视网膜动脉平滑肌细胞中大电导钙离子激活钾通道电流和钙离子浓度变化对糖尿病视网膜动脉收缩的影响  

Effect of large conductance Ca2＋-activated K＋ channel current and cytosolic calcium concentrations in retinalartery smooth muscle cells on diabetic retinal artery tension

作　　者：邵珺[1] 姚勇[1] 孙尉[3] 王如兴[2] 

机构地区：[1]南京医科大学附属无锡人民医院眼科,无锡214023 [2]南京医科大学附属无锡人民医院心内科,无锡214023 [3]苏州市立医院眼科,215000

出　　处：《中华实验眼科杂志》2016年第4期312-316,共5页Chinese Journal Of Experimental Ophthalmology

基　　金：江苏省自然科学基金项目（SBK201222073）;无锡市医管中心联合攻关项目（YGZX1206）

摘　　要：背景糖尿病视网膜病变（DR）是常见的视网膜微血管并发症，视网膜血管平滑肌细胞大电导钙激活钾离子通道（BK）是调节血管舒缩和血液动力的主要因素。目前关于视网膜动脉平滑肌细胞（RASMCs）上BK通道的功能变化在DR形成中的作用鲜有研究报道。目的研究正常及糖尿病模型大鼠RASMCs中BK通道电流、钙离子浓度及不同钙离子浓度下BK通道开放概率（NP0）的变化，探讨DR早期的血管损伤机制。方法采用随机数字表法将50只SPF级8～12周龄SD大鼠随机分为正常对照组和糖尿病模型组，糖尿病模型组40只大鼠腹腔内注射60mg／kg链脲佐菌素（STZ）法制作1型糖尿病模型，正常对照组大鼠10只同法注射枸橼酸钠溶液。采用酶消化法分离大鼠RASMCs，应用全细胞膜片钳技术记录大鼠RASMCs中BK通道电流；采用荧光探针法测定大鼠RASMCs内钙离子浓度；采用膜内向膜外型单通道膜片钳技术记录不同钙离子浓度条件下RASMCs中BK单通道NP。的变化。结果36只大鼠糖尿病造模成功，造模成功率为90％。刺激电压大于60mV时，糖尿病模型组大鼠RASMCs中BK通道电流密度明显下降，刺激电压为100mV时，正常对照组和糖尿病模型组大鼠RASMCsBK通道电流分别为（100±23）PA／PF和（50＋7）PA／PF，差异有统计学意义（t=19．80，P〈0．05）。当加入BK通道特异性阻滞剂非洲蝎毒素100nmol后，正常对照组的BK通道电流明显减弱，而糖尿病模型组BK通道电流无明显变化；正常对照组和糖尿病模型组大鼠RASMCs内钙离子浓度分别为（123±11）nmol／L和（255±10）nmol／L，差异有统计学意义（t=32．50，P〈0．05）。在刺激电位为60mV条件下，随着钙离子浓度增加，BK通道NP。增加，差异有统计学意义（F=15．28，P〈0．05）。结论糖尿病模型大鼠RASMCs中BK通道电流下降，细胞内钙离子浓度升高，BK单通道NP0下降。BK�Background Diabetic retinopathy （DR） is a common microvascular complications of the retina, retinal vascular smooth muscle ceils of large conductance calcium-activated potassium channels （BK） is a major factor in regulating vasomotor and hemodynamic. Currently,functional changes of BK channel in retinal artery smooth muscle ceils （RASMCs） and its role in DR were rarely reported. Objective This study was to investigate the early vascular damage mechanisms in DR by detecting the changes of BK channels current,calcium concentration and open probability （NP0） of BK channel with different calcium concentration in RASMCs of normal and diabetic rats.Method Fifty SPF SD 8-12 weeks old rats were randomly divided into normal control group and diabetic model group. Forty diabetic rats was intraperitoneally injected with 60 mg/kg streptozotocin to form type 1 diabetic model, 10 rats （the normal control group） were injected sodium citrate solution with the same manner. Fluorescent probe was applied to detect calcium concentration in rat RASMCs;RASMCs were isolated by using enzyme digestion, and BK- channel electric currents and calcium concentrations in the RASMCs were measured by whole-cell patch clamp technique and fluorescence assay,respectively. The NP0 of BK channel was measured by single patch clamp technique. Results Diabetic models were successfully established in 36 rats with the success rate 90%. When stimulation voltage is greater than 60 mV,the current density of BK channel in RASMCs of diabetic model group decreased;when stimulating voltage was 100 mV ,the BK channel currents of RASMCs in the normal control group and diabetic model group were （100±23） PA,/PF and （ 50 ± 7 ） PA/PF, the difference was statistically significant （ t = 19.80, P 〈 0.05 ）. After adding specific BK channel blocker African scorpion toxin 100 nmol, the BK channel current in the normal control group significantly reduced, and that in the diabetes model group was not significantly changed; the calcium ion c

关 键 词：实验性糖尿病 视网膜小动脉 血管平滑肌/代谢 大电导钙离子激活钾离子通道 钙/生 理 动物 大鼠 膜片钳 

分 类 号：R587.2[医药卫生—内分泌] R774.1[医药卫生—内科学]