Gerace E, Masi A, Resta F, Felici R, Landucci E, Mello T, Pellegrini-Giampietro D E, Mannaioni G, Moroni F
Department of Health Sciences, Section of Clinical Pharmacology and Oncology, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.
Department of Neuroscience, Section of Pharmacology, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.
Neurobiol Dis. 2014 Oct;70:43-52. doi: 10.1016/j.nbd.2014.05.023. Epub 2014 Jun 20.
An excessive activation of poly(ADP-ribose) polymerases (PARPs) may trigger a form of neuronal death similar to that occurring in neurodegenerative disorders. To investigate this process, we exposed organotypic hippocampal slices to N-methyl-N'-nitro-N'-nitrosoguanidine (MNNG, 100μM for 5min), an alkylating agent widely used to activate PARP-1. MNNG induced a pattern of degeneration of the CA1 pyramidal cells morphologically similar to that observed after a brief period of oxygen and glucose deprivation (OGD). MNNG exposure was also associated with a dramatic increase in PARP-activity and a robust decrease in NAD(+) and ATP content. These effects were prevented by PARP-1 but not PARP-2 inhibitors. In our experimental conditions, cell death was not mediated by AIF translocation (parthanatos) or caspase-dependent apoptotic processes. Furthermore, we found that PARP activation was followed by a significant deterioration of neuronal membrane properties. Using electrophysiological recordings we firstly investigated the suggested ability of ADP-ribose to open TRPM2 channels in MNNG-induced cells death, but the results we obtained showed that TRPM2 channels are not involved. We then studied the involvement of glutamate receptor-ion channel complex and we found that NBQX, a selective AMPA receptor antagonist, was able to effectively prevent CA1 neuronal loss while MK801, a NMDA antagonist, was not active. Moreover, we observed that MNNG treatment increased the ratio of GluA1/GluA2 AMPAR subunit expression, which was associated with an inward rectification of the IV relationship of AMPA sEPSCs in the CA1 but not in the CA3 subfield. Accordingly, 1-naphthyl acetyl spermine (NASPM), a selective blocker of Ca(2+)-permeable GluA2-lacking AMPA receptors, reduced MNNG-induced CA1 pyramidal cell death. In conclusion, our results show that activation of the nuclear enzyme PARP-1 may change the expression of membrane proteins and Ca(2+) permeability of AMPA channels, thus affecting the function and survival of CA1 pyramidal cells.
聚（ADP - 核糖）聚合酶（PARPs）的过度激活可能引发一种神经元死亡形式，类似于神经退行性疾病中发生的情况。为了研究这一过程，我们将器官型海马切片暴露于N - 甲基 - N'- 硝基 - N'- 亚硝基胍（MNNG，100μM，处理5分钟），这是一种广泛用于激活PARP - 1的烷基化剂。MNNG诱导CA1锥体细胞的退化模式在形态上类似于短暂缺氧缺糖（OGD）后观察到的情况。暴露于MNNG还与PARP活性的显著增加以及NAD(+)和ATP含量的显著降低有关。这些效应可被PARP - 1抑制剂而非PARP - 2抑制剂所阻断。在我们的实验条件下，细胞死亡不是由AIF易位（parthanatos）或半胱天冬酶依赖性凋亡过程介导的。此外，我们发现PARP激活后神经元膜特性显著恶化。使用电生理记录，我们首先研究了ADP - 核糖在MNNG诱导的细胞死亡中打开TRPM2通道的推测能力，但我们得到的结果表明TRPM2通道不参与其中。然后我们研究了谷氨酸受体 - 离子通道复合物的参与情况，发现选择性AMPA受体拮抗剂NBQX能够有效预防CA1神经元损失，而NMDA拮抗剂MK801则无活性。此外，我们观察到MNNG处理增加了GluA1/GluA2 AMPAR亚基表达的比例，这与CA1区而非CA3区AMPA微小兴奋性突触后电流（sEPSCs）的电流 - 电压关系的内向整流有关。因此，1 - 萘基乙酰精胺（NASPM），一种缺乏GluA2的Ca(2+)通透性AMPA受体的选择性阻滞剂，减少了MNNG诱导的CA1锥体细胞死亡。总之，我们的结果表明，核酶PARP - 1的激活可能会改变膜蛋白的表达和AMPA通道的Ca(2+)通透性，从而影响CA1锥体细胞 的功能和存活。
