鉴定大鼠膀胱平滑肌间质细胞中的 T 型钙通道。
Identification of T-type calcium channels in the interstitial cells of Cajal in rat bladder.
机构信息
Department of Urology, First Affiliated Hospital, Third Military Medical University, Chongqing, China.
出版信息
Urology. 2012 Dec;80(6):1389.e1-7. doi: 10.1016/j.urology.2012.07.031. Epub 2012 Sep 18.
OBJECTIVE
To investigate the expression and function of T-type calcium channels in the interstitial cells of Cajal in rat bladders.
METHODS
Bladders were harvested from Sprague-Dawley rats. The expression of T-type calcium channels subtypes (α1G, α1H, and α1I) in interstitial cells of Cajal were identified by double-labeled immunofluorescence analysis and reverse transcription-polymerase chain reaction analysis in whole mount preparations of rat bladders. The function of T-type calcium channels in freshly isolated interstitial cells of Cajal was assessed by detecting the changes of intracellular calcium (Ca(2+)) with preloading fluo-3 AM, and by evaluating the changes of the phasic contractions of rat bladder strips after treating with mibefradil and glivec.
RESULTS
Three T-type calcium channels subtypes, α1G, α1H, and α1I, colocalized with c-kit in bladder interstitial cells of Cajal by double-labeled immunofluorescence analysis, and this was confirmed using reverse transcription-polymerase chain reaction. The T-type calcium channels selective blocker, mibefradil (1 μM), significantly decreased the intracellular calcium concentration (Ca(2+)) in isolated interstitial cells of Cajal (P < .01) and inhibited the spontaneous phasic contraction of bladder strips (P < .01). Moreover, the c-kit receptor blocker, glivec, significantly decreased the Ca(2+) of interstitial cells of Cajal further (P < .01) and the spontaneous phasic contraction of bladder strips.
CONCLUSION
T-type calcium channel subtypes were confirmed to colocalize in interstitial cells of Cajal in rats bladders, which might participate in the spontaneous activity of interstitial cells of Cajal and phasic contractions of bladder strips by modulating Ca(2+) in interstitial cells of Cajal.
目的
研究 T 型钙通道在大鼠膀胱平滑肌间质细胞中的表达和功能。
方法
取 Sprague-Dawley 大鼠的膀胱。通过双重免疫荧光分析和膀胱全层铺片的逆转录聚合酶链反应分析，鉴定 T 型钙通道亚型（α1G、α1H 和 α1I）在 Cajal 间质细胞中的表达。通过检测细胞内钙离子浓度(Ca(2+))的变化，评估米贝地尔和格列卫处理后大鼠膀胱条带的相位收缩变化，来评估 T 型钙通道在新分离的 Cajal 间质细胞中的功能。
结果
通过双重免疫荧光分析发现，三种 T 型钙通道亚型α1G、α1H 和 α1I 与膀胱 Cajal 间质细胞中的 c-kit 共定位，并通过逆转录聚合酶链反应得到证实。T 型钙通道选择性阻滞剂米贝地尔（1 μM）显著降低了分离的 Cajal 间质细胞中的细胞内钙离子浓度(Ca(2+))（P<0.01），并抑制了膀胱条带的自发性相位收缩（P<0.01）。此外，c-kit 受体阻滞剂格列卫进一步降低了 Cajal 间质细胞的Ca(2+)（P<0.01），并降低了膀胱条带的自发性相位收缩。
结论
T 型钙通道亚型被证实与大鼠膀胱 Cajal 间质细胞共定位，可能通过调节 Cajal 间质细胞中的Ca(2+)参与 Cajal 间质细胞的自发性活动和膀胱条带的相位收缩。
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