大电导钙激活钾通道对平滑肌的调控作用研究进展

钾通道是组织器官中的一种重要通道,几乎所有的组织中都有该通道的分布,并且它在调节细胞功能方面起着极其重要的作用,例如动作电位的形成和信号传导等。大电导钙激活钾通道(BKca/Maxi K)以其广泛的分布,以及参与调节多种细胞功能吸引了更多研究者的关注。BKca/Maxi K的激活可导致细胞膜的超极化,从而抑制电压依赖性钙通道的激活,抑制钙离子内流,引起平滑肌舒张。近年来研究发现,BKca/Maxi K的激活、失活和变异与多种疾病的发病有关,BKca/Maxi K对心血管平滑肌、子宫平滑肌、呼吸道平滑肌和阴茎勃起等具有调控作用,尤其是其基因疗法对阴茎勃起障碍的治疗逐渐显现出较大的优势。     

Carbon monoxide activates large-conductance calcium-activated potassium channels of human cardiac fibroblasts through various mechanisms

Carbon monoxide (CO) is a cardioprotectant and potential cardiovascular therapeutic agent. Human cardiac fibroblasts (HCFs) are important determinants of myocardial structure and function. Large-conductance Ca²⁺-activated K⁺ (BK) channel is a potential therapeutic target for cardiovascular disease. We investigated whether CO modulates BK channels and the signaling pathways in HCFs using whole-cell mode patch-clamp recordings. CO-releasing molecules (CORMs; CORM-2 and CORM-3) significantly increased the amplitudes of BK currents (IBK). The CO-induced stimulating effects on IBK were blocked by pre-treatment with specific nitric oxide synthase (NOS) blockers (L-N^G-monomethyl arginine citrate and L-N^G-nitroarginine methyl ester). 8-bromo-cyclic GMP increased IBK. KT5823 (inhibits PKG) or ODQ (inhibits soluble guanylate cyclase) blocked the CO-stimulating effect on IBK. Moreover, 8-bromo-cyclic AMP also increased IBK, and pre-treatment with KT5720 (inhibits PKA) or SQ22536 (inhibits adenylate cyclase) blocked the CO effect. Pre-treatment with N-ethylmaleimide (a thiol-alkylating reagent) also blocked the CO effect on IBK, and DL-dithiothreitol (a reducing agent) reversed the CO effect. These data suggest that CO activates IBK through NO via the NOS and through the PKG, PKA, and S-nitrosylation pathways.     

Pharmacological roles of the large-conductance calcium-activated potassium channel

The gating of large-conductance Ca(2+)-activated K(+) (BK(Ca)) channel is primarily controlled by intracellular Ca(2+) and/or membrane depolarization. These channels play a role in the coupling of excitation-contraction and stimulus-secretion. A variety of structurally distinct compounds may influence the activity of these channels. Squamocin, an Annonaceous acetogenin, could interact with the BK(Ca) channel to increase the amplitude of Ca(2+)-activated K(+) current in coronary smooth muscle cells. Its stimulatory effect is related to intracellular Ca(2+) concentrations. In inside-out patches, application of ceramide to the bath suppressed the activity of BK(Ca) channels recorded from pituitary GH(3) cells and from retinal pigment epithelial cells. ICI-182,780, an estrogen receptor antagonist, was found to modulate BK(Ca)-channel activity in cultured endothelial cells and smooth muscle cells in a mechanism unlinked to the inhibition of estrogen receptors. Caffeic acid phenethyl ester (CAPE) and its analogy, cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate, could directly increase the activity of BK(Ca) channels in GH(3) cells. CAPE also reduced the frequency and amplitude of intracellular Ca(2+) oscillations in these cells. The CAPE-stimulated activity in BK(Ca) channels is thought to be unassociated with its inhibition of NF-kappaB activation. Cilostazol, an inhibitor of cyclic nucleotide phosphodiesterase, could stimulate BK(Ca) channel-activity and reduce the firing of action currents simultaneously in GH(3) cells. Therefore, the regulation by these compounds of BK(Ca) channels may in part be responsible for their regulatory actions on cell functions.     

High-conductance calcium-activated potassium channels

High-conductance, calcium-activated potassium (Maxi-K) channels represent a group of proteins with diverse physiologic functions. Although the role of Maxi-K channels in the CNS is complex and still an area of active academic research, there appears to be better agreement concerning the contribution of these channels to the regulation of smooth muscle tone and, thus the expectations remain high for Maxi-K channel openers having use in the treatment of hypertension, overactive bladder, asthma or erectile dysfunction. Despite this consensus view, at the present time, there is only one compound that targets Maxi-K channels in clinical development for overactive bladder conditions. In the present review, the latest developments in the identification of potent and selective Maxi-K channel openers and their utility in the treatment of smooth muscle disorders will be discussed.     

Methamphetamine inhibits voltage-gated potassium currents in NG108-15 cells: Possible contribution of large-conductance calcium-activated potassium channels

Methamphetamine (MA), a highly abused amphetamine-like psychostimulant, has surged in popularity worldwide in the last decade. Repeated MA exposure has been shown to affect the alternative splice variant expression of large conductance Ca²⁺-activated K⁺ (BK) channels. It remains unclear whether MA affects BK channel activity. The present study investigated the effects of MA on BK channels in NG108-15 mouse neuroblastoma × rat glioma hybrid cells using whole-cell and cell-attached patch clamp techniques. In whole-cell recordings, the macroscopic K⁺ outward currents were inhibited by MA with an EC₅₀ of 146 μM, but not affected by dopamine (DA). It implies that DA is not involved in the effects of MA on K⁺ outward currents. In cell-attached patches, MA significantly decreased BK channel activity. Moreover, MA significantly decreased the BK channel opener NS1619-evoked whole-cell K⁺ outward currents and BK channel activity. Finally, the effect of MA on membrane potential was examined by current-clamp configuration. MA caused membrane depolarization and application of NS1619 returned the depolarized potential to resting value. These findings suggest that MA might act as an inhibitor of BK channels, and thereby increase the neuronal excitability and enhance neurotransmitter release.     

Carbon monoxide releasing molecule-2 suppresses stretch-activated atrial natriuretic peptide secretion by activating large-conductance calcium-activated potassium channels

Carbon monoxide (CO) is a known gaseous bioactive substance found across a wide array of body systems. The administration of low concentrations of CO has been found to exert an anti-inflammatory, anti-apoptotic, anti-hypertensive, and vaso-dilatory effect. To date, however, it has remained unknown whether CO influences atrial natriuretic peptide (ANP) secretion. This study explores the effect of CO on ANP secretion and its associated signaling pathway using isolated beating rat atria. Atrial perfusate was collected for 10 min for use as a control, after which high atrial stretch was induced by increasing the height of the outflow catheter. Carbon monoxide releasing molecule-2 (CORM-2; 10, 50, 100 µM) and hemin (HO-1 inducer; 0.1, 1, 50 µM), but not CORM-3 (10, 50, 100 µM), decreased high stretch-induced ANP secretion. However, zinc porphyrin (HO-1 inhibitor) did not affect ANP secretion. The order of potency for the suppression of ANP secretion was found to be hemin > CORM-2 >> CORM-3. The suppression of ANP secretion by CORM-2 was attenuated by pretreatment with 5-hydroxydecanoic acid, paxilline, and 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one, but not by diltiazem, wortmannin, LY-294002, or NG-nitro-L-arginine methyl ester. Hypoxic conditions attenuated the suppressive effect of CORM-2 on ANP secretion. In sum, these results suggest that CORM-2 suppresses ANP secretion via mitochondrial K_ATP channels and large conductance Ca²⁺-activated K⁺ channels.     

阻塞性睡眠呼吸暂停低通气综合征患者脑血管运动反应性及大电导钙激活钾通道β1亚单位表达的改变

目的 初步探讨阻塞性睡眠呼吸暂停低通气综合征患者(OSAHS)脑血管运动反应性及外周血白细胞大电导钙激活钾通道β1(BKcaβ1)亚单位表达的改变.方法 选OSAHS患者216例为OSAHS组,根据呼吸暂停低通气指数(AHI)和夜间最低血氧饱和度(LSaO2)将OSAHS组分为轻度(76例)、中度(87例)、重度(53例)OSAHS组;选择同期单纯性打鼾(AHI＜5次/h)的健康体检者65例作为健康对照组.通过过度换气、屏气及倾斜试验评价患者脑血管运动反应性和自动调节能力,并检测白细胞中BKCaβ1mRNA表达.结果 与健康对照组比较,中、重度OSAHS组患者低碳酸血症期、高碳酸血症期脑血管运动反应性降低,由卧位至立位平均动脉压和脑血流速度恢复90％所用时间增加,差异均有统计学意义(均P＜0.05);中、重度OSAHS组卧位和立位的平均收缩压分别为(117 ±15)和(108 ±14)mm Hg(1 mm Hg=0.13 kPa),平均舒张压分别为(72±16)和(64±12)mm Hg,平均脑血流速度分别为(41±9)和(33±13)cm/s,不同体位间指标的差异均有统计学意义(均P＜0.05).健康对照组和中、重度OSAHS组的BKCaβ1mRNA表达值分别为1.13 ±0.41和0.63±0.38、0.48 ±0.29,中、重度OSAHS与健康对照组的差异有统计学意义(t值分别为7.838、9.588,P＜0.05),中、重度OSAHS组间差异有统计学意义(t=2.391,P＜0.05).脑血管运动反应性与BKcaβ1mRNA呈正相关,与血管的舒张能力关系较强(r=0.685,P＜0.05).结论 OSAHS患者尤其是中、重度患者脑血管运动反应性减弱,BKcaβ1mRNA表达降低,BKcaβ1导致了血管的功能失调.     

Ethylbromide tamoxifen, a membrane-impermeant antiestrogen, activates smooth muscle calcium-activated large-conductance potassium channels from the extracellular side

Smooth-muscle calcium-activated large-conductance potassium channels (BK channels) are activated by tamoxifen and 17-beta-estradiol. This increase in NP(o), the number of channels, N, multiplied by open probability, depends on the presence of the regulatory beta1-subunit. Furthermore, a previous study indicated that 17-beta-estradiol might bind an extracellular site on the beta1-subunit. Because tamoxifen and 17-beta-estradiol may share a common binding site, we hypothesized that tamoxifen activates BK channels through a site on the extracellular surface of the membrane. A membrane-impermeant analog of tamoxifen, ethylbromide tamoxifen, was synthesized and used to test this hypothesis in whole-cell, outside-out, cell-attached, and inside-out patches from canine colonic smooth muscle cells. Ethylbromide tamoxifen is positively charged and is therefore membrane-impermeant. In whole-cell experiments, ethylbromide tamoxifen increased K(+) current at potentials positive to +40 mV, which has previously been attributed to BK channels. Unlike tamoxifen, ethylbromide tamoxifen did not inhibit delayed rectifier current. In outside-out patches, ethylbromide tamoxifen increased BK channel NP(o) with an EC(50) value of 1 microM. Ethylbromide tamoxifen did not increase BK channel NP(o) in cell-attached or inside-out patches; however, subsequent addition of equimolar tamoxifen did. Both drugs diminished BK channel unitary conductance to a degree that paralleled the effect on NP(o), suggesting an additional interaction with the pore-forming alpha-subunit. An interaction of tamoxifen with the pore was supported by a right shift in the concentration-response curve for tetraethylammonium; similar results were evident with iberiotoxin and charybdotoxin block. Our data suggest that ethylbromide tamoxifen does not easily traverse the plasma membrane and that tamoxifen binding responsible for activation of BK channels is at an extracellular site. The tamoxifen binding site may be within the extracellular loop of the BK channel beta1-subunit or, alternatively, on an as-yet-unidentified mediator that has an extracellular binding site.     

Intermediate-conductance calcium-activated potassium channels participate in neurovascular coupling

BACKGROUND AND PURPOSE

Controlling vascular tone involves K⁺ efflux through endothelial cell small- and intermediate-conductance calcium-activated potassium channels (K_Ca2.3 and K_Ca3.1, respectively). We investigated the expression of these channels in astrocytes and the possibility that, by a similar mechanism, they might contribute to neurovascular coupling.

EXPERIMENTAL APPROACH

Transgenic mice expressing enhanced green fluorescent protein (eGFP) in astrocytes were used to assess K_Ca2.3 and K_Ca3.1 expression by immunohistochemistry and RT-PCR. K_Ca currents in eGFP-positive astrocytes were determined in situ using whole-cell patch clamp electrophysiology. The contribution of K_Ca3.1 to neurovascular coupling was investigated in pharmacological experiments using electrical field stimulation (EFS) to evoke parenchymal arteriole dilatation in FVB/NJ mouse brain slices and whisker stimulation to evoke changes in cerebral blood flow in vivo, measured by laser Doppler flowmetry.

KEY RESULTS

K_Ca3.1 immunoreactivity was restricted to astrocyte processes and endfeet and RT-PCR confirmed astrocytic K_Ca2.3 and K_Ca3.1 mRNA expression. With 200 nM [Ca²⁺]_i, the K_Ca2.1-2.3/K_Ca3.1 opener NS309 increased whole-cell currents. CyPPA, a K_Ca2.2/K_Ca2.3 opener, was without effect. With 1 µM [Ca²⁺]_i, the K_Ca3.1 inhibitor TRAM-34 reduced currents whereas apamin (K_Ca2.1-2.3 blocker) had no effect. CyPPA also inhibited currents evoked by NS309 in HEK293 cells expressing K_Ca3.1. EFS-evoked Fluo-4 fluorescence confirmed astrocyte endfoot recruitment into neurovascular coupling. TRAM-34 inhibited EFS-evoked arteriolar dilatation by 50% whereas charybdotoxin, a blocker of K_Ca3.1 and the large-conductance K_Ca channel, K_Ca1.1, inhibited dilatation by 82%. TRAM-34 reduced the cortical hyperaemic response to whisker stimulation by 40%.

CONCLUSION AND IMPLICATIONS

Astrocytes express functional K_Ca3.1 channels, and these contribute to neurovascular coupling.

LINKED ARTICLES

This article is part of a themed issue on Vascular Endothelium in Health and Disease. To view the other articles in this issue visit http://dx.doi.org/10.1111/bph.2011.164.issue-3     

Functional contribution of the endothelial component to the vasorelaxing effect of resveratrol and NS 1619, activators of the large-conductance calcium-activated potassium channels

Large-conductance calcium-activated potassium channels (BK) of smooth muscle play a role in the relevant modulation of vascular tone, due to their calcium- and voltage-dependent mechanisms of activation. A potential role of endothelial BK channels has also been suggested by approaches on endothelial cell cultures. However, no functional study, aimed at evaluating the contribution of endothelial BK channels to the effect of BK-openers, has been reported. Resveratrol and NS 1619, BK-openers, have been tested on endothelium-intact and -denuded aortic rings. Furthermore, the effects of high depolarisation of potassium channel blockers TEA (Tetraethylammonium), 4-AP ( 4-Aminopyridine) and IbTX (Iberiotoxin) and of inhibitors of NO-pathway (L-NAME and ODQ) have been evaluated. The presence of endothelium increased the vasorelaxing potency of BK-openers. This potentiation was eliminated by L-NAME and ODQ. TEA, 4-AP, IbTX and high depolarisation had modest or no antagonist influence on resveratrol in endothelium-denuded aortic rings. The effects of NS 1619 on endothelium-denuded aortic rings were not affected by IbTX, and were modestly antagonised by TEA, 4-AP and high depolarisation. In intact endothelium vessels, TEA, IbTX and 4-AP antagonised the vasorelaxing effect of the two BK-activators. A BK-mediated release of endothelial NO seems a very important factor, determining a strong influence on vasodilator profile of BK-openers. Therefore, an eventual therapy with a BK-opener could promote a series of cardiovascular impacts not confined to the only direct vasorelaxing effects, but also due to a significant contribution of endothelial NO.     

Modulators of small- and intermediate-conductance calcium-activated potassium channels and their therapeutic indications

Calcium-activated potassium channels modulate calcium signaling cascades and membrane potential in both excitable and non-excitable cells. In this article we will review the physiological properties, the structure activity relationships of the existing peptide and small molecule modulators and the therapeutic importance of the three small-conductance channels KCa2.1-KCa2.3 (a.k.a. SK1-SK3) and the intermediate-conductance channel KCa3.1 (a.k.a. IKCa1). The apamin-sensitive KCa2 channels contribute to the medium afterhyperpolarization and are crucial regulators of neuronal excitability. Based on behavioral studies with apamin and on observations made in several transgenic mouse models, KCa2 channels have been proposed as targets for the treatment of ataxia, epilepsy, memory disorders and possibly schizophrenia and Parkinson's disease. In contrast, KCa3.1 channels are found in lymphocytes, erythrocytes, fibroblasts, proliferating vascular smooth muscle cells, vascular endothelium and intestinal and airway epithelia and are therefore regarded as targets for various diseases involving these tissues. Since two classes of potent and selective small molecule KCa3.1 blocker, triarylmethanes and cyclohexadienes, have been identified, several of these postulates have already been validated in animal models. The triarylmethane ICA-17043 is currently in phase III clinical trials for sickle cell anemia while another triarylmethane, TRAM-34, has been shown to prevent vascular restenosis in rats and experimental autoimmune encephalomyelitis in mice. Experiments showing that a cyclohexadiene KCa3.1 blocker reduces infarct volume in a rat subdural hematoma model further suggest KCa3.1 as a target for the treatment of traumatic and possibly ischemic brain injury. Taken together KCa2 and KCa3.1 channels constitute attractive new targets for several diseases that currently have no effective therapies.     

阿托伐他汀对自发性高血压大鼠肠系膜动脉平滑肌细胞大电导钙激活钾通道的影响

目的:探讨阿托伐他汀对自发性高血压大鼠(SHR)动脉血压及肠系膜动脉平滑肌细胞大电导钙激活钾通道(large-conductance calcium-activated potassium channel,BKCa,MaxiK)的影响。方法:选取雄性9周龄SHR12只,随机分为阿托伐他汀组(AVT组)、蒸馏水组(DW组),每组6只,以Wistar-Kyoto大鼠为对照组(WKY组)(n=6)。AVT组以AVT加适量蒸馏水灌胃10周(50 mg.kg-1.d-1)。观察给药前后大鼠腹主动脉血压的变化,测定大鼠血清TC、TG及高密度脂蛋白胆固醇(HDL-C)含量。利用膜片钳全细胞模式记录肠系膜动脉平滑肌细胞钾电流、用四乙胺阻断大电导钙激活钾通道后的电流、膜电容,以计算BKCa电流值、电流密度。结果:AVT组腹主动脉血压明显低于DW组[(171±8)mmHgvs(190±10)mmHg,P<0.01](1 mmHg=0.133 kPa);与DW组相比,AVT组肠系膜动脉平滑肌细胞BKCa电容减低[(23.8±2.6)pFvs(30.0±2.5)pF,P<0.01]而电流密度则显著高于DW组[(13.2±1.2)pA...     

Vascular large conductance calcium-activated potassium channels: Functional role and therapeutic potential

Large-conductance Ca²⁺-activated K⁺ channels (BK_Ca or maxiK channels) are expressed in different cell types. They play an essential role in the regulation of various cell functions. In particular, BK_Ca channels have been extensively studied in vascular smooth muscle cells, where they contribute to the control of vascular tone. They facilitate the feedback regulation against the rise of intracellular Ca²⁺, membrane depolarization and vasoconstriction. BK_Ca channels promote a K⁺ outward current and lead to membrane hyperpolarization. In endothelial cells expression and function of BK_Ca channels play an important role in the regulation of the vascular smooth muscle activity. Endothelial BK_Ca channels modulate the biosyntheses and release of various vasoactive modulators and regulate the membrane potential. Because of their regulatory role in vascular tone, endothelial BK_Ca channels have been suggested as therapeutic targets for the treatment of cardiovascular diseases. Hypertension, atherosclerosis, and diabetes are associated with altered current amplitude, open probability, and Ca²⁺-sensing of BK_Ca channels. The properties of BK_Ca channels and their role in endothelial and vascular smooth muscle cells would address them as potential therapeutic targets. Further studies are necessary to identify the detailed molecular mechanisms of action and to investigate selective BK_Ca channels openers as possible therapeutic agents for clinical use.     

Role of calcium-activated potassium channels in impaired acetylcholine vasodilatory responses in diabetic rats

Muscarinic agonists produce endothelium-dependent vasodilatation in the presence of nitric oxide synthase (NOS) inhibition. The importance of this mechanism was assessed in the methoxamine-preconstricted perfused mesenteric vascular bed (MVB) of streptozotocin diabetic Sprague-Dawley rats. At 9 weeks of age, male rats were treated with streptozotocin (55 mg/kg in citrate buffer) or with citrate buffer alone. The superior mesenteric artery was cannulated and the MVB was detached from its intestinal borders. Concentration-response curves to acetylcholine were determined in the presence and in the absence of indomethacin, tetrabutylammonium (a calcium-activated potassium channel blocker), high extracellular potassium, or NOS inhibition with Nomega-nitro-l-arginine and l-NG-nitro-l-arginine. There was a rightward shift in the concentration-response curve with an increase in median inhibitory concentration (p < 0.05) and a reduction in acetylcholine IMAX (p < 0.05) values in 14-week streptozotocin rats. The ability of NOS inhibition to attenuate vasodilatation was reduced in the 14-week streptozotocin group relative to the 2-week streptozotocin treatment group (p < 0.05). However, the ability of tetrabutylammonium to block acetylcholine-mediated vasodilatation remained consistent in streptozotocin rats at both stages. The results demonstrate that an alternate pathway involving calcium-activated potassium channels may compensate for diminished nitric oxide bioactivity. This effect is contingent on the duration of diabetes. This study provides insight into the development and progression of altered diabetic vascular responses.     

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

The effects of aminopyridine analogs on Ca2+-activated K+ channels in GH3 clonal anterior pituitary cells were studied using whole-cell voltage-clamp and single-channel recording techniques. Step depolarization from a holding potential of -50 mV activated a noninactivating, tetraethylammonium- and Cd2+-sensitive outward current. Tail current analysis indicated that this sustained outward current is carried predominantly by K+ ions. Extracellular perfusion with 4-aminopyridine and 3,4-diaminopyridine (0.05-5 mM) caused a dose-dependent enhancement of the outward current by up to 100 and 170%, respectively. This effect typically occurred with prolonged depolarizations of greater than 1-2 sec. Patch-clamp recordings in the cell-attached configuration demonstrated that 4-aminopyridine (2 mM) promotes the activity of a large-conductance (150-175 pS; 50-135 mM external K+), tetraethylammonium-sensitive, Ca2+-activated K+ channel; the drug had no effect on these channels in excised patches. These results indicate that aminopyridines enhance the opening of Ca2+-activated K+ channels in GH3 cells. Several lines of evidence suggest that this effect may occur indirectly, possibly as a result of an increase in the effective intracellular free Ca2+ level.     

Effects of calmodulin antagonists on calcium-activated potassium channels in pregnant rat myometrium.

1. The effects of W-7, trifluoperazine, and W-5 on Ca2(+)-activated K(+)-channels were investigated with the inside-out patch-clamp method in smooth muscle cells freshly dispersed from pregnant rat myometrium. These drugs are known to have different potencies as calmodulin antagonists. 2. In the presence of 1 microM Ca2+ on the cytoplasmic side ([Ca2+]i), the fraction of time the channel was open (open probability, Po) was about 0.9 and the calmodulin antagonists (1-30 microM) applied to the cytoplasmic face reduced Po to 0.65-0.55 dose-dependently. In the presence of 0.1-0.16 microM Ca2+, when Po was very low (0.02), calmodulin antagonists increased Po. All antagonists used produced almost identical effects at the same concentration. 3. The probability density function of the open time distribution could be described by the sum of two exponentials. W-7 decreased the time constant of slow component of distribution and at 30 microM the slow component disappeared both at 1 and 0.25 microM [Ca2+]i, reflecting the appearance of flickering channel activity. The probability density function of the closed time distribution could be fitted with three exponentials. The time constants of these components were not significantly altered by W-7. 4. Internally applied calmodulin (1-5 microM) did not produce any significant effect on channel activity. 5. The effects of calmodulin antagonists are considered to be due to a direct action of these compounds on the channel, and suggest that channel activation by Ca2+ is not mediated by calmodulin.     

Relative importance of calcium-activated potassium channels in nipradilol-induced aortic relaxation in rats.

Nipradilol (CAS 81486-22-8), a vasodilatory beta-blocker, has been shown to dilate smaller vessels than nitroglycerin does, and the vasodilative effects of nipradilol have been reported to be less mediated by cyclic GMP (guanosine monophosphate) than those of nitroglycerin. To test the hypothesis that cyclic GMP-independent potassium channels have a larger role in nipradilol-induced aortic relaxation than cyclic GMP-dependent mechanisms, the effects of a potassium channel blocker, tetraethylammonium (TEA, CAS 56-34-8), and of a guanylate cyclase inhibitor, methylene blue (MB, CAS 61-73-4), on nipradilol-induced aortic relaxation were investigated and compared with those on nitroglycerin-induced aortic relaxation in isolated rat aortic rings. Relaxation response was expressed as percent relaxation, which is a percentage of the tension developed by 10(-7) mol/l norepinephrine. Nitroglycerin and nipradilol similarly relaxed the aortic ring in a concentration-dependent manner (10(-9)-10(-4) mol/l). In contrast, desnitronipradilol, a nipradilol analogue which has no nitroxy group, induced almost no aortic relaxation. TEA at 10(-3) mol/l, which is selective for calcium-activated potassium channels, inhibited the aortic relaxation induced by nipradilol (10(-5) mol/l) to a significantly greater extent than that induced by nitroglycerin (10(-5) mol/l) (% relaxation: 30.0 +/- 6.8 vs. 51.1 +/- 6.1%, p < 0.05). MB (10(-5) mol/l) suppressed the relaxation by nitroglycerin slightly but not significantly more than that by nipradilol. (% relaxation: 54.7 +/- 9.9 vs. 64.6 +/- 5.7%). The combination of TEA and MB almost completely eliminated the relaxation induced by nipradilol as well as by nitroglycerin. Thus, cyclic GMP-independent calcium activated potassium channels may be more involved in the aortic relaxation by nipradilol than that by nitroglycerin in rats.     

Role of calcium-activated potassium channels in acetylcholine-induced vasodilation of rat retinal arterioles in vivo

The vascular endothelium plays an important role in regulating retinal blood flow via actions of several vasodilators, including nitric oxide (NO), prostaglandin I?, and an endothelium-derived hyperpolarizing factor (EDHF). Our previous in vivo studies demonstrated that acetylcholine (ACh) dilates the rat retinal arteriole partly through NO- and prostaglandin-independent pathway, possibly the EDHF-mediated pathway, but the underlying mechanism(s) remains to be elucidated. It has been suggested that activation of Ca2+-activated K+ (K(Ca)) channels contributes to the EDHF-mediated responses; therefore, the roles of K(Ca) channels in ACh-induced vasodilation of retinal arterioles were examined in rats. The retinal vascular responses were assessed by determining changes in diameters of retinal arterioles in ocular fundus images that were captured with an original fundus camera system. Intravitreal injection of charybdotoxin, an inhibitor of intermediate- and large-conductance K(Ca) (I/BK(Ca)) channels, or iberiotoxin, an inhibitor of large-conductance K(Ca) (BK(Ca)) channels, significantly reduced the ACh-induced vasodilation of retinal arterioles, whereas neither apamin, an inhibitor of small-conductance K(Ca) (SK(Ca)) channels, nor TRAM-34, an inhibitor of intermediate-conductance K(Ca) (IK(Ca)) channels, altered the response. The vasodilator response to ACh observed under the combined blockade of NO synthase and cyclooxygenase with N(G)-nitro-L-arginine methyl ester plus indomethacin was also diminished by iberiotoxin. Iberiotoxin did not affect the NO donor NOR3-induced vasodilation of retinal arterioles, whereas it significantly reduced the BK(Ca) channel opener BMS-191011-induced responses. These results suggest that activation of BK(Ca) channels is involved in the EDHF-mediated component of the vasodilator response to ACh in the rat retinal arterioles in vivo.     

Mechanisms underlying the cardiac pacemaker: the role of SK4 calcium-activated potassium channels

The proper expression and function of the cardiac pacemaker is a critical feature of heart physiology. The sinoatrial node (SAN) in human right atrium generates an electrical stimulation approximately 70 times per minute, which propagates from a conductive network to the myocardium leading to chamber contractions during the systoles. Although the SAN and other nodal conductive structures were identified more than a century ago, the mechanisms involved in the generation of cardiac automaticity remain highly debated. In this short review, we survey the current data related to the development of the human cardiac conduction system and the various mechanisms that have been proposed to underlie the pacemaker activity. We also present the human embryonic stem cell-derived cardiomyocyte system, which is used as a model for studying the pacemaker. Finally, we describe our latest characterization of the previously unrecognized role of the SK4 Ca²⁺-activated K⁺ channel conductance in pacemaker cells. By exquisitely balancing the inward currents during the diastolic depolarization, the SK4 channels appear to play a crucial role in human cardiac automaticity.     

钩藤碱对大鼠肺动脉平滑肌细胞钙激活钾通道的影响

用膜片钳单通道记录法研究钩藤碱(Rhy)对大鼠肺动脉平滑肌细胞钙激活钾通道(K_Ca)的影响.结果：Rhy 30, 45和60 μmol·L^-1缩短通道的开放时间, 但浓度依赖性增加K_Ca开放概率,Rhy 15, 30, 45和60 μmol·L^-1使开放概率由加药前的0.085±0.005分别增加到0.176±0.011, 0.315±0.009, 0.485±0.016和0.761±0.012(x±s, 均P<0.01).说明Rhy有增加肺动脉平滑肌细胞K_Ca开放作用.