Monoamineoxidase and cholinesterase activity in neurons and neuropil from the rat cerebral cortex
Monoamineoxidase and cholinesterase activity in neurons and neuropil from the rat cerebral cortex
复制标题
大鼠大脑皮层神经元和神经纤维中的单胺氧化酶和胆碱酯酶活性
作者：
A. K. Sinha;S. Rose
在最近的一系列出版物中，我们报道了从大鼠大脑皮层中大量分离的神经元和神经元组分的代谢和酶性质。在这些研究中，我们检测了葡萄糖和氨基酸代谢、蛋白质合成和许多酸水解酶（ROSE，1968，1970; ROSE和SINHA，1969; SINHA和ROSE，1971; TIPLADY和ROSE，1971）。这种交流是指细胞组分中另外三种酶的活性，即单胺氧化酶（MAO）、乙酰胆碱酯酶（AChE）和丁酰胆碱酯酶（BChE）。我们之所以选择研究这些酶，部分是因为它们作为参与递质代谢系统的puerectin而感兴趣，部分是因为它们优先定位于一种或另一种细胞类型的报道。因此，使用我们自己改进的细胞分离方法，BLOMSTRAND、HAMBERGER和LEHNINGER（1970）发现，从富含神经毡的组分中分离的线粒体中MA 0的活性高于从神经元中分离的线粒体。在组织化学证据，乙酰胆碱酯酶被广泛认为是优先本地化的神经元，而BChE声称是更普遍的分布。(For综述和参考文献见FRIEDE，1966年。）AChE和BChE都浓缩在由脑匀浆制备的线粒体中（McCAMAN、DE LORES ARNAIZ和DE ROBERTIS，1965年），AChE也存在于突触体中（DE ROBERTIS、DE IRALDI、DE LORES ARNAIZ和SALGANICOFF，1962年）。完全按照以前的方法（ROSE，1967），通过密度梯度程序从大鼠皮质制备神经元和神经胶质部分，该程序产生四种部分：A（未破坏的组织，髓磷脂）; B（神经胶质，包括神经胶质细胞、树突和轴突突起以及突触体）; C（神经元）和D（细胞核，红细胞）。分级分离后，通过制粒收集样品，将颗粒重悬于10 ml蒸馏水中并冷冻储存直至测定。该过程通过渗透压休克破坏细胞。酶活性测定如下：MA 0（EC 1.4.3.4）采用KRAJL（1965）的方法;总胆碱酯酶（TChE）、AChE（EC 3.1.1.7）和BChE（EC 3.1.1.8）采用ELLMAN、COURTNEY、ANDRES和费瑟斯通（1961）的方法。蛋白质的测定采用LOWRY、ROSEBELT、FARR和RANDALL（1951）的方法。酶活性以任意单位/g蛋白/h表示。对从梯度中回收的合并级分进行三次试验，出于计算目的，认为每个此类试验组代表一个实验。
IN A SERIES of recent publications, we have reported on aspects of the metabolic and enzymic properties of bulk-isolated neuronal and neuropil fractions from rat cerebral cortex. In these studies we have examined glucose and amino acid metabolism, protein synthesis and a number of acid hydrolase enzymes (ROSE, 1968,1970; ROSE and SINHA, 1969; SINHA and ROSE, 1971; TIPLADY and ROSE, 1971). This communication refers to the activity of three further enzymes in the cell fractions, monoamine oxidase (MAO), acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). We have chosen to study these enzymes partly because of their interest as putatively involved in systems of transmitter metabolism and partly because of reports as to their preferential localization in one or another cell type. Thus, using a cell separation modified from our own, BLOMSTRAND, HAMBERGER and LEHNINGER (1970) have found a higher activity of MA0 in mitochondria isolated from fractions enriched in neuropil than from neurons. On histochemical evidence, AChE is widely believed to be preferentially localised in neurons whilst BChE is claimed to be more generally distributed. (For review and references see FRIEDE, 1966.) Both AChE and BChE are concentrated in mitochondria prepared from brain homogenates (MCCAMAN, DE LORES ARNAIZ and DE ROBERTIS, 1965) and AChE is also present in synaptosomes (DE ROBERTIS, DE IRALDI, DE LORES ARNAIZ and SALGANICOFF, 1962). Neuronal and neuropil fractions were prepared from rat cortex exactly as previously (ROSE, 1967), by a density gradient procedure which produces four fractions :A (undisrupted tissue, myelin); B (neuropil, including glial cells, dendrites and axonal processes and synaptosomes) ; C (neuronal) and D (nuclei, red cells). Following fractionation, the samples were collected by pelletting, the pellets resuspended in 10 ml of distilled water and stored frozen until assay. This procedure disrupts the cells by osmotic shock. Enzyme activities were determined as follows:M A 0 (EC 1.4.3.4) by the method of KRAJL (1965); total cholinesterase (TChE), AChE (EC 3.1.1.7) and BChE (EC 3.1.1.8) by the method of ELLMAN, COURTNEY, ANDRES and FEATHERSTONE (1961). Protein was determined by the method of LOWRY, ROSEBROUGH, FARR and RANDALL (1951). Enzyme activities were expressed in arbitrary units/g proteinlh. Assays were performed in tripilcate on pooled fractions recovered from the gradient, each such assay set being considered as representing one experiment for the purpose of calculation.