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）是神经营养因子之一，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BMSC衍生的神经元是否获得电兴奋性，或者暴露于NGF是否诱导或扩大神经元电兴奋性，我们使用细胞外记录技术研究了NGF对BMSC衍生的神经元的影响。经神经元特异性标记Nestin、Neurosilament和Glial Acidic Protein染色证实，小鼠骨髓基质干细胞分化为神经元样细胞。大多数骨髓基质细胞源性神经元显示自发放电，幅度可达40 mV。当浓度为100 ng/ml的NGF作用于BMSC来源的神经元时，电位的幅度在1 min内增大，并在应用NGF后3 min达到最大。而应用NGF后5 min，电位幅值的增大消失。第二次应用NGF产生了类似的效果，但第三次应用没有。另一方面，在注意到细胞-细胞接触很少的培养条件下，即使施加浓度为100 ng/ml的NGF几次，也没有观察到细胞外电位振幅的增大。这些结果表明，神经生长因子具有诱导来自小鼠骨髓基质细胞的神经细胞的瞬时但非持续的电兴奋性的作用。

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.