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1.
目的和方法 本研究采用离子探针Fura-2/AM结合计算机图象分析技术,并通过施加NO合酶抑制剂L-NNA和NO的作用靶--鸟苷酸环化酶(GC)的抑制剂美兰(Methylene Blue;MB),观察经培养的大鼠大脑皮层微血管内皮细胞和平滑肌细胞中的〖Ca^2+〗i在低氧作用后的变化以及与有关血管舒张因子NO和cGMP之间的关系。结果 低氧时大脑微血管内皮细胞和平滑肌细胞内的Ca^2+浓度有下降,  相似文献   

2.
血管平滑肌细胞上的自发内向阳离子通道   总被引:1,自引:0,他引:1  
采用膜片箝技术的细胞贴附式(cel-atached),在酶法分离的大鼠尾动脉平滑肌细胞上记录到一种自发的内向阳离子通道。结果发现:1、在实验条件下(池子液:Krebs,电极液:高钾),该通道电导为26.5±4.1pS,且具有明显的电压依赖、时间依赖和Ca2+依赖的特性;2、该电流具有极显著的内向整流特性;3、离子替换实验表明,该通道对Na+和K+具有很好的通透性,对Cl-的通透则很差;4、胞外加入4-AP(2mmol/L或5mmol/L),BaCl2(1mmol/L)和CsCl(20mmol/L)均不能抑制该电流;5、内向阳离子电流可与Ca2+-激活K+电流在同一细胞上交替活动,提示这两种电流可能在调控平滑肌细胞的基本活动方面起关键作用。  相似文献   

3.
目的:研究多巴胺对血管平滑肌细胞大电流、钙激活钾(BKca)通道的影响及其信息传递机制。方法用膜片钳细胞贴附式技术,记录细胞能液内灌流多巴胺受体激动剂、阻断剂以及第二信使及相关蛋白激酶拮机剂对猪冠状动脉血管平滑肌细胞BKca 爱道活动的影响。结果:多巴胺增加BKca通道活性(P〈0.01),并可被CA-1受体阻断剂SCH23390完全阻断,但不受β2受体阻断剂普藉洛尔的影响。腺苷酸环化酶抑制剂SO  相似文献   

4.
大鼠胰腺β细胞离子通道的一些特性   总被引:1,自引:1,他引:0  
Zeng XH  Lou XL  Qu AL  Wu HX  Zhou Z 《生理学报》2000,52(2):98-102
实验以单个Wistar大鼠胰腺β细胞为对象,用穿孔膜片箝和细胞贴附式记录技术研究ATP敏感K^+通道(KATP)、延迟整流型K^+通道(KDR)、Ca^2+通道和Na^+通道的有关特性。结果表明:⑴KATP通道的内流电导约65pS,外流电导约31pS,反转电位在-60mV左右;⑵KDR通道在延迟20ms后达到最大激活,KDR电流约为KATP的1/3;⑶钙电流在0mV左右达到40-60pA的峰值,L  相似文献   

5.
万勤  王福庄 《生理学报》1997,49(5):545-550
实验用Fluo-3负载细胞,在激光扫描共聚焦显微镜下直接监测缺氧后分散培养的大鼠海马CA1区神经元内游离Ca^2+浓度([Ca^2+]i)的变化,观察腺苷对这种变化的影响并初步探讨其作用机制。结果发现,急性缺氧使海马神经元[Ca^2+]i显著升高;腺苷(100μmol/L)明显抑制缺氧引起的[Ca^2+]i增高,腺苷A1受体拮抗剂CPT以及K^+通道阻断剂4-AP和ATP敏感性K^+通道阻断剂gl  相似文献   

6.
Xu WH  Li W  Wang XL 《生理学报》1998,50(1):75-81
本文用膜片箝全细胞技术比较了研究了单个兔肺动脉血管平滑肌细胞上延迟整流钾通道与克隆Kv1.5通道的电生理及药理学特性。将平滑肌细胞箝制在-40mV,以10mV的步跨阶跃去极化(0 ̄60mV)可产生一系列快速上升的外向电流,几无衰减,其激活曲线的V1/2为27.2mV。灌流液中加入100mmol/L和TEA 1mmol/L 4AP,电流幅度均明显减小,细胞外Ca^2+水平由1.5mmol/L降至0.  相似文献   

7.
G蛋白在亮啡肽诱导心肌细胞内钙释放中的作用   总被引:1,自引:0,他引:1  
魏振宇  谈世进 《生理学报》1995,47(2):173-178
本实验采用分离的SD大鼠心室肌细胞,以Fura-2AM荧光指示剂负载,检测心肌细胞内游离钙浓度(Ca^2+)变化。探讨亮啡肽(LEK)对(Ca^2+)的作用及其机制。实验结果:LEK(60μmol/L)能升高(Ca^2+)移去细胞外液钙此效应仍能出现,用caffeine (5mmol/L)耗竭细胞内钙池的钙,该效应消失,纳洛酮(100μmol/L),百日咳毒素(200ng/L)处理8-10h及pr  相似文献   

8.
鸭肫平滑肌肌球蛋白的提纯及其ATP酶性质的研究   总被引:1,自引:1,他引:0  
从鸭肫肌肉中分离纯化了平滑肌肌球蛋白,并对平滑肌肌球蛋白分子的亚单位组成和平滑肌肌球蛋白的ATP酶性质进行了分析研究。鸭肫平滑肌肌蛋白的Ca^2+-ATP酶活力与溶液的离子强度有关。在低于0.20mol/L的KCL浓度时,酶活力很低;大于0.30mol/L的KCL浓度时,酶活力较高,Ca^2+-ATP酶有两个最适pH值。K^+/EDTA-ATP酶活力随KCL浓度升高而增加。肌动蛋白激活的磷酸化肌球  相似文献   

9.
JNK/SAPK信号传递途径与细胞应激反应   总被引:2,自引:0,他引:2  
c-JunNH2-末端激酶(JNK)又称为应激活化蛋白激酶(SAPK),是有丝分裂原活化蛋白激酶(MAPK)家族成员之一。大量研究证实,JNK/SAPK信号传递途径在细胞应激反应中起重要作用。JNK/SAPK信号传递途径的激活促进细胞凋亡发生,其机制与诱导FasL表达、调控凋亡相关基因差异表达和改变细胞内Ca^2+环境与激活caspases家族在关。在某些情况下,JNK/SAPK信号传递途径的激活  相似文献   

10.
为测定植物细胞质内[Ca^2+]i,对胡萝卜(Daucuscartavar.sativaDC.)原生质体制备介质做了改进,并在正常生理条件,用温和的、非损伤性的方法将Ca^2+荧光指示剂indo-1K^+和fura-2K^+地入该原生体,能很好地标记细胞质的游离Ca^2+。  相似文献   

11.
Nitric oxide (NO) released from the endothelium or from NO-donors is a powerful vasodilator. Its effect is mediated partly by vascular smooth muscle high conductance calcium-activated potassium (Kca) channels. Contradictory data exist as to whether NO activated the KCa channel directly or indirectly via protein kinase G (PKG). Thus the hypothesis that NO-donors can activate the KCa directly was investigated using the patch-clamp technique and freshly isolated smooth muscle cells from the rat tail artery. In inside-out experiments, the activity of KCa-channels was increased 1.61 +/- 0.20-fold (n = 10) by 10 microM SNP and 1.45 +/- 0.17-fold (n = 8) by 10 microM SNAP. However, the activity of KCa channels was also increased 1.46 +/- 0.20-fold (n = 8) by addition of the experimental bath solution. Thus these results suggest that NO released from NO-donors cannot activate KCa channel of the rat tail artery smooth muscle cells directly.  相似文献   

12.
In smooth muscle cells, localized intracellular Ca2+ transients, termed "Ca2+ sparks," activate several large-conductance Ca2+-activated K+ (KCa) channels, resulting in a transient KCa current. In some smooth muscle cell types, a significant proportion of Ca2+ sparks do not activate KCa channels. The goal of this study was to explore mechanisms that underlie fractional Ca2+ spark-KCa channel coupling. We investigated whether membrane depolarization or ryanodine-sensitive Ca2+ release (RyR) channel activation modulates coupling in newborn (1- to 3-day-old) porcine cerebral artery myocytes. At steady membrane potentials of -40, 0, and +40 mV, mean transient KCa current frequency was approximately 0.18, 0.43, and 0.26 Hz and KCa channel activity [number of KCa channels activated by Ca2+ sparksxopen probability of KCa channels at peak of Ca2+ sparks (NPo)] at the transient KCa current peak was approximately 4, 12, and 24, respectively. Depolarization between -40 and +40 mV increased KCa channel sensitivity to Ca2+ sparks and elevated the percentage of Ca2+ sparks that activated a transient KCa current from 59 to 86%. In a Ca2+-free bath solution or in diltiazem, a voltage-dependent Ca2+ channel blocker, steady membrane depolarization between -40 and +40 mV increased transient KCa current frequency up to approximately 1.6-fold. In contrast, caffeine (10 microM), an RyR channel activator, increased mean transient KCa current frequency but did not alter Ca2+ spark-KCa channel coupling. These data indicate that coupling is increased by mechanisms that elevate KCa channel sensitivity to Ca2+ sparks, but not by RyR channel activation. Overall, KCa channel insensitivity to Ca2+ sparks is a prominent factor underlying fractional Ca2+ spark uncoupling in newborn cerebral artery myocytes.  相似文献   

13.
We investigated the mechanisms involved in the nitric oxide (NO)-induced inhibitory effects on longitudinal smooth muscle of mouse ileum, using organ bath technique. Exogenously applied NO, delivered as sodium nitroprusside (SNP; 0.1-100 micromol/L) induced a concentration-dependent reduction of the ileal spontaneous contractions. 1H-[1,2,4]oxadiazolol[4,3,a]quinoxalin-1-one (ODQ; 1 micromol/L), a guanilyl cyclase inhibitor, reduced the SNP-induced effects. Tetraethylammonium chloride (20 mmol/L), a non-selective K+ channel blocker, and charybdotoxin (0.1 micromol/L), blocker of large conductance Ca2+-dependent K+ channels, significantly reduced SNP-induced inhibitory effects. In contrast, apamin (0.1 micromol/L), blocker of small conductance Ca2+-dependent K+ channels, was not able to affect the response to SNP. Ciclopiazonic acid (10 micromol/L) or thapsigargin (0.1 micromol/L), sarcoplasmatic reticulum Ca2+-ATPase inhibitors, decreased the SNP-inhibitory effects. Ryanodine (10 micromol/L), inhibitor of Ca2+ release from ryanodine-sensitive intracellular stores, significantly reduced the SNP inhibitory effects. The membrane permeable analogue of cGMP, 8-bromoguanosine 3',5'-cyclic monophosphate (100 micromol/L), also reduced spontaneous mechanical activity, and its effect was antagonized by ryanodine. The present study suggests that NO causes inhibitory effects on longitudinal smooth muscle of mouse ileum through cGMP which in turn would activate the large conductance Ca2+-dependent K+ channels, via localized ryanodine-sensitive Ca2+ release.  相似文献   

14.
To clarify the mechanism by which lactate affects insulin secretion, we investigated the effect of lactate on insulin secretion, cytosolic free Ca2+ ([Ca2+](i), the ATP sensitive K+ channel (K(ATP)) and the Ca2+-activated K+ channel (K(Ca)) in HIT-T15 cells, and the results were compared with those of glucose and glibenclamide. All three agents caused insulin secretion and increased [Ca2+](i), but the effects on the K+ channels were different. In cell-attached patch configurations, 10 mmol/l glucose blocked both the K(ATP) and KCa channels, while 100 nmol/l glibenclamide had no effect on KCa channels, but blocked K(ATP) channels. Lactate at a concentration of 10 mmol/l activated both the K(ATP) and KCa channels, not only in cell-attached, but also in inside-out patch configurations, indicating that the increase in [Ca2+](i) and secretion of insulin by lactate cannot be explained by the blocking of the K+ channels. Lactate, at concentrations of 10 mmol/l and 50 mmol/l decreased 45Ca2+ efflux, while glibenclamide increased the efflux. These results suggest that the lactate-induced Ca2+ increase is not due to the closing of K+ channels, but at least in part, to the suppression of Ca2+ efflux from HIT cells.  相似文献   

15.
The role of inducible nitric oxide synthase (iNOS) in the acute activation of large-conductance, Ca(2+)-dependent K(+) channels (BK channels) by Escherichia coli endotoxin (lipopolysaccharide, LPS) was studied in murine vascular smooth muscle cells. Confocal laser scanning microscopy and patch clamp recordings were utilised. Within 2 h of donor rat sacrifice, iNOS-like immunoreactivity could be detected in cerebrovascular smooth muscle cells (CVSMCs) enzymatically dispersed from rat cerebral arteries. This staining was absent in cells fixed immediately after donor rat sacrifice. LPS was then applied to the cytoplasmic face of inside-out membrane patches excised from rat CVSMCs within 2-4 h of donor rat sacrifice. It was found that LPS (10-100 microg/ml) rapidly and reversibly increased the open probability of BK channels in these patches. This LPS response was not altered in the presence of the non-isoform specific NOS inhibitor N(omega)-nitro-L-arginine. LPS responses were then compared in aortic smooth muscle (ASMC) BK channels derived from wild-type and iNOS-knockout (iNOS-KO) mice. LPS activated BK channels in inside-out patches of ASMC membrane derived from both wild-type and iNOS-knockout mice. These studies establish that LPS can activate BK channels by a mechanism quite independent of the well-established pathway mediated by iNOS in vascular smooth muscle cells.  相似文献   

16.
Tamoxifen has been reported to directly activate large conductance calcium-activated potassium (KCa) channels through the KCa beta1 subunit, suggesting a cardio-protective role of this compound. The present study using knock-out (KO) mice for the KCa channel beta1 subunit was aimed at understanding the molecular mechanisms of the effects of tamoxifen on arterial smooth muscle KCa channels. Single channel studies were conducted in excised patches from cerebral artery myocytes from both wild-type and KO animals. The present data demonstrated that tamoxifen can inhibit arterial KCa channels due to a major decrease in channel open probability (P(o)), a mechanism different from the reduction in single channel amplitude reported previously and also observed in the present work. A tamoxifen-induced decrease in P(o) was present in arterial KCa channels from both wild-type and beta1 KO animals. This inhibition was concentration-dependent and partially reversible with a half-maximal concentration constant IC(50) of 2.6 microm. The effect of tamoxifen was actually dual Single channel kinetic analysis showed that tamoxifen shortens both mean closed time and mean open time; the latter is probably due to an intermediate duration voltage-independent blocking mechanism. Thus, tamoxifen block would predominate when KCa channel P(o) is >0.1-0.2, limiting the maximum P(o), whereas a leftward shift in voltage or Ca(2+) activation curves can be observed for P(o) values lower than those values. This dual effect of tamoxifen appears to be independent of the beta1 subunit. The molecular specificity of tamoxifen, or eventually other xenoestrogen derivatives, for the KCa channel beta1 subunit is uncertain.  相似文献   

17.
Although ketamine and Ca2+-activated K+ (KCa) channels have been implicated in the contractile activity regulation of cerebral arteries, no studies have addressed the specific interactions between ketamine and the KCa channels in cerebral arteries. The purpose of this study was to examine the direct effects of ketamine on KCa channel activities using the patch-clamp technique in single-cell preparations of rabbit middle cerebral arterial smooth muscle. We tested the hypothesis that ketamine modulates the KCa channel activity of the cerebral arterial smooth muscle cells of the rabbit. Vascular myocytes were isolated from rabbit middle cerebral arteries using enzymatic dissociation. Single KCa channel activities of smooth muscle cells from rabbit cerebral arteries were recorded using the patch-clamp technique. In the inside-out patches, ketamine in the micromolar range inhibited channel activity with a half-maximal inhibition of the ketamine concentration value of 83.8 +/- 12.9 microM. The Hill coefficient was 1.2 +/- 0.3. The slope conductance of the current-voltage relationship was 320.1 +/- 2.0 pS between 0 and +60 mV in the presence of ketamine and symmetrical 145 mM K+. Ketamine had little effect on either the voltage-dependency or open- and closed-time histograms of KCa channel. The present study clearly demonstrates that ketamine inhibits KCa channel activities in rabbit middle cerebral arterial smooth muscle cells. This inhibition of KCa channels may represent a mechanism for ketamine-induced cerebral vasoconstriction.  相似文献   

18.
川芎嗪对猪冠状动脉平滑肌细胞大电导钙激活钾通道的作用   总被引:14,自引:0,他引:14  
Yang YY  Yang Y  Zeng XR  Liu ZF  Cai F  Li ML  Zhou W  Pei J 《生理学报》2006,58(1):83-89
本工作旨在研究川芎嗪对猪冠状动脉平滑肌细胞钾通道的作用,为阐明其扩张冠状动脉血管的机制提供实验依据。采用膜片钳细胞贴附式和内面向外式记录方式观察川芎嗪对猪冠状动脉平滑肌细胞大电导钙激活钾通道(large-conductance Ca2+- activated potassium channels,BKCa channels)的作用,分别用蛋白激酶A(protein kinase A,PKA)抑制剂H-89和蛋白激酶G (protein kinase G,PKG)抑制剂KT-5823处理细胞,再观察川芎嗪对BKCa通道作用的变化。结果表明在研究的0.73-8.07 mmol/L浓度范围,川芎嗪可以剂量依赖性地激活BKCa通道,使通道的开放概率从(0.01±0.003)增加到(0.03±0.01)-(.21± 0.18)(P<0.01,n=10),使通道平均关闭时间从(732.33±90.67)ms降低到(359.67±41.30)-(2.96±0.52)ms(P<0.01, n=10)。川芎嗪的这种激活作用在浴液游离钙离子浓度接近0 mmol/L时也存在。PKA的特异性抑制剂H-89(3 μmol/L)和 PKG的特异性抑制剂KT-5823(1 μmol/L)对川芎嗪激活BKCa通道的作用无影响。以上结果提示:川芎嗪能直接激活冠状动脉平滑肌BKCa通道,这种作用可能是川芎嗪扩张冠状动脉血管的一种重要机制。  相似文献   

19.
Pulmonary artery smooth muscle cell (PASMC) relaxation at birth results from an increase in cytosolic cGMP, cGMP-dependent and kinase-mediated activation of the Ca2+-sensitive K+ channel (KCa), and closure of voltage-operated Ca2+ channels (VOCC). How chronic intrauterine pulmonary hypertension compromises perinatal pulmonary vasodilation remains unknown. We tested the hypothesis that chronic intrauterine pulmonary hypertension selectively modifies gene expression to mitigate perinatal pulmonary vasodilation mediated by the cGMP kinase-KCa-VOCC pathway. PASMC were isolated from late-gestation fetal lambs that had undergone either ligation of the ductus arteriosus (hypertensive) or sham operation (control) at 127 days of gestation and were maintained under either hypoxic (approximately 25 Torr) or normoxic (approximately 120 Torr) conditions in primary culture. We studied mRNA levels for cGMP kinase Ialpha (PKG-1alpha), the alpha-chain of VOCC (Cav1.2), and the alpha-subunit of the KCa channel. Compared with control PASMC, hypertensive PASMC had decreased VOCC, KCa, and PKG-1alpha expression. In response to sustained normoxia, expression of VOCC and KCa channel decreased and expression of PKG-1alpha increased. In contrast, sustained normoxia had no effect on PKG-1alpha levels and an attenuated effect on VOCC and KCa channel expression in hypertensive PASMC. Protein expression of PKG-1alpha was consistent with the mRNA data. We conclude that chronic intrauterine pulmonary hypertension decreases PKG expression and mitigates the genetic effects of sustained normoxia on pulmonary vasodilation, because gene expression remains compromised even after sustained exposure to normoxia.  相似文献   

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