Department of Neuroscience, University of Rome Tor Vergata, Via Montpellier, 1-00133 Rome, Italy.
Neurobiol Dis. 2011 Oct;44(1):92-101. doi: 10.1016/j.nbd.2011.06.008. Epub 2011 Jun 25.
Substance P (SP), a neuropeptide member of the tachykinin (TK) family, has a functional role both in physiological and pathological conditions, including Amyotrophic Lateral Sclerosis disease. One hypothesis of the selective motor neuron death in ALS involves the excitatory neurotransmitter glutamate, because these neurons are extremely susceptible to excessive stimulation of AMPA receptors. It has been reported that SP exerts its action against a variety of insults including excitotoxicity, and that altered levels of SP have been observed in the cerebrospinal fluid (CSF) of patients with ALS. Here we have analyzed the interaction between SP and AMPA receptor functionality, both in Control cortical neurons in culture and in those obtained from a genetic mouse model of ALS (G93A). Our studies demonstrate that SP reduces the kainate-activated currents in Control and G93A neurons and that this reduction is significantly higher in the mutated neurons. SP effect is mediated by its receptor NK1 because GR 82334 (5 μM), a NK1 competitive antagonist, is able to suppress the current reduction. Analysis of miniature excitatory postsynaptic currents (mEPSCs) in Control and G93A neurons indicates that SP (200 nM) is able to significantly decrease the mEPSC amplitudes in G93A neurons, whereas it is ineffective on Control mEPSCs. Western blotting experiments in cultures and cortical tissues show a higher NK1 expression level in G93A mice compared to that of Control. This is also confirmed by immunocytochemistry experiments in cultured neurons. In addition, the amount of GluR1 subunit AMPA receptors is not modified following SP exposure, indicating a non internalization of the AMPA receptors. Finally, toxicity experiments have revealed that SP is able to rescue G93A cortical cells whereas it is ineffective on those of Control. These findings provide the first evidence of SP having a physiological and protective role in the G93A mouse model of ALS, and may suggest the possible use of SP as a clinical therapeutic treatment.
P 物质（SP）是速激肽（TK）家族的一种神经肽，在生理和病理条件下都具有功能作用，包括肌萎缩侧索硬化症（ALS）疾病。ALS 中选择性运动神经元死亡的一个假设涉及兴奋性神经递质谷氨酸，因为这些神经元极易受到 AMPA 受体过度刺激。有报道称 SP 对抗多种刺激（包括兴奋性毒性）发挥作用，并且在 ALS 患者的脑脊液（CSF）中观察到 SP 水平改变。在这里，我们分析了 SP 与 AMPA 受体功能之间的相互作用，既包括在培养的对照皮质神经元中，也包括在 ALS 的遗传小鼠模型（G93A）中获得的神经元。我们的研究表明，SP 减少了对照和 G93A 神经元中的海人藻酸激活电流，并且这种减少在突变神经元中显著更高。SP 作用是由其受体 NK1 介导的，因为 NK1 竞争性拮抗剂 GR 82334（5 μM）能够抑制电流减少。对照和 G93A 神经元中小型兴奋性突触后电流（mEPSC）的分析表明，SP（200 nM）能够显著降低 G93A 神经元中的 mEPSC 幅度，而对对照 mEPSC 无效。培养物和皮质组织中的 Western 印迹实验显示 G93A 小鼠中的 NK1 表达水平明显高于对照。在培养神经元中的免疫细胞化学实验也证实了这一点。此外，暴露于 SP 后 AMPA 受体的 GluR1 亚基数量没有改变，表明 AMPA 受体没有内化。最后，毒性实验表明 SP 能够挽救 G93A 皮质细胞，而对对照细胞无效。这些发现为 SP 在 G93A 肌萎缩侧索硬化症小鼠模型中具有生理和保护作用提供了第一个证据，并可能表明 SP 作为临床治疗的可能用途。
