Establishment of Primary Culture of Primate Brain Neurons.

非人类的原始材料在生物医学研究中是有用的，因为它们与人类的高度相似。如果要在中央神经系统（CNS）的体外研究中取得最大的优势，那么初级神经元的初级文化应该得到证实。在这项研究中，一种成功的低温保留和胚胎大鼠的脑细胞的初级培养的最佳方法首次被证实。一个成功的协议就像遵循一样。在文化介质补充中，由18天-旧的粪便分离的小Cerebral组织碎片，10%的DMSO是第一次冻结到？80 ° C时的缓慢速率，然后储存在液体硝酸盐中。从低温保留的半胱氨酸中恢复的活细胞占那些新鲜的半胱氨酸的20- 30%。由于免疫细胞化学，没有对脑细胞的低温保留的影响。下一步，本协议被用于低温保留和主基质的脑细胞（Cynomolgus猴子）的初级培养，并带有缓慢修改。Cynomolgus猴子在适宜的初级文化后的最佳生育年龄约为80天。“从80天旧的80岁的胎儿中恢复的可再生细胞对超过80%的新生细胞的反应。”来自新鲜和低温保留的初级半胱氨酸的形态特征是几乎可以独立的。通过添加细胞素花生苷，选择性的初级神经元文化已被证实。Cultured neurons from the cryopreserved primate cerebral tissues spontaneously formed networks， and randomly selected neurons in culture showed synchronous oscillations of [CaイイD12+イエD1]イイD2 inイエD2 without any stimulation。因此，初级神经元在文化中形成了功能性综合症，即使在低温保留之后。我们结合了一种主要的Cerebral神经元的初级文化，用于中枢神经系统的生物医学研究的有用和有效的方法。

Non-human primates are useful in biomedical studies because of their high similarity to humans. In order to make the most of this advantage in in vitro studies on central nervous system (CNS), primary culture of primate neurons should be established. In this study, the optimal procedure for the successful cryopreservation and primary culture of brain cells of fetal rats was first established. The successful protocol was as follows. Small cerebral tissue pieces isolated from 18-day-old fetuses in culture medium supplemented with 10% DMSO were first frozen to ?80℃ at slow rate, and then stored in liquid nitrogen. The recovery of viable cells from cryopreserved cerebral tissues was 20-30% of those from fresh tissues. By immunocytochemistry, no influence of cryopreservation on brain cells was observed. Next, this protocol was employed for cryopreservation and primary culture of brain cells of primate (Cynomolgus monkey) with slight modification. The optimal fetal age of cynomolgus monkey for appropriated primary culture after cryopreservation was around 80-day-old. The recovery of viable cells from the cryopreserved primate fetal cerebral tissues of the 80-day-old fetus corresponded to more than 80% of that from the fresh tissues. Morphological characteristics of neurons from the fresh and cryopreserved primate cerebral tissues were almost indistinguishable. By addition of cytosine arabinoside, selective culture of primate neurons was established. Cultured neurons from the cryopreserved primate cerebral tissues spontaneously formed networks, and randomly selected neurons in culture showed synchronous oscillations of [CaィイD12+ィエD1]ィイD2inィエD2 without any stimulation. Thus, primate neurons made functional synapses in culture even after cryopreservation. We concluded that primary culture of primate cerebral neuron was useful and effective method for biomedical study on CNS.