodium ions across the cell membrane and, in excitable cells, are responsible for propagating action potentials. One of the nine human NaV isoforms, NaV1.8, is a promising target for analgesics, and selective inhibitors are of interest as therapeutics. One such inhibitor, the gating-modifier peptide Protoxin-I derived from tarantula venom, blocks channel opening by shifting the activation voltage threshold to more depolarised potentials, but the structural basis for this inhibition has not previously been determined. Using monolayer graphene grids, we report the cryogenic electron microscopy structures of full-length human apo-NaV1.8 and the Protoxin-I-bound complex at 3.1 Å and 2.8 Å resolution, respectively. The apo structure shows an unexpected movement of the Domain I S4-S5 helix, and VSDI was unresolvable. We find that Protoxin-I binds to and displaces the VSDII S3-S4 linker, hindering translocation of the S4II helix during activation.

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We studied changes of pulmonary microhemodynamics when modeling pulmonary artery thromboembolism on perfused isolated rabbit lungs after pretreatment with ranolazine and ivabradine. The increase in pulmonary artery pressure, pulmonary vascular resistance, and pre- and postcapillary resistance was less pronounced than in control animals, but was close to that in case of pulmonary thromboembolism after pretreatment with voltage-gated Na+ channel blockers lidocaine and ropivacaine. The increase of capillary filtration coefficient inversely correlated with values of capillary hydrostatic pressure. Thus, ranolazine and ivabradine exhibit the properties of voltage-gated Na+ channel blockers mainly in smooth muscles of pulmonary arterial vessels and promote the decrease in endothelial permeability.

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OBJECTIVES: The antiepileptic phenytoin has a narrow therapeutic window, nonlinear pharmacokinetics, and can cross the placenta causing apathy and jitteriness in postpartum newborns. Further, the sudden decay of phenytoin concentration can cause withdrawal seizures. This work aimed to assess the brain toxic exposure to phenytoin in newborns after transplacental transfer using neonatal saliva-brain correlations. METHODS: The phenytoin dose that the newborn receives transplacentally at birth was estimated using verified physiologically based pharmacokinetic (PBPK) model simulations in third-trimester pregnancy (pregnancy T3). Such doses were used as an input to the newborn PBPK model to estimate the neonatal levels of phenytoin and their correlations in brain extracellular fluid (bECF), plasma, and saliva. RESULTS: The PBPK model-estimated neonatal plasma and bECF concentrations of phenytoin were below the necessary thresholds for anticonvulsant and toxic effects. The neonatal salivary thresholds for phenytoin anticonvulsant and toxic effects were estimated to be 1.3 and 2.5 mg/L, respectively using the plasma-saliva-bECF correlations established herein. CONCLUSIONS: The salivary TDM of phenytoin can be a more convenient option for avoiding phenytoin brain toxicity in newborns of epileptic mothers. Still, the appropriateness of using the same adult values of phenytoin anticonvulsant and toxic effects for infants needs investigation.

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Homozygous variants of Calcium Voltage-Gated Channel Subunit Alpha1 S (CACNA1S) gene mutation were previously identified as causes of periodic paralysis and congenital early-onset myopathy, while it could be manifested as a late-onset congenital core myopathy. Abstract: Calcium Voltage-Gated Channel Subunit Alpha1 S (CACNA1S) gene mutation has been linked to various neuromuscular conditions in recent years. Congenital myopathy with core-like features is one of the cardinal associations reported previously, causing severe respiratory insufficiency and death in neonates. Informed consent was received from the patients. Subsequently, peripheral blood leukocytes were utilized to extract genomic DNA. Moreover, exome enrichment was implemented through the Twist Human Core Exome Kit (Twist Bioscience) and exome sequenced using Illumina NovaSeq 6000 platform (Illumina, San Diego, CA, USA). Sanger sequencing using BIG Dye Terminators confirmed the presence of the final variant. Finally, the candidate variants were classified based on the American College of Medical Genetics and Genomics (ACMG) guidelines. In this report, we describe two siblings, who presented with childhood and late-onset progressive muscle weakness, and had a homozygous variant in exon 2 of the CACNA1S gene defined as c.188C > A (p.Ala63Asp) (NM_000069.3). The SIFT, Polyphen2, CADD PHRED, and Mutation Taster analysis tools classified the variant as pathogenic/damaging. The muscle biopsy of the younger brother revealed intermyofibrillar network pattern disruption as cytoplasmic core-like lesions. The muscle magnetic resonance imaging (MRI) reported grade IIa and IIb fatty changes. Finally, the electromyography (EMG) findings suggested a myopathic change pattern. This report illustrates the clinical variability in CACNA1S-related myopathy by reviewing prior reports and adding newly found aspects, additionally expanding the gene defects associated with core myopathy.

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The case of a 72-year-old female patient with arrhythmogenic syncope associated with a combination therapy of flecainide and lacosamide is presented. The authors believe in an additive effect of both drugs on myocardial voltage-gated sodium channels with extraordinary QRS widening, exit block with temporary pacing and complete reversibility through infusion of sodium bicarbonate as bail-out therapy.

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Genetic loss-of-function mutations of Nav1.7 channel, abundantly expressed in peripheral nociceptive neurons, cause congenital insensitivity to pain (CIP) in humans, indicating that selective inhibition of the channel may lead to potential therapy of pain disorders. In this study, we investigated a novel compound, 5-chloro-N-(cyclopropylsulfonyl)-2-fluoro-4-(2-(8-(furan-2-ylmethyl)-8-azaspiro [4.5] decan-2-yl) ethoxy) benzamide (QLS-278) that inhibits Nav1.7 channel and exhibits anti-nociceptive activity. Compound QLS-278 exhibits inactivation- and concentration-dependent inhibition of macroscopic currents of Nav1.7 channels stably expressed in HEK293 cells with an IC50 of 1.2 {plus minus} 0.2 µM. QLS-278 causes a hyperpolarization shift of the channel inactivation and delays recovery from inactivation, without an obvious effect on voltage-dependent activation. In mouse DRG neurons, QLS-278 suppresses native TTX-sensitive Nav currents and also reduces neuronal firing. Moreover, QLS-278 dose-dependently relieves neuropathic pain induced by spared nerve injury and inflammatory pain induced by formalin without significant alteration of spontaneous locomotor activity in mice. Altogether, our identification of the novel compound QLS-278 may hold developmental potential for the treatment of chronic pain. Significance Statement QLS-278, a novel voltage-gated sodium Nav1.7 channel blocker, inhibits native TTX-S Na+ current and reduces action potential firings in DRG sensory neurons. QLS-278 also exhibits antinociceptive activity in mouse models of pain, thus demonstrating potential for the development of a treatment for chronic pain.

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Natural ion channels are proteins embedded in the cell membrane that control many aspects of cell and human physiology by acting as gatekeepers, regulating the flow of ions in and out of cells. Advances in nanotechnology have influenced the methods for studying ion channels in vitro, as well as ways to unlock the delivery of therapeutics by modulating them in vivo. This review provides an overview of nanotechnology-enabled approaches for ion channel research with a focus on the synthesis and applications of synthetic ion channels. Further, the uses of nanotechnology for therapeutic applications are critically analyzed. Finally, we provide an outlook on the opportunities and challenges at the intersection of nanotechnology and ion channels. This work highlights the key role of nanoscale interactions in the operation and modulation of ion channels, which may prompt insights into nanotechnology-enabled mechanisms to study and exploit these systems in the near future.

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Melatonin is synthesized in and secreted from the pineal glands, and regulates circadian rhythms. Although melatonin has been reported to modulate the activity of ion channels in several tissues, its effects on pineal ion channels remain unclear. In the present study, the effects of melatonin on voltage-gated K+ (KV) channels, which play a role in regulating the resting membrane potential, were examined in rat pinealocytes. The application of melatonin reduced pineal KV currents in a concentration-dependent manner (IC50=309 mM). An expression analysis revealed that KV4.2 channels were highly expressed in rat pineal glands. Melatonin-sensitive currents were abolished by the small interfering RNA knockdown of KV4.2 channels in rat pinealocytes. In human embryonic kidney 293 (HEK293) cells expressing KV4.2 channels, melatonin decreased outward currents (IC50=479 mM). Inhibitory effects were mediated by a shift in voltage dependence from steady-state inactivation to a hyperpolarizing direction. This inhibition was observed even in the presence of 100 nM luzindole, an antagonist of melatonin receptors. Melatonin also blocked the activity of KV4.3, KV1.1, and KV1.5 channels in reconstituted HEK293 cells. The application of 1 mM melatonin caused membrane depolarization in rat pinealocytes. Furthermore, KV4.2 channel inhibition by 5 mM 4-aminopyridine attenuated melatonin secretion induced by 1 mM noradrenaline in rat pineal glands. These results strongly suggest that melatonin directly inhibited KV4.2 channels and caused membrane depolarization in pinealocytes, resulting in a decrease in melatonin secretion through parasympathetic signaling pathway. This mechanism may function as a negative-feedback mechanism of melatonin secretion in pineal glands.

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Ion channels are membrane proteins that allow the passage of ions across the membrane. They characteristically contain a pore where the selectivity of certain ion species is determined and gates that open and close the pore are found. The pore is often connected to additional domains or subunits that regulate its function. Channels are grouped into families based on their selectivity for specific ions and the stimuli that control channel opening and closing, such as voltage or ligands. Ion channels are fundamental to the electrical properties of excitable tissues. Dysfunction of channels can lead to abnormal electrical signaling of neurons and muscle cells, accompanied by clinical manifestations, known as channelopathies. Many naturally occurring toxins target ion channels and affect excitable cells where the channels are expressed. Furthermore, ion channels, as membrane proteins and key regulators of a number of physiologic functions, are an important target for drugs in clinical use. In this chapter, we give a general overview of the classification, genetics and structure-function features of the main ion channel families, and address some pharmacologic aspects relevant to neurologic channelopathies.

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Introduction: The KCNQ2/KCNQ3 genes encode the voltage-gated K channel underlying the neuronal M-current, regulating neuronal excitability. Loss-of-function (LoF) variants cause neonatal epilepsy, treatable with the M-current-opener retigabine, which is no longer marketed due to side effects. Gain-of-function (GoF) variants cause developmental encephalopathy and autism that could be amenable to M-current, but such therapies are not clinically available. In this translational project, we investigated whether donepezil, a cholinergic drug used in Alzheimer's, suppresses M currents in vitro and improves cognitive symptoms in patients with GoF variants. Methods: (1) The effect of 1 µM donepezil on the amplitude of the M-current was measured in excitatory and inhibitory neurons of mouse primary cultured hippocampal cells. M-current was measured using the standard deactivation protocol (holding at 0 mV and deactivation at -60 mV) in the voltage-clamp configuration of the whole-cell patch clamp technique. The impact of donepezil was also examined on the spontaneous firing activity of hippocampal neurons in the current-clamp configuration. (2) Four children with autism, aged 2.5-8 years, with the following GoF variants were enrolled: KCNQ2 (p. Arg144Gln) and KCNQ 3 (p.Arg227Gln, p.Arg230Cys). Patients were treated off-label with donepezil 2.5-5 mg/d for 12 months and assessed with: clinical Global Impression of Change (CGI-c), Childhood Autism Rating Scale 2 (CARS-2), Adaptive Behavior Assessment System-II (ABAS-II), and Child Development Inventory (CDI). Results: (1) Application of donepezil for at least 6 min produced a significant inhibition of the M-current with an IC50 of 0.4 µM. At 1 µM, donepezil reduced by 67% the M-current density of excitatory neurons (2.4 ± 0.46 vs. 0.89 ± 0.15 pA/pF, p < 0.05*). In inhibitory neurons, application of 1 µM donepezil produced a lesser inhibition of 59% of the M-current density (1.39 ± 0.43 vs. 0.57 ± 0.21, p > 0.05). Donepezil (1 µM) potently increased by 2.6-fold the spontaneous firing frequency, which was prevented by the muscarinic receptor antagonist atropine (10 µM). (2) The CARS-2 decreased by 3.8 ± 4.9 points (p > 0.05), but in two patients with KCNQ3 variants, the improvement was over the 4.5 clinically relevant threshold. The global clinical change was also clinically significant in these patients (CGI-c = 1). The CDI increased by 65% (p < 0.05*), while the ABAS-II remained unchanged. Discussion: Donepezil should be repurposed as a novel alternative treatment for GoF variants in KCNQ2/KCNQ3 encephalopathy.

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Electrical excitability-the ability to fire and propagate action potentials-is a signature feature of neurons. How neurons become excitable during development and whether excitability is an intrinsic property of neurons remain unclear. Here, we demonstrate that Schwann cells, the most abundant glia in the peripheral nervous system, promote somatosensory neuron excitability during development. We find that Schwann cells secrete prostaglandin E2, which is necessary and sufficient to induce developing somatosensory neurons to express normal levels of genes required for neuronal function, including voltage-gated sodium channels, and to fire action potential trains. Inactivating this signaling pathway in Schwann cells impairs somatosensory neuron maturation, causing multimodal sensory defects that persist into adulthood. Collectively, our studies uncover a neurodevelopmental role for prostaglandin E2 distinct from its established role in inflammation, revealing a cell non-autonomous mechanism by which glia regulate neuronal excitability to enable the development of normal sensory functions.

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Neuropathic pain (NP) is a chronic pain caused by injury or disease of the somatosensory nervous system, or it can be directly caused by disease. It often presents with clinical features like spontaneous pain, hyperalgesia, and dysesthesia. At present, voltage-gated calcium ion channels (VGCCs) are known to be closely related to the development of NP, especially the α2δ subunit. The α2δ subunit is a regulatory subunit of VGCCs. It exists mainly in the brain and peripheral nervous system, especially in nerve cells, and it plays a crucial part in regulating presynaptic and postsynaptic functions. Furthermore, the α2δ subunit influences neuronal excitation and pain signaling by promoting its expression and localization through binding to VGCC-related subunits. The α2δ subunit is widely used in the management of NP as a target of antiepileptic drugs gabapentin and pregabalin. Although drug therapy is one of the treatments for NP, its clinical application is limited due to the adverse reactions caused by drug therapy. Therefore, further research on the therapeutic target α2δ subunit is needed, and attempts are made to obtain an effective treatment for relieving NP without side effects. This review describes the current associated knowledge on the function of the α2δ subunit in perceiving and modulating NP.