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Depression is a common and complex psychiatric illness with multiple clinical symptoms, even leading to the disability and suicide. Owing to the partial understanding of the pathogenesis of depressive-like disorders, available pharmacotherapeutic strategies are developed mainly based on the "monoamine hypothesis", resulting in a limited effectiveness and a number of adverse effects in the clinical practice. The concept of multiple pathogenic factors be helpful for clarifying the etiology of depression and developing the antidepressants. It is well documented that K + channels serve crucial roles in modulating the neuronal excitability and neurotransmitter release in the brain, and abnormality of these channels participated in the pathogenic process of diverse central nervous system (CNS) pathologies, such as seizure and Alzheimer's disease (AD). The clinical and preclinical evidence also delineates that the involvement of several types of K + channels in depressive-like behaviors appear to be evident, suggesting these channels being one of the multiple factors in the etiology of this debilitating disorder. Emerging data manifest that diverse antidepressants impact distinct K + channels, such as Kv, Kir and K 2P , meaning the functioning of these drug via a "multi-target" manner. On the other hand, the scenario of antidepressants impinging K + channels could render an alternative interpretation for the pharmacological effectiveness and numerous side effects in clinical trials. Furthermore, these channels serve to be considered as a "druggable target" to develop novel therapeutic compound to antagonize this psychiatry., (© 2024. The Author(s).)

Background & Aims: The treatment of cardiovascular diseases (CVD) could greatly benefit from using nitric oxide (NO) donors. This study aimed to investigate the mechanisms of action of NONO2P that contribute to the observed responses in the mesenteric artery. The hypothesis was that NONO2P would have similar pharmacological actions to sodium nitroprusside (SNP) and NO., Methods: Male Wistar rats were euthanized to isolate the superior mesenteric artery for isometric tension recordings. NO levels were measured using the DAF-FM/DA dye, and cyclic guanosine monophosphate (cGMP) levels were determined using a cGMP-ELISA Kit., Results: NONO2P presented a similar maximum efficacy to SNP. The free radical of NO (NO • ) scavengers (PTIO; 100 μM and hydroxocobalamin; 30 μM) and nitroxyl anion (NO - ) scavenger (L-cysteine; 3 mM) decreased relaxations promoted by NONO2P. The presence of the specific soluble guanylyl cyclase (sGC) inhibitor (ODQ; 10 μM) nearly abolished the vasorelaxation. The cGMP-dependent protein kinase (PKG) inhibition (KT5823; 1 μM) attenuated the NONO2P relaxant effect. The vasorelaxant response was significantly attenuated by blocking inward rectifying K + channels (K ir ), voltage-operated K + channels (K V ), and large conductance Ca 2+ -activated K + channels (BK Ca ). NONO2P-induced relaxation was attenuated by cyclopiazonic acid (10 μM), indicating that sarcoplasmic reticulum Ca2+-ATPase (SERCA) activation is involved in this relaxation. Moreover, NONO2P increased NO levels in endothelial cells and cGMP production., Conclusions: NONO2P induces vasorelaxation with the same magnitude as SNP, releasing NO • and NO - . Its vasorelaxant effect involves sGC, PKG, K + channels opening, and SERCA activation, suggesting its potential as a therapeutic option for CVD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Discovered just over 25 years ago in cerebellar Purkinje neurons, resurgent Na current was originally described operationally as a component of voltage-gated Na current that flows upon repolarization from relatively depolarized potentials and speeds recovery from inactivation, increasing excitability. Its presence in many excitable cells and absence from others has raised questions regarding its biophysical and molecular mechanisms. Early studies proposed that Na channels capable of generating resurgent current are subject to a rapid open-channel block by an endogenous blocking protein, which binds upon depolarization and unblocks upon repolarization. Since the time that this mechanism was suggested, many physiological and structural studies of both Na and K channels have revealed aspects of gating and conformational states that provide insights into resurgent current. These include descriptions of domain movements for activation and inactivation, solution of cryo-EM structures with pore-blocking compounds, and identification of native blocking domains, proteins, and modulatory subunits. Such results not only allow the open-channel block hypothesis to be refined but also link it more clearly to research that preceded it. This review considers possible mechanisms for resurgent Na current in the context of earlier and later studies of ion channels and suggests a framework for future research., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Competing Interests: Declaration of interests The authors declare no competing interests.

A variety of equilibrium and non-equilibrium methods have been used in a multidisciplinary approach to study the conformational landscape associated with the binding of different cations to the pore of potassium channels. These binding processes, and the conformational changes resulting therefrom, modulate the functional properties of such integral membrane properties, revealing these permeant and blocking cations as true effectors of such integral membrane proteins. KcsA, a prototypic K + channel from Streptomyces lividans, has been extensively characterized in this regard. Here, we revise several fluorescence-based approaches to monitor cation binding under different experimental conditions in diluted samples, analyzing the advantages and disadvantages of each approach. These studies have contributed to explain the selectivity, conduction, and inactivation properties of K + channels at the molecular level, together with the allosteric communication between the two gates that control the ion channel flux, and how they are modulated by lipids., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Jose Antonio Poveda Larrosa reports administrative support, article publishing charges, and equipment, drugs, or supplies were provided by Spain Ministry of Science and Innovation., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The relationships between parathyroid hormone (PTH) secretion and parathyroid cell membrane potential, including the identities and roles of K + channels that regulate and/or modulate membrane potential are not well defined. Here we have used Western blot/immunohistochemistry as well as patch-clamp and perifusion techniques to identify and localize specific K + channels in parathyroid cells and to investigate their roles in the control of membrane potential and PTH secretion. We also re-investigated the relationship between membrane potential and exocytosis. We showed that in single human parathyroid cells K + current is dependent on at least two types of Ca 2+ -activated K + channels: a small-conductance Ca 2+ -activated K + channel (K SK ) and a large-conductance voltage and Ca 2+ -activated K + channel (K BK ). These channels were sensitive to specific peptide blocking toxins including apamin, charybdotoxin, and iberiotoxin. These channels confer sensitivity of the membrane potential in single cells to high extracellular K + , TEA, and peptide toxins. Blocking of K BK potently inhibited K + channel current, and K BK was shown to be expressed in the plasma membrane of parathyroid cells. In addition, when using the capacitance technique as an indicator of exocytosis, clamping the parathyroid cell at -60 mV prevented exocytosis, whereas holding the membrane potential at 0 mV facilitated it. Taken together, the results show that human parathyroid cells have functional K BK and K SK channels but the data presented herein suggest that K BK /K SK channels likely contribute to the maintenance of the membrane potential, and that membrane potential, per se, modulates exocytosis independently of [Ca 2+ ] i ., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Mutations of the Na + -activated K + channel Slack (KCNT1) are associated with terrible epilepsy syndromes that already begin in infancy. Here we report increased severity of acute kainic acid-induced seizures in adult and juvenile Slack knockout mice (Slack -/- ) in vivo. Fittingly, we find exacerbation of cell death following kainic acid exposure in organotypic hippocampal slices as well as dissociated hippocampal cultures from Slack -/- in vitro. Furthermore, in cultured Slack -/- neurons, kainic acid-triggered Ca 2+ influx and K + efflux as well as depolarization-induced tetrodotoxin-sensitive inward currents are higher compared to the respective controls. This apparent changes in ion homeostasis could possibly explain altered action potential kinetics of Slack -/- neurons: steeper rise slope, decreased threshold, and duration of afterhyperpolarization, which ultimately lead to higher action potential frequencies during kainic acid application or injection of depolarizing currents. Based on our data, we propose Slack as crucial gatekeeper of neuronal excitability to acutely limit seizure severity., (© 2023. Springer Nature Limited.)

We have experimented with isolated cardiomyocytes of mollusks Helix. During the whole-cell patch-clamp recordings of K + currents a considerable decrease in amplitude was observed upon repeated voltage steps at 0.96 Hz. For these experiments, ventricular cells were depolarized to identical + 20 mV from a holding potential of - 50 mV. The observed spontaneous inhibition of outward currents persisted in the presence of 4-aminopyridine, tetraethylammonium chloride or E-4031, the selective class III antiarrhythmic agent that blocks HERG channels. Similar tendency was retained when components of currents sensitive to either 4-AP or TEA were mathematically subtracted. Waveforms of currents sensitive to 1 and 10 micromolar concentration of E-4031 were distinct comprising prevailingly those activated during up to 200 ms pulses. The outward current activated by a voltage ramp at 60 mV x s -1 rate revealed an inward rectification around + 20 mV. This feature closely resembles those of the mammalian cardiac delayed rectifier I Kr ., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Irisin, an exercise-induced myokine, has been shown to have a peripheral vasodilator effect. However, little is known about the mechanisms underlying its effects. In this study, it was aimed to investigate the vasoactive effects of irisin on rat thoracic aorta, and the hypothesis that voltage-gated potassium (K V ) channels, ATP-sensitive potassium (K ATP ) channels, small-conductance calcium-activated potassium (SK Ca ) channels, large-conductance calcium-activated potassium (BK Ca ) channels, intermediate-conductance calcium-activated potassium (IK Ca ) channels, inward rectifier potassium (K ir ) channels, and two-pore domain potassium (K 2P ) channels may have roles in these effects. Isometric contraction-relaxation responses of isolated thoracic aorta rings were measured with an organ bath model. The steady contraction was induced with both 10 -5 M phenylephrine and 45 mM KCl, and then the concentration-dependent responses of irisin (10 -9 -10 -6 M) were examined. Irisin exerted the vasorelaxant effects in both endothelium-intact and -denuded aortic rings at concentrations of 10 -8 , 10 -7 , and 10 -6 M (p < 0.001). Besides, K V channel blocker 4-aminopyridine, K ATP channel blocker glibenclamide, SK Ca channel blocker apamin, BK Ca channel blockers tetraethylammonium and iberiotoxin, IK Ca channel blocker TRAM-34, and K ir channel blocker barium chloride incubations significantly inhibited the irisin-induced relaxation responses. However, incubation of K 2P TASK-1 channel blocker anandamide did not cause a significant decrease in the relaxation responses of irisin. In conclusion, the first physiological findings were obtained regarding the functional relaxing effects of irisin in rat thoracic aorta. Furthermore, this study is the first to report that irisin-induced relaxation responses are associated with the activity of K V , K ATP , SK Ca , BK Ca , IK Ca , and K ir channels., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The field of mitochondrial ion channels underwent a rapid development during the last decade, thanks to the molecular identification of some of the nuclear-encoded organelle channels and to advances in strategies allowing specific pharmacological targeting of these proteins. Thereby, genetic tools and specific drugs aided definition of the relevance of several mitochondrial channels both in physiological as well as pathological conditions. Unfortunately, in the case of mitochondrial K + channels, efforts of genetic manipulation provided only limited results, due to their dual localization to mitochondria and to plasma membrane in most cases. Although the impact of mitochondrial K + channels on human diseases is still far from being genuinely understood, pre-clinical data strongly argue for their substantial role in the context of several pathologies, including cardiovascular and neurodegenerative diseases as well as cancer. Importantly, these channels are druggable targets, and their in-depth investigation could thus pave the way to the development of innovative small molecules with huge therapeutic potential. In the present review we summarize the available experimental evidence that mechanistically link mitochondrial potassium channels to the above pathologies and underline the possibility of exploiting them for therapy., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest and the paper has not been published elsewhere., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Potassium ions have widespread roles in cellular homeostasis and activation as a consequence of their large outward concentration gradient across the surface membrane and ability to rapidly move through K + -selective ion channels. In platelets, the predominant K + channels include the voltage-gated K + channel Kv1.3, and the intermediate conductance Ca 2+ -activated K + channel KCa3.1, also known as the Gardos channel. Inwardly rectifying potassium GIRK channels and KCa1.1 large conductance Ca 2+ -activated K + channels have also been reported in the platelet, although they remain to be demonstrated using electrophysiological techniques. Whole-cell patch clamp and fluorescent indicator measurements in the platelet or their precursor cell reveal that Kv1.3 sets the resting membrane potential and KCa3.1 can further hyperpolarize the cell during activation, thereby controlling Ca 2+ influx. Kv1.3 -/- mice exhibit an increased platelet count, which may result from an increased splenic megakaryocyte development and longer platelet lifespan. This review discusses the evidence in the literature that Kv1.3, KCa3.1. GIRK and KCa1.1 channels contribute to a number of platelet functional responses, particularly collagen-evoked adhesion, procoagulant activity and GPCR function. Putative roles for other K + channels and known accessory proteins which to date have only been detected in transcriptomic or proteomic studies, are also discussed.

The aim of this study was to examine if the peripheral antinociception of α-bisabolol involves the participation of nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) synthesis followed by K + channel opening in the formalin test. Wistar rats were injected in the dorsal surface of the right hind paw with formalin (1%). Rats received a subcutaneous injection into the dorsal surface of the paw of vehicles or increasing doses of α-bisabolol (100-300 µg/paw). To determine whether the peripheral antinociception induced by α-bisabolol was mediated by either the opioid receptors or the NO-cGMP-K + channels pathway, the effect of pretreatment (10 min before formalin injection) with the appropriate vehicles, naloxone, naltrexone, N G -nitro-l-arginine methyl ester (L-NAME), 1H-[1,2,4]-oxadiazolo[4,2-a]quinoxalin-1-one (ODQ), glibenclamide, glipizide, apamin, charybdotoxin, tetraethylammonium, or 4-aminopyridine on the antinociceptive effects induced by local peripheral α-bisabolol (300 µg/paw) were assessed. α-Bisabolol produced antinociception during both phases of the formalin test. α-Bisabolol antinociception was blocked by L-NAME, ODQ, and all the K + channels blockers. The peripheral antinociceptive effect produced by α-bisabolol was not blocked by the opioid receptor inhibitors. α-Bisabolol was able to active the NO-cGMP-K + channels pathway to produce its antinoceptive effect. The participation of opioid receptors in the peripheral local antinociception induced by α-bisabolol is excluded.

Potassium (K + ) channels are highly conserved proteins found in all domains of life, that allow for selective movement of K + ions across membranes. Despite their broad distribution, the physiological roles of individual members of this diverse channel family have only been thoroughly explored in eukaryotic systems, where they have critical functions in a variety of cellular processes. Recent studies have demonstrated that bacterial K + channels have integral roles in electrical signaling, information propagation, and intercellular communication. We discuss how these novel findings impact our understanding of bacterial physiology and the need to continue to explore the native roles of ion channels in microbes., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The large K + channel functional diversity in the pulmonary vasculature results from the multitude of genes expressed encoding K + channels, alternative RNA splicing, the post-transcriptional modifications, the presence of homomeric or heteromeric assemblies of the pore-forming α-subunits and the existence of accessory β-subunits modulating the functional properties of the channel. K + channels can also be regulated at multiple levels by different factors controlling channel activity, trafficking, recycling and degradation. The activity of these channels is the primary determinant of membrane potential (Em) in pulmonary artery smooth muscle cells (PASMC), providing an essential regulatory mechanism to dilate or contract pulmonary arteries (PA). K + channels are also expressed in pulmonary artery endothelial cells (PAEC) where they control resting Em, Ca 2+ entry and the production of different vasoactive factors. The activity of K + channels is also important in regulating the population and phenotype of PASMC in the pulmonary vasculature, since they are involved in cell apoptosis, survival and proliferation. Notably, K + channels play a major role in the development of pulmonary hypertension (PH). Impaired K + channel activity in PH results from: 1) loss of function mutations, 2) downregulation of its expression, which involves transcription factors and microRNAs, or 3) decreased channel current as a result of increased vasoactive factors (e.g., hypoxia, 5-HT, endothelin-1 or thromboxane), exposure to drugs with channel-blocking properties, or by a reduction in factors that positively regulate K + channel activity (e.g., NO and prostacyclin). Restoring K + channel expression, its intracellular trafficking and the channel activity is an attractive therapeutic strategy in PH., Competing Interests: Declaration of Competing Interest The authors declare that there is no conflict of interest., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Neuronal ankyrins cluster and link membrane proteins to the actin and spectrin-based cytoskeleton. Among the three vertebrate ankyrins, little is known about neuronal Ankyrin-R (AnkR). We report AnkR is highly enriched in Pv + fast-spiking interneurons in mouse and human. We identify AnkR-associated protein complexes including cytoskeletal proteins, cell adhesion molecules (CAMs), and perineuronal nets (PNNs). We show that loss of AnkR from forebrain interneurons reduces and disrupts PNNs, decreases anxiety-like behaviors, and changes the intrinsic excitability and firing properties of Pv + fast-spiking interneurons. These changes are accompanied by a dramatic reduction in Kv3.1b K + channels. We identify a novel AnkR-binding motif in Kv3.1b, and show that AnkR is both necessary and sufficient for Kv3.1b membrane localization in interneurons and at nodes of Ranvier. Thus, AnkR regulates Pv + fast-spiking interneuron function by organizing ion channels, CAMs, and PNNs, and linking these to the underlying β1 spectrin-based cytoskeleton., Competing Interests: SS, CL, YO, LT, AA, SN, JO, AB, MC, MX, MR No competing interests declared, (© 2021, Stevens et al.)

Sinus node (SAN) dysfunction (SND) manifests as low heart rate (HR) and is often accompanied by atrial tachycardia or atrioventricular (AV) block. The only currently available therapy for chronic SND is the implantation of an electronic pacemaker. Because of the growing burden of SND in the population, new pharmacological therapies of chronic SND and heart block are desirable. We developed a collection of genetically modified mouse strains recapitulating human primary SND associated with different degrees of AV block. These mice were generated with genetic ablation of L-type Ca v 1.3 (Ca v 1.3 -/- ), T-type Ca v 3.1 (Ca v 3.1 -/- ), or both (Ca v 1.3 -/- /Ca v 3.1 -/- ). We also studied mice haplo-insufficient for the Na + channel Na v 1.5 (Na v 1.5 +/ ) and mice in which the cAMP-dependent regulation of hyperpolarization-activated f-(HCN4) channels has been abolished (HCN4-CNBD). We analysed, by telemetric ECG recording, whether pharmacological inhibition of the G-protein-activated K + current (I KACh ) by the peptide tertiapin-Q could improve HR and AV conduction in these mouse strains. Tertiapin-Q significantly improved the HR of Ca v 1.3 -/- (19%), Ca v 1.3 -/- /Ca v 3.1 -/- (23%) and HCN4-CNBD (14%) mice. Tertiapin-Q also improved cardiac conduction of Na v 1.5 +/- mice by 24%. Our data suggest that the development of pharmacological I KACh inhibitors for the management of SND and conduction disease is a viable approach.

Potassium channels are present in every living cell and essential to setting up a stable, non-zero transmembrane electrostatic potential which manifests the off-equilibrium livelihood of the cell. They are involved in other cellular activities and regulation, such as the controlled release of hormones, the activation of T-cells for immune response, the firing of action potential in muscle cells and neurons, etc. Pharmacological reagents targeting potassium channels are important for treating various human diseases linked to dysfunction of the channels. High-resolution structures of these channels are very useful tools for delineating the detailed chemical basis underlying channel functions and for structure-based design and optimization of their pharmacological and pharmaceutical agents. Structural studies of potassium channels have revolutionized biophysical understandings of key concepts in the field - ion selectivity, conduction, channel gating, and modulation, making them multi-modality targets of pharmacological regulation. In this chapter, I will select a few high-resolution structures to illustrate key structural insights, proposed allostery behind channel functions, disagreements still open to debate, and channel-lipid interactions and co-evolution. The known structural consensus allows the inference of conserved molecular mechanisms shared among subfamilies of K + channels and makes it possible to develop channel-specific pharmaceutical agents., (© 2021. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

Resistance towards known antimalarial drugs poses a significant problem, urging for novel drugs that target vital proteins in the malaria parasite Plasmodium falciparum. However, recombinant production of malaria proteins is notoriously difficult. To address this, we have investigated two putative K + channels, PfKch1 and PfKch2, identified in the P. falciparum genome. We show that PfKch1 and PfKch2 and a C-terminally truncated version of PfKch1 (PfKch1 1-1094 ) could indeed be functionally expressed in vivo, since a K + -uptake deficient Saccharomyces cerevisiae strain was complemented by the P. falciparum cDNAs. PfKch1 1-1094 -GFP and GFP-PfKch2 fusion proteins were overexpressed in yeast, purified and reconstituted in lipid bilayers to determine their electrophysiological activity. Single channel conductance amounted to 16 ± 1 pS for PfKch1 1-1094 -GFP and 28 ± 2 pS for GFP-PfKch2. We predicted regulator of K + -conductance (RCK) domains in the C-terminals of both channels, and we accordingly measured channel activity in the presence of Ca 2+.

For twenty years, ion channels have been studied in cancer progression. Several information have been collected about their involvement in cancer cellular processes like cell proliferation, motility and their participation in tumour progression using in-vivo models. Tumour microenvironment is currently the focus of many researches and the highlighting of the relationship between cancer cells and surrounding elements, is expanding. One of the major physic-chemical parameter involved in tumour progression is the hypoxia conditions observed in solid cancer. Due to their position on the cell membrane, ion channels are good candidates to transduce or to be modulated by environmental modifications. Until now, few reports have been interested in the modification of ion channel activities or expression in this context, compared to other pathological situations such as ischemia reperfusion. The aim of our review is to summarize the current knowledge about the calcium and potassium channels properties in the context of hypoxia in tumours. This review could pave the way to orientate new studies around this exciting field to obtain new potential therapeutic approaches., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Rapid conduction of nerve impulses is critical in life and relies on action potential (AP) leaps through the nodes of Ranvier (NRs) along myelinated nerves. While NRs are the only sites where APs can be regenerated during nerve conduction on myelinated nerves, ion channel mechanisms underlying the regeneration and conduction of APs at mammalian NRs remain incompletely understood. Here, we show that TREK-1 and TRAAK, the thermosensitive and mechanosensitive two-pore-domain potassium (K2P) channels, are clustered at NRs of rat trigeminal Aβ-afferent nerves with a density over 3,000-fold higher than that on their somas. These K2P channels, but not voltage-gated K + channels as in other parts of nerves, are required for rapid AP repolarization at the NRs. Furthermore, these channels permit high-speed and high-frequency AP conduction along the myelinated afferent nerves, and loss of function of these channels at NRs retards nerve conduction and impairs sensory behavioral responses in animals., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Chronic hyperglycemia and diabetes lead to impaired cardiac repolarization, K + channel remodeling and increased arrhythmia risk. However, the exact signaling mechanism by which diabetic hyperglycemia regulates cardiac K + channels remains elusive. Here, we show that acute hyperglycemia increases inward rectifier K + current (I K1 ), but reduces the amplitude and inactivation recovery time of the transient outward K + current (I to ) in mouse, rat, and rabbit myocytes. These changes were all critically dependent on intracellular O-GlcNAcylation. Additionally, I K1 amplitude and I to recovery effects (but not I to amplitude) were prevented by the Ca 2+ /calmodulin-dependent kinase II (CaMKII) inhibitor autocamtide-2-related inhibitory peptide, CaMKIIδ-knockout, and O-GlcNAc-resistant CaMKIIδ-S280A knock-in. I to reduction was prevented by inhibition of protein kinase C (PKC) and NADPH oxidase 2 (NOX2)-derived reactive oxygen species (ROS). In mouse models of chronic diabetes (streptozotocin, db/db, and high-fat diet), heart failure, and CaMKIIδ overexpression, both I to and I K1 were reduced in line with the downregulated K + channel expression. However, I K1 downregulation in diabetes was markedly attenuated in CaMKIIδ-S280A. We conclude that acute hyperglycemia enhances I K1 and I to recovery via CaMKIIδ-S280 O-GlcNAcylation, but reduces I to amplitude via a NOX2-ROS-PKC pathway. Moreover, chronic hyperglycemia during diabetes and CaMKII activation downregulate K + channel expression and function, which may further increase arrhythmia susceptibility.

Previous studies indicate that sex-related differences exist in the regulation of cutaneous vasodilation, however, the mechanisms remain unresolved. We assessed if sex-differences in young adults exist for cholinergic, nicotinic, and β-adrenergic cutaneous vasodilation with a focus on nitric oxide synthase (NOS), cyclooxygenase (COX), and K + channel mechanisms. In twelve young men and thirteen young women, four intradermal forearm skin sites were perfused with the following: 1) lactated Ringer's solution (control), 2) 10 mM N ω -nitro-l-arginine, a non-selective NOS inhibitor, 3) 10 mM ketorolac, a non-selective COX inhibitor, or 4) 50 mM BaCl 2 , a nonspecific K + channel blocker. At all four sites, cutaneous vasodilation was induced by 1) 10 mM nicotine, a nicotinic receptor agonist, 2) 100 μM isoproterenol, a nonselective β-adrenergic receptor agonist, and 3) 2 mM and 2000 mM acetylcholine, an acetylcholine receptor agonist. Nicotine and isoproterenol were administered for 3 min, whereas each acetylcholine dose was administered for 25 min. Regardless of treatment site, cutaneous vasodilation in response to nicotine and a high dose of acetylcholine (2000 mM) were lower in women than men. By contrast, isoproterenol induced cutaneous vasodilation was greater in women vs. men. Irrespective of sex, NOS inhibition or K + channel blockade attenuated isoproterenol-mediated cutaneous vasodilation, whereas K + channel blockade decreased nicotine-induced cutaneous vasodilation. Taken together, our findings indicate that while the mechanisms underlying cutaneous vasodilation are comparable between young men and women, sex-related differences in the magnitude of cutaneous vasodilation do exist and this response differs as a function of the receptor agonist., Competing Interests: Declaration of competing interest None., (Copyright © 2020. Published by Elsevier Inc.)

Cl - -sensitive with-no-lysine kinase (WNK) plays a key role in regulating the thiazide-sensitive Na + -Cl - cotransporter (NCC) in the distal convoluted tubule (DCT). Cl - enters DCT cells through NCC and leaves the cell across the basolateral membrane via the Cl - channel ClC-K2 or K + -Cl - cotransporter (KCC). While KCC is electroneutral, Cl - exit via ClC-K2 is electrogenic. Therefore, an alteration in DCT basolateral K + channel activity is expected to influence Cl - movement across the basolateral membrane. Although a role for intracellular Cl - in the regulation of WNK and NCC has been established, intracellular Cl - concentrations ([Cl - ] i ) have not been directly measured in the mammalian DCT. Therefore, to measure [Cl - ] i in DCT cells, we generated a transgenic mouse model expressing an optogenetic kidney-specific Cl-Sensor and measured Cl - fluorescent imaging in the isolated DCT. Basal measurements indicated that the mean [Cl - ] i was ~7 mM. Stimulation of Cl - exit with low-Cl - hypotonic solutions decreased [Cl - ] i , whereas inhibition of KCC by DIOA or inhibition of ClC-K2 by NPPB increased [Cl - ] i , suggesting roles for both KCC and ClC-K2 in the modulation of [Cl - ] i . Blockade of basolateral K + channels (Kir4.1/5.1) with barium significantly increased [Cl - ] i . Finally, a decrease in extracellular K + concentration transiently decreased [Cl - ] i , whereas raising extracellular K + transiently increased [Cl - ] i , further suggesting a role for Kir4.1/5.1 in the regulation of [Cl - ] i . We conclude that the alteration in ClC-K2, KCC, and Kir4.1/5.1 activity influences [Cl - ] i in the DCT.

TOKs are outwardly rectifying K + channels in fungi with two pore-loops and eight transmembrane spans. Here, we describe the TOKs from four pathogens that cause the majority of life-threatening fungal infections in humans. These TOKs pass large currents only in the outward direction like the canonical isolate from Saccharomyces cerevisiae (ScTOK), and distinct from other K + channels. ScTOK, AfTOK1 (Aspergillus fumigatus), and H99TOK (Cryptococcus neoformans grubii) are K + -selective and pass current above the K + reversal potential. CaTOK (Candida albicans) and CnTOK (Cryptococcus neoformans neoformans) pass both K + and Na + and conduct above a reversal potential reflecting the mixed permeability of their selectivity filter. Mutations in CaTOK and ScTOK at sites homologous to those that open the internal gates in classical K + channels are shown to produce inward TOK currents. A favored model for outward rectification is proposed whereby the reversal potential determines ion occupancy, and thus, conductivity, of the selectivity filter gate that is coupled to an imperfectly restrictive internal gate, permitting the filter to sample ion concentrations on both sides of the membrane., (© 2020 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

The TMEM175 family constitutes recently discovered K + channels that are important for autophagosome turnover and lysosomal pH regulation and are associated with the early onset of Parkinson Disease. TMEM175 channels lack a P-loop selectivity filter, a hallmark of all known K + channels, raising the question how selectivity is achieved. Here, we report the X-ray structure of a closed bacterial TMEM175 channel in complex with a nanobody fusion-protein disclosing bound K + ions. Our analysis revealed that a highly conserved layer of threonine residues in the pore conveys a basal K + selectivity. An additional layer comprising two serines in human TMEM175 increases selectivity further and renders this channel sensitive to 4-aminopyridine and Zn 2+ . Our findings suggest that large hydrophobic side chains occlude the pore, forming a physical gate, and that channel opening by iris-like motions simultaneously relocates the gate and exposes the otherwise concealed selectivity filter to the pore lumen., Competing Interests: JB, RJ, TS, YN, AM, GT, TM, SS No competing interests declared, (© 2020, Brunner et al.)

KcsA, a prokaryote tetrameric potassium channel, was the first ion channel ever to be structurally solved at high resolution. This, along with the ease of its expression and purification, made KcsA an experimental system of choice to study structure-function relationships in ion channels. In fact, much of our current understanding on how the different channel families operate arises from earlier KcsA information. Being an integral membrane protein, KcsA is also an excellent model to study how lipid-protein and protein-protein interactions within membranes, modulate its activity and structure. In regard to the later, a variety of equilibrium and non-equilibrium methods have been used in a truly multidisciplinary effort to study the effects of lipids on the KcsA channel. Remarkably, both experimental and "in silico" data point to the relevance of specific lipid binding to two key arginine residues. These residues are at non-annular lipid binding sites on the protein and act as a common element to trigger many of the lipid effects on this channel. Thus, processes as different as the inactivation of channel currents or the assembly of clusters from individual KcsA channels, depend upon such lipid binding.

Substantial experimental evidences support the hypothesis that dietary flavonoid intake has a favourable impact on cardiovascular diseases such as systemic, arterial hypertension and coronary artery diseases, which represent the leading cause of morbidity and mortality worldwide. The biological effects of flavonoids involve complex biochemical interactions with numerous, specific, cellular and molecular targets. K + channels, fine modulators of both cardiac action potential and vascular cell membrane potential, represent one of these targets. Overexpression, downregulation or dysfunction of these channel proteins are the cause of many cardiovascular diseases. Therefore, it appears of particular interest a detailed analysis of the flavonoid potential, direct/indirect modulation of cardiovascular K + channels as these natural compounds ingested with the diet, despite extensive gut metabolism, may accumulate at cellular level in the form of the parent aglycones. The present review will portray their effects on cardiovascular K + channels. Molecular docking was used to strengthen experimental evidences and describe flavonoid-channel interactions at molecular level., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Ion permeation, selectivity, and the behavior of the K + channel selectivity filter have been studied intensively in the previous two decades. The agreement among multiple approaches used to study ion flux in K + channels suggests a consensus mechanism of ion permeation across the selectivity that has been put to the test in recent years with the proposal of an alternative way by which ions can cross the selectivity filter of K + channels via direct Coulomb repulsion between contacting cations. Past experimental work by Zhou and MacKinnon ( J. Mol. Biol. 2004 , 839) showed that mutation of the site S4 reduces the total occupancy of the selectivity filter to less than two ions on average by lowering the occupancy of the S2-S4 configuration without changing the S1-S3 configuration much, and this reduction of occupancy means that ion configurations different from the ones involved in the canonical mechanism are likely to be involved. At that time, calculations using complicated kinetic networks to relate occupancy to conduction did not provide deeper insight into the conduction mechanism. Here, to help solve this enigma, umbrella sampling simulations have been performed to evaluate the potential of mean force of two KcsA mutant channels where the S4 site is substituted. Our new results provide insights into the significance of threonine in this position, revealing the effect of substitution on the alternate mechanisms of conduction proposed, involving either water or vacant sites.338 , 839) showed that mutation of the site S4 reduces the total occupancy of the selectivity filter to less than two ions on average by lowering the occupancy of the S2-S4 configuration without changing the S1-S3 configuration much, and this reduction of occupancy means that ion configurations different from the ones involved in the canonical mechanism are likely to be involved. At that time, calculations using complicated kinetic networks to relate occupancy to conduction did not provide deeper insight into the conduction mechanism. Here, to help solve this enigma, umbrella sampling simulations have been performed to evaluate the potential of mean force of two KcsA mutant channels where the S4 site is substituted. Our new results provide insights into the significance of threonine in this position, revealing the effect of substitution on the alternate mechanisms of conduction proposed, involving either water or vacant sites.

This study sought to identify and localize SLO1 channels in boar spermatozoa by immunoblotting and immunofluorescence, and to determine their physiological role during in vitro sperm capacitation. Sperm samples from 14 boars were incubated in a capacitation medium for 300 min in the presence of paxilline (PAX), a specific SLO1-channel blocker, added either at 0 min or after 240 min of incubation. Negative controls were incubated in capacitation medium, and positive controls in capacitation medium plus tetraethyl ammonium (TEA), a general K + -channel blocker, also added at 0 min or after 240 min of incubation. In all samples, acrosome exocytosis was triggered with progesterone after 240 min of incubation. Sperm motility and kinematics, integrity of plasma and acrosome membranes, membrane lipid disorder, intracellular calcium levels and acrosin activity were evaluated after 0, 60, 120, 180, 240, 250, 270 and 300 min of incubation. In boar spermatozoa, SLO1 channels were found to have 80 kDa and be localized in the anterior postacrosomal region and the mid and principal piece of the tail; their specific blockage through PAX resulted in altered calcium levels and acrosome exocytosis. As expected, TEA blocker impaired in vitro sperm capacitation, by altering sperm motility and kinematics and calcium levels. In conclusion, SLO1 channels are crucial for the acrosome exocytosis induced by progesterone in in vitro capacitated boar spermatozoa., Competing Interests: The authors declare no conflict of interest.

Sex hormones, such as testosterone, progesterone, and 17β-estradiol, control various physiological functions. This review focuses on the sex hormonal regulation of K + channels and the effects of such regulation on electrophysiological and contractile functions of muscles. In the cardiac tissue, testosterone and progesterone shorten action potential, and estrogen lengthens QT interval, a marker of increased risk of ventricular tachyarrhythmias. We have shown that testosterone and progesterone in physiological concentration activate KCNQ1 channels via membrane-delimited sex hormone receptor/eNOS pathways to shorten the action potential duration. Mitochondrial K + channels are also involved in the protection of cardiac muscle. Testosterone and 17β-estradiol directly activate mitochondrial inner membrane K + channels (Ca 2+ activated K + channel (K Ca channel) and ATP-sensitive K + channel (K ATP channel)) that are involved in ischemic preconditioning and cardiac protection. During pregnancy, uterine blood flow increases to support fetal growth and development. It has been reported that 17β-estradiol directly activates large-conductance Ca 2+ -activated K + channel (BK Ca channel) attenuating arterial contraction. Furthermore, 17β-estradiol increases expression of BK Ca channel β1 subunit which enhances BK Ca channel activity by DNA demethylation. These findings are useful for understanding the mechanisms of sex or generation-dependent differences in the physiological and pathological functions of muscles, and the mechanisms of drug actions., (Copyright © 2019 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Cholinergic-activated sweating depends on an influx of Ca 2+ from extracellular fluid. It is thought that the opening of K + channels on secretory epithelial cells facilitates Ca 2+ entry. We examined the hypothesis that tetraethylammonium (TEA)-sensitive K + channels participate in sweat production. We used a pre-post experimental design and initiated cholinergic-mediated sweating with intradermal electrical stimulation, monitored local sweat rate (SR) with a small sweat capsule mounted on the skin, and delivered 50 mM TEA via intradermal microdialysis. Local SR was activated by intradermal stimulation frequencies of 0.2-64 Hz, and we generated a sigmoid-shaped stimulus-response curve by plotting the area under the SR-time curve versus log 10 stimulus frequency. Peak local SR was reduced from 0.372 ± 0.331 to 0.226 ± 0.190 mg·min -1 ·cm -2 ( P = 0.0001) during application of 50 mM TEA, whereas the EC 50 and Hill slopes were not altered. The global sigmoid-shaped stimulus-response curves for control and 50 mM TEA were significantly different ( P < 0.0001), and the plateau region was significantly reduced ( P = 0.0023) with the TEA treatment. The effect of TEA on peak local SR was similar in male and female subjects. However, we did note a small effect of sex on the shape of the stimulus-response curves during intradermal electrical stimulation. Overall, these data support the hypothesis that cholinergic control of sweat gland activity is modulated by the presence of TEA-sensitive K + channels in human sweat gland epithelial cells. NEW & NOTEWORTHY The contribution of various potassium channels to the process of cholinergic-mediated human eccrine sweat production is unclear. Using a novel model for cholinergic-mediated sweating in humans, we provide evidence that tetraethylammonium-sensitive K + channels (K Ca 1.1 and K v channels) contribute to eccrine sweat production.

We examined mechanisms of ammonia handling in the anterior, mid, and posterior intestine of unfed and fed freshwater rainbow trout (Oncorhynchus mykiss), with a focus on the Na + :K + :2Cl - co-transporter (NKCC), Na + :K + -ATPase (NKA), and K + channels. NKCC was localized by immunohistochemistry to the mucosal (apical) surface of enterocytes, and NKCC mRNA was upregulated after feeding in the anterior and posterior segments. NH 4 was equally potent to K + was equally potent to K + in supporting NKA activity in all intestinal sections. In vitro gut sac preparations were employed to examine mucosal ammonia flux rates (Jm amm , disappearance from the mucosal saline), serosal ammonia flux rates (Js amm , appearance in the serosal saline), and total tissue ammonia production rates (Jt amm  = Js amm - Jm amm ). Bumetanide (10 -4 mol L -1 ), a blocker of NKCC, inhibited Js amm in most preparations, but this was largely due to reduction of Jt amm ; Jm amm was significantly inhibited only in the anterior intestine of fed animals. Ouabain (10 -4 mol L -1 ), a blocker of NKA, generally reduced both Jm amm and Js amm without effects on Jt amm in most preparations, though the anterior intestine was resistant after feeding. Barium (10 -2 ), a blocker of K -1 channels, inhibited Jm + channels, inhibited Jm amm in most preparations, and Js amm in some, without effects on Jt amm and K + and K + appears to be taken up through barium-sensitive K 4 channels on the mucosal surface. Mucosal NH + uptake via both NKCC and K + channels is energized by basolateral NKA, which plays an additional role in scavenging NH 4 + uptake via both NKCC and K + channels is energized by basolateral NKA, which plays an additional role in scavenging NH 4 + on the serosal surface to possibly minimize blood toxicity or enhance ion uptake and amino acid synthesis following feeding. Together with recent findings from other studies, we have provided an updated model to describe the current understanding of intestinal ammonia transport in teleost fish.

Mural cells in precapillary arterioles (PCAs) generate spontaneous Ca 2+ transients primarily arising from the periodic release of Ca 2+ from sarcoendoplasmic reticulum (SR/ER). The Ca 2+ release induces Ca 2+ -activated chloride channel (CaCC)-dependent depolarisations that spread to neighbouring mural cells to develop the synchrony of their Ca 2+ transients. Here, we explored the roles of K + channels in maintaining the synchrony of spontaneous Ca 2+ transients. Intracellular Ca 2+ dynamics in mural cells were visualised by Cal-520 fluorescence Ca 2+ imaging in the submucosal PCAs of rat rectum. Increasing extracellular K + concentration ([K + ] o ) from 5.9 to 29.7 mM converted synchronous spontaneous Ca 2+ transients into asynchronous, high-frequency Ca 2+ transients. Similarly, the blockade of inward rectifier K + (K ir ) channels with Ba 2+ (50 μM) or K v 7 voltage-dependent K + (K v 7) channels with XE 991 (10 μM) disrupted the synchrony of spontaneous Ca 2+ transients, while the blockers for large-, intermediate- or small-conductance Ca 2+ -activated K + channels had no effect. K ir 2.1 immunoreactivity was detected in the arteriolar endothelium but not mural cells. In the PCAs that had been pretreated with XE 991 or Ba 2+ , nifedipine (1 μM) attenuated the asynchronous Ca 2+ transients but failed to restore their synchrony. In contrast, levcromakalim, an ATP-sensitive K + channel opener, restored the synchronous Ca 2+ transients. Thus, constitutively active K v 7 and K ir channels appear to be involved in maintaining the relatively hyperpolarised membrane of mural cells. The hyperpolarised membrane prevents depolarisation-induced 'premature' Ca 2+ transients to ensure sufficient SR/ER Ca 2+ refilling that is required for regenerative Ca 2+ release resulting in synchronous Ca 2+ transients amongst the mural cells.

Spontaneous activity shifts at constant experimental conditions represent a widespread regulatory mechanism in ion channels. The molecular origins of these modal gating shifts are poorly understood. In the K + channel KcsA, a multitude of fast activity shifts that emulate the native modal gating behaviour can be triggered by point-mutations in the hydrogen bonding network that controls the selectivity filter. Using solid-state NMR and molecular dynamics simulations in a variety of KcsA mutants, here we show that modal gating shifts in K + channels are associated with important changes in the channel dynamics that strongly perturb the selectivity filter equilibrium conformation. Furthermore, our study reveals a drastically different motional and conformational selectivity filter landscape in a mutant that mimics voltage-gated K + channels, which provides a foundation for an improved understanding of eukaryotic K + channels. Altogether, our results provide a high-resolution perspective on some of the complex functional behaviour of K + channels.

Guard cells integrate various hormone signals and environmental cues to balance plant gas exchange and transpiration. The wounding-associated hormone jasmonic acid (JA) and the drought hormone abscisic acid (ABA) both trigger stomatal closure. In contrast to ABA however, the molecular mechanisms of JA-induced stomatal closure have remained largely elusive. Here, we identify a fast signaling pathway for JA targeting the K + efflux channel GORK. Wounding triggers both local and systemic stomatal closure by activation of the JA signaling cascade followed by GORK phosphorylation and activation through CBL1-CIPK5 Ca 2+ sensor-kinase complexes. GORK activation strictly depends on plasma membrane targeting and Ca 2+ binding of CBL1-CIPK5 complexes. Accordingly, in gork, cbl1, and cipk5 mutants, JA-induced stomatal closure is specifically abolished. The ABA-coreceptor ABI2 counteracts CBL1-CIPK5-dependent GORK activation. Hence, JA-induced Ca 2+ signaling in response to biotic stress converges with the ABA-mediated drought stress pathway to facilitate GORK-mediated stomatal closure upon wounding., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Potassium channels have the unique ability to allow the selective passage of potassium ions at near diffusion-free rates while inhibiting the passage of more abundant sodium ions. Local interactions between chemical functional groups and the ions are responsible for both selectivity and transport. As an initial step in characterizing these interactions, the structures of Na+ and K+ complexed to the Ac-Tyr-NHMe peptide have been determined from infrared laser spectroscopy and supporting ab initio calculations. Ac-Tyr-NHMe, a termini-protected peptide sequence, replicates the GYG portion of one of the four peptide chains comprising the selectivity filter of a K+ channel. This peptide contains two carbonyl groups, among the eight C[double bond, length as m-dash]O groups forming the S1 binding site of the selectivity filter. Three conformations have been identified for both ions by laser IR-IR double resonance methods. Two conformations have the ion bound to the two C[double bond, length as m-dash]O groups. The third conformation has, in addition, a cation-π interaction with the aromatic ring of tyrosine, i.e. tridentate binding. The relative contributions of the three conformers are approximately the same for K+Ac-Tyr-NHMe, while the tridentate conformer is preferred for Na+Ac-Tyr-NHMe. These differences will be discussed in the context of ion mobility and selectivity.

The goal of the present study was to evaluate vascular potassium channels and Na + -K + -ATPase activity in estrogen deficient female rats. Female rats that underwent ovariectomy were assigned to receive daily treatment with placebo (OVX) or estrogen replacement (OVX+E2, 1mg/kg, once a week, i.m.). Aortic rings were used to examine the involvement of K + channels and Na + -K + -ATPase in vascular reactivity. Acetylcholine (ACh)-induced relaxation was analyzed in the presence of L-NAME (100μM) and K + channels blockers: tetraethylammonium (TEA, 5mM), 4-aminopyridine (4-AP, 5mM), iberiotoxin (IbTX, 30nM), apamin (0.5mM), charybdotoxin (ChTX, 0.1mM) and iberiotoxin plus apamin. When aortic rings were pre-contracted with KCl (60mM) or pre-incubated with TEA (5mM), 4-aminopyridine (4-AP, 5mM) and iberiotoxin (IbTX, 30nM) plus apamin (0.5μM), the ACh-induced relaxation was less effective in the ovariectomized group. Additionally, 4-AP and IbTX decreased the relaxation by sodium nitroprusside in all groups but this reduction was greater in the ovariectomized group. Estrogen deficiency also increased aortic functional Na + -K + ATPase activity evaluated by K + -induced relaxation. L-NAME or endothelium removal were not able to block the increase in aortic functional Na + -K + ATPase activity, however, TEA (5mM) restored this increase to the control level. We also found that estrogen deficiency increased superoxide anion production and reduced nitric oxide release in aortic ring from ovariectomized animals. In summary, our results emphasize that the process underlying ACh-induced relaxation is preserved in ovariectomized animals due to the activation of K + channels and increased Na + -K + ATPase activity., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The aim of the present study was to investigate the acute effect and mechanism of tumor necrosis factor (TNF) on basolateral 50 pS K channels in the thick ascending limb (TAL) of the rat kidney. The TAL tubules were isolated from the rat kidney, and the activity of the 50 pS K channels was recorded using the patch‑clamp technique. The results indicated that the application of TNF (10 nM) significantly activated the 50 pS K channels and the TNF effect was concentration‑dependent. Inhibition of protein kinase A, phospholipase A2 and protein tyrosine kinase using pathway inhibitors (H89, AACOCF3 and Herbimycin A, respectively) did not abolish the stimulatory effect of TNF, indicating that none of these pathways mediated the TNF effect. By contrast, the phenylarsine oxide inhibitor against protein tyrosine phosphatase (PTP) decreased the activity of the 50 pS K channels and blocked the stimulatory effect of TNF on these channels. Furthermore, western blot analysis demonstrated that the application of TNF (10 nM) in the TAL increased the phosphorylation of PTP, an indication of PTP activity stimulation. Thus, it was concluded that the acute application of TNF may stimulate the basolateral 50 pS K channel in the TAL and the stimulatory effect of TNF may be mediated by the PTP‑dependent pathway.

Potassium ion channels utilize a highly selective filter to rapidly transport K + ions across cellular membranes. This selectivity filter is composed of four binding sites which display almost equal electron density in crystal structures with high potassium ion concentrations. This electron density can be interpreted to reflect a superposition of alternating potassium ion and water occupied states or as adjacent potassium ions. Here, we use single wavelength anomalous dispersion (SAD) X-ray diffraction data collected near the potassium absorption edge to show experimentally that all ion binding sites within the selectivity filter are fully occupied by K + ions. These data support the hypothesis that potassium ion transport occurs by direct Coulomb knock-on, and provide an example of solving the phase problem by K-SAD.

Self-assembled alkyl-ureido-benzo-15-crown-5-ethers are selective ionophores for K + cations, which are preferred to Na + cations. The transport mechanism is determined by the optimal coordination rather than classical dimensional compatibility between the crown ether hole and the cation diameter. Herein, we demonstrate that systematic changes of the structure lead to unexpected modifications in the cation-transport activity and suffice to produce adaptive selection. We show that the main contribution to performance arises from optimal constraints on the conformational freedom, which are determined by the binding macrocycles, the nature of the hydrogen-bonding groups, and the hydrophobic tails. Simple changes to the flexible 15-crown-5-ether lead to selective carriers for Na + . Hydrophobic stabilization of the channels through mutual interactions between lipids and variable hydrophobic tails appears to be an important cause of increased activity. Oppositely, restricted translocation is achieved when constrained hydrogen-bonded macrocyclic relays are less dynamic in a pore superstructure., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The human ether-a-go-go-related gene (hERG) encodes a voltage-gated potassium channel that controls repolarization of cardiac action potentials. Accumulating evidence suggests that most disease-related hERG mutations reduce the function of the channel by disrupting protein biogenesis of the channel in the endoplasmic reticulum (ER). However, the molecular mechanism underlying the biogenesis of ERG K + channels is largely unknown. By forward genetic screening, we identified an ER-located chaperone CNX-1, the worm homologue of mammalian chaperone Calnexin, as a critical regulator for the protein biogenesis of UNC-103, the ERG-type K + channel in Caenorhabditis elegans Loss-of-function mutations of cnx-1 decreased the protein level and current density of the UNC-103 K + channel and suppressed the behavioral defects caused by a gain-of-function mutation in unc-103 Moreover, CNX-1 facilitated tetrameric assembly of UNC-103 channel subunits in a liposome-assisted cell-free translation system. Further studies showed that CNX-1 act in parallel to DNJ-1, another ER-located chaperone known to regulate maturation of UNC-103 channels, on controlling the protein biogenesis of UNC-103. Importantly, Calnexin interacted with hERG proteins in the ER in HEK293T cells. Deletion of calnexin reduced the expression and current densities of endogenous hERG K + channels in SH-SY5Y cells. Collectively, we reveal an evolutionarily conserved chaperone CNX-1/Calnexin controlling the biogenesis of ERG-type K + channels., (© 2018 Bai et al.)

The seeming contradiction that K + channels conduct K + ions at maximal throughput rates while not permeating slightly smaller Na + ions has perplexed scientists for decades. Although numerous models have addressed selective permeation in K + channels, the combination of conduction efficiency and ion selectivity has not yet been linked through a unified functional model. Here, we investigate the mechanism of ion selectivity through atomistic simulations totalling more than 400 μs in length, which include over 7,000 permeation events. Together with free-energy calculations, our simulations show that both rapid permeation of K + and ion selectivity are ultimately based on a single principle: the direct knock-on of completely desolvated ions in the channels' selectivity filter. Herein, the strong interactions between multiple 'naked' ions in the four filter binding sites give rise to a natural exclusion of any competing ions. Our results are in excellent agreement with experimental selectivity data, measured ion interaction energies and recent two-dimensional infrared spectra of filter ion configurations.

To understand the molecular mechanisms underlying the beneficial effects of sildenafil in animal models of neurological disorders, we investigated the effects of sildenafil on the mitochondrial toxicity induced by β-amyloid (Aβ) peptide. Treatment of HT-22 hippocampal neuronal cells with Aβ 25∼35 results in increased mitochondrial Ca 2+ load, which is subsequently suppressed by sildenafil as well as by diazoxide, a selective opener of the ATP-sensitive K + channels (K ATP ). However, the suppressive effects of sildenafil and diazoxide are significantly attenuated by 5-hydroxydecanoic acid (5-HD), a K ATP inhibitor. The increased mitochondrial Ca 2+ overload is accompanied by decrease in the intracellular ATP concentration, increase in intracellular ROS generation, occurrence of mitochondrial permeability transition, and activation of caspase-9 and cell death. Exposure to sildenafil inhibited the mitochondria-associated changes and cell death induced by Aβ. However, the inhibitory effects of sildenafil are abolished or weakened in the presence of 5-HD, suggesting that opening of the mitochondrial K ATP is required for sildenafil to exert these effects. Taken together, these results indicate that at the mitochondrial levels, sildenafil plays a protective role towards neuronal cell in an environment rich in Aβ, and exerts its effects via the mitochondrial K ATP channels-dependent mechanisms., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The selectivity filter and the activation gate in potassium channels are functionally and structurally coupled. An allosteric coupling underlies C-type inactivation coupled to activation gating in this ion-channel family (i.e., opening of the activation gate triggers the collapse of the channel's selectivity filter). We have identified the second Threonine residue within the TTVGYGD signature sequence of K + channels as a crucial residue for this allosteric communication. A Threonine to Alanine substitution at this position was studied in three representative members of the K + -channel family. Interestingly, all of the mutant channels exhibited lack of C-type inactivation gating and an inversion of their allosteric coupling (i.e., closing of the activation gate collapses the channel's selectivity filter). A state-dependent crystallographic study of KcsA-T75A proves that, on activation, the selectivity filter transitions from a nonconductive and deep C-type inactivated conformation to a conductive one. Finally, we provide a crystallographic demonstration that closed-state inactivation can be achieved by the structural collapse of the channel's selectivity filter., Competing Interests: The authors declare no conflict of interest.

Electrical dynamics of freshly isolated cerebral endothelium have not been determined independently of perivascular nerves and smooth muscle. We tested the hypothesis that endothelium of cerebral and skeletal muscle arteries differentially utilizes purinergic and muscarinic signaling pathways to activate endothelium-derived hyperpolarization. Changes in membrane potential ( V m ) were recorded in intact endothelial tubes freshly isolated from posterior cerebral and superior epigastric arteries of male and female C57BL/6 mice (age: 3-8 months). V m -activated K 3 ) receptors in addition to small- and intermediate-conductance Ca 2+ -activated K + ) and inward rectifying K C (K a ) channels using ATP (100 μmol·L C a ) and inward rectifying K + (K IR ) channels using ATP (100 μmol·L -1 ), acetylcholine (ACh; 10 μmol·L -1 ), NS309 (0.01-10 μmol·L -1 ), and 15 mmol·L -1 activity is preserved across brain and skeletal muscle. As vascular reactivity decreases with aging and cardiovascular disease, endothelial K V m ~-30 mV to maximum ~-80 mV). Hyperpolarization (~8 mV) occurred during 15 mmol·L -1 KCl treatment in cerebral but not skeletal muscle endothelial tubes. Despite weaker hyperpolarization during endothelial GPCR stimulation in cerebral vs skeletal muscle endothelium, the capability for robust SK C a /IK C a activity is preserved across brain and skeletal muscle. As vascular reactivity decreases with aging and cardiovascular disease, endothelial K + channel activity may be calibrated to restore blood flow to respective organs regardless of gender.

To explore the roles of various K + channels in regulating the spontaneous activity of bladder muscularis mucosae (MM) that is considered to play an important role in maintaining mucosal function. Effects of K + channel modulators on electrical and contractile activity in the guinea-pig bladder MM were examined using intracellular microelectrode and isometric tension recording. The MM predominately generated bursting spontaneous action potentials (SAPs) and phasic contractions (SPCs) that were blocked by nifedipine (1µM). NS309 (10µM), a small-conductance Ca 2+ -activated K + (SK) channel opener, dramatically prolonged after-hyperpolarisation (AHP) and converted bursting SAPs into individually action potentials in an apamin (100nM)-sensitive manner. Apamin alone increased the number of SAPs during bursts. NS1619 (10µM), a large-conductance Ca 2+ -activated K + (BK) channel opener, abolished SAPs in a manner reversed by iberiotoxin (IbTX, 100nM), a BK channel blocker. IbTX alone enlarged SAPs and abolished their AHPs. Flupirtine (10µM), a voltage-dependent K + channel (K v 7) opener, diminished SAPs in a manner reversed by XE991 (10µM), a K v 7 channel blocker. XE991 alone exerted modest excitatory effects on SAPs. These K + channel modulators had corresponding effects on SPCs. Bursting SAP firing appears to result from a lower level activation of SK channels in MM than that DSM. BK channels play a predominant role in regulating SAP configuration, while K v 7 channels have only a marginal role. The prevention of bursting SAPs and associated reduction in SPCs upon the pharmacological activation of a reserved population of SK channels may well have a considerable therapeutic potential., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Mechanical force sensation is fundamental to a wide breadth of biology from the classic senses of touch, pain, hearing, and balance to less conspicuous sensations of proprioception, blood pressure, and osmolarity and basic aspects of cell growth, differentiation, and development. These diverse and essential systems use force-gated (or mechanosensitive) ion channels that convert mechanical stimuli into cellular electrical signals. TRAAK, TREK1, and TREK2 are K + -selective ion channels of the two-pore domain K + (K2P) family that are mechanosensitive: they are gated open by increasing membrane tension. TRAAK and TREK channels are thought to play roles in somatosensory and other mechanosensory processes in neuronal and non-neuronal tissues. Here, we present protocols for three assays to study mechanical activation of these channels in cell membranes: (1) cell swelling, (2) cell poking, and (3) patched membrane stretching. Patched membrane stretching is also applicable to the study of mechanosensitive K2P channel activity in a cell-free system and a procedure for proteoliposome reconstitution and patching is also presented. These approaches are also readily applicable to the study of other mechanosensitive ion channels.