大鼠耳蜗一氧化氮合酶的分布和表达

Distribution and Expression of NOS in Rat Cochlea

目的 本实验先用组织化学法 ,通过观察还原型辅酶Ⅱ -黄递酶 (NADPH -黄递酶 )的活性了解一氧化氮合酶 (nitricoxidesynthase ,NOS)在大鼠耳蜗内分布。再用亲合免疫细胞组织化学技术 ,研究大鼠耳蜗内神经元型NOS(neuronalNOS ,nNOS)与内皮型NOS(endothelialNOS ,eNOS)的表达。方法 组化组大鼠耳蜗切片用辅酶Ⅱ孵育液在 37℃条件下孵育 1小时。免疫组化组大鼠耳蜗切片经消除内源性过氧化物酶 ,3%山羊正常血清封闭非正常结合点后 ,用兔抗nNOS抗体、兔抗eNOS抗体 ,室温下孵育 6 0分钟 ,再用生物素标记的山羊抗兔第二抗体孵育、滴加ABC试剂 ,以DAB试剂显色。结果 大鼠耳蜗血管球内皮细胞有明显NADPH -黄递酶活性 ,血管纹及螺旋神经节细胞也有NADPH -黄递酶活性反应。大鼠耳蜗内、外毛细胞、螺旋神经节细胞nNOS、eNOS的表达呈阳性。血管纹细胞处有阳性nNOS、eNOS的表达。耳蜗血管球的内皮细胞无nNOS的表达 ,但eNOS的表达呈强阳性。

Objective The purpose of the investigation was to identify the distribution and the expression of nitric oxide synthases(NOS) in the cochlea of the rat.Methods The cochlear sections of rat were incubated with NADPH incubation solution for 60 mins at 37℃. Sections of the Immunohistochemistry group were incubated with antibodies to nNOS and eNOS. This was followed by an incubation in secondary biotinylated antibody. At last, diaminobenzidine was used as the substrate. The primary antibody incubation was omited in the negative control.Results The staining for NADPH -diaphorase in frozen sections of inner ear showed a strong reactivity in the stria vascularis, spiral ganglion cells and glomeruli. The staining for nNOS in frozen sections of inner ear showed a strong reactivity in the inner and outer hair cells, stria vascularis and spiral ganglion cells. Especially the strong expression was seen in the outer hair cells-cochlear nerve synapse. The eNOS reactivity was seen in the glomeruli, stria vascularis and spiral ganglion cells.Conclusion Nitric xide(No) produced by nNOS is involved in neurotransmission according to the intense staining in the hair cells-cochlear nerve synapse. The intense reactivity of endothelial cells of glomeruli indicates the vasodilation action of NO produced by eNOS.