Otto Frauke, Görtz Philipp, Fleischer Wiebke, Siebler Mario
Department of Neurology, Heinrich-Heine-University, Moorenstr. 5, 40225 Düsseldorf, Germany.
J Neurosci Methods. 2003 Sep 30;128(1-2):173-81. doi: 10.1016/s0165-0270(03)00186-9.
Neurons growing on microelectrode arrays (MEAs) are promising tools to investigate principal neuronal network mechanisms and network responses to pharmaceutical substances. However, broad application of these tools, e.g. in pharmaceutical substance screening, requires neuronal cells that provide stable activity on MEAs. Cryopreserved cortical neurons (CCx) from embryonic rats were cultured on MEAs and their immunocytochemical and electrophysiological properties were compared with acutely dissociated neurons (Cx). Both cell types formed neuritic networks and expressed the neuron-specific markers microtubule associated protein 2, synaptophysin, neurofilament and gamma-aminobutyric acid (GABA). Spontaneous spike activity (SSA) was recorded after 9 up to 74 days in vitro (DIV) in CCx and from 5 to 30 DIV in Cx, respectively. Cx and CCx exhibited synchronized burst activity with similar spiking characteristics. Tetrodotoxin (TTX) abolished the SSA of both cell types reversibly. In CCx SSA-inhibition occurred with an IC50 of 1.1 nM for TTX, 161 microM for magnesium, 18 microM for D,L-2-amino-5-phosphonovaleric acid (APV) and 1 microM for GABA. CCx cells were easy to handle and developed long living, stable and active neuronal networks on MEAs with similar characteristics as Cx. Thus, these neurochips seem to be suitable for studying neuronal network properties and screening in pharmaceutical research.
在微电极阵列（MEA）上生长的神经元是研究主要神经网络机制以及网络对药物物质反应的有前景的工具。然而，这些工具的广泛应用，例如在药物物质筛选中，需要能在MEA上提供稳定活性的神经元细胞。将来自胚胎大鼠的冷冻保存的皮质神经元（CCx）培养在MEA上，并将其免疫细胞化学和电生理特性与急性解离的神经元（Cx）进行比较。两种细胞类型都形成了神经突网络，并表达了神经元特异性标记物微管相关蛋白2、突触素、神经丝和γ-氨基丁酸（GABA）。分别在体外培养9至74天（DIV）的CCx和5至30 DIV的Cx中记录到自发尖峰活动（SSA）。Cx和CCx表现出具有相似尖峰特征的同步爆发活动。河豚毒素（TTX）可逆地消除了两种细胞类型的SSA。在CCx中，SSA抑制发生时，TTX的IC50为1.1 nM，镁为161 microM，D，L-2-氨基-5-磷酸戊酸（APV）为18 microM，GABA为1 microM。CCx细胞易于处理，并在MEA上形成了寿命长、稳定且活跃的神经网络，其特征与Cx相似。因此，这些神经芯片似乎适用于研究神经网络特性以及药物研究中的筛选。
