Nakamura Y, Iga K, Shibata T, Shudo M, Kataoka K
Department of Physiology, Ehime University, School of Medicine, Japan.
Glia. 1993 Sep;9(1):48-56. doi: 10.1002/glia.440090107.
By a Percoll density-gradient centrifugation of rat hippocampal homogenate, we found a novel subcellular fraction (specific gravity approximately 1.046 g/ml), besides synaptosomes (approximately 1.060 g/ml), which showed a high activity of Na(+)-dependent glutamate uptake. The initial rate of the glutamate uptake in this fraction was as high as twice that in synaptosomes. Activities of choline acetyltransferase and high affinity choline uptake were, on the other hand, much lower. gamma-Aminobutyric acid uptake activity was nearly equivalent in both fractions. Electron microscopic observations revealed that the fraction was morphologically different from synaptosomal or myelin fractions, but mainly consisted of two different types of empty membrane vesicles; irregular (0.3-0.8 micron in diameter) and spheroid type (0.2 micron). The immunoreactivity to glial fibrillary acidic protein was appreciably high in this fraction. The marker enzyme analysis showed the fraction was rich in plasma membranes. On the basis of these results, the fraction is termed glial plasmalemmal vesicles (GPV). We analyzed kinetically the reaction of Na(+)-dependent glutamate uptake by GPV comparing with that by synaptosomes. Km values for glutamate in GPV was 4.7 microM and Vmax was 33 nmol/mg/min, while in synaptosomes 11 microM and 17 nmol/mg/min, respectively. Hill coefficients of Na+ activation in GPV and synaptosomes were 1.1 and 2.0, respectively. Thus, the mechanism or transporter molecule in glial cells for Na(+)-dependent glutamate transport is likely to be different from that in neurons.
通过对大鼠海马匀浆进行Percoll密度梯度离心，我们发现除了突触体（比重约为1.060 g/ml）外，还有一种新的亚细胞组分（比重约为1.046 g/ml），其显示出高活性的钠依赖性谷氨酸摄取。该组分中谷氨酸摄取的初始速率高达突触体中的两倍。另一方面，胆碱乙酰转移酶活性和高亲和力胆碱摄取则低得多。γ-氨基丁酸摄取活性在这两个组分中几乎相当。电子显微镜观察表明，该组分在形态上与突触体或髓鞘组分不同，但主要由两种不同类型的空膜囊泡组成；不规则型（直径0.3 - 0.8微米）和球型（0.2微米）。该组分对胶质纤维酸性蛋白的免疫反应性明显较高。标记酶分析表明该组分富含质膜。基于这些结果，该组分被称为胶质细胞质膜囊泡（GPV）。我们通过动力学分析比较了GPV和突触体对钠依赖性谷氨酸摄取的反应。GPV中谷氨酸的Km值为4.7 microM，Vmax为33 nmol/mg/min，而突触体中分别为11 microM和17 nmol/mg/min。GPV和突触体中钠激活的希尔系数分别为1.1和2.0。因此，胶质细胞中钠依赖性谷氨酸转运的机制或转运分子可能与神经元中的不同。
