MINIMUM NEURON DENSITY FOR SYNCHRONIZED BURSTS IN A RAT CORTICAL CULTURE ON MULTI-ELECTRODE ARRAYS
MINIMUM NEURON DENSITY FOR SYNCHRONIZED BURSTS IN A RAT CORTICAL CULTURE ON MULTI-ELECTRODE ARRAYS
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DOI：
10.1016/j.neuroscience.2010.08.038
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发表时间：
2010-11-24
期刊：
影响因子：
3.3
通讯作者：
Gohara, K.
中科院分区：
文献类型：
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作者：
Ito, D.;Tamate, H.;Gohara, K.
为了研究同步爆发所需的最小神经元和神经突密度，我们使用两种培养基在平面多电极阵列（MEA）上以五种平板密度（2500、1000、500、250和100个细胞/mm 2）培养大鼠皮层神经元：神经元培养基和补充有血清的Dulbecco改良Eagle培养基（DMEM/血清）。自发电活动的长期记录表明，显示同步爆发的培养物需要神经元培养基的初始接种密度至少为250个细胞/mm（2），DMEM/血清的初始接种密度至少为500个细胞/mm（2）。电记录后，立即在MEA上直接进行微管相关蛋白2（MAP 2）和神经丝200 kD（NF 200）的免疫细胞化学，以研究形成网络的神经元和神经突的实际密度。免疫荧光观察显示，复杂的神经元网络的建设需要相同的初始铺板密度同步爆发，过于稀疏的文化表现出显着减少的神经元和神经突。我们还发现，在1个月的存活神经元的最终密度大大降低，与初始铺板密度相比，并成为饱和在密集的培养。此外，神经突起的面积和细胞核的数量在密度较高的培养物中达到饱和。通过比较电生理记录和免疫细胞化学观察的结果，我们发现，有一个最低阈值的神经元密度，必须满足展览的同步爆发。有趣的是，MAP 2阳性最终神经元的最小密度在两种培养基之间没有差异;密度约为50个神经元/mm 2。该值是在神经元培养基初始接种密度为250个细胞/mm（2）和DMEM/血清初始接种密度为500个细胞/mm（2）的培养物中获得的。(C)2010年IBRO。由爱思唯尔有限公司出版。保留所有权利。
To investigate the minimum neuron and neurite densities required for synchronized bursts, we cultured rat cortical neurons on planar multi-electrode arrays (MEAs) at five plating densities (2500, 1000, 500, 250, and 100 cells/mm(2)) using two culture media: Neuron Culture Medium and Dulbecco's Modified Eagle Medium supplemented with serum (DMEM/serum). Long-term recording of spontaneous electrical activity clarified that the cultures exhibiting synchronized bursts required an initial plating density of at least 250 cells/mm(2) for Neuron Culture Medium and 500 cells/mm(2) for DMEM/serum. Immediately after electrical recording, immunocytochemistry of microtubule-associated protein 2 (MAP2) and Neurofilament 200 kD (NF200) was performed directly on MEAs to investigate the actual densities of neurons and neurites forming the networks. Immunofluorescence observation revealed that the construction of complicated neuronal networks required the same initial plating density as for synchronized bursts, and that overly sparse cultures showed significant decreases of neurons and neurites. We also found that the final densities of surviving neurons at 1 month decreased greatly compared with the initial plating densities and became saturated in denser cultures. In addition, the area of neurites and the number of nuclei were saturated in denser cultures. By comparing both the results of electrophysiological recording and immunocytochemical observation, we revealed that there is a minimum threshold of neuron densities that must be met for the exhibition of synchronized bursts. Interestingly, these minimum densities of MAP2-positive final neurons did not differ between the two culture media; the density was approximately 50 neurons/mm(2). This value was obtained in the cultures with the initial plating densities of 250 cells/mm(2) for Neuron Culture Medium and 500 cells/mm(2) for DMEM/serum. (C) 2010 IBRO. Published by Elsevier Ltd. All rights reserved.