ABSTRACT
BACKGROUND:Primary culture in vitro of neurons plays an important role in the development, regeneration, signal transduction mechanisms, neuropharmacology and gene expressions of the nervous system. OBJECTIVE:To establish a simple method for primary culture of high-purity cortical neurons in neonatal Sprague-Dawley rats. METHODS:Cortical tissues were acquired from neonatal Sprague-Dawley rats born 1 day. In traditional experimental group, the whole cortex was removed;in improved experimental group, the cortical tissues, 2-3 mm thick on the brain surface were removed. Single cel suspensions were prepared after papain digestion and centrifugation and were then seeded onto 24-wel culture plates containing neuron solutions for primary culture (1×105 per wel ). Cel s were identified by neuronal specific markers MAP-2 and Tuj1 after 3-day culture. The number of neurons and neurite length were observed under inverted phase contrast microscope and recorded at 6, 24, 48 and 72 hours, 5 and 7 days of culture, resprctively. RESULTS AND CONCLUSION:The cultured cel s expressing MAP-2 and Tuj1 were neurons that could be used in the fol owing experiments. The purity of neurons in the improved experimental group was 92%at 3 days, while only 51%in the traditional experimental group. Cel s in both two groups had attached to the wal presenting with smal processes at 6 hours, and a simple neural network formed at the 3rd day until dense neural networks could be found at the 5th day. To conclude, our culture method herein is simple and convenient, and can be used to produce neurons with high purity, which wil be helpful for the experimental studies on cortical neurons from Sprague-Dawley rats.