Parallel solid-state nanopore measurements are expected to improve not only throughput but also molecular discrimination performance. Here, we report the development of a 16-channel solid-state nanopore array measurement system that enables parallel nanopore fabrication by dielectric breakdown and subsequent ionic current recording. The system supports two fabrication modes: pulsed-voltage-induced dielectric breakdown (MPVI) and controlled dielectric breakdown (CBD) under constant-voltage bias, allowing nanopore formation over a wide diameter range. To demonstrate the operation of the developed system, two experimental studies were conducted. In the first study, nanopores were fabricated in 5 nm-thick SiN membranes using MPVI, followed by detection of single-stranded DNA (ssDNA) translocation. Arrays of nanopores with diameters ranging from 1 to 2 nm were formed, and the pore-diameter dependence of dwell time and current blockade amplitude during ssDNA translocation could be evaluated at sub-nanometer resolution. In the second study, nanopores were fabricated in 14 nm-thick SiN membranes using CBD, and double-stranded DNA (dsDNA) translocation was detected. Analysis of current traces during CBD suggested that pore enlargement remained limited for a period after dielectric breakdown, followed by accelerated pore growth accompanied by a rapid increase in current. The resulting pore diameters were distributed within 8-10 nm for 13 of the 16 channels and within 8-9 nm for 12 channels, demonstrating tight size control across most channels. Using these fabricated nanopores, dsDNA translocation events were observed in all channels.

Plants respond to mechanical stimuli by a rapid increase in cytosolic calcium. The intensity and kinetics of the calcium changes define calcium signatures important for biological responses. In this study, we determine the properties of a calcium-permeable force-gated channel localized at the plasma membrane called Rapid Mechanically Activated (RMA). Using patch clamp and pressure clamp, we characterized the kinetics of the Arabidopsis thaliana RMA channel upon stimulation by pressure pulses applied onto the plasma membrane. Combining pressure pulse protocols at different frequencies with modeling, we investigated the channel's capacity to transduce high frequency mechanical stimuli. The RMA channel rapidly activates in response to membrane tension, then it inactivates during prolonged stimulation. Upon repeated stimulations, the RMA current amplitude decreases irreversibly indicating that it undergoes attenuation. The channel kinetics were modeled with four chemical states and the model predicts that it behaves as a pass band filter in the 10 Hz-1 kHz range. In conclusion, due to its activation/inactivation, the RMA channel is a candidate for mediating cytosolic calcium signaling in response to mechanostimulation. Its attenuation and filtering properties suggest its involvement in the transduction of high frequency mechanical stimulation, such as those produced by insects' vibrations.

Large conductance calcium-activated potassium channels (BKCa) play an important role in the regulation of vascular tone. However, the properties of BKCa channels in smooth muscle of pulmonary arteries are poorly understood. Previous experimental studies demonstrated that pulmonary hypoxic vasoconstriction as a normal physiological response to decreased oxygen levels was impaired in diabetic animals due to abnormal activation of BKCa channels. The aim of this study was to identify mechanisms of diabetes-induced activation of BKCa channels in freshly isolated smooth muscle cells from rat pulmonary arteries. Type 1 diabetes was induced by streptozotocin (STZ). Whole-cell potassium currents were recorded using the patch-clamp method. Expression levels of BK-α and BK-β1 subunits were measured by real-time PCR. Our results demonstrate that the amplitude of whole-cell current through BKCa channels in rat pulmonary artery smooth muscle cells is significantly increased during STZ-induced diabetes without altering the expression of BK-α and BK-β1 subunits and channel calcium sensitivity. The slow component of BKCa current deactivation time constant and spontaneous transient outward current amplitude were increased in diabetic animals compared to healthy animals. We conclude that abnormal activation of the BKCa channel in pulmonary arterial smooth muscle during diabetes is associated with alterations in the local control mechanism of the BKCa pore-gate domain.

Integrated electrophysiological, cellular, and pharmacological profiling reveals variant-specific mechanisms in SCN4A-related myotonia.

A comprehensive first-principles investigation of tunable quantum transport in pyrrole-based molecular junctions is performed by using density functional theory combined with the nonequilibrium Green's function approach. When coupled to zigzag graphene nanoribbon electrodes, the pyrrole monomer/oligomer-based devices exhibit three essential transport characteristics, namely a pronounced negative differential resistance (NDR) effect, nonlinear gate-controlled modulation, and destructive quantum interference (DQI)-induced switching. The current amplitude decreases systematically with increasing oligomer length, while the peak-to-valley ratio increases, reaching a maximum of 17.36. Gate modulation effectively preserves and enhances the NDR effect, where a negative gate voltage shifts the HOMO toward the Fermi level, broadening the HOMO-dominated transmission peak and improving the stability of NDR. Furthermore, conformational rotations disrupt the π-conjugated pathway, inducing strong DQI that drastically suppresses conductance. This mechanism enables a robust molecular switching behavior, achieving an on/off ratio reaching as high as 6.48 × 103. These results establish clear structure-transport correlations and demonstrate the potential of pyrrole-based molecular junctions for highly tunable functional components in future molecular electronic devices.

Developmental and epileptic encephalopathies (DEEs) are severe neurodevelopmental disorders associated with genetic mutations, including some in GRIN genes encoding NMDA receptor (NMDA-R) subunits. Despite affecting the same receptor, each mutation may lead to distinct neurological disorders, emphasizing the necessity of understanding receptor dysfunction to tailor treatments effectively. In a genetic screen of DEEs patients, we identified a de novo pathogenic GRIN2B nonsense mutation, p.Glu839Ter (called GluN2B-E839*), which truncates the C-terminal domain (CTD) of the GluN2B subunit, a poorly characterized region critical for receptor function. This variant was prioritized for functional study due to the limited knowledge about the CTD's role. We fully characterized the clinical presentation of the patient, who displayed intellectual disability, epilepsy, and hyperkinetic behavioral disorders. Molecular and cellular analyses in heterologous systems and patient-derived neurons revealed that GluN2B-E839* subunit assembles correctly with other subunits to form NMDA-Rs but exhibits reduced surface expression, impaired interactions with PSD95, and altered biophysical properties, including reduced current amplitudes, increased magnesium sensitivity, and diminished calcium influx. These dysfunctions likely contribute to impaired synaptic plasticity and DEEs pathophysiology. Our findings highlight the critical role of the GluN2B CTD in NMDA-R function and neuronal signaling and underscore the need for systematic characterization of GRIN variants to improve diagnostic precision and therapeutic targeting for DEEs. This study establishes a robust framework combining complementary experimental approaches with patient-derived preclinical models to link molecular dysfunctions to clinical phenotypes.

Insulin, an important regulator of peripheral blood glucose levels, can also act in the brain to influence energy metabolism. Insulin receptors are expressed in the dorsomedial hypothalamus (DMH), but nothing is known about the effects of insulin on synaptic function in this region. To determine the effect of insulin on glutamate transmission in the DMH, we used whole-cell patch clamp electrophysiology to examine glutamatergic currents in DMH neurons of young male and female Sprague-Dawley rats. Insulin decreased evoked glutamate current amplitude in both sexes and variance analysis suggested this was due to a postsynaptic change in quantal size. Blocking insulin receptors failed to remove the effect of insulin in both sexes, and further experiments revealed that both insulin receptors (IRs) and insulin-like growth factor-1 receptors (IGF-1Rs), and mechanistic target of rapamycin (mTOR), were necessary to completely block the effects of insulin. Tonic insulin significantly decreased glutamate transmission in the DMH through an IR and mTOR-mediated pathway. Overall, our data suggest that insulin acts through IRs and IGF-1Rs to alter synaptic transmission in the DMH and tonic insulin controls the activity of DMH neurons. This research contributes to our understanding of how insulin acts in the hypothalamus to alter synaptic function, with potential implications for diabetes and obesity research.

Images captured in low-light environments suffer from insufficient luminance and lost details, which severely restrict their application in scenarios such as surveillance and aerial photography. Conventional sigmoid-based luminance adjustment relies on software computation and is limited by issues including response latency and poor hardware compatibility. In this work, we propose a low-light image enhancement scheme that leverages magnetic tunnel junctions (MTJs) to implement hardware-based fitting of the sigmoid function. The scheme transforms low-light images from the RGB to the HSV color space and decouples the luminance component from chrominance components, enabling independent luminance adjustment. Utilizing the sigmoid-like relationship between the switching probability of an MTJ and the applied current, the scheme maps low pixel values in the luminance channel to corresponding current amplitudes and directly accomplishes nonlinear fitting of the sigmoid function through the intrinsic physical response of MTJ, which enables efficient enhancement of the luminance component. Experimental results demonstrate that the proposed method effectively improves the luminance and detail clarity of low-light images, achieving an improvement of 3.82 dB in PSNR and 9.91% in SSIM compared with the original sigmoid method and a performance gain of 13.46% in SSIM over the lightweight learning-based scheme. Furthermore, the proposed approach supports the efficient processing of low-light images with different resolutions and is well-suited for resource-constrained edge terminals.This work contributes both theoretical insights and engineering value to the practical deployment of low-light image enhancement technologies.

Tumor necrosis factor-alpha (TNF-α), a pleiotropic cytokine, modulates neuronal functions under both physiological and pathological conditions. In the auditory system, it is required for refinement of the cortical frequency map during early development. In adulthood, TNF-α upregulation following noise trauma contributes to synaptic imbalance and central auditory processing deficits. The effects of TNF-α deficiency on adult auditory cortical circuits and function have not been examined. Here, we report that compared to wild-type (WT) control mice (an equal number of males and females per group), adult TNF-α knockout (KO) mice had reduced PV neuron density and PV expression levels. Pyramidal neurons in the auditory cortex of TNF-α KO mice had larger miniature excitatory postsynaptic current (mEPSC) amplitude and lower miniature inhibitory postsynaptic current (mIPSC) frequency, suggesting a shift in synaptic E/I balance. Cortical multiunit had increased spontaneous and evoked activity and broadened tuning bandwidth consistent with the increased synaptic E/I ratio. Importantly, unlike WT mice, TNF-α KO mice exhibited persistent critical period-like plasticity into adulthood. Following exposure to a single-frequency tone, the representation of the tone was enlarged in adult TNF-α KO mice, but not in WT mice. Together with existing literature, our results suggest that TNF-α has a bell-shaped influence on adult auditory cortical circuits, with both elevated and deficient TNF-α expression leading to PV neuron dysfunction, increased synaptic E/I ratio, and enhanced cortical frequency map plasticity.Tumor necrosis factor-α (TNF-α) is a pleiotropic cytokine that contributes to the regulation of synaptic excitation–inhibition (E/I) balance. Sensory restriction is associated with increased TNF-α expression and an elevated synaptic E/I ratio. Here, we show that TNF-α-deficient mice have reduced parvalbumin-positive (PV) inhibitory interneuron density, an increased E/I ratio, impaired sensory restriction-induced homeostatic plasticity, and persistent critical period-like plasticity in adulthood. Together, these findings suggest a nonlinear, bell-shaped relationship between TNF-α signaling and cortical circuit function, in which both excessive and deficient TNF-α levels are associated with impaired PV interneuron function and increased E/I ratio.

Overexpression effects of 4.1B protein on epilepsy and excitatory synaptic transmission in mice.

Cerebral ischemia-reperfusion (I/R) injury remains a major therapeutic challenge, primarily due to complex mechanisms involving oxidative stress and apoptosis. Growth arrest and DNA damage-inducible protein α (Gadd45α), a stress sensor linked to cellular stress responses, has been implicated in I/R injury, yet its precise role in ischemic stroke is incompletely understood. This study aimed to elucidate the function and underlying mechanisms of Gadd45α in cerebral I/R injury using both in vivo and in vitro models. In rats subjected to middle cerebral artery occlusion (MCAO) and in neurons exposed to oxygen-glucose deprivation/reperfusion (OGD/R), Gadd45α expression was significantly upregulated. Lentivirus-mediated knockdown of Gadd45α (sh-Gadd45α) reduced infarct volumes, improved neurological function and increased miniature excitatory postsynaptic current (mEPSC) amplitude. In primary cortical neurons exposed to OGD/R, Gadd45α knockdown decreased reactive oxygen species (ROS) production, DNA damage, and apoptosis, while Gadd45α overexpression exacerbated these effects. Mechanistically, Gadd45α directly interacts with forkhead box O1 (FOXO1) and positively regulates its transcriptional activity. Gadd45α knockdown attenuated the ischemia-induced upregulation of both total FOXO1 and its phosphorylated form (p-FOXO1), thereby suppressing FOXO1 signaling and mitigating cerebral I/R injury. Furthermore, FOXO1 overexpression reversed the neuroprotective effects of Gadd45α silencing, confirming that FOXO1 acts as a critical downstream mediator. These findings demonstrate that Gadd45α silencing alleviates cerebral I/R injury by suppressing FOXO1 signaling, suggesting the Gadd45α/FOXO1 axis as a promising therapeutic target for ischemic stroke.

Learning broadly alters neocortical synapses, although the input and target specificity for this plasticity has not been well-defined. Feedforward synapses into sensory cortex have early critical periods for plasticity after which they are resistant to experience-dependent changes. Whether these synapses are altered during learning has not been investigated, particularly in a setting where animals must identify causal relationships between sensory stimuli and rewards. Here, we examined whether these feedforward synapses can be altered by training mice in a freely-moving and whisker-dependent association task. Pathway-specific optogenetic stimulation and analysis of quantal excitatory postsynaptic currents in layer 2/3 (L2/3) pyramidal neurons from barrel cortex revealed a rapid and transient potentiation of layer 4 (L4) inputs at the onset of training, without any change in thalamocortical inputs onto L4 neurons. In contrast, pseudotraining—where stimuli and rewards were decoupled—drove depression of L4–L2/3 quantal excitatory postsynaptic currents. Because environmental enrichment did not influence quantal excitatory postsynaptic current amplitude, these data suggest that reward-prediction accuracy is a key driver of feedforward plasticity in primary sensory cortex.Significance statementAlthough it is well accepted that sensory learning can alter cortical synapses, the pathways that are modified and the specific cues that drive this synaptic change have not been systematically investigated. By manipulating stimulus–reward probabilities, we identified discrete and opposite changes in the strength of L4–L2/3 synapses depending on the predictive accuracy of the stimulus. These data suggest that feedforward sensory circuits are exquisitely sensitive to the predictive value of sensory input in a goal-directed task.

Pyridine- and quinoline-based piperazine nitriles: Potent and selective 5-HT3 receptor agonists and efficacious desensitizers.

Epilepsy is a common neurological disorder with a strong genetic basis, most frequently arising from ion channel dysfunction. Although multiple inwardly rectifying potassium (Kir) channels have been implicated in epileptogenesis, the contribution of KCNJ4, which encodes the Kir2.3 channel, has not previously been established in human epilepsy. The present study aimed to identify pathogenic KCNJ4 variants and to elucidate their functional consequences in the context of epilepsy. Trio whole exome sequencing was performed in four unrelated individuals with refractory epilepsy and neurodevelopmental abnormalities. Identified KCNJ4 variants were evaluated for rarity and inheritance patterns. Functional consequences were assessed using two-electrode voltage-clamp recordings in Xenopus laevis oocytes coexpressing wild-type or mutant Kir2.3 together with Kir2.1. Protein expression levels were examined by Western blot analysis to exclude effects attributable to altered channel expression or trafficking. We identified four rare heterozygous missense variants in KCNJ4 (Gly136Ser, Val206Met, Met293Lys, and Glu384Lys), all of which were absent from public population databases. Clinically, affected individuals exhibited a broad phenotypic spectrum ranging from isolated epilepsy to severe developmental and epileptic encephalopathy. Electrophysiological analyses revealed variant-specific functional alterations; the Gly136Ser and Glu384Lys variants significantly increased inwardly rectifying potassium currents, consistent with gain-of-function effects, whereas the Val206Met and Met293Lys variants markedly reduced current amplitudes, indicating loss of function. These functional changes were independent of channel protein expression levels. Our findings establish KCNJ4 as a novel epilepsy-associated gene and demonstrate that both gain- and loss-of-function mechanisms of Kir2.3 can contribute to epileptogenesis. This study expands the genetic landscape of epilepsy and highlights the critical role of inward-rectifier potassium channel regulation in neuronal excitability, with potential implications for mechanism-based therapeutic strategies.

Neuroexcitotoxicity mediated by NMDA receptor is a central contributor to ischemic stroke pathology. Neuroprotective effect and mechanism of the new compound GW201, which targets NMDA receptor, were studied. We evaluated the neuroprotective effects of GW201 using in vivo and in vitro models of ischemic stroke, including the middle cerebral artery occlusion/reperfusion (MCAOi/r) model, permanent MCAO (pMCAO) model and primary neuronal oxygen-glucose deprivation (OGD) model. Pharmacodynamic studies examined the dose-response relationship and therapeutic time window of GW201 neuroprotective effects. Electrophysiological membrane clamp experiments assessed the NMDA receptor subunit selectivity of GW201 and its influence on neuronal currents under ischemia-mimicking conditions. The mechanism of its action was further analysed using molecular docking technology. Live-cell fluorescence imaging was employed to evaluate the impact of GW201 on intracellular calcium levels in simulated ischemic stroke. Furthermore, bioinformatics analysis of the GEO database identified 16 significant calcium-related genes, with their regulation by GW201 validated in the MCAOi/r model. Across all models tested, GW201 exhibited marked neuroprotective effects. Membrane clamp studies identified GW201 as an allosteric modulator of the NMDA receptor GluN2A subunit reducing current amplitude under high glutamate/NMDA + glycine conditions and subsequently lowering intracellular calcium levels. Bioinformatics analysis revealed 16 calcium-related genes, with GW201 significantly modulating Ccl3, Stat3, Anxa1, Anxa2, Mgp, S100A8, S100A9 and Cacna1a. These results suggest that the neuroprotective effects of GW201 are mediated by its ability to allosterically modulate NMDA receptor activity, reducing calcium overload, influence downstream gene expression and suppress inflammatory responses.

Water plasma generated from in-situ adsorbent-released vapor inactivates bacterial spores and damages DNA and proteins via OH-radical-rich chemistry

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) have gained interest as a pharmacological modelbut their immaturity leads to uncertainty regarding translation. We studied the key player in maintaining ionic homeostasis, the Na+/K+-ATPase (NKA), in hiPSC-CM. Atrial and ventricular engineered heart tissues (EHTs) were produced from hiPSC-CMs. For comparison, adult atrial and ventricular tissues were obtained from patients undergoing open heart surgery. We measured NKA gene expression, NKA pump currents (INKA) and ouabain effects on action potentials and contractility. Computational modelling was used to further investigate the direct and indirect impacts of NKA inhibition. The mRNA abundance of the major NKA isoforms was higher in ventricular than atrial cardiomyocytes (adult and hiPSC-CM). Consistently, INKA was also higher in ventricular than in atrial hiPSC-CM and higher in ventricular than in atrial adult cardiomyocytes. Ouabain potency to block INKA did not differ between hiPSC-derived and adult cardiomyocytes. Ouabain shortened plateau phase in EHT and adult tissue. However, lower ouabain concentrations depolarized diastolic potential and depressed force more in EHTs, demonstrating a higher integrated sensitivity to NKA inhibition. Computational modelling indicated that weaker IK1 increases the susceptibility to depolarisation by NKA block. HiPSC-EHTs express NKA with biophysical characteristics and ouabain-sensitivity that are not different from human tissue. Action potential duration responses in EHT reproduces chamber-specific response pattern as seen in human heart. The higher sensitivity of EHTs to depolarize under ouabain-induced block of NKA needs to be considered when employing hiPSC-CM in drug research.

Anorexia nervosa is a severe psychiatric disorder characterized by persistent food restriction and often excessive physical activity, implicating dysfunction in neural circuits governing motivation, reward, and behavioral persistence. The nucleus accumbens (NAc) is a central component of these circuits, yet synaptic and cellular adaptations within this region during anorexia‐like states remain poorly defined. Using the activity‐based anorexia (ABA) paradigm in adult female mice, we examined glutamatergic transmission and intrinsic neuronal properties in the NAc shell. ABA exposure produced rapid weight loss, reduced food intake, and progressively increased running‐wheel activity. Biochemical analyses of NAc shell tissue revealed elevated membrane‐associated GluA2 AMPA receptor protein. Consistent with this finding, whole‐cell patch‐clamp recordings from medium spiny neurons showed increased amplitude of spontaneous excitatory postsynaptic currents. ABA also enhanced intrinsic neuronal excitability, reflected by greater firing in response to depolarizing current injections. Together, these convergent biochemical and electrophysiological results demonstrate that ABA induces coordinated postsynaptic strengthening and increased intrinsic excitability in NAc shell medium spiny neurons. These adaptations suggest a sustained increase in accumbal output that may bias motivational circuit function and contribute to excessive activity and suppressed feeding during anorexia‐like conditions, paralleling glutamatergic plasticity observed in other compulsive disorders, including substance use disorder.

Disease variants in GABR genes encoding γ-Aminobutyric acid type A receptor (GABAaR) subunits are major causes of developmental and epileptic encephalopathies (DEEs). There is no effective treatment for these DEEs although the GABAaR is a major target for antiseizure drugs. We previously identified the therapeutic effect of 4-phenylbutyrate (PBA) in Gabrg2+/Q390X knockin DEE mice and now test the effect of the drug in GABRA1 variants that encode the α1 subunit. We used a multidisciplinary approach including in silico structural modeling, flow cytometry, patch clamp recordings and bio-chemistry in conjunction with differential tagging of the wild-type and the mutant alleles to evaluate the effect of PBA on rescue of GABAaR subunit expression, surface trafficking, and function in vitro in heterologous HEK293T cell model and in vivo in Gabra1+/A322D mice. We found that both total and cell surface α1 expression was reduced when the variant α1 protein was present; suggesting reduced functional receptor on the cell membrane and synapse. Patch clamp recordings identified α1 variants reduced GABA-evoked current amplitude. In silico prediction indicated reduced protein stability for GABRA1 variants indicated by negative ΔΔG values. PBA increased both total and surface expression of wildtype α1 and α1 variants; and improved expression of both wildtype and variant α1 alleles when these were co-expressed. Importantly, PBA also increased the GABAaR expression in the thalamus of the Gabra1+/A322D mice. This study indicates that PBA is a promising treatment option for DEEs associated with GABRA1 mutations. Our previous work has demonstrated that PBA improves proteostasis by enhancing expression of the wildtype allele, repairing the mutant allele, and reducing endoplasmic reticulum stress. Therefore, it can mitigate seizures and improve neurobehavioral phenotypes at behavioral levels. Based on this and our previous work on GABRG2 and SLC6A1 mutations, we propose that PBA holds promise as a common medicine for multiple genetic neurologic disorders that share the proteostasis pathology with a broad clinical application in DEEs.

Neurogenic lower urinary tract dysfunction (NLUTD) impairs bladder control and remains difficult to treat. We aim to define how electrical stimulation (ES) parameters of the external urethral sphincter (EUS) affect urinary leakage thresholds to guide neuromodulation strategies for NLUTD. We performed direct EUS stimulation in anesthetized rats using charge-balanced biphasic pulses while systematically varying current amplitude (0.5-3.0 mA), frequency (20-100 Hz), and pulse duration (0.5-3 ms). Urine leakage thresholds were mapped across the multidimensional parameter space. Stimulation parameters exhibited strong nonlinear interdependence in determining leakage onset. At a fixed pulse duration, higher current amplitudes required lower stimulation frequencies to evoke leakage. Increasing pulse duration substantially reduced both current and frequency thresholds. Age and sex caused modest shifts in absolute thresholds but did not alter the fundamental parameter-response relationships. Pulse duration, current amplitude, and frequency jointly govern urinary leakage thresholds, with pulse duration serving as the dominant modulator of stimulation efficiency. This work establishes a quantitative framework for charge-efficient stimulation parameter selection, enabling the design of energy-aware, precision neuromodulation protocols and implantable systems for NLUTD rehabilitation.