Vaid R R, Yee B K, Rawlins J N, Totterdell S
Department of Pharmacology, Oxford University, UK.
Brain Res. 1996 Sep 9;733(1):31-40. doi: 10.1016/0006-8993(96)00530-6.
NADPH-diaphorase histochemistry has been shown to stain cells which contain nitric oxide synthase, an enzyme responsible for the biosynthesis of the freely diffusable gas nitric oxide. A number of studies have mapped the distribution of NADPH-diaphorase-reactive neurons in the hippocampal formation but they have failed to yield consistent data. The major point of controversy concerns the presence of NADPH-diaphorase-reactive pyramidal cells in the CA1 subfield of the rat hippocampal formation. The present results show that CA1 pyramidal neurons do contain nitric oxide synthase (NOS) which can be reliably demonstrated with the appropriate histochemical procedure. One of the critical determinants of CA1 pyramidal cell NADPH-diaphorase activity is shown to be incubation of brains in sucrose solution prior to histochemical processing. Subicular pyramidal cells were also found to contain NOS and to possess NADPH-diaphorase activity. These results explain a number of contradictory reports in the literature relating to the presence of NADPH-diaphorase activity in hippocampal principal cells. Additionally, densitometric analysis carried out on 20 microns thick sections, from brains incubated in sucrose solution, indicated that there were characteristic gradients. The intensity of NADPH-diaphorase activity in pyramidal cells located in the ventral subiculum was found to be greater than those in the dorsal subiculum. A similar, yet marginal, trend was apparent for pyramidal cells in CA1 and CA3, as well as nonpyramidal cells in CA1. At both dorsal and ventral levels, NADPH-diaphorase-positive subicular pyramidal cells and CA1 nonpyramidal cells also show a greater intensity than CA1 or CA3 reactive pyramidal neurons. This study also showed that tissue incubation in sucrose solution prior to immunocytochemistry, enhanced immunoreactivity of the endothelial isoform of NOS whilst having little effect on neuronal NOS reactivity.
已证实，还原型辅酶Ⅱ黄递酶组织化学可使含有一氧化氮合酶的细胞着色，一氧化氮合酶是一种负责生物合成可自由扩散气体一氧化氮的酶。许多研究已绘制出还原型辅酶Ⅱ黄递酶反应性神经元在海马结构中的分布，但未能得出一致的数据。主要争议点在于大鼠海马结构CA1亚区中是否存在还原型辅酶Ⅱ黄递酶反应性锥体细胞。目前的结果表明，CA1锥体细胞确实含有一氧化氮合酶（NOS），通过适当的组织化学方法可可靠地证明这一点。研究表明，CA1锥体细胞还原型辅酶Ⅱ黄递酶活性的关键决定因素之一是在组织化学处理之前将大脑置于蔗糖溶液中孵育。还发现海马下托锥体细胞含有NOS并具有还原型辅酶Ⅱ黄递酶活性。这些结果解释了文献中一些关于海马主要细胞中还原型辅酶Ⅱ黄递酶活性存在情况的相互矛盾的报道。此外，对来自蔗糖溶液孵育大脑的20微米厚切片进行的光密度分析表明存在特征性梯度。发现位于腹侧海马下托的锥体细胞中还原型辅酶Ⅱ黄递酶活性强度大于背侧海马下托中的锥体细胞。CA1和CA3中的锥体细胞以及CA1中的非锥体细胞也有类似但不明显的趋势。在背侧和腹侧水平，还原型辅酶Ⅱ黄递酶阳性的海马下托锥体细胞和CA1非锥体细胞的强度也高于CA1或CA3反应性锥体细胞。这项研究还表明，在免疫细胞化学之前将组织置于蔗糖溶液中孵育，可增强NOS内皮亚型的免疫反应性，而对神经元NOS反应性影响不大。
