Interaction of hydrogen sulfide with ion channels

1. Hydrogen sulfide (H₂S) is a signalling gasotransmitter. It targets different ion channels and receptors, and fulfils its various roles in modulating the functions of different systems. However, the interaction of H₂S with different types of ion channels and underlying molecular mechanisms has not been reviewed systematically. 2. H₂S is the first identified endogenous gaseous opener of ATP‐sensitive K⁺ channels in vascular smooth muscle cells. Through the activation of ATP‐sensitive K⁺ channels, H₂S lowers blood pressure, protects the heart from ischemia and reperfusion injury, inhibits insulin secretion in pancreatic β cells, and exerts anti‐inflammatory, anti‐nociceptive and anti‐apoptotic effects. 3. H₂S inhibited L‐type Ca²⁺ channels in cardiomyocytes but stimulated the same channels in neurons, thus regulating intracellular Ca²⁺ levels. H₂S activated small and medium conductance K_Ca channels but its effect on BK_Ca channels has not been consistent. 4. H₂S‐induced hyperalgesia and pro‐nociception seems to be related to the sensitization of both T‐type Ca²⁺ channels and TRPV₁ channels. The activation of TRPV₁ and TRPA₁ by H₂S is believed to result in contraction of nonvascular smooth muscles and increased colonic mucosal Cl^? secretion. 5. The activation of Cl^? channel by H₂S has been shown as a protective mechanism for neurons from oxytosis. H₂S also potentiates N‐methyl‐d ‐aspartic acid receptor‐mediated currents that are involved in regulating synaptic plasticity for learning and memory. 6. Given the important modulatory effects of H₂S on different ion channels, many cellular functions and disease conditions related to homeostatic control of ion fluxes across cell membrane should be re‐evaluated.     

The real life of voltage-gated K channels: more than model behaviour

The past ten years have provided an embarrassment of riches for those interested in cloned voltage-gated K⁺ (Kv) channels. Details of their physiology and pharmacology in expression systems, and their precise cellular location abound, making them excellent targets for pharmacologists. However, there is still a considerable and important gap in our knowledge between the behaviour of expressed Kv channels and K⁺ currents in vivo. In this review Brian Robertson focuses on a few of the recent developments in the field of Kv channels, namely modulation of their behaviour by accessory subunits, their control, and localization of identified Kv subunits.     

Arachidonic acid and ion channels: an update

Arachidonic acid (AA), a polyunsaturated fatty acid with four double bonds, has multiple actions on living cells. Many of these effects are mediated by an action of AA or its metabolites on ion channels. During the last 10 years, new types of ion channels, transient receptor potential (TRP) channels, store-operated calcium entry (SOCE) channels and non-SOCE channels have been studied. This review summarizes our current knowledge about the effects of AA on TRP and non-SOCE channels as well as classical ion channels. It aims to distinguish between effects of AA itself and effects of AA metabolites. Lipid mediators are of clinical interest because some of them (for example, leukotrienes) play a role in various diseases, others (such as prostaglandins) are targets for pharmacological therapeutic intervention.     

罗哌卡因对豚鼠心室肌细胞钠电流、钙电流和钾电流的影响

目的:研究罗哌卡因(Rop)对豚鼠心室肌细胞钠电流(Ⅰ_(Na))、L-型钙电流(Ⅰ_(Ca-L)、内向整流钾电流(Ⅰ_(Kl)及延迟整流钾电流(I_K)的影响.方法:全细胞膜片箝技术.结果:罗哌卡因10,50与100μmol/L使Ⅰ_(Na)的峰电流分别减小8.3％、33.3 ％和62.5％(P<0.01),使失活时间常数分别延长8.2％、24.7％和64.1％(P<0.05);罗哌卡因50与100μmol/L使Ⅰ_(Ca-L)的峰电流分别减小7.6％和22.5％(P<0.05),使慢失活时间常数分别延长15.5％和33.0％(P<0.01);罗哌卡因50与100μmol/L对Ⅰ_(Kl)和Ⅰ_K的峰电流无明显影响.结论:罗哌卡因抑制Ⅰ_(Na)和Ⅰ_(Ca-L),可能与其心脏毒性作用有关.     

An assessment of the present and future roles of non-ligand gated ion channel modulators as CNS therapeutics

Few approved drugs have, as their primary known mechanism of action, modulation of non-ligand gated ion channels. However, these proteins are important regulators of neuronal function through their control of sodium, potassium, calcium and chloride flux, and are ideal candidates as drug discovery targets. Recent progress in the molecular biology and pharmacology of ion channels suggests that many will be associated with specific pharmacological profiles that will include both activators and inhibitors. Ion channels, through their regulation by G-proteins, are a major component of the final common pathway of many drugs acting at classical neuronal receptors. Thus, targeting of the ion channels themselves may confer different profiles of efficacy and specificity to drug action in the brain and spinal cord. Three areas for drug discovery are profiled that the authors consider prime targets for ion channel based therapies, anticonvulsant drugs, cognition enhancing drugs and drugs for improving neurone survival following ischaemia.     

Pharmacological activation and inhibition of Slack (Slo2.2) channels

The Slack (Sequence like a calcium-activated K channel) (Slo2.2) gene is abundantly expressed in the mammalian brain and encodes a sodium-activated K+ (KNa) channel. Although the specific roles of Slack channel subunits in neurons remain to be identified, they may play a role in the adaptation of firing rate and in protection against ischemic injury. In the present study, we have generated a stable cell line expressing the Slack channel, and have analyzed the pharmacological properties of these channels in these cells and in Xenopus oocytes. Two known blockers of KNa channels, bepridil and quinidine, inhibited Slack currents in a concentration-dependent manner and decreased channel activity in excised membrane patches. The inhibition by bepridil was potent, with an IC50 of 1.0 microM for inhibition of Slack currents in HEK cells. In contrast, bithionol was found to be a robust activator of Slack currents. When applied to the extracellular face of excised patches, bithionol rapidly induced a reversible increase in channel opening, suggesting that it acts on Slack channels relatively directly. These data establish an important early characterization of agents that modulate Slack channels, a process essential for the experimental manipulation of Slack currents in neurons.     

高血压发展过程中脑血管平滑肌细胞离子通道的变化

脑血管重构是高血压发展过程中最重要的病理生理改变,由此引起的脑卒中更日益危害人类的健康。在高血压发展过程中,脑血管平滑肌细胞上分布的多种离子通道,如钾钙、氯等均发生变化,导致细胞内离子浓度异常,在脑血管重构的发生发展过程中发挥了重要作用。     

Pharmacological modulation of voltage-gated potassium channels as a therapeutic strategy

Importance of the field: The human genome encodes at least 40 distinct voltage-gated potassium channel subtypes, which vary in regional expression, pharmacological and biophysical properties. Voltage-dependent potassium (Kv) channels help orchestrate many of the physiological and pathophysiological processes that promote and sometimes hinder the healthy functioning of our bodies.

Areas covered in this review: This review summarizes patent and scientific literature reports from the past decade highlighting the opportunities that Kv channels offer for the development of new therapeutic interventions for a wide variety of disorders.

What the reader will gain: The reader will gain an insight from an analysis of the associations of different Kv family members with disease processes, summary and evaluation of the development of therapeutically relevant pharmacological modulators of these channels, particularly focusing on proprietary agents being developed.

Take home message: Development of new drugs that target Kv channels continue to be of great interest but is proving to be challenging. Nevertheless, opportunities for Kv channel modulators to have an impact on a wide range of disorders in the future remain an exciting prospect.     

Functional and pharmacological properties of human and rat NaV1.8 channels

The aim of this work is to characterise the functional properties of human and rat Na_V1.8 channels and to investigate the action of anti-nociceptive agents. Na_V1.8 α-subunits were expressed in mammalian sensory neuron-derived ND7/23 cells, and sodium currents were recorded using whole-cell patch clamp. The current-voltage curves for activation were similar for human and rat Na_V1.8 channels. However, for inactivation, human Na_V1.8 showed more hyperpolarised voltage-dependence than for the rat channel, faster development of inactivation, slower recovery from the fast component of inactivation, and faster recovery from the slow component. Thus, this would imply that the human channel is more inactivated at normal resting potentials. Compounds 227c89, A-803467, V102862, ralfinamide and tetracaine all showed greater affinity for the inactivated state than for the resting state. Compounds A-803467 and V102862 were the most potent, and A-803467 showed greater inactivated state affinity for human than for rat channels. Surprisingly, during recovery from inactivation, an increase in current was observed for V102862 and A-803467, probably due to disinhibition of resting block. Rather than the use-dependent inhibition normally seen with inactivated state blockers, for A-803467 this disinhibition led to an increase in current during repetitive stimulation, while V102862 showed less inhibition than otherwise expected at lower frequencies. Thus the data supports the suggestion that, while both V102862 and A-803467 are potent inhibitors of Na_V1.8, the compound V102862, rather than A-803467, may be useful as an analgesic where physiological firing frequencies are higher.     

cAMP-dependent kinase does not modulate the Slack sodium-activated potassium channel

The Slack gene encodes a Na⁺-activated K⁺ channel and is expressed in many different types of neurons. Like the prokaryotic Ca²⁺-gated K⁺ channel MthK, Slack contains two ‘regulator of K⁺ conductance’ (RCK) domains within its carboxy terminal, domains likely involved in Na⁺ binding and channel gating. It also contains multiple consensus protein kinase C (PKC) and protein kinase A (PKA) phosphorylation sites and although regulated by protein kinase C (PKC) phosphorylation, modulation by PKA has not been determined. To test if PKA directly regulates Slack, nystatin-perforated patch whole-cell currents were recorded from a human embryonic kidney (HEK-293) cell line stably expressing Slack. Bath application of forskolin, an adenylate cyclase activator, caused a rapid and complete inhibition of Slack currents however, the inactive homolog of forskolin, 1,9-dideoxyforskolin caused a similar effect. In contrast, bath application of 8-bromo-cAMP did not affect the amplitude nor the activation kinetics of Slack currents. In excised inside-out patch recordings, direct application of the PKA catalytic subunit to patches did not affect the open probability of Slack channels nor was open probability affected by direct application of protein phosphatase 2B. Preincubation of cells with the protein kinase A inhibitor KT5720 also did not change current density. Finally, mutating the consensus phosphorylation site located between RCK domain 1 and domain 2 from serine to glutamate did not affect current activation kinetics. We conclude that unlike PKC, phosphorylation by PKA does not acutely modulate the function and gating activation kinetics of Slack channels.     

Effects of KATP openers on the QT prolongation induced by HERG‐blocking drugs in guinea‐pigs

Objectives This work evaluated the potential usefulness of pharmacological activation of cardiac ATP‐sensitive potassium channels (K_ATP) in the prevention of drug‐induced QT prolongation in anaesthetised guinea‐pigs. Prolongation of cardiac repolarisation and QT interval is an adverse effect of many drugs blocking HERG potassium channels. This alteration can be dangerously arrhythmogenic and has been associated with the development of a particular form of ventricular tachyarrhythmia known as torsade de pointes. Methods The well‐known K_ATP openers aprikalim, cromakalim and pinacidil were used. Moreover, three benzothiazine derivatives, which have been reported as potent activators of K_ATP channels, were also used. Key findings Pharmacological activation of K_ATP channels caused a reduction of the QT prolongation, induced by astemizole, cisapride, quinidine and thioridazine. In contrast, the QT prolongation induced by haloperidol, sotalol and terfenadine, which are known to block HERG channels but also K_ATP channels, was not influenced by K_ATP activation. Glibenclamide and tolbutamide (non‐selective blockers of K_ATP channels expressed both in sarcolemmal and in mitochondrial membranes) antagonised the effects of K_ATP openers, whereas 5‐hydroxydecanoic acid (selective blocker of the mitochondrial K_ATP channels) failed to antagonise the effects of K_ATP openers, indicating that only the sarcolemmal K_ATP is involved in the cardioprotective activity. Conclusions The data suggest that pharmacological K_ATP activation might represent an option for treatment of patients exposed to QT‐prolonging drugs.     

药物对hERG钾通道作用机制研究进展

人ether-a-go-go-related gene(hERG)钾通道表达了延迟整流钾电流的快激活成分,对动作电位的复极至关重要。hERG钾电流不仅是抗心律失常作用的主要靶点,也是诸多药物增加尖端扭转型室速和心源性猝死风险的关键位点,而该电流的降低和(或)升高与基因突变或药物阻滞作用密切相关。随着对药物与hERG钾通道相互作用机制研究的深入,药物与通道孔道区蛋白结合位点的作用及其对通道转运的影响逐步被揭示,但这些药物对hERG作用的临床应用仍有待评价。     

POTASSIUM CHANNELS IN VASCULAR SMOOTH MUSCLE

1. Regulation of smooth muscle membrane potential through changes in K⁺ channel activity and subsequent alterations in the activity of voltage-dependent calcium channels is a major mechanism of vasodilation and vasoconstriction, both in normal and pathophysiological conditions. The contribution of a given K⁺ channel type to this mechanism of vascular regulation depends on the vascular bed and species examined. 2. Multiple K⁺ channels are present in most vascular smooth muscle cells and these different K⁺ channels play unique roles in regulating vascular tone. Voltage-dependent K⁺ (Kv) channels are activated by depolarization, may contribute to steady state resting membrane potential and are inhibited by certain vasoconstrictors. Calcium-activated K⁺ (K_Ca) channels oppose the depolarization associated with intrinsic vascular tone and are activated by some endogenous vasodilators. Small-conductance, apamin-sensitive K_Ca channels may be activated by endothelium-derived hyperpolarizing factor. ATP-sensitive K⁺ (K_ATP) channels are activated by pharmacological and endogenous vasodilators. Inward rectifier K⁺ (K_ir) channels are activated by slight changes in extracellular K⁺ and may contribute to resting membrane potential. 3. Membrane potential and diameter are determined, in part, by the integrated activity of several K⁺ channels, which are regulated by multiple dilator and constrictor signals in vascular smooth muscle.     

积分法:一种药理学中分析电压依赖性瞬时外向钾电流的方法(英文)

目的　评价一种药理学中分析电压依赖性瞬时外向钾电流 (Ito)的积分方法。方法　Ito的失活相可被 2指数或 3指数方程拟合。原始电流曲线下面积 (AUC)可通过指数方程积分获得。以细胞膜电容标准化后的曲线下面积表示除极化时程中钾离子的净通量 ,作为比较指标。钙调磷酸酶过表达转基因鼠心肌细胞表现Ito下调 ,此数据用于验证Ito的积分分析方法的可靠性。结果　AUC可通过 3指数或2指数方程的积分公式获得 :AUC =A1τ1+A2 τ2 +A3τ3+A0 t -A1τ1e-t/τ1-A2 τ2 e-t/τ2 -A3τ3e-t/τ3或AUC =A1τ1+A2 τ2 +A0 t-A1τ1e-t/τ1-A2 τ2 e-t/τ2 。小鼠心室肌 5 0 %和 90 %动作电位时程分别约为 10ms和 30ms。将Ito0～ 10ms (AUC50 )和 0～ 30ms(AUC90 )的曲线下面积以细胞膜电容进行标准化。野生型鼠心肌细胞的AUC50 和AUC90 明显高于钙调磷酸酶过表达转基因鼠 ,此结果与转基因鼠心室肌细胞动作电位时程延长相一致 ,与前期发表结果一致 (钙调磷酸酶过表达转基因鼠心肌细胞Ito各成分下调 )。结论　积分法是药理学中一种简便准确的分析Ito的方法。     

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Blockade and enhancement of glutamate receptor responses in Xenopus oocytes by methylated arsenicals

Pentavalent and trivalent organoarsenic compounds belong to the major metabolites of inorganic arsenicals detected in humans. Recently, the question was raised whether the organic arsenicals represent metabolites of a detoxification process or methylated species with deleterious biological effects. In this study, the effects of trivalent arsenite (AsO₃ ³⁻; iA^III), the pentavalent organoarsenic compounds monomethylarsonic acid (CH₃AsO(OH)₂; MMA^V) and dimethylarsinic acid ((CH₃)₂AsO(OH); DMA^V) and the trivalent compounds monomethylarsonous acid (CH₃As(OH)₂, MMA^III) and dimethylarsinous acid ((CH₃)₂As(OH); DMA^III) were tested on glutamate receptors and on voltage-operated potassium and sodium channels heterologously expressed in Xenopus oocytes. Membrane currents of ion channels were measured by conventional two-electrode voltage-clamp techniques. The effects of arsenite were tested in concentrations of 1–1,000 μmol/l and the organic arsenical compounds were tested in concentrations of 0.1–100 μmol/l. We found no significant effects on voltage-operated ion channels; however, the arsenicals exert different effects on glutamate receptors. While MMA^V and MMA^III significantly enhanced ion currents through N-methyl-d-aspartate (NMDA) receptor ion channels with threshold concentrations <10 μmol/l, DMA^V and DMA^III significantly reduced NMDA-receptor mediated responses with threshold concentrations <0.1 μmol/l; iA^III had no effects on glutamate receptors of the NMDA type. MMA^III and DMA^V significantly reduced ion currents through α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-receptor ion channels with threshold concentrations <10 μmol/l (MMA^III) and <1 μmol/l (DMA^V). MMA^V and iA^III had no significant effects on glutamate receptors of the AMPA type. The effects of MMA^V, MMA^III, DMA^V and DMA^III on glutamate receptors point to a neurotoxic potential of these substances.Katharina Krüger and Janina Gruner contributed equally to this work.     

二乙酰单肟抑制豚鼠和鸡胚心肌细胞离子通道电导

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Gambierol, a toxin produced by the dinoflagellate Gambierdiscus toxicus, is a potent blocker of voltage-gated potassium channels

In this study, we pharmacologically characterized gambierol, a marine polycyclic ether toxin which is produced by the dinoflagellate Gambierdiscus toxicus. Besides several other polycyclic ether toxins like ciguatoxins, this scarcely studied toxin is one of the compounds that may be responsible for ciguatera fish poisoning (CFP). Unfortunately, the biological target(s) that underlies CFP is still partly unknown. Today, ciguatoxins are described to specifically activate voltage-gated sodium channels by interacting with their receptor site 5. But some dispute about the role of gambierol in the CFP story shows up: some describe voltage-gated sodium channels as the target, while others pinpoint voltage-gated potassium channels as targets. Since gambierol was never tested on isolated ion channels before, it was subjected in this work to extensive screening on a panel of 17 ion channels: nine cloned voltage-gated ion channels (mammalian Na_v1.1–Na_v1.8 and insect Para) and eight cloned voltage-gated potassium channels (mammalian K_v1.1–K_v1.6, hERG and insect ShakerIR) expressed in Xenopus laevis oocytes using two-electrode voltage-clamp technique. All tested sodium channel subtypes are insensitive to gambierol concentrations up to 10 μM. In contrast, K_v1.2 is the most sensitive voltage-gated potassium channel subtype with almost full block (>97%) and an half maximal inhibitory concentration (IC₅₀) of 34.5 nM. To the best of our knowledge, this is the first study where the selectivity of gambierol is tested on isolated voltage-gated ion channels. Therefore, these results lead to a better understanding of gambierol and its possible role in CFP and they may also be useful in the development of more effective treatments.     

Ionotropic receptors and ion channels in ischemic neuronal death and dysfunction

Loss of energy supply to neurons during stroke induces a rapid loss of membrane potential that is called the anoxic depolarization. Anoxic depolarizations result in tremendous physiological stress on the neurons because of the dysregulation of ionic fluxes and the loss of ATP to drive ion pumps that maintain electrochemical gradients. In this review, we present an overview of some of the ionotropic receptors and ion channels that are thought to contribute to the anoxic depolarization of neurons and subsequently, to cell death. The ionotropic receptors for glutamate and ATP that function as ligand-gated cation channels are critical in the death and dysfunction of neurons. Interestingly, two of these receptors (P2X7 and NMDAR) have been shown to couple to the pannexin-1 (Panx1) ion channel. We also discuss the important roles of transient receptor potential (TRP) channels and acid-sensing ion channels (ASICs) in responses to ischemia. The central challenge that emerges from our current understanding of the anoxic depolarization is the need to elucidate the mechanistic and temporal interrelations of these ion channels to fully appreciate their impact on neurons during stroke.