BACKGROUND: Multiple Sclerosis (MS) is an autoimmune neurodegenerative disease, whose primary hallmark is the occurrence of inflammatory lesions in white and grey matter structures. Increasing evidence in MS patients and respective murine models reported an impaired ionic homeostasis driven by inflammatory-demyelination, thereby profoundly affecting signal propagation. However, the impact of a focal inflammatory lesion on single-cell and network functionality has hitherto not been fully elucidated.
OBJECTIVE: In this study, we sought to determine the consequences of a localized cortical inflammatory lesion on the excitability and firing pattern of thalamic neurons in the auditory system. Moreover, we tested the neuroprotective effect of Retigabine (RTG), a specific Kv7 channel opener, on disease outcome.
METHODS: To resemble the human disease, we focally administered pro-inflammatory cytokines, TNF-α and IFN-γ, in the primary auditory cortex (A1) of MOG35-55 immunized mice. Thereafter, we investigated the impact of the induced inflammatory milieu on afferent thalamocortical (TC) neurons, by performing ex vivo recordings. Moreover, we explored the effect of Kv7 channel modulation with RTG on auditory information processing, using in vivo electrophysiological approaches.
RESULTS: Our results revealed that a cortical inflammatory lesion profoundly affected the excitability and firing pattern of neighboring TC neurons. Noteworthy, RTG restored control-like values and TC tonotopic mapping.
CONCLUSIONS: Our results suggest that RTG treatment might robustly mitigate inflammation-induced altered excitability and preserve ascending information processing.

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The growing interest in Kv7.2/7.3 agonists originates from the involvement of these channels in several brain hyperexcitability disorders. In particular, Kv7.2/7.3 mutants have been clearly associated with epileptic encephalopathies (DEEs) as well as with a spectrum of focal epilepsy disorders, often associated with developmental plateauing or regression. Nevertheless, there is a lack of available therapeutic options, considering that retigabine, the only molecule used in clinic as a broad-spectrum Kv7 agonist, has been withdrawn from the market in late 2016. This is why several efforts have been made both by both academia and industry in the search for suitable chemotypes acting as Kv7.2/7.3 agonists. In this context, in silico methods have played a major role, since the precise structures of different Kv7 homotetramers have been only recently disclosed. In the present review, the computational methods used for the design of Kv.7.2/7.3 small molecule agonists and the underlying medicinal chemistry are discussed in the context of their biological and structure-function properties.

The KCNQ family is comprised of five genes and the expression products form voltage-gated potassium channels (Kv7.1-7.5) that have a major impact upon cellular physiology in many cell types. Each functional Kv7 channel forms as a tetramer that often associates with proteins encoded by the KCNE gene family (KCNE1-5) and is critically reliant upon binding of phosphatidylinositol bisphosphate (PIP2) and calmodulin. Other modulators like A-kinase anchoring proteins, ubiquitin ligases and Ca-calmodulin kinase II alter Kv7 channel function and trafficking in an isoform specific manner. It has now been identified that for Kv7.4, G protein βγ subunits (Gβγ) can be added to the list of key regulators and is paramount for channel activity. This article provides an overview of this nascent field of research, highlighting themes and directions for future study.

Despite the increasing availability of genomic data and enhanced data analysis procedures, predicting the severity of associated diseases remains elusive in the absence of clinical descriptors. To address this challenge, we have focused on the KV7.2 voltage-gated potassium channel gene (KCNQ2), known for its link to developmental delays and various epilepsies, including self-limited benign familial neonatal epilepsy and epileptic encephalopathy. Genome-wide tools often exhibit a tendency to overestimate deleterious mutations, frequently overlooking tolerated variants, and lack the capacity to discriminate variant severity. This study introduces a novel approach by evaluating multiple machine learning (ML) protocols and descriptors. The combination of genomic information with a novel Variant Frequency Index (VFI) builds a robust foundation for constructing reliable gene-specific ML models. The ensemble model, MLe-KCNQ2, formed through logistic regression, support vector machine, random forest and gradient boosting algorithms, achieves specificity and sensitivity values surpassing 0.95 (AUC-ROC > 0.98). The ensemble MLe-KCNQ2 model also categorizes pathogenic mutations as benign or severe, with an area under the receiver operating characteristic curve (AUC-ROC) above 0.67. This study not only presents a transferable methodology for accurately classifying KCNQ2 missense variants, but also provides valuable insights for clinical counseling and aids in the determination of variant severity. The research context emphasizes the necessity of precise variant classification, especially for genes like KCNQ2, contributing to the broader understanding of gene-specific challenges in the field of genomic research. The MLe-KCNQ2 model stands as a promising tool for enhancing clinical decision making and prognosis in the realm of KCNQ2-related pathologies.

The mechanisms behind renal vasodilatation elicited by stimulation of β-adrenergic receptors are not clarified. As several classes of K channels are potentially activated, we tested the hypothesis that KV7 and BKCa channels contribute to the decreased renal vascular tone in vivo and in vitro. Changes in renal blood flow (RBF) during β-adrenergic stimulation were measured in anaesthetized rats using an ultrasonic flow probe. The isometric tension of segmental arteries from normo- and hypertensive rats and segmental arteries from wild-type mice and mice lacking functional KV7.1 channels was examined in a wire-myograph. The β-adrenergic agonist isoprenaline increased RBF significantly in vivo. Neither activation nor inhibition of KV7 and BKCa channels affected the β-adrenergic RBF response. In segmental arteries from normo- and hypertensive rats, inhibition of KV7 channels significantly decreased the β-adrenergic vasorelaxation. However, inhibiting BKCa channels was equally effective in reducing the β-adrenergic vasorelaxation. The β-adrenergic vasorelaxation was not different between segmental arteries from wild-type mice and mice lacking KV7.1 channels. As opposed to rats, inhibition of KV7 channels did not affect the murine β-adrenergic vasorelaxation. Although inhibition and activation of KV7 channels or BKCa channels significantly changed baseline RBF in vivo, none of the treatments affected β-adrenergic vasodilatation. In isolated segmental arteries, however, inhibition of KV7 and BKCa channels significantly reduced the β-adrenergic vasorelaxation, indicating that the regulation of RBF in vivo is driven by several actors in order to maintain an adequate RBF. Our data illustrates the challenge in extrapolating results from in vitro to in vivo conditions.

KCNQ (Kv7) channels are voltage-gated, phosphatidylinositol 4,5-bisphosphate- (PIP2-) modulated potassium channels that play essential roles in regulating the activity of neurons and cardiac myocytes. Hundreds of mutations in KCNQ channels are closely related to various cardiac and neurological disorders, such as long QT syndrome, epilepsy, and deafness, which makes KCNQ channels important drug targets. During the past several years, the application of single-particle cryo-electron microscopy (cryo-EM) technique in the structure determination of KCNQ channels has greatly advanced our understanding of their molecular mechanisms. In this review, we summarize the currently available structures of KCNQ channels, analyze their special voltage gating mechanism, and discuss their activation mechanisms by both the endogenous membrane lipid and the exogenous synthetic ligands. These structural studies of KCNQ channels will guide the development of drugs targeting KCNQ channels.

K+ channels play potent roles in the process of neurotransmitter release by influencing the action potential waveform and modulating neuronal excitability and release probability. These diverse effects of K+ channel activation are ensured by the wide variety of K+ channel genes and their differential expression in different cell types. Accordingly, a variety of K+ channels have been implicated in regulating neurotransmitter release, including the Ca2+- and voltage-gated K+ channel Slo1 (also known as BK channel), voltage-gated K+ channels of the Kv3 (Shaw-type), Kv1 (Shaker-type), and Kv7 (KCNQ) families, G-protein-gated inwardly rectifying K+ (GIRK) channels, and SLO-2 (a Ca2+-. Cl-, and voltage-gated K+ channel in C. elegans). These channels vary in their expression patterns, subcellular localization, and biophysical properties. Their roles in neurotransmitter release may also vary depending on the synapse and physiological or experimental conditions. This chapter summarizes key findings about the roles of K+ channels in regulating neurotransmitter release.

Cannabidiol (CBD) is used clinically as an anticonvulsant. Its precise mechanism of action has remained unclear. CBD was recently demonstrated to enhance the activity of the neuronal KV 7.2/7.3 channel, which may be one important contributor to CBD anticonvulsant effect. Curiously, CBD inhibits the closely related cardiac KV 7.1/KCNE1 channel. Whether and how CBD affects other KV 7 subtypes remains uninvestigated and the CBD interaction sites mediating these diverse effects remain unknown.
Here, we used electrophysiology, molecular dynamics simulations, molecular docking and site-directed mutagenesis to address these questions.
We found that CBD modulates the activity of all human KV 7 subtypes and that the effects are subtype dependent. CBD enhanced the activity of KV 7.2-7.5 subtypes, seen as a V50 shift towards more negative voltages or increased maximum conductance. In contrast, CBD inhibited the KV 7.1 and KV 7.1/KCNE1 channels, seen as a V50 shift towards more positive voltages and reduced conductance. In KV 7.2 and KV 7.4, we propose a CBD interaction site at the subunit interface in the pore domain that overlaps with the interaction site of other compounds, notably the anticonvulsant retigabine. However, CBD relies on other residues for its effects than the conserved tryptophan that is critical for retigabine effects. We propose a similar, though not identical CBD site in KV 7.1, with a non-conserved phenylalanine being important.
We identify novel targets of CBD, contributing to a better understanding of CBD clinical effects and provide mechanistic insights into how CBD modulates different KV 7 subtypes.

KV7 channels exert a pivotal role regulating vascular tone in several vascular beds. In this context, KV7 channel agonists represent an attractive strategy for the treatment of pulmonary arterial hypertension (PAH). Therefore, in this study, we have explored the pulmonary vascular effects of the novel KV7 channel agonist URO-K10. Consequently, the vasodilator and electrophysiological effects of URO-K10 were tested in rat and human pulmonary arteries (PA) and PA smooth muscle cells (PASMC) using myography and patch-clamp techniques. Protein expression was also determined by Western blot. Morpholino-induced knockdown of KCNE4 was assessed in isolated PA. PASMC proliferation was measured by BrdU incorporation assay. In summary, our data show that URO-K10 is a more effective relaxant of PA than the classical KV7 activators retigabine and flupirtine. URO-K10 enhanced KV currents in PASMC and its electrophysiological and relaxant effects were inhibited by the KV7 channel blocker XE991. The effects of URO-K10 were confirmed in human PA. URO-K10 also exhibited antiproliferative effects in human PASMC. Unlike retigabine and flupirtine, URO-K10-induced pulmonary vasodilation was not affected by morpholino-induced knockdown of the KCNE4 regulatory subunit. Noteworthy, the pulmonary vasodilator efficacy of this compound was considerably increased under conditions mimicking the ionic remodelling (as an in vitro model of PAH) and in PA from monocrotaline-induced pulmonary hypertensive rats. Taking all together, URO-K10 behaves as a KCNE4-independent KV7 channel activator with much increased pulmonary vascular effects compared to classical KV7 channel activators. Our study identifies a promising new drug in the context of PAH.