Search Results (572)

Autosomal-dominant sleep-related hypermotor epilepsy (ADSHE) was the first distinct genetic epilepsy proven to be caused by mutation of the CHRNA4 gene, originally reported in 1994. In the past three decades, pathomechanisms of ADSHE associated with mutant nicotinic acetylcholine receptors (nAChRs) have been explored via various studies, including in vitro experiments and genetic rodent models. However, findings emphasize that functional abnormalities of ADSHE-mutant nAChRs alone cannot generate ictogenesis; rather, development of abnormalities in various other transmission systems induced by ADSHE-mutant nAChRs during the neurodevelopmental process before the ADSHE onset is involved in development of epileptogenesis/ictogenesis. Intra-thalamic GABAergic disinhibition induced by loss-of-function of S284L-mutant nAChRs (S286L-mutant nAChRs in rat ADSHE models) contributes to enhancing propagation of physiological ripple-burst high-frequency oscillation (HFO) and Erk signaling during sleep, leading to enhancement of the trafficking of pannexin1, connexin43, and P2X7 purinergic receptor to the astroglial plasma membrane. The combination of activation of physiological ripple-HFO and upregulation of astroglial hemichannels under the GABAergic disinhibition plays an important role in generation of epileptogenic fast-ripple-HFO during sleep. Therefore, loss-of-function of the S284L-mutation alone cannot drive ictogenesis but contributes to the development of epileptogenesis as an initial abnormality. Based on these recent findings using genetic rat ADSHE models, harboring the rat S286L-mutant Chrna4 corresponding to the human S284L-mutant CHRNA4, this report proposes hypothetical pathomechanisms of ADSHE. Full article

Polyamine (PA) spermine (SPM) (i) plays an essential role in the function of neurons, while (ii) accumulating predominantly in glial cells by an unknown mechanism. In addition, the translocation of SPM synthesis and redistribution in the developing and maturating retinas remains unclear. Therefore, the expression of the SPM-synthesizing enzyme, spermine synthase (SpmS), was compared in rat retinas on postnatal days 3, 21, and 120 using immunocytochemistry, Western blot (WB), and ImageJ analyses. The anti-glutamine synthetase (GS) antibody identified glial cells, and DAPI labeled the cell nuclei. At postnatal day 3 (P3), the neonatal retina shows widespread SpmS expression throughout most neuroblast cells, but absent in the developing synaptic layers and Müller cell (MCs) processes. By day 21 (P20), SpmS becomes strongly expressed in neurons, and not in glia. On day 120 (P120), SpmS was observed in synaptic areas, with significantly less presence in neuronal soma and still none in MCs. WBs showed a decrease in SpmS expression during maturation. Therefore, glial cells do not synthesize SPM, and the accumulation of SPM in MCs found earlier suggests that glial cells take up SPM via a hypothetical high-affinity SPM transporter. In glia, SPM regulates glial connexin (Cx43) and potassium (Kir4.1) channels, being a key player in CNS diseases and aging. Full article

Neonatal rodent cardiomyocytes (CMs) are a mainstay of in vitro cardiac research, yet their immature phenotype limits the study of key physiological processes such as excitation–contraction coupling (ECC) and sympathetic modulation. Here, we present a defined low-glucose, serum-free (LGSF) culture protocol that drives the structural and functional maturation of neonatal CMs and supports their integration into functional neuro-cardiac co-cultures. After 15 days in LGSF conditions, CMs exhibit elongated morphology, organized sarcomeres, polarized connexin-43, mitochondrial redistribution, and sarcoplasmic reticulum (SR) development, all closely resembling features of adult cells. These structural hallmarks were paralleled by enhanced Ca²⁺ handling, with increased SR contribution and reduced spontaneous activity, indicative of a mature ECC phenotype. When co-cultured with sympathetic neurons (SN), CMs established anatomically distinct neuro-cardiac junctions. Notably, nicotine stimulation triggered spatially restricted, reversible increases in CM Ca²⁺ transients, confined to varicosity-contacted cells. Pharmacological analysis revealed subtype-specific roles for β₁- and β₂-adrenergic receptors, and uncovered evidence of functional crosstalk between them. Our study defines a reproducible culture framework that advances CM maturation and enables the high-resolution interrogation of synaptic-like sympathetic modulation. This approach opens new avenues for mechanistic studies and drug testing in developmentally relevant neuro-cardiac systems. Full article

In forensic medicine, spotting signs of an acute myocardial infarction (AMI) right after it happens is still a tough call, especially in sudden-death cases. Standard histology often misses changes in those critical first hours because the tissue damage is too subtle to see. To tackle this, we reviewed research (1990–2023) from PubMed and Web of Science, following PRISMA guidelines. We focused on studies that used immunohistochemistry to identify markers of early AMI in both human autopsies and animal models, specifically in the first six hours post-event. Our selection process narrowed 418 records to 37 key papers. We screened 49 markers in total, but only a handful stood out for reliable diagnosis: C5b-9, cardiac troponins, dystrophin, and H-FABP—all showing high specificity. Markers like S100A1 and IL-15 also showed promise, whereas JunB and connexin-43 appeared less dependable. We believe immunohistochemistry can add real value in early AMI identification, especially when using combinations of markers chosen for complementary strengths. Still, to make this approach practical in forensic settings, we need more studies on human samples and agreement on standardized lab protocols. Full article

Traumatic brain injury (TBI) is one of the most common forms of neurotrauma, accompanied by significant disruptions in neuronal homeostasis and intercellular communication. A key protein involved in these processes is connexin 43 (Cx43), which facilitates the formation of gap junctions in the astrocytic network. In this study, using confocal and immunofluorescence microscopy, ultrastructural analysis, and molecular modeling, we investigated the dynamics of Cx43 expression and structural changes in neuroglia during various post-traumatic periods following TBI. It was shown that in the acute phase, 24 h post-injury, there is a reduction in Cx43 expression, accompanied by apoptotic neuronal degradation, disruption of nuclear NeuN localization, and destruction of cellular ultrastructure. By 7 days post-injury, a significant increase in Cx43 levels was observed, along with the formation of protein aggregates associated with pronounced reactive astrogliosis. Peripheral blood analysis revealed persistent neutrophilia, lymphopenia, and reduced monocyte levels, reflecting a systemic inflammatory response and immunosuppression, which was corroborated by a custom-trained neural network-based computer vision model. Linear regression and correlation analyses further identified a strong positive association between normalized monocyte levels and Cx43 expression, a moderate negative correlation with lymphocytes, and no significant correlation with neutrophils. Using a custom-built computer vision model, we confirmed these hematological trends and detected subtle changes, such as early increases in platelet counts, that were not captured by manual evaluation. The model demonstrated strong performance in classifying common blood cell types and proved to be a valuable tool for monitoring dynamic post-traumatic shifts in blood. Molecular dynamics modeling of Cx43 identified a pH-dependent mechanism of conformational reorganization under post-traumatic acidosis, mediated by the interaction between protonated His142 and Glu103. This mechanism mimics the structural consequences of the pathogenic E103K mutation and may play a critical role in the neurotoxic effects of Cx43 in TBI. These findings highlight the complexity of Cx43 regulation under traumatic conditions and its potential significance as a target for neuroprotective therapy. Full article

Hearing loss is a genetically heterogeneous sensory defect for which biallelic pathogenic variants in the GJB2 gene are a frequent cause. Here, we report a novel intragenic large deletion in GJB2 in a Mayan family with several members affected by congenital non-syndromic hearing loss. The analysis of the GJB2 gene in the proband was performed through Sanger sequencing. A novel homozygous 1259 bp deletion in GJB2 was identified, starting at nucleotide 248 of the coding region and ending at nucleotide 825 of the 3′-UTR (g.20188077_20189335del). Bioinformatic tools were used to predict the structural impact of the variant. This deletion would result in a truncated protein of 86 amino acids, p.(Phe83Cysfs*5), disrupting several critical domains of the connexin-26 protein. We developed an endpoint-PCR assay to test for the deletion. It was present homozygously in all affected siblings and was absent in 153 ethnically matched controls with normal hearing. Both parents and two unaffected siblings were heterozygous carriers, consistent with an autosomal recessive inheritance pattern. The identification of this novel large deletion expands the spectrum of GJB2 pathogenic variants causing non-syndromic hearing loss, and it is of concern to GJB2 screening methods that rely primarily on Sanger sequencing for its coding region. Full article

Hearing loss is often caused by genetic and environmental factors, with inherited mutations responsible for 50–60% of cases. The GJB2 gene, encoding connexin 26, is a major contributor to nonsyndromic sensorineural hearing loss (NSHL) due to its role in cellular communication critical for auditory function. In Taiwan, common deafness-associated genes include GJB2, SLC26A4, OTOF, MYO15A, and MTRNR1, which were similar to those found in other populations. The most common pathogenic genes is GJB2 mutations and the hearing level in children with GJB2 p.V37I/p.V37I or p.V37I/c.235delC was estimated to deteriorate at approximately 1 decibel hearing level (dB HL)/year. We found another common mutation in Taiwan Biobank, GJB2 p.I203T, which were identified in our data and individuals carrying this mutation experienced more severe hearing loss, suggesting a synergistic effect of these mutations on auditory impairment. We suggest GJB2 whole genetic screening is recommended for clinical management and prevention strategies in Taiwan. This study used data from the Taiwan Biobank to analyze allele frequencies of GJB2 gene variants. Predictive software (PolyPhen-2 version 2.2, SIFT for missense variants 6.2.1, MutationTaster Ensembl 112 and Alphamissense CC BY-NC-SA 4.0) assessed the pathogenicity of specific mutations. Additionally, 82 unrelated NSHL patients were screened for mutations in these genes using PCR and DNA sequencing. The study explored the correlation between genetic mutations and the severity of hearing loss in patients. Several common GJB2 mutation sites were identified from the Taiwan Biobank, including GJB2 p.V37I (7.7%), GJB2 p.I203T (6%), GJB2 p.V27I (31%), and GJB2 p.E114G (22%). Bioinformatics analysis classified GJB2 p.I203T as pathogenic, while GJB2 p.V27I and GJB2 p.E114G were considered polymorphisms. Patients with GJB2 p.I203T mutation experienced more severe hearing loss, emphasizing the potential interaction between the gene in auditory impairment. The mutation patterns of GJB2 in the Taiwanese population are similar to other East Asian regions. Although GJB2 mutations represent the predominant genetic cause of hereditary hearing loss, the corresponding mutant proteins exhibit detectable aggregation, particularly at cell–cell junctions, suggesting at least partial trafficking to the plasma membrane. Genetic screening for these mutations—especially GJB2 p.I203T (6%), GJB2 p.V27I (31%), and GJB2 p.E114G (22%)—is essential for the effective diagnosis and management of non-syndromic hearing loss (NSHL) in Taiwan. We found GJB2 p.I203T which were identified in our data and individuals carrying this mutation experienced more severe hearing loss, suggesting a synergistic effect of these mutations on auditory impairment. We suggest whole GJB2 gene sequencing in genetic screening is recommended for clinical management and prevention strategies in Taiwan. These findings have significant clinical and public health implications for the development of preventive and therapeutic strategies. Full article

Perinatal hypoxic–ischemic encephalopathy (HIE) is a condition resulting from oxygen deprivation around the time of birth and may be associated with death, brain damage, and disability. Alongside this, studies have shown that HIE may result in visual impairment. Previously, this was thought to be due to damage to the visual pathways in the brain, in a condition known as cerebral visual impairment. However, recent studies suggest that direct injury to the retina may occur after HIE. Of note, the nucleotide-binding domain, leucine-rich-containing family, and pyrin domain-containing-3 (NLRP3) inflammasome is thought to play a role in perpetuating inflammatory damage in the brain after hypoxia–ischemia (HI). As such, this study aimed to characterize retinal inflammation and the role of the NLRP3 inflammasome after HI using a modified Rice-Vannucci model in postnatal day 10 (P10) rat. Eighteen Sprague-Dawley rats were allocated evenly to three groups. Two groups received surgery to ligate the right common-carotid artery and induce HI, while another group received only sham surgery. Rats exposed to HI received subcutaneous injections of tonabersat (HI + Ton) or saline (HI + vehicle) at 1, 24 and 48 h after HI, and were culled at P17 for analysis. The results showed that the protein expression of GFAP, Iba-1, NLRP3, caspase-1 and connexin43 increased in the retina at 7 d after HI-vehicle compared with sham surgery, much more so in the ipsilateral = than the contralateral retina. Furthermore, = inflammasome components NLRP3, cleaved caspase-1 and connexin43 were significantly upregulated in the ipsilateral retina following HI-vehicle compared to the sham surgery group. Treatment with a connexin43 hemichannel blocker, tonabersat, significantly decreased the expression of the inflammasome markers, cleaved caspase-1 and connexin43, and diminished Iba-1+ cell infiltration in the ipsilateral retina. These findings suggest that direct retinal damage and inflammation may occur after HI. Furthermore, these inflammatory changes are likely mediated and propagated by activation of the NLRP3 inflammasome. Importantly, inhibition of the inflammasome by tonabersat may be able to inhibit retinal inflammation and damage, potentially preventing visual impairment after HI. Further investigation in humans is required to determine the efficacy of tonabersat in treating hypoxic–ischemic injuries to the brain and eye. Full article

Over the past four decades, cardioprotective research has revealed an extraordinary complexity of cellular and molecular mechanisms capable of mitigating ischemia/reperfusion injury (IRI). Among these, ischemic conditioning has emerged as one of the most influential discoveries: brief episodes of ischemia followed by reperfusion activate protective programs that reduce myocardial damage. These effects can be elicited locally (pre- or postconditioning) or remotely (remote conditioning), acting mainly through paracrine signaling and mitochondria-linked kinase pathways, with both early and delayed windows of protection. We have contributed to clarifying the roles of mitochondria, oxidative stress, prosurvival kinases, connexins, extracellular vesicles, and sterile inflammation, particularly via activation of the NLRP3 inflammasome. Despite robust preclinical evidence, clinical translation of these approaches has remained disappointing. The challenges largely stem from experimental models that poorly reflect real-world clinical settings—such as advanced age, comorbidities, and multidrug therapy—as well as the reliance on surrogate endpoints that do not reliably predict clinical outcomes. Nevertheless, interest in multi-target protective strategies remains strong. New lines of investigation are focusing on emerging mediators—such as gasotransmitters, extracellular vesicles, and endogenous peptides—as well as targeted modulation of inflammatory responses. Future perspectives point toward personalized cardioprotection tailored to patient metabolic and immune profiles, with special attention to high-risk populations in whom IRI continues to represent a major clinical challenge. Full article

Heart-specific overexpression of transcriptional regulator JDP2 (jun dimerization protein 2) for 5 weeks provokes paroxysmal atrial fibrillation (AF) in mice. We now investigated whether AF and atrial remodeling will be reversible upon termination of JDP2 overexpression, and whether paroxysmal AF converts to permanent AF in the presence of maintained JDP2 overexpression. Cardiac-specific JDP2 overexpression for 5 weeks, resulting in paroxysmal AF, was either continued or repressed via a tet-off system for another 5 weeks. ECGs were recorded weekly. Thereafter, heart and lung weights, and atrial mRNA and protein expression were determined. Extending JDP2 overexpression did not aggravate the AF phenotype, still paroxysmal AF, prolongation of PQ intervals, and atrial hypertrophy were present. This phenotype was completely reversible upon cessation of JDP2 overexpression. A massive downregulation of connexin40 and calcium handling proteins, including SERCA2a, calsequestrin, and ryanodine receptor, was observed in atria after prolonged JDP2 overexpression. In conclusion, atrial remodeling and paroxysmal AF under JDP2 overexpression are not sufficient to maintain or aggravate AF in the absence of JDP2. The comparison of the two groups indicates that the downregulation of calcium proteins and connexins is an important factor in the maintenance of the disease. Full article

Patients with ulcerative colitis exhibit an increased risk for supraventricular arrhythmia during the active disease phase of the disease and show signs of atrial electrophysiological remodeling in remission. The goal of this study was to determine the basis for colitis-induced changes in atrial excitability. In a mouse model (C57BL/6; 3 months) of dextran sulfate sodium (DSS)-induced active colitis (3.5% weight/volume, 7 days), electrocardiograms (ECG) revealed altered atrial electrophysiological properties with a prolonged P-wave duration and PR interval. ECG changes coincided with a decreased atrial conduction velocity in Langendorff perfused hearts. Action potentials (AP) recorded from isolated atrial myocytes displayed an attenuated maximal upstroke velocity and amplitude during active colitis, as well as a prolonged AP duration (APD). Voltage clamp analysis revealed a colitis-induced shift in the voltage-dependent activation of the Na-current (I_Na) to more depolarizing voltages. In addition, protein levels of Na_v1.5 protein and connexin isoform Cx43 were reduced. APD prolongation depended on a reduction in the transient outward K-current (I_to) mostly generated by K_v4.2 channels. The changes in ECG, atrial conductance, and APD were reversible upon remission. The change in conduction velocity predominantly depended on the reversibility of the reduced Cx43 and Na_v1.5 expression. Treatment of mice with inhibitors of Angiotensin-converting enzyme (ACE) or Angiotensin II (AngII) receptor type 1 (AT1R) prevented the colitis-induced atrial electrophysiological remodeling. Our data support a colitis-induced increase in AngII signaling that promotes atrial electrophysiological remodeling and puts colitis patients at an increased risk for atrial arrhythmia. Full article

Background: Kölliker’s organ (KO), a transient structure in the cochlea, plays a critical role in the auditory maturation of mammals, particularly during embryonic and early postnatal development. This organ is essential for the proper differentiation and function of cochlear cells, acting as a pivotal source of signalling molecules that influence hair cell development and synaptic connectivity. Methods: This study systematically analyses the literature according to the PRISMA statement in order to evaluate the function roles of KO during cochlea development, reporting the molecular mechanisms and signalling pathways involved. Results: From our study, it emerged that KO supporting cells release adenosine triphosphate (ATP) through connexin hemichannels, initiating a cascade of intracellular calcium (Ca²⁺) signalling in adjacent inner hair cells (IHCs). This signalling promotes the release of glutamate, facilitating synaptic excitation of afferent nerve fibres and enhancing auditory neuron maturation prior to the onset of hearing. Additionally, the spontaneous electrical activity generated within KO supports the establishment of essential neural connections in the auditory pathway. The dynamic interplay between ATP release, Ca²⁺ signalling, and morphological changes in KO is crucial for cochlear compartmentalisation and fluid regulation, contributing to the formation of endolymph and perilymph. Furthermore, KO supports cellular plasticity and may provide a reservoir of precursor cells capable of trans-differentiating into hair cells under specific conditions. Conclusions: Dysregulation of KO function or delayed degeneration of its supporting cells has been implicated in auditory disorders, underscoring the importance of this organ in normal cochlear development and auditory function. Despite its identification over a century ago, further investigation is necessary to elucidate the molecular mechanisms underlying KO’s contributions to auditory maturation, particularly in human physiology. Full article

A part of autosomal dominant sleep-related hypermotor epilepsy (ADSHE) is caused by mutant CHRNA4. The pathomechanisms underlying motor seizures followingly brief/sudden awakening (paroxysmal arousal) in ADSHE seizures remain to be clarified. This study determined extracellular levels of ACh and L-glutamate in the pedunculopontine nucleus (PPN) and its projection regions, including the thalamus and basal ganglia, during wakefulness, slow-wave sleep (SWS) and paroxysmal arousal of transgenic rats bearing rat S286L-mutant Chrna4 (S286L-TG), corresponding to human S284L-mutant CHRNA4, using microdialysis. The expression of connexin43 and pannexin1 in the plasma membrane of the PPN was determined using capillary immunoblotting. The expressions of connexin43 and pannexin1 in the PPN plasma membrane of S286L-TG were larger than the wild type. The extracellular L-glutamate levels in the PPN and projection regions of S286L-TG consistently increased during both wakefulness and SWS compared to the wild type. The extracellular levels of ACh and L-glutamate in the PPN and projection regions decreased accompaning SWS in the wild type. In S286L-TG, this decreasing extracellular ACh level was observed, whereas decreasing L-glutamate level was impaired. Both extracellular levels of ACh and L-glutamate in the PPN and projection regions drastically increased during paroxysmal arousal. Hemichannel inhibitors suppressed the increasing releases of ACh and L-glutamate induced by paroxysmal arousal but decreased and did not affect extracellular levels of L-glutamate and ACh during wakefulness and SWS, respectively. In particular, under hemichannels inhibition, decreasing L-glutamate release accompanying SWS was observed in S286L-TG. This study elucidated that enhanced hemichannels are predominantly involved in the dysfunction of glutamatergic transmission compared to AChergic transmission during the interictal stage in S286L-TG, whereas the hyperactivation of hemichannels contributes to the generation of paroxysmal arousal. Therefore, the hyperactivated excitatory tripartite synaptic transmission associated with hemichannels in the PPN and projection regions plays important roles in epileptogenesis/ictogenesis in S286L-TG. Full article

Endothelial progenitor cells (EPC) are considered to support neovascularization and endothelial repair by being incorporated into newly formed or injured vessels and by improving vascularization in a paracrine manner by secreting proangiogenic factors. Here, we studied the role of gap junctional communication between EPC and endothelial cells in long-term co-cultures in vitro. The cultivation of endothelial cells together with mouse embryonic EPC (E 7.5) induced the spontaneous formation of angiogenic networks after 3–6 days consisting of both cell types, but not in the respective monocultures, whereas their respective cultivation on a basement matrix induced the formation of tube-like structures, as expected. The angiogenic network formation could not be mimicked by the incubation of endothelial cells with supernatants of EPC only. We therefore hypothesized that direct interaction and cell-cell communication is required to induce the angiogenic network formation in co-cultures with endothelial cells. Expression analysis demonstrated expression of the gap junctional protein connexin 43 (Cx43) in EPC. Moreover, dye injection studies as well as FACS analysis identified gap junctional communication between endothelial cells and EPC. The inhibition of gap junctions by pharmacological blockers significantly reduced the angiogenic network formation, confirming that gap junctional communication between both cell types is required for this process. Full article