尼莫地平对小鼠齿状回诱导型一氧化氮合酶的表达及其活性的影响

Effects of nimodipine on expression and activity of inducible nitric oxide synthase in adult mouse dentate gyrus

目的探讨阻断L型电压门控式Ca2+通道(LVGCC)对成年动物齿状回诱导型一氧化氮合酶(Induciblenitricoxidesynthase,iNOS)的表达及其活性的影响。方法通过阻塞大脑中动脉(MCAO)90min制备局灶性脑缺血动物模型。缺血前15min静脉单次注射尼莫地平(2.0mg·kg-1),24h处死动物,采用RTPCR和Westernblot的方法研究尼莫地平对齿状回iNOS的mRNA和蛋白水平的影响,并测定iNOS酶活性和NO含量的变化。用LPS和TNFα诱导培养的星型胶质细胞表达iNOS,同时给予尼莫地平,培养24h后,检测各组细胞的iNOSmRNA水平和iNOS酶活性。结果与假手术组相比,MCAO后小鼠缺血侧齿状回iNOSmRNA和蛋白水平明显升高,iNOS活性增强,NOx含量增加。给予尼莫地平的动物,缺血侧齿状回无论是iNOSmRNA水平、iNOS蛋白含量,还是iNOS活性及NOx含量,与假手术组相比均显著下降。尼莫地平还可以降低培养的星型胶质细胞的mRAN水平。结论尼莫地平阻断LVGCC,可以明显下调缺血动物的齿状回和培养细胞的iNOS表达,降低iNOS的酶活性。

Aim To study the effect of nimodipine, a L-type voltage-gated Ca 2+ channel (L-VGCC) antagonist,on iNOS expression in adult mouse dentate gyrus after focal cerebral ischemia. Methods Focal cerebral ischemia was induced by intraluminal middle cerebral artery occlusion (MCAO). Nimodipine was administered as a single intravenous injection (2.0 mg·kg -1 body weight) 15 min before MCAO. To determine the effects of nimodipine on iNOS mRNA and protein levels, iNOS enzymatic activity and NO content, the animals were killed at 24 h after MCAO. To determine the effect of nimodipine on iNOS mRNA content of cultured astrocytes, LPS and TNFα were added into the culture flasks for 24 h to induce the expression of iNOS. At the same time, nimodipine was added into the culture medium.Results The mRNA level and protein content in the ipsilateral dentate gyrus were increased after ischemia, compared with those of sham-operated animals. The enzymatic activity of iNOS was stimulated and NOx content was elevated. In nimodipine-treated animals, there were marked decreases in the mRNA level, iNOS protein content, the enzymatic activity of iNOS and NOx content in the ipsilateral dentate gyrus compared with vehicle-treated animals. Nimodipine also significantly reduced iNOS mRNA level in cultured astrocytes.Conclusions Nimodipine inhibits the enzymatic acitivity of iNOS and reduces NO production by prohibiting iNOS expression both in adult mouse dentate gyrus after focal cerebral ischemia and cultured astrocytes.