Biochemical studies on the mechanism of the action of nerve growth factor

我们研究了神经生长因子（NGF）对培养的P3基底前脑胆碱能神经元中低亲和力NGF受体（LNGFR）基因表达的影响。基于RT-PCR（逆转录-聚合酶链反应）方法的结果，在添加100 ng/ml NGF后1天，与对照培养物相比，在NGF处理的培养物中发现LNGFR mRNA水平增加超过2倍。这种增加在添加NGF后维持长达3天。即使在1 ng/ml NGF（=40 pM）下也发现LNGFR mRNA的增加，表明在培养的胆碱能神经元中LNGFR mRNA水平的上调发生在接近高亲和力NGF受体的Kd值的NGF浓度下。此外，免疫组织化学染色显示，与对照组相比，NGF处理的培养物中的胆碱能神经元的单克隆抗LNGFR抗体染色更强。我们的研究结果表明，神经生长因子可以上调LNGFR在mRNA和蛋白水平上在培养的P3基底前脑胆碱能神经元的表达，这一机制可能发挥了重要作用，在增强神经生长因子对这些神经元的影响。

We examined the effect of nerve growth factor (NGF) on the expression of the low-affinity NGF receptor (LNGFR) gene in cultured P3 basal forebrain cholinergic neurons, on which NGF acts to promote differentiation. Based on the results of the RT-PCR (reverse transcriptase-polymerase chain reaction) method, over a 2-fold increase in the LNGFR mRNA level was found in NGF-treated cultures compared with control cultures at one day after the addition of 100 ng/ml NGF. This increase was maintained for up to 3 days after the addition of NGF. The increase in LNGFR mRNA was found even at 1 ng/ml NGF (=40 pM), indicating that the upregulation of the LNGFR mRNA level in cultured cholinergic neurons occurred at an NGF concentration near the Kd value for the high-affinity NGF receptor. In addtion, immunohistochemical staining showed stronger staining with a monoclonal anti-LNGFR antibody of the cholinergic neurons in NGF-treated cultures than those in control cultures. Our results suggest that NGF can up-regulate LNGFR expression at the mRNA and protein levels in cultured P3 basal forebrain cholinergic neurons and that this mechanism may play an important role in potentiating the effect of NGF on these neurons.