Background： Disrupted Ca2＋ homeostasis contributes to the development of colonic dysmotility in ulcerative colitis （UC）, but the underlying mechanisms are unknown. This study aimed to examine the alteration of col...Background： Disrupted Ca2＋ homeostasis contributes to the development of colonic dysmotility in ulcerative colitis （UC）, but the underlying mechanisms are unknown. This study aimed to examine the alteration of colonic smooth muscle （SM） Ca2＋ signaling and Ca2＋ handling proteins in a rat model of dextran sulfate sodium （DSS）-induced UC. Methods： Male Sprague-Dawley rats were randomly divided into control （n = 18） and DSS （n = 17） groups. Acute colitis was induced by 5% DSS in the drinking water for 7 days. Contractility of colonic SM strips （controls, n = 8 and DSS, n = 7） was measured in an organ bath. Cytosolic resting Ca2＋ levels （n = 3 in each group） and Ca2＋ transients （n = 3 in each group） were measured in single colonic SM cells. Ca2＋ handling protein expression was determined by Western blotting （n = 4 in each group）. Differences between control and DSS groups were analyzed by a two-sample independent t-test. Results： Average tension and amplitude of spontaneous contractions of colonic muscle strips were significantly enhanced in DSS-treated rats compared with controls （1.25 ± 0.08 g vs. 0.96 - 0.05 g, P = 0.007; and 2.67 - 0.62 g vs. 0.52 ±0.10 g, P= 0.013）. Average tensions of carbachol-evoked contractions were much weaker in the DSS group （1.08 ±0.10 g vs. 1.80 ±0.19 g, P = 0.006）. Spontaneous Ca2＋ transients were observed in more SM cells from DSS-treated rats （15/30 cells） than from controls （5/36 cells）. Peak caffeine-induced intracellular Ca2＋ release was lower in SM cells of DSS-treated rats than controls （0.413 ±0.046 vs. 0.548 ±0.041, P = 0.033）. Finally, several Ca2＋ handling proteins in colonic SM were altered by DSS treatment, including sarcoplasmic reticulum calcium-transporting ATPase 2a downregulation and phospholamban and inositol 1,4,5-trisphosphate receptor 1 upregulation. Conclusions： Impaired intracellular Ca2＋ signaling of colonic SM, caused by alteration of Ca2＋ handing proteins, contribute to展开更多
目的观察小鼠HCN1通道基因敲除对其膀胱Cajal间质细胞(interstitial cells of Cajal,ICCs)中BK通道的表达和功能的影响,探讨这种影响对膀胱兴奋性调控的意义。方法健康清洁成年的野生型C57BL/6J小鼠和HCN1通道基因敲除的C57BL/6J小鼠各4...目的观察小鼠HCN1通道基因敲除对其膀胱Cajal间质细胞(interstitial cells of Cajal,ICCs)中BK通道的表达和功能的影响,探讨这种影响对膀胱兴奋性调控的意义。方法健康清洁成年的野生型C57BL/6J小鼠和HCN1通道基因敲除的C57BL/6J小鼠各48只(雌雄各半)分别记为正常组和敲基因组,反转录PCR(RT-PCR)、荧光定量PCR(Q-PCR)、Western blot和免疫荧光双标检测其膀胱ICCs中BK通道各亚基的表达变化,离体逼尿肌肌条实验检测加入BK通道阻滞剂IBTX前后肌条收缩的变化,激光共聚焦显微镜下检测分别加入BK通道激动剂NS1619、阻滞剂IBTX前后小鼠膀胱ICCs内钙荧光的变化。结果 Q-PCR显示敲基因组小鼠膀胱中BK通道α、β1、β2、β3、β4各亚基表达均降低(P<0.01);Western blot显示敲基因组小鼠膀胱中BK通道α、β1、β2、β3、β4亚基表达均降低(其中α、β3、β4:P<0.01;β1、β2:P<0.05);免疫荧光双标显示敲基因组小鼠膀胱ICCs中BK通道α亚基表达降低(P<0.01);离体逼尿肌肌条实验显示加入IBTX后敲基因组和正常组肌条收缩幅度均变大(P<0.01,P<0.05),且敲基因组肌条收缩幅度的变化值小于正常组(P<0.05);激光共聚焦显微镜下可见加入NS1619后两组ICCs内钙荧光均降低(P<0.05)、加入IBTX后两组ICCs内钙荧光均增强(P<0.01),且不论加入激动剂或阻滞剂,敲基因组加药前后ICCs内钙荧光的变化值均小于正常组(P<0.01)。结论小鼠HCN1通道基因敲除下调了其膀胱ICCs中BK通道的表达及功能,这种下调可能是对HCN1通道基因敲除后膀胱收缩减弱的一种代偿。展开更多
旨在明确棉铃虫Helicoverpa armigera(Hübner)神经细胞上是否存在大电导钙激活钾通道(Largeconductance calcium-activated potassium channels,BKCa),进而探究BKCa通道是否为七氟菊酯(I型)和溴氰菊酯(II型)的作用靶标.应用全细胞...旨在明确棉铃虫Helicoverpa armigera(Hübner)神经细胞上是否存在大电导钙激活钾通道(Largeconductance calcium-activated potassium channels,BKCa),进而探究BKCa通道是否为七氟菊酯(I型)和溴氰菊酯(II型)的作用靶标.应用全细胞膜片钳技术首次记录了棉铃虫中枢神经细胞BKCa通道电流并分析了七氟菊酯和溴氰菊酯对BKCa通道的影响.结果显示,棉铃虫神经细胞膜上表达BKCa通道,其电流为一串快速激活、有快失活成分的电流,在-40 m V左右激活,电流幅值随刺激电位的增加而增强.七氟菊酯和溴氰菊酯均能显著抑制BKCa通道的峰值电流,使BKCa通道激活的电压依赖性发生改变,使激活曲线向去极化方向移动,其中七氟菊酯使半数激活电压(V1/2)向去极化方向移动约8 m V,溴氰菊酯使V1/2向去极化方向移动约16 m V.七氟菊酯和溴氰菊酯均能显著缩短BKCa通道快失活时间常数.上述结果表明BKCa通道是七氟菊酯和溴氰菊酯的作用靶标.展开更多
文摘Background： Disrupted Ca2＋ homeostasis contributes to the development of colonic dysmotility in ulcerative colitis （UC）, but the underlying mechanisms are unknown. This study aimed to examine the alteration of colonic smooth muscle （SM） Ca2＋ signaling and Ca2＋ handling proteins in a rat model of dextran sulfate sodium （DSS）-induced UC. Methods： Male Sprague-Dawley rats were randomly divided into control （n = 18） and DSS （n = 17） groups. Acute colitis was induced by 5% DSS in the drinking water for 7 days. Contractility of colonic SM strips （controls, n = 8 and DSS, n = 7） was measured in an organ bath. Cytosolic resting Ca2＋ levels （n = 3 in each group） and Ca2＋ transients （n = 3 in each group） were measured in single colonic SM cells. Ca2＋ handling protein expression was determined by Western blotting （n = 4 in each group）. Differences between control and DSS groups were analyzed by a two-sample independent t-test. Results： Average tension and amplitude of spontaneous contractions of colonic muscle strips were significantly enhanced in DSS-treated rats compared with controls （1.25 ± 0.08 g vs. 0.96 - 0.05 g, P = 0.007; and 2.67 - 0.62 g vs. 0.52 ±0.10 g, P= 0.013）. Average tensions of carbachol-evoked contractions were much weaker in the DSS group （1.08 ±0.10 g vs. 1.80 ±0.19 g, P = 0.006）. Spontaneous Ca2＋ transients were observed in more SM cells from DSS-treated rats （15/30 cells） than from controls （5/36 cells）. Peak caffeine-induced intracellular Ca2＋ release was lower in SM cells of DSS-treated rats than controls （0.413 ±0.046 vs. 0.548 ±0.041, P = 0.033）. Finally, several Ca2＋ handling proteins in colonic SM were altered by DSS treatment, including sarcoplasmic reticulum calcium-transporting ATPase 2a downregulation and phospholamban and inositol 1,4,5-trisphosphate receptor 1 upregulation. Conclusions： Impaired intracellular Ca2＋ signaling of colonic SM, caused by alteration of Ca2＋ handing proteins, contribute to
基金supported by the Science Foundation of Health Bureau of Xiamen Municipality(No.WZK213502z20077077)+1 种基金the Science Foundation of Health Department of Fujian ProvinceChina(No.2007-1-45)
文摘旨在明确棉铃虫Helicoverpa armigera(Hübner)神经细胞上是否存在大电导钙激活钾通道(Largeconductance calcium-activated potassium channels,BKCa),进而探究BKCa通道是否为七氟菊酯(I型)和溴氰菊酯(II型)的作用靶标.应用全细胞膜片钳技术首次记录了棉铃虫中枢神经细胞BKCa通道电流并分析了七氟菊酯和溴氰菊酯对BKCa通道的影响.结果显示,棉铃虫神经细胞膜上表达BKCa通道,其电流为一串快速激活、有快失活成分的电流,在-40 m V左右激活,电流幅值随刺激电位的增加而增强.七氟菊酯和溴氰菊酯均能显著抑制BKCa通道的峰值电流,使BKCa通道激活的电压依赖性发生改变,使激活曲线向去极化方向移动,其中七氟菊酯使半数激活电压(V1/2)向去极化方向移动约8 m V,溴氰菊酯使V1/2向去极化方向移动约16 m V.七氟菊酯和溴氰菊酯均能显著缩短BKCa通道快失活时间常数.上述结果表明BKCa通道是七氟菊酯和溴氰菊酯的作用靶标.
