瑞芬太尼对人肠系膜小动脉平滑肌细胞大电导钙激活钾通道的影响

目的 观察瑞芬太尼对人肠系膜小动脉平滑肌细胞大电导钙激活钾通道(BKCa)的影响,探讨其扩张血管的机制.方法 酶消化法急性分离人肠系膜小动脉平滑肌细胞,采用全细胞膜片钳技术,+80 mV钳制电压下记录不同浓度瑞芬太尼(1.2、4.8、19.4、77.4和310.0 nmol/L)给药后人肠系膜小动脉平滑肌细胞BKCa电流及达峰时间,计算BKCa激活率.结果 瑞芬太尼可激活BKCa,使电流密度-电压曲线上移,激活电压不变;随瑞芬太尼浓度升高,BKCa激活率逐渐升高(P＜0.01),19.4 nmol/L时BKCa激活率趋于稳定;各浓度瑞芬太尼给药后BKCa电流达峰时间比较差异无统计学意义(P＞0.05);瑞芬太尼浓度与BKCa激活率成对数曲线关系,其半数最大激活效应浓度为(118±7)nmol/L.结论 瑞芬太尼浓度依赖性地激活人肠系膜小动脉平滑肌细胞BKCa,该作用可能是其产生扩张血管作用的机制.     

蛋白激酶C和线粒体ATP敏感性钾通道在未成熟心肌预处理中的作用

目的探讨蛋白激酶C(PKC)和线粒体三磷酸腺苷敏感性钾通道(mitoKATP)在未成熟心肌预处理保护中的作用。方法采用Langendorff离体心脏灌注模型,30只新生日本长耳大白兔分为5组:缺血/再灌注组(I/R组),心脏缺血预处理组(E1组),蛋白激酶C(PKC)阻滞剂chelerythrine(CLT) 心脏缺血预处理(E2组),mitoKATP阻滞剂5-hydroxydecanoate(5-HD) 心脏缺血预处理(E3组),mitoKATP通道开放剂Diazoxide(Diaz)预处理组(E4组)。以血流动力学、生化指标、心肌超微结构等作为观察指标。结果E1和E4组心功能恢复、心肌含水量优于I/R、E2和E4组(P<0.05),三磷酸腺苷含量、超氧化物歧化酶活性、心肌线粒体Ca2 -ATPase活性、心肌线粒体合成三磷酸腺苷(ATP)的能力优于I/R、E2和E4组(P<0.01),丙二醛含量、血清肌酸激酶和乳酸脱氢酶漏出率、心肌细胞内Ca2 含量、心肌线粒体Ca2 含量低于I/R、E2和E4组(P< 0.01),心肌超微结构损伤较I/R、E2和E4组明显减轻。结论心肌缺血预处理对未成熟心肌具有明显的保护作用,其机制可能是通过PKC的激活和mitoKATP通道的开放起作用。     

蛋白激酶C在未成熟心肌预处理中的作用

目的为进一步加强未成熟心肌保护和临床应用的可行性,探讨蛋白激酶C(PKC)在未成熟心肌预处理保护中的作用。方法建立兔Langendorff灌注模型,将24只幼兔随机分为4组:缺血再灌注组(I/R组)、心脏缺血预处理组(MIP组)、PKC阻滞组(CLT组)和PKC激活剂组(PKC组),观察4组幼兔血流动力学、生化、心肌超微结构等指标。结果 MIP组和PKC组心功能恢复、心肌含水量优于I/R组和CLT组(P0.01),三磷酸腺苷(ATP)含量、超氧化物歧化酶活性、心肌线粒体Ca2+-ATPase活性、心肌线粒体合成ATP的能力优于I/R组和CLT组(P0.01),丙二醛含量、血清肌酸激酶和乳酸脱氢酶漏出率、心肌细胞内Ca2+含量、心肌线粒体Ca2+含量低于I/R组和CLT组(P0.01),心肌超微结构损伤较I/R组和CLT组明显减轻。结论心肌缺血预处理对未成熟心肌具有明显的保护作用,其机制可能是通过PKC的激活起作用。     

丙泊酚对Ca2+诱导大鼠离体心肌线粒体损伤的影响

目的 探讨丙泊酚对Ca2+诱导大鼠离体心肌线粒体损伤的影响.方法 Wistar大鼠35只,断头处死迅速取出心脏制备线粒体悬液后,随机均分为五组:空白对照组(C组);CaCl2组(Ca2组),加入CaCl2100 nmol/mg prot;P25组,加入丙泊酚25 μp.mol/L)P50组,加入丙泊酚50 μmol/L;P100组,加入丙泊酚100 μmol/L.P25、P50、P100组也加入CaCl2 100 nmol/mg prot.20 min后采用Western blot法测定线粒体释放细胞色素C(Cyt C)和凋亡诱导因子(AIF)的水平.结果 与C组比较,其他四组Cyt C和AIF释放增加(P<0.01);与Ca2+组比较,P25组、P50组、P100组Cyt C、AIF释放减少(P<0.01);P50组、P100组又较P25组Cyt-C释放减少(P<0.01).结论 丙泊酚25、50、100μmol/L均可减轻Ca2+诱导大鼠离体心肌线粒体损伤,从而产生心肌保护作用.     

七氟烷后处理的心肌保护作用及其机制

背景 大量实验证据表明缺血后处理和药物后处理对心肌再灌注损伤具有确切的保护作用.七氟烷是一种新型的、理想的吸入性麻醉药,被广泛应用于全身麻醉.实验证明七氟烷后处理可以保护心肌对抗缺血/再灌注损伤(ischemia/reperfusion injury,I/RI).目的 通过对近年研究进展的总结对七氟烷后处理的心肌保护作用及机制予以阐述.内容七氟烷后处理可以减少再灌注心肌的梗死面积、线粒体损害和再灌注室性心律失常的发生,改善心脏的血流动力学.七氟烷后处理心肌保护作用复杂且涉及多个方面,如阻断线粒体通透性转运孔(mitochondrial permeability transition pore,mPTP)、激活线粒体ATP敏感性K+通道(mitochondrial KATP-channel,mKATP),激活细胞外信号调节激酶1/2(extracellular signal-regulated kinase 1/2,ERK1/2)以及磷酯酰肌醇-3激酶-丝氨酸/苏氨酸激酶(phosphatidylin ositol-3-kinase-serine/threonine,PI3K-Akt)信号通道等. 趋势 未来的研究除进一步探究七氟烷后处理的心肌保护机制,同时应加强七氟烷后处理的临床应用,为实际工作提供可靠依据.     

异丙酚预先给药对外源性Ca2+致大鼠离体脑线粒体损伤的影响

线粒体不仅合成ATP供给细胞能量,而且在细胞氧化还原状态调节、Ca2+稳态及细胞信号转导方面起重要作用.器官组织缺血再灌注可导致细胞钙超载,为维持Ca2+稳态,线粒体从细胞浆摄取大量Ca2+,使线粒体积聚超量的Ca2+,诱发线粒体膜通透性转换孔开放而发生线粒体通透性转换(MPT)[1],导致线粒体功能障碍,引起神经元的坏死或凋亡.异丙酚对脑组织缺血再灌注时具有脑保护作用[2],其机制是否与线粒体有直接关系有待进一步探讨.     

线粒体ATP敏感性钾通道在利多卡因预先给药减轻大鼠离体心脏缺血再灌注损伤中的作用

目的 评价线粒体ATP敏感性钾通道在利多卡因预先给药减轻大鼠离体心脏缺血再灌注损伤中的作用.方法 雌性成年Wistar大鼠,体重220～250 g,采用Langendorff装置建立大鼠离体心脏缺血再灌注模型.取模型制备成功的大鼠心脏24个,随机分为3组(n=8):缺血再灌注组(IR组)、利多卡因组(L组)和利多卡因+格列苯脲组(LG组).K-H液平衡灌注10 min后,C组、L组和LG组分别灌注K-H液、含2.5 mg/L利多卡因的K-H液、含2.5 mg/L利多卡因+10μmol/L格列苯脲(线粒体ATP敏感性钾通道阻断剂)的K-H液20 min,然后缺血30 min,再灌注60 min.分别于平衡灌注末(T0)、再灌注15 min(T1)、30 min(T2)、45 min(T3)和60 min(T4)时,记录HR、左心室发展压(LVDP)、左心室内压最大上升速率(+dp/dtmax)和左心室内压最大下降速率(-dp/dtmax).于T0和T4时,收集冠状动脉流出液,测定乳酸脱氢酶(LDH)和肌酸激酶(CK)的活性.于T4时取心尖周围心肌组织,测定Na+-K+-ATP酶和SOD的活性、MDA和Ca2+的含量.结果 与IR组比较,L组HR、LVDP、+dp/dtmax和-dp/dtmax升高,CK和LDH的活性降低,心肌Na+-K+-ATP酶和SOD的活性升高,Ca2+和MDA的含量降低(P＜0.05),LG组上述各指标差异无统计学意义(P＞0.05).与L组比较,LG组HR、LVDP、+dp/dtmax和-dp/dmax.降低,CK和LDH的活性升高,心肌Na+-K+-ATP酶和SOD的活性降低,Ca2+和MDA的含量升高(P＜0.05).结论 利多卡因预先给药减轻大鼠离体心脏缺血再灌注损伤与促进线粒体ATP敏感性钾通道的开放有关.     

BKCa和PKG在氯胺酮舒张哮喘大鼠离体气管平滑肌中的作用

目的 评价大电导钙激活钾通道(BKCa)和蛋白激酶G(PKG)在氯胺酮舒张哮喘大鼠离体气管平滑肌中的作用.方法 健康SD大鼠,体重250 ～ 300 g,采用卵蛋白致敏法建立哮喘模型,取哮喘大鼠15只,每只大鼠制备2～3条离体气管环.取哮喘大鼠离体气管环36条,采用随机数字表法,将其分为3组(n=12):氯胺酮处理组(AK组)、BKCa阻断剂IBTX+氯胺酮处理组(AKI组)和PKG抑制剂KT-58232+氯胺酮处理组(AKK组);AK组采用0.1 mmol/L乙酰胆碱预收缩大鼠气管环达稳态后,用0.4 g/L氯胺酮孵育15 min; AKI组用乙酰胆碱和氯胺酮孵育前,用3 μmol/L IBTX孵育30 min;AKK组用入乙酰胆碱和氯胺酮孵育前,用2μmol/L KT-5823孵育30 min;采用气管环相连的力-位移换能器测定气管环舒张幅度.结果 与AK组比较,AKI组和AKK组大鼠离体气管平滑肌舒张幅度降低(P＜0.05).结论 氯胺酮可通过激活BKCa和PKG信号通路舒张哮喘大鼠离体气管平滑肌.     

热休克蛋白70表达对未成熟心肌和心肌间质的保护作用

目的 观察热休克蛋白70(HSP70)对未成熟心肌和心肌间质的保护作用.方法 健康新生长耳大白兔30只随机分为5组.对照组:腹腔注射生理盐水0.4 ml 24 h后取离体心脏,建立Langendorff离体心脏灌注模型;E4h、E12h、E24h、E48h组,腹腔注射去甲肾上腺素,4、12、24、48 h后分别取离体心脏,方法同对照组.测定心肌细胞中HSPT0含量、血流动力学指标、心肌含水量(MWC)、心肌肌酸激酶(CK)和乳酸脱氢酶(LDH)漏出率、三磷酸腺苷(ATP)含量、超氧化物歧化酶(SOD)和丙二醛(MDA)含量、心肌组织羟脯氨酸(HP)含量、内皮素(ET)含量、心肌细胞内Ca2+含量、心肌线粒体Ca2+-ATPase活性及其Ca2+含量、心肌线粒体合成ATP能力[ATP]m,心肌超微结构.结果 E24h组与其他各组比较,HSPT0含量明显增高(P＜0.01),MWC(72.48±1.36)低于其他各组(P＜0.05),ATP含量(11.64±1.87)、SOD活性(235.83±12.30)、心肌线粒体Ca2+-AT-Pase活性(18.46±1.95)、[ATP]m(106.26±9.42),HP含量(6.45±1.53)优于其他各组(P＜0.01),MDA含量(1.17±0.12)、CK(57.38±4.75),LDH漏出率(37.28±3.26)、心肌细胞内Ca2+含量(2.54±0.34)、心肌线粒体Ca2+含量(38.37±3.61)、ET含量(76.84±10.37)低于其他各组(P＜0.01),心肌超微结构损伤较其他各组明显减轻.结论 腹腔注射去甲肾上腺素24 h后可诱导未成熟心肌HSP70高表达,一定量的HSP70表达可明显减轻未成熟心肌和心肌间质的缺血再灌注损伤.     

线粒体KATP通道在脑啡肽预处理体外循环心肌保护中的作用

目的 通过建立猪体外循环(CPB)心肌缺血再灌注(MI/R)模型对δ阿片受体介导的CPBMI/R损伤早期时相的心肌保护效果、作用机制以及线粒体KATP通道的角色进行探讨,以期为临床心肌保护提供理论和实验依据.方法 24只成年家猪(30～35kg)随机分为对照组(C组)、δ阿片样受体激动剂脑啡肽组(D组)和脑啡肽+线粒体KATP通道阻滞剂组(D+K组),每组8只,常规建立CPB模型,主动脉阻断同时灌注改良St.Thomas停搏液.每组于CPB前、主动脉开放时、CPB结束时、停机后1、2h检测冠状静脉窦血肌钙蛋白(TnT)含量,相应时间点监测左室收缩压(LVSP),左室舒张末期压(LVEDP)和左室内压最大变化速率(±dp/dtmax),分别于CPB前和停机后2h取左心室心肌,作心肌组织Gia蛋白的表达、PKC的表达和ATP含量的测定.并应用透射电镜观察心肌再灌注损伤超微结构的变化.结果 (1)D组的心功能改变明显优于C和D+K组.(2)C和D+K组TnT的升高比D组明显.(3)D组心肌ATP含量较C和D+K组高.(4)透射电镜下C和D+K组呈现出明显的MI/R损伤表现,而D组的心肌超微结构损伤较轻.(5)和C和D+K组以及正常心肌相比,D组Gia蛋白和PKC的表达更为明显.结论 (1)CPB前1h应用一次δ阿片受体激动剂脑啡肽(DADLE)能产生明显的早期时相心肌保护作用.(2)Gi蛋白-PKC-线粒体KATP通道途径是δ阿片受体介导的猪CPB MI/R损伤早期时相心肌保护中一条重要的信号传导途径.     

Muscarinic M2 receptors inhibit Ca2+-activated K+ channels in rat bladder smooth muscle.

PURPOSE: To clarify the functional relationship between M2 muscarinic receptor and Ca2+-activated K+ channel, we investigated the effect of carbachol (CCh) on the membrane current of rat bladder smooth muscle cells. METHODS: Rat bladder single smooth muscle cells were patch clamped with whole-cell configuration. RESULTS: CCh (10 micro mol/L) transiently induced an outward current in the presence of K+ in the pipette solution. A high Ca2+ concentration in the pipette solution persistently induced an outward current, which was inhibited by CCh. In the presence of M2 inhibitor, AFDX-384, CCh induced the outward current persistently, indicating that M2 was involved in the current inhibition. In pertussis toxin pretreated cells, CCh did not apparently inhibit the outward current. The CCh-induced outward current was inhibited by iberiotoxin, a selective inhibitor of large-conductance Ca2+-activated K+ channels (BKCa). CONCLUSION: CCh induces BKCa, which is inhibited by M2- and Gi-mediated signal transduction pathway. This M2-mediated pathway may enhance contraction which is initiated by M3-stimulation in rat bladder smooth muscle.     

Modulation of Ca2+-activated K+ channel in renal artery endothelium in situ by nitric oxide and reactive oxygen species

BACKGROUND: Endothelium-derived nitric oxide and reactive oxygen species (ROS) have been proposed to regulate vascular tone by complex mechanisms, including the modulation of ion channel function. In endothelial function itself, activation of Ca2+-activated K+ channels (KCa) plays a crucial role by inducing hyperpolarization, which promotes membrane potential-driven Ca2+ influx and Ca2+-dependent synthesis of vasodilatory factors. In the present study, we tested whether nitric oxide and ROS modulate endothelial KCa function. METHODS: By employing the patch-clamp technique in endothelium of porcine renal arteries in situ, we identified a large-conductance Ca2+-activated K+ channel (big K+ channel, BKCa) with a conductance of 297 +/- 6 pS. RESULTS: Channel activity was strongly controlled by the membrane potential and the cytosolic Ca2+ concentration (EC50 3.1 +/- 0.5 micromol/L Ca2+ at 0 mV). Channel activity was inhibited by Ba2+ and iberiotoxin. At submicromolar [Ca2+]i, nitric oxide induced a dose-dependent stimulation of BKCa activity with a 10-fold increase at the highest dose tested (1 micromol/L). A similar stimulation was achieved by the nitric oxide donors, sodium nitroprusside (SNP), and diethylamine nitric oxide complex (DEA-NO). In contrast, ROS and, in particular, hydrogen peroxide (H2O2) led to dose-dependent inactivation of BKCa with an IC50 of 80 +/- 6 nmol/L and 1.1 +/- 0.4 micromol/L, respectively. In isolated porcine renal arteries, bradykinin-induced vasodilation was significantly reduced by either iberiotoxin or H2O2. CONCLUSION: Direct stimulation of endothelial BKCa by nitric oxide might represent a novel mechanism of autocrine regulation of endothelial function and points to a positive feedback mechanism by promoting hyperpolarization and nitric oxide production itself. The ROS-induced inhibition of BKCa could be part of the cellular mechanisms by which ROS impairs endothelium-dependent vasodilation.     

Synergism between prostanoids and other vasoactive agents

BACKGROUND: Prostaglandin E2 is usually considered to be a vasodilator, but some vascular beds respond with weak vasoconstriction mediated by prostanoid EP3-receptors. We have used the guinea pig isolated thoracic aorta to examine the nature of the synergism between the EP3 agonist sulprostone and other vasoactive agents. METHODS: Muscle tension was recorded from endothelium-denuded rings of aorta suspended in conventional organ baths. Indomethacin and the TP-receptor antagonist GR 32191 were usually present. RESULTS: Sulprostone (0.1-300 nM) showed two profiles: low-responder preparations (maximum response = 15-35% of 100 nM U-46619 response) were insensitive to the L-type Ca2+ blocker nifedipine, whereas high-responders (maximum = 35-70%) showed a nifedipine-sensitive component at higher sulprostone concentrations only. Charybdotoxin (CTX), a blocker of large-conductance Ca2+-activated K+ channels (BKCa), slightly enhanced threshold sulprostone responses and markedly enhanced larger responses; the enhancements were abolished by nifedipine. In contrast, threshold sulprostone responses were dramatically enhanced in the presence of established small contractions to phenylephrine (alpha1-adrenoceptor agonist), U-46619 (TP agonist), cyclopiazonic acid (sarcoplasmic Ca2+ pump inhibitor), and 4-aminopyridine (4-AP, Kv channel blocker). Nifedipine had no effect on enhancements of threshold sulprostone responses, and partially inhibited the enhancements of larger responses. CONCLUSIONS: BKCa channel activation appears to increase progressively as sulprostone-induced contraction increases. CTX removes this "BKCa brake," thereby providing an "L-type channel" Ca2+ flux to prime the EP3-receptor-driven Ca2+-sensitization mechanism (via Rhokinase activation, unpublished observations). In contrast, the other agents, including 4-AP, direct a non-L-type channel source of Ca2+ to the calmodulin-myosin light chain arm.     

Potassium channel openers as potential therapeutic weapons in ion channel disease

The opening of potassium (K+) channels, causing hyperpolarization of the cell membrane, is a physiological means of decreasing cell excitability. Thus, drugs with this property will demonstrate a broad clinical potential. The identification of synthetic molecules that evoke physiological responses (for example smooth muscle relaxation) by the opening of K+ channels led to a new direction in the pharmacology of ion channels. The term "potassium channel openers" was initially associated with a group of chemically diverse agents (for example, cromakalim, pinacidil, nicorandil) that evoke K+ efflux through adenosine 5'-triphosphate (ATP)-sensitive K+ channels (KATP). This finding initiated a search to identify molecules that specifically open other K+ channel subtypes (for example large conductance calcium-activated K+ channels [BKCa]). K+ channel opening properties have been demonstrated in a diverse range of synthetic chemical structures and endogenous substances. Second generation KATP channel openers (KATPCOs) demonstrate heterogeneous pharmacology indicative of independent sites of action for the different agents. Successful cloning of the KATP channel has shed light on the heterogeneity of the structure targeted by KATPCOs. Expression of the actions of KATPCOs involves three isoforms of the sulfonylurea (SUR) receptor (which forms the beta subunit of the KATP channel). The distribution of the SUR isoforms (and potential of identifying new isoforms) provides unique targets for the development of selective KATPCOs giving focused therapeutic approaches to clinical conditions for example cardiac ischemia, urinary incontinence, neurodegeneration, obesity and autoimmune diseases. BKCa channels are found in a diverse array of tissues and due to voltage and Ca sensitivity may work as a negative feedback process. A variety of small synthetic molecules (for example, NS004, fenamates) and natural product-derived compounds (DHS-I, maxikdiol) have been identified as selective BKCa channel openers which should have a profound impact in controlling diseases. The discovery of numerous variants of the alpha subunit (ion conductance pore) and beta subunit (contributes biophysical and pharmacological properties) complex of the BKCa channel gives potential to target specific tissues with selective openers. Little is known, however, about the site(s) of interaction of openers of these channels. The discovery of K+ channel subtype-specific openers and their evaluation in different diseases will determine the degree to which these channels (KATP, BKCa), or their isoforms, represent realistic therapeutic targets. Drugs already marketed that open K+ channels were discovered empirically, and most have serious safety and efficacy problems. New scientific methods, utilizing molecular insight, are implicating K+ channel dysfunction in numerous disease states and are identifying new targets for the future generation of K+ channel opening drugs.     

A description of Ca2+ channels in human detrusor smooth muscle

OBJECTIVE: To characterize the Ca2+ channels in human detrusor smooth muscle and to investigate their contribution to spontaneous electrical activity. MATERIALS AND METHODS: Isolated human detrusor smooth muscle myocytes were used to measure ionic currents under voltage-clamp or membrane potential under current-clamp. Membrane potential oscillations were analysed in terms of oscillation frequency and amplitude using fast Fourier transforms. RESULTS: Under voltage-clamp an inward current dependent on extracellular Ca2+ was recorded using Cs+-filled patch electrodes. The current could be separated into two components on the basis of their sensitivity to Ni2+, verapamil or nicardipine, and their dependence on holding and clamp potential. A Ni2+-sensitive component activated over a relatively negative range of potentials (-60 to -20 mV) comprised about a third of the total current and was designated a T-type Ca2+ current. A verapamil/nicardipine-sensitive component, activated at more positive potentials, was designated an l-type Ca2+ current. Using K+-based filling solutions spontaneous transient outward currents were recorded that had the characteristics of current flow through BK channels. Membrane potential oscillations, under current-clamp increased in frequency but not amplitude as the mean membrane potential was made less negative. The voltage-dependence of oscillation frequency was similar to that of the l-type, but not T-type, Ca2+ current activation curve. Furthermore oscillation frequency was slowed by verapamil but not Ni2+. CONCLUSION: The study showed, for the first time, the presence of both T- and L-type Ca2+ channels in human detrusor smooth muscle; we propose a role for these channels in spontaneous activity. The results suggest that the L-type Ca2+ current can control membrane potential oscillation frequency. The significance of this finding for spontaneous contractions is discussed.     

Adrenergic regulation of the intracellular [Ca2+] and voltage-operated Ca2+ channel currents in the rat prostate neuroendocrine cells

BACKGROUND: The prostate gland contains numerous neuroendocrine cells (PNECs) innervated by adrenergic neurons. PNECs are believed to influence the growth and physiological function of the prostate gland via paracrine release of hormones. MATERIALS AND METHODS: Using fura-2 fluorescence measurement and patch-clamp techniques, we investigated the effects of adrenergic stimulation on cytosolic concentration of Ca2+ ([Ca2+]c) and high voltage-activated Ca2+ channel currents (HVA-I(Ca)) of the putative rat prostate neuroendocrine cells (RPNECs) freshly isolated by an enzymic digestion. RESULTS: Noradrenaline (NA, 1 microM) induced a sharp, transient increase of [Ca2+]c measured by the fura-2 fluorescence. Pharmacological studies showed that alpha1-adrenoceptors (alpha1-ARs) coupled with PLC/IP3 signaling pathway induce the release of stored Ca2+, which subsequently recruits store-operated Ca2+ entry pathways. In the whole-cell voltage clamp experiment, NA decreased the amplitude of HVA-I(Ca) by 40%, which was mimicked by an alpha2-AR agonist (UK14304) but not by an alpha1-AR agonist (phenyleprine). After selective blockade of N-type Ca2+ channels by omega-conotoxin GVIA, the addition of NA showed no further inhibition on the remaining L-type Ca2+ channel currents. The adrenergic inhibition of HVA-I(Ca) was partially prevented by the pretreatment with pertussis toxin (PTX) (5 microg/ml, 4 hr, 37 degrees C). CONCLUSIONS: RPNECs express both alpha1- and alpha2-ARs, signaling the release of stored Ca2+ and the inhibition of N-type Ca2+ channels, respectively.     

Isoflurane Alters Angiotensin II-Induced Ca2+ Mobilization in Aortic Smooth Muscle Cells from Hypertensive Rats: Implication of Cytoskeleton

Background: Angiotensin II (AngII) is a potent vasoconstrictor involved in the short-term control of arterial blood pressure. Isoflurane was reported to decrease vascular tone through an alteration of vascular smooth muscle cell vasomotor response to several agonists, but its effect on AngII signaling is not known. On the other hand, vascular response to AngII is altered in hypertension. In this study, the authors tested the hypothesis that (1) isoflurane alters AngII-induced intracellular Ca2+ mobilization in aortic vascular smooth muscle cell from Wistar Kyoto and spontaneously hypertensive rats, and (2) this effect could be associated with an alteration of the organization of microtubular network, reported to be involved in AngII signaling.

Methods: The effect of 0.5-3% isoflurane was studied (1) on AngII (10-6 m)-induced intracellular Ca2+ mobilization, intracellular Ca2+ release from internal stores, and Ca2+ influx in Fura-2 loaded cultured aortic vascular smooth muscle cell isolated from 6-week-old Wistar Kyoto and spontaneously hypertensive rats, using fluorescent imaging microscopy; and (2) on the organization of cytoskeletal elements, using immunofluorescence labeling.

Results: In both stains, isoflurane decreased in a concentration-dependent manner AngII-induced intracellular Ca2+ mobilization, Ca2+ release from internal stores, and Ca2+ influx through nifedipine-insensitive Ca2+ channels. This effect occurred at a lower concentrations of isoflurane in Wistar Kyoto rats than in spontaneously hypertensive rats. In both strains, the effect of isoflurane on AngII- Ca2+ mobilization was abolished by impairment with nocodazole, vinblastine, or paclitaxel of microtubules polymerization. Isoflurane directly altered tubular network organization in a concentration-dependent and reversible manner.     

七氟醚后处理对大鼠心肌缺血再灌注时Na+-K+-ATP酶和Ca2+-Mg2+-ATP酶活性的影响

目的 评价七氟醚后处理对大鼠心肌缺血再灌注时Na+-K+-ATP酶和Ca2+-Mg2+-ATP酶活性的影响,探讨其减轻心肌缺血再灌注损伤的机制.方法 健康雄性Wistar大鼠45只,体重250～280 g,随机分为3组(n=15):假手术组(S组)、缺血再灌注组(I/R组)和七氟醚后处理组(Spo组).I/R组和Spo组采用结扎左冠状动脉前降支30 min时进行再灌注的方法制备心肌缺血再灌注模型,S组仅在左冠状动脉前降支下穿线.Spo组再灌注前1 min开始吸入2.5%七氟醚持续5 min.于再灌注2 h时取心肌组织,测定心肌梗死体积、Na+-K+-ATP酶活性和Ca2+-Mg2+-ATP酶活性.结果 与S组比较,I/R组心肌梗死体积扩大,心肌组织Na+-K+-ATP酶和Ca2+-Mg2+-ATP酶的活性降低(P＜0.05);与I/R组比较,Spo组心肌梗死体积缩小,心肌组织Na+-K+-ATP酶和Ca2+-Mg2+-ATP酶的活性升高(P＜0.05).结论七氟醚后处理可提高Na+-K+-ATP酶和Ca2+-Mg2+-ATP酶的活性,从而减轻大鼠心肌缺血再灌注损伤.     

Ionic currents in single smooth muscle cells of the human vas deferens

PURPOSE: Smooth muscle cells of the vas deferens have an important role in carrying sperms to the exterior but little is known of their electrophysiological properties. We characterized the voltage-gated ion channel currents in single smooth muscle cells of the human vas deferens (HVSMCs). MATERIALS AND METHODS: We observed contractile responses of 8 circular smooth muscle strips of the human vas deferens to a high concentration (10 mM) of tetraethylammonium. HVSMCs were isolated using proteolytic enzymes (collagenase and papain), and were used for an electro-physiological study using whole cell and inside-out patch clamp configurations. RESULTS: The application of 10 mM tetraethylammonium induced rhythmic contractions of the strips. When HVSMCs were dialyzed with a KCl solution, step depolarizations of membrane potential evoked oscillatory outward K currents that were not inactivated. The large conductance Ca activated K (BKCa) and delayed rectifier components of the outward current were identified. The BKCa channel showed a large single channel conductance (162.7 +/- 13.2 pS with 5 mM K in the patch pipette). Two types of Ca currents were identified in the whole cell configuration. With a cell held at -50 mV an L-type Ca current was present during a depolarizing step pulse. From a holding potential of -90mV L-type and T-type Ca currents were elicited by depolarizing step pulses. CONCLUSIONS: HVSMCs have 2 (L and T) types of Ca channels and 2 types of K (BKCa and delayed rectifier) channels. Voltage dependent changes of these ion channels and their interactions may be important in regulating vas contractility.