Electrophysiological analysis for neurons derived from bone marrow stromal cells

骨髓基质细胞(BMSCs)最近被证明具有分化为神经元的能力。神经生长因子(NGF)是神经营养因子的一种，具有触发神经元长时程兴奋性的作用。在本研究中，为了确定BMSCs来源的神经元是否获得电兴奋性，或者NGF暴露是否诱导或扩大了神经元的电兴奋性，我们利用细胞外记录技术检测了NGF对BMSCs来源的神经元的影响。经鉴定，小鼠骨髓间充质干细胞来源的神经元样细胞表达特异性神经元标志物巢蛋白、神经细丝和胶质纤维酸性蛋白。大部分BMSCs来源的神经元表现为自发放电，放电幅度可达40 mV。在BMSCs来源的神经元中加入100 ng/ml的NGF，1min内电位幅值增大，3min后最大。但在注射NGF后5min，电位幅值的增大消失。第二次使用NGF也产生了类似的效果，但第三次使用则没有。另一方面，在细胞与细胞接触很少的培养条件下，即使多次施加100 ng/ml的NGF，也没有观察到细胞外电位的幅度增大。这些结果表明，NGF对小鼠骨髓基质细胞来源的神经细胞具有诱导短暂性电兴奋性的作用，但不具有持续性电兴奋性。

Bone marrow stromal cells (BMSCs) have recently been shown to have the capacity to differentiate into neurons. Nerve growth factor (NGF) which is one of neurotrophins has the effects to trigger long-term neuronal excitability. In the present study, to determine whether BMSCs-derived neurons acquire the electrical excitability, or whether an exposure to NGF induces or enlarges neuronal electrical excitability, we examine the effect of NGF for BMSCs-derived neurons using extracellular recording techniques. The neuron-like cells derived from murine BMSCs were confirmed to be stained with the specific neuronal marker Nestin, Neurofilament and Glial fibrillary acidic protein. Most BMSCs-derived neurons showed spontaneous discharges which amplitudes were up to 40 mV. When NGF at a concentration of 100 ng/ml was applied to BMSCs-derived neurons, the amplitudes of potentials were enlarged within 1 min and became greatest 3 min after NGF application. However, the enlargement of the amplitudes of potentials disappeared 5 min after NGF application. The similar effect by NGF was produced by the second application, but not by the third application. On the other hand, in the culture condition that little cell-cell contact were noted, enlargement of the amplitude of extracellular potentials was not observed even when NGF at a concentration of 100 ng/ml was applied a few times. These results indicate that NGF has the effect to induce transient but not sustained electrical excitability of neural cells derived from murine bone marrow stromal cells.