Ohkuma S, Tomono S, Tanaka Y, Kuriyama K, Mukainaka T
Department of Pharmacology, Kyoto Prefectural University of Medicine, Japan.
Int J Dev Neurosci. 1986;4(4):383-95. doi: 10.1016/0736-5748(86)90056-0.
Developmental patterns of taurine biosynthesizing system were investigated using primary cultured neurons prepared from the neopallium of 15-day-old fetal mice by a trypsin treatment in comparison with those in cerebral cortices obtained from age-matched fetal and neonatal mice. The morphological observations by phase contrast and scanning electron micrographies indicated that the cells in primary culture used in the present study possessed typical features of neurons. In addition, the immunohistochemical studies using the antibody to glial fibrillary acidic protein (GFAP), a specific marker for astroglia, revealed that the contamination of astroglias was negligible. The contents of taurine and metabolic intermediates in taurine biosynthesis, cysteine sulfinic acid and cysteic acid, in primary cultured neurons showed decreases during their development, especially during the first week after the inoculation. Similar developmental patterns of these amino acids were observed in cerebral cortices in vivo during perinatal stage, which corresponded to the first week of neuronal growth in vitro. On the other hand, the activities of cysteine sulfinic acid decarboxylase and cysteine dioxygenase, both of which are involved in the biosynthesis of taurine, were found to be increased progressively both in primary cultured neurons and in cerebral cortices in vivo during their growth. The immunohistochemical study using antitaurine antibody obtained from rabbit clearly demonstrated that immunoreactive materials were localized in cell bodies and the processes of neurons, and the intensity of the immunoreactivity in primary cultured neurons also showed a reduction with time of culture. These results indicate that primary cultured neurons used in this study possess a similar capacity to synthesize taurine from cysteine as developing brains in vivo. The present results also strongly suggest the well known decrease in cerebral taurine content in vivo during neonatal stages may be predominantly due to the decrease of taurine in neuronal cells.
利用胰蛋白酶处理从15日龄胎鼠新皮层制备的原代培养神经元，研究牛磺酸生物合成系统的发育模式，并与同龄胎鼠和新生鼠大脑皮层中的发育模式进行比较。相差显微镜和扫描电子显微镜的形态学观察表明，本研究中使用的原代培养细胞具有典型的神经元特征。此外，使用针对星形胶质细胞特异性标志物胶质纤维酸性蛋白（GFAP）的抗体进行的免疫组织化学研究表明，星形胶质细胞的污染可忽略不计。原代培养神经元中牛磺酸及其生物合成的代谢中间体半胱氨酸亚磺酸和半胱氨酸磺酸的含量在其发育过程中，尤其是接种后的第一周内有所下降。在围产期体内大脑皮层中观察到这些氨基酸类似的发育模式，这与体外神经元生长的第一周相对应。另一方面，发现参与牛磺酸生物合成的半胱氨酸亚磺酸脱羧酶和半胱氨酸双加氧酶的活性在原代培养神经元和体内大脑皮层生长过程中均逐渐增加。使用从兔获得的抗牛磺酸抗体进行的免疫组织化学研究清楚地表明，免疫反应性物质定位于神经元的细胞体和突起中，并且原代培养神经元中免疫反应性的强度也随培养时间而降低。这些结果表明，本研究中使用的原代培养神经元具有与体内发育中的大脑从半胱氨酸合成牛磺酸相似的能力。目前的结果还强烈表明，新生期体内大脑牛磺酸含量众所周知的下降可能主要是由于神经元细胞中牛磺酸的减少。
