Opening Of Calcium-activated Potassium Channels Research Articles

Pathological role of the calcium-sensing receptor in sepsis-induced hypotensive shock: Therapeutic possibilities and unanswered questions.

Sepsis is a life-threatening disease involving multiorgan dysfunction, prompted by an unregulated host response to infection. Shock is a complication of sepsis in which the circulatory and cellular metabolism anomalies are significant enough to raise the risk of death. Calcium dyshomeostasis occurs during sepsis condition due to imbalance between calcium uptake and excessive release induced by inflammatory cytokines. This calcium imbalance can cause activation of calcium-sensing receptors (CaSRs) located on the surface of T cells and thereby promote release of reactive oxygen species (ROS). The elevated ROS and inflammatory cytokines during sepsis condition have been reported to directly damage the endothelial cells, disrupt the barrier functions that might result in leakage of fluids, and inflammatory cells in tissues Moreover, several evidence have revealed that the calcium mediated activation of CaSR could produce systemic vasodilatory response by stimulating the nitric oxide production and opening of calcium-activated potassium channels, while infusion of its antagonist elevated the blood pressure. These evidence indicate that activation of CaSR during sepsis conditions results in release of ROS and inflammatory cytokines, which could produce an endothelial barrier damage, cardiomyocyte apoptosis. These pathological events could produce loss of fluid in tissues and cardiac dysfunction. Further the direct vasodilatory effects of CaSR activation might add to the shock-like condition. Thus, we hereby propose that inhibition of CaSR could suppress the release of ROS, inflammatory mediators, and thereby prevent the endothelial damage, cardiac dysfunction, and maintain systemic vascular tone.

Possible Mechanisms Involved in the Vasorelaxant Effect Produced by Anorexigenic Drugs in Rat Aortic Rings

Anorexigenics are compounds capable of reducing or suppressing appetite. Their three main types act on different neurotransmitters, either norepinephrine, serotonin or a combination of both. Among the drugs that act on norepinephrine are fenproporex, amfepramone and clobenzorex. Derivatives of the thyroid hormone triiodothyronine have also been associated with weight loss and used as a controversial treatment for obesity, despite their known cardiovascular side effects. Recent data suggest a possible vasodilating effect for these four substances that might be beneficial in a subset of patients. Herein we performed a systematic review of the literature (with emphasis on recent reports) to determine the implications and mechanisms of the vasodilating effects of some anorectics, specifically fenproporex, clobenzorex, amfepramone and triiodothyronine. Data analysis showed these four drugs to be vasodilating agents for rat aortic rings. The different mechanisms of action include endothelium-dependent vasodilation via activation of the NO-cGMP-PKG pathway and the opening of calcium-activated potassium channels. The finding of vasodilating activity indicates a potential role for some anorexigenic drugs in the treatment of obesity in hypertensive patients. Further in vivo studies are needed to test the clinical benefits of these four drugs.

Small and Intermediate Calcium-Activated Potassium Channel Openers Improve Rat Endothelial and Erectile Function.

Modulation of endothelial calcium-activated potassium (KCa) channels has been proposed as an approach to restore endothelial function. The present study investigated whether novel openers of KCa channels with small (KCa2.x) and intermediate (KCa3.1) conductance, NS309 and NS4591, improve endothelium-dependent relaxation and erectile function. Rat corpus cavernosum (CC) strips were mounted for isometric tension recording and processed for immunoblotting. Mean arterial pressure (MAP), intracavernosal pressure (ICP), and electrocardiographic (ECG) measurements were conducted in anesthetized rats. Immunoblotting revealed the presence of KCa2.3 and large KCa conductance (KCa1.1) channels in the corpus cavernosum. NS309 and NS4591 increased current in CC endothelial cells in whole cell patch clamp experiments. Relaxation induced by NS309 (<1 μM) was inhibited by endothelial cell removal and high extracellular potassium. An inhibitor of nitric oxide (NO) synthase, and blockers of KCa2.x and KCa1.1 channels, apamin and iberiotoxin also inhibited NS309 relaxation. Incubation with NS309 (0.5 μM) markedly enhanced acetylcholine relaxation. Basal erectile function (ICP/MAP) increased during administration of NS309. Increases in ICP/MAP after cavernous nerve stimulation with NS309 were unchanged, whereas NS4591 significantly improved erectile function. Administration of NS309 and NS4591 caused small changes in the electrocardiogram, but neither arrhythmic events nor prolongation of the QTc interval were observed. The present study suggests that openers of KCa2.x and KCa3.1 channels improve endothelial and erectile function. The effects of NS309 and NS4591 on heart rate and ECG are small, but will require additional safety studies before evaluating whether activation of KCa2.3 channels has a potential for treatment of erectile dysfunction.

Opening of calcium-activated potassium channels improves long-term left-ventricular function after coronary artery occlusion in mice

BackgroundOpening of mitochondrial calcium-activated potassium channels (BKCa) reduces infarct size after myocardial ischemia/reperfusion injury (I/R). It is unknown if targeting BKCa-channels improves cardiac performance in the long-term after I/R. MethodsExperiments were conducted in compliance with institutional and national guidelines in C57BL/6 mice (n=7–8/group). Animals were randomized into two groups. Preconditioning was induced by intraperitoneal application of NS1619 (NS, 1μg/g bw) 10min before ischemia, control animals (Con) received the vehicle. All animals underwent 45min of myocardial ischemia and four weeks of reperfusion. Transthoracal Echocardiography (TTE) was conducted one and four weeks after ischemia (TTEW1/TTEW4) and additionally a cardiac MRI was done in week four. At the end of experiments the infarction scar was determined by AZAN staining. ResultsTTE revealed that NS1619 improved ejection fraction one week (Con: 36±4%, NS: 45±4%; P<0.05) and four weeks after I/R (Con: 33±11%, NS: 46±8%; P<0.05). Preconditioning with NS1619 reduced end-diastolic volume at both time points (TTEW1: Con: 60±12μl, NS: 45±8μl; TTEW4: Con: 82±31μl, NS: 44±8μl; each P<0.05) and increased fractional shortening after four weeks (TTEW4: Con: 12±6%, NS: 24±8%; P<0.05). MRI-analysis after four weeks confirmed the echocardiographic results. NS1619 increased ejection fraction by 45% (MRI: Con: 29±6%, NS: 42±9%; P<0.05 vs. Con) and reduced end-diastolic and -systolic volume (EDV, ESV) compared to control (MRI: EDV: Con: 110±19μl, NS: 88±16μl; ESV: Con: 79±19μl, NS: 53±18μl; each P<0.05). Preconditioning reduced infarction scar after four weeks by 25% (Con: 12±3%, NS: 9±2%; P<0.05). ConclusionsPreconditioning by opening of BKCa-channels with NS1619 improves cardiac performance after four weeks of reperfusion and reduces myocardial infarction scar.

Antihypertensive drugs

Antihypertensive drugs

Enantioresolution, stereochemical characterization and biological activity of a chiral large-conductance calcium-activated potassium channel opener

A number of large-conductance calcium-activated potassium (BK) channel openers based on the 2-aryl-1,4-benzothiazine scaffold were previously synthesized, and 2-(5-bromo-2-methoxyphenyl)-6-trifluoromethyl-2H-1,4-benzothiazin-3(4H)-one (1) was identified as the most active compound. Since a stereoselective activation of BK channels was demonstrated for arylindolone derivatives, the effect of the absolute configuration at the C-2 position on the vasorelaxing potency of 2-aryl-1,4-benzothiazines is investigated in this article. Compound 1 was initially evaluated as a racemate: subsequently, the “racemic approach” was used to isolate its enantiomers. The excellent enantioresolution obtained using the Sepapak-4 column (CSP 4, cellulose tris(4-chloro-3-metylphenylcarbamate); RS=8.36; α=2.03) allowed to collect highly pure enantiomeric fractions, with enantiomeric excess (e.e.) values higher than 97% and 98% for the first- and second-eluted enantiomer, respectively. Electronic circular dichroism (ECD) studies on the two isolated enantiomers, combined with time-dependent density functional theory (TD-DFT) calculations allowed to characterize the configuration of the enantiomers and determine a (R), (S) elution order. Results from biological assays indicated that the racemate and the isolated enantiomers are endowed with comparable vasorelaxing potency.

The therapeutic future in headache

There are many categories and individual types of headache and most have a variety of treatment protocols, while a few are best treated by just one medication. This paper will concentrate on acute care medications for migraine and discuss some new and future acute care treatments. There is not much to discuss about prevention, except that onabotulinumtoxinA has been approved for prevention of chronic migraine. Cluster headache will also be discussed, as there are some future treatments for acute care and prevention being studied at present. For the acute care of migraine in the US, we have seven triptans by tablet plus other routes and one non steroidal anti-inflammatory medication approved by the FDA that is currently available (Cambia brand of buffered diclofenac potassium for oral solution). There are several other acute care medications in various stages of development and there are three new methods of administering a triptan and others under investigation. The optimal acute care therapy for migraine should be faster, easier to use and more efficient with fewer adverse events than what is currently available. What follows is a brief review of the status in development for five of the many new acute care medications being investigated: the CGRP antagonist tablet telcagepant, the sumatriptan iontophoretic patch Zelrix, sumatriptan powder for use in the OptiNose apparatus, dihydroergotamine for oral inhalation (Levadex), civamide nasal solution for prevention of episodic cluster headache (Civanex) and sphenopalatine ganglion stimulation for acute cluster attacks in chronic cluster headaches. Other future treatments that will not be discussed include transcranial magnetic stimulation, a 5-HT(1F) agonist named alniditan, large conductance calcium-activated potassium channel openers, glial modulators or other medications and devices in early stages of development.

New frontiers in headache therapy

There are numerous headache therapies available for our patients, more for migraine than for any of the other primary headache disorders. Only four medications have been approved for migraine prevention in the last few decades in the US and onabotulinumtoxinA was recently approved in the UK and the US for chronic migraine. We have been more fortunate in the acute care arena where in the US we have had seven triptans and one nonsteroidal anti-inflammatory medication approved by the FDA and currently available. There are several other acute care medications in various stages of development and there are two new methods of administering a triptan and others under investigation. We are always looking for faster, easier and more efficient administration of medications with fewer adverse events, as optimal migraine therapy requires these characteristics. What follows is a brief review of the progress in development for four of the many new acute care medications being investigated: the CGRP antagonist tablet telcagepant, the sumatriptan iontophoretic patch, sumatriptan powder for use in the OptiNose apparatus and the dihydroergotamine oral inhaler. I will not include transcranial magnetic stimulation, a 5-HT(1F) agonist, large conductance calcium-activated potassium channel openers, glial modulators or other medications and devices in early stages of development [1].

Endothelium as target for large-conductance calcium-activated potassium channel openers.

The endothelium is a highly active organ responsible for vasculatory tone and structure, angiogenesis, as well as hemodynamic, humoral, and inflammatory responses. The endothelium is constantly exposed to blood flow, sheer stress and tension. Endothelial cells are present as a vasculature in every tissue of the body and react to and control its microenvironment. A variety of ion channels are present in the plasma membranes of endothelial cells. These include potassium channels such as inwardly rectifying potassium (K(ir)) channels, voltage-dependent (K(v)) channels, ATP-regulated potassium (K(ATP)) channels and three types of calcium-activated potassium channels (K(Ca)), the large (BK(Ca)), intermediate (IK(Ca)), and small (SK(Ca)) -conductance potassium channels. Potassium current plays a critical role in action potentials in excitable cells, in setting the resting membrane potential, and in regulating neurotransmitter release. Mitochondrial isoforms of potassium channel contribute to the cytoprotection of endothelial cells. Prominent among potassium channels are families of calcium-activated potassium channels, and especially large-conductance calcium-activated potassium channels. The modulation of BK(Ca) channels, which are voltage- and calcium-dependent, has been intensively studied. The BK(Ca) channels show large expression dynamics in endothelial cells and tissue-specific expression of large numbers of alternatively spliced isoforms. In this review, a few examples of the modulatory mechanisms and physiological consequences of the expression of BK(Ca) channels are discussed in relation to potential targets for pharmacological intervention.

Activation of muscarinic receptors by a hydroalcoholic extract of Dicksonia sellowiana Presl. HooK (Dicksoniaceae) induces vascular relaxation and hypotension in rats

Dicksonia sellowiana (Presl.) Hook is a native plant from the Central and South Americas that contain high levels of polyphenols, antioxidant compounds involved in protection against inflammation, cancer and cardiovascular risk. A phytomedicinal preparation obtained from aerial parts of D. sellowiana is currently under clinical evaluation in Brazil against asthma, and has been associated with several other beneficial effects. This study demonstrates that a hydroalcoholic extract obtained from D. sellowiana leaves (HEDS) fully relax, in a concentration-dependent manner, rat aortic rings precontracted with phenylephrine. Moreover, administration of HEDS (10, 20 and 40 mg/kg, i.v.) in anaesthetized rats resulted in a strong but reversible hypotension. Aortic relaxation induced by HEDS was abolished by endothelium removal, by incubation of the nitric oxide synthase inhibitor L-NAME, or the soluble guanylate cyclase inhibitor ODQ. In addition, this effect was partially inhibited by indomethacin (a cyclooxygenase inhibitor) and KT 5730 (a PKA inhibitor). The potassium channels blockade by either tetraethylammonium or charybdotoxin also resulted in a potent inhibition of HEDS-induced aortic relaxation, whereas apamine only slightly reduced it. In addition HEDS-induced relaxation was unchanged by 4-amynopiridine and glibenclamide. The selective muscarinic receptor antagonist atropine counteracted both aortic relaxation and blood pressure reduction generated by HEDS. Experiments using HPLC revealed the presence of high amounts of phenolic compounds in this extract. Taken together, our results reveal that the D. sellowiana possess substances with both in vivo and in vitro activities and that the vascular effect of HEDS involves activation of muscarinic receptors, stimulation of the nitric oxide pathway and opening of calcium-activated potassium channels.

Bradykinin induces microvascular preconditioning through the opening of calcium-activated potassium channels

This study was designed to investigate whether the activation of calcium-activated potassium (K(Ca)) or adenosine triphosphate sensitive potassium (K(ATP)) channels are required for bradykinin-induced microvascular preconditioning. Isolated rabbit hearts underwent retrograde perfusion with Krebs-Henseleit buffer (KHB) followed by 60 minutes of ischemic arrest with cold crystalloid cardioplegia (CCCP). Eight CCCP hearts received no pretreatment. Six bradykinin-preconditioned hearts received a 10-minute coronary infusion of 10(-8) mol/L bradykinin-enriched KHB followed by a 5-minute recovery period before CCCP. Six hearts received both 10(-8) mol/L charybdotoxin (a K(Ca) channel blocker) and bradykinin preconditioning. Finally, 6 other hearts received 10(-5 degrees ) mol/L glibenclamide (a K(ATP) channel blocker) to bradykinin-enriched KHB. All hearts were reperfused for 30 minutes with KHB. Bradykinin preconditioning significantly improved the recovery of left ventricular and microvascular function, as compared with control. On the other hand, bradykinin preconditioning significantly reduced the contractile responses to U46619, a thromboxane A2 analogue. Charybdotoxin significantly inhibited the improved recovery of bradykinin-induced left ventricular and microvascular function. Glibenclamide tended to diminish the bradykinin preconditioning-enhanced recovery of left ventricular function, but failed to affect bradykinin preconditioning-improved recovery of microvascular function. Both K(Ca) and K(ATP) channels were involved partially in bradykinin-induced myocardial preconditioning. However, bradykinin induces microvascular preconditioning through the opening of K(Ca) channels rather than K(ATP) channels.

Endothelium-Derived Hyperpolarizing Factor

The endothelium controls vascular tone not only by releasing nitric oxide (NO) and prostacyclin but also by other pathways causing hyperpolarization of the underlying smooth muscle cells. This characteristic was at the origin of the denomination endothelium-derived hyperpolarizing factor (EDHF). We know now that this acronym includes different mechanisms. In general, EDHF-mediated responses involve an increase in the intracellular calcium concentration, the opening of calcium-activated potassium channels of small and intermediate conductance and the hyperpolarization of the endothelial cells. This results in an endothelium-dependent hyperpolarization of the smooth muscle cells, which can be evoked by direct electrical coupling through myo-endothelial junctions and/or the accumulation of potassium ions in the intercellular space. Potassium ions hyperpolarize the smooth muscle cells by activating inward rectifying potassium channels and/or Na+/K(+)-ATPase. In some blood vessels, including large and small coronary arteries, the endothelium releases arachidonic acid metabolites derived from cytochrome P450 monooxygenases. The epoxyeicosatrienoic acids (EET) generated are not only intracellular messengers but also can diffuse and hyperpolarize the smooth muscle cells by activating large conductance calcium-activated potassium channels. Additionally, the endothelium can produce other factors such as lipoxygenases derivatives or hydrogen peroxide (H2O2). These different mechanisms are not necessarily exclusive and can occur simultaneously.

Hydrochlorothiazide exerts no direct vasoactivity in the human forearm

Recently, hydrochlorothiazide has been shown to relax vascular smooth muscle in vitro in clinically relevant concentrations by the opening of calcium-activated potassium channels, leading to hyperpolarization and consequent closing of voltage-operated calcium channels. Long-term administration of hydrochlorothiazide reduces peripheral vascular resistance in vivo in man. These results indicate that hydrochlorothiazide has hemodynamic activity, and we therefore examined the direct vascular action of this drug in vivo. Forearm vasodilator responses to the infusion of a placebo and five increasing doses of hydrochlorothiazide into the brachial artery were recorded by venous occlusion strain-gauge plethysmography (perfused forearm technique) in eight normotensive male volunteers. Venous samples were taken from an ipsilateral antecubital vein at the end of each infusion period to measure the hydrochlorothiazide concentration. Plasma concentrations of hydrochlorothiazide averaged 3.5 +/- 0.3 mu g/ml at the highest infusion rate. This concentration leads to a 60 +/- 10% relaxation in vitro and is more than 10 times the therapeutic plasma concentration. Despite these supratherapeutic levels, we were unable to demonstrate a change in forearm blood flow and vascular resistance. Also, no significant changes were observed in blood pressure and heart rate. In contrast to in vitro results, hydrochlorothiazide does not exert any direct vasoactivity in the forearm vascular bed of healthy normotensive male volunteers at (supra)therapeutic plasma concentrations.

5-HT2A receptor-mediated outward current in C6 glioma cells is mimicked by intracellular IP3 release.

The C6 glioma cell line possesses 5-HT2A receptors that have been shown to increase intracellular calcium levels. We have studied the electrophysiological response of these cells to 5-HT using the whole-cell recording method. Under voltage-clamp, 5-hydroxytryptamine (5-HT) produced an outward current in these cells which was inhibited by extracellularly applied ketanserin and spiperone and by EGTA (10 mM) in the recording electrode. The 5-HT induced response could be mimicked by intracellular photolytic release of inositol (1,4,5) trisphosphate (IP3) from caged molecules. The reversal potentials for the IP3- and 5-HT-induced responses were closely matched. The data indicates that the outward current is likely to be mediated by 5-HT2A receptors stimulating IP3 production which increases intracellular calcium leading to the opening of calcium-activated potassium channels.

Role of G‐proteins in muscarinic receptor inward and outward currents in rabbit jejunal smooth muscle.

1. Single smooth muscle cells obtained by enzymic dispersion of the longitudinal muscle layer of rabbit jejunum were held under voltage clamp using patch pipettes and membrane currents measured. The effects of carbachol or caffeine applied externally were examined in cells dialysed with normal pipette solutions or with a solution containing heparin (which blocks receptors for D-myo-inositol 1,4,5-trisphosphate, InsP3), guanosine 5-O-(gamma-thio)triphosphate (GTP gamma S) or guanosine 5-O-(beta-thio)diphosphate (GDP beta S). 2. Outward current in response to application of carbachol or caffeine was considered to represent the opening of calcium-activated potassium channels in response to a localized rise in the free ionized calcium concentration occasioned by the rapid discharge of stored calcium (Ca) by these agents. 3. Heparin included in the pipette solution blocked outward current to muscarinic receptor activation by carbachol but not that to caffeine, suggesting that receptor-evoked discharge of stored cellular Ca is caused by InsP3 action. However, heparin did not affect muscarinic-receptor inward current. 4. After dialysis with 0.1-0.5 mM-GTP gamma S, carbachol inward current was evoked in two out of three of the cells; after dialysis with 0.1-0.2 mM-GTP gamma S for an average of 7.7 min it was 80% of the normal response; after dialysis for an average of 8.6 min with 0.5 mM-GTP gamma S it was 31% of the normal response. In contrast, 0.1 mM-GTP gamma S reduced caffeine outward current by 93% after an average 4.5 min dialysis and spontaneous transient outward currents (STOCs) were abolished in 2.9 min on average. 5. Carbachol inward current (at -40 or -50 mV) and carbachol outward current (at 0 mV) in responding cells were reduced only by half after 8-10 min dialysis with 1 mM-GDP beta S which has been shown in portal vein cells to antagonize the depletion of Ca stores by intracellular GTP gamma S (Komori & Bolton, 1989). After 8-10 min dialysis with 5 mM-GDP beta S outward current was 27% of normal. However, if GDP beta S was present, outward current generally could not be evoked by a second application of carbachol. 6. The discharge of Ca stores by dialysis with 0.1 mM-GTP gamma S was prevented completely by heparin included in the pipette solution, suggesting that activation of a G-protein associated with phospholipase C (PLC) enzyme accelerates PLC activity. InsP3 production and depletion of Ca stores.(ABSTRACT TRUNCATED AT 400 WORDS)

Aminopyridines enhance opening of calcium-activated potassium channels in GH3 anterior pituitary cells.

Aminopyridines enhance opening of calcium-activated potassium channels in GH3 anterior pituitary cells.

A model for exocytosis based on the opening of calcium-activated potassium channels in vesicles

It is proposed that the role of calcium in calcium-induced exocytosis is to open Ca-activated K channels present in vesicle membranes. The opening of these channels coupled with anion transport across the vesicle membranes would result in an influx of K and anions, increasing the osmotic pressure of the vesicles. For those vesicles situated very close to the cell plasma membrane, this would lead to fusion with the membrane and exocytosis of the vesicle contents. This model can account for facilitation and other key properties of transmitter release. In addition, the model predicts that vesicles with a higher transmitter content, and hence higher initial osmotic pressure, would be preferentially discharged. The model also predicts that a faster response can be obtained for small vesicles than for large vesicles, providing a rationaleas to why neurotransmitters, which must be released quickly, are packaged in small vesicles.