海马神经元PSD-95 PDZ在幼期异氟醚麻醉致长期认知功能损害中的作用

发布时间：2018-05-03 01:32

本文选题：突触后致密蛋白-95 + 长时程增强　；参考：《郑州大学》2014年博士论文

【摘要】：每年约150万胎儿或新生儿接受麻醉。研究显示,幼期(出生后早期或胎儿期)麻醉暴露对大脑学习记忆能力的发育是一个危险因素。异氟醚吸入麻醉新生啮齿类动物可导致长期的神经病理学改变,这些改变可能会持续至成年。但异氟醚对神经元在发育过程中急性的影响研究较少,而且幼期经历异氟醚麻醉对神经元发育和突触形成的损害机制尚不清楚。基于这些研究现状,促使更多研究者重新审视异氟醚在儿科领域应用的安全性,并开展更多的关于麻醉药对大脑发育影响的研究。海马神经元发育在大脑的学习和记忆能力发育中起着十分关键的作用。海马神经元学习和记忆能力在神经元发育和突触快速形成期极易受到伤害。研究提示,新生大鼠异氟醚麻醉所致的长期神经认知功能损害可能与异氟醚导致的海马区神经元发育抑制有关。 N-甲基-D-天冬氨酸(N-methyl-D-aspartate receptors, NMDA)受体在海马神经元学习和记忆能力发育过程中起关键作用。NMDA受体调节海马神经元发育和突触的形成。幼期抑制或阻断NMDA受体将导致发育期大脑神经元发生退行性病变,并可能持续至成年。突触后膜致密蛋白-95(postsynaptic density-95,PSD-95)通过调节谷氨酸释放、并通过结合NMDA受体亚基NR2形成PSD-95/NR2复合物调节NMDA受体活性,参与NMDA受体介导的突触信号传递和突触可塑性调节等。我们的预实验结果显示异氟醚可剂量依赖性地改变PSD-95PDZ的化学结合位点结构,阻碍PSD-95/NR2复合物形成,并抑制NMDA受体活性。因此我们推测PSD-95PDZ介导的蛋白间的相互作用(PSD-95PDZ-mediated proteininteractions, PSD-95PDZ MPI)很可能参与了幼期麻醉暴露对海马认知功能损害的机制。为了验证探究PSD-95PDZ结构域是否参与异氟醚这一临床常用的吸入麻醉药物幼期暴露引起的长期认知功能损害机制,本课题从四个部分探讨PSD-95PDZ参与异氟醚幼期麻醉致长期认知功能损害的机制：1.异氟醚麻醉对PSD-95PDZ功能的破坏作用。通过异氟醚对出生后7天(PND7)大鼠进行的麻醉,应用Co-IP观察异氟醚对PSD-95/NR2相互作用的干扰和破坏。2. PSD-95PDZ在异氟醚对海马神经元LTP影响中的作用。通过构建PSD-95PDZ结构域抑制剂(Tat-PSD-95)观察异氟醚对海马神经元LTP的影响。3. PSD-95PDZ在幼期异氟醚麻醉对认知功能损害机制中的作用。通过上调表达PSD-95和应用PSD-95PDZ结构域抑制剂探讨幼期异氟醚麻醉致长期认知功能损害的机制。4.PSD-95PDZ在大鼠幼期异氟醚麻醉对海马神经元发育和突触形成影响机制中的作用。通过上调表达PSD-95和应用Tat-PSD-95探讨异氟醚麻醉对大鼠海马神经元发育和突触形成的影响。结果表明：1Co-IP结果显示,异氟醚剂量依赖性的抑制PND7和PND60大鼠的PSD-95与NR2之间的相互作用；2.1.5%异氟醚麻醉、Tat-PSD-95均能够明显抑制PND7大鼠海马脑片的兴奋性突触后场电位(field excitatory postsynaptic potential, fEPSP) LTP。1.5%异氟醚麻醉对PND60大鼠海马脑片fEPSP LTP无明显影响；3.异氟醚麻醉可导致PND60大鼠短期的海马依赖的认知功能损害(短于7天),但对PND7大鼠可导致长期的海马依赖的认知功能损害(长于7天,持续至出生后60天以后),Tat-PSD-95可导致与异氟醚相同的PND7大鼠长期的海马依赖的认知功能损害,上调海马神经元PSD-95可以减轻异氟醚麻醉对PND7大鼠长期的海马依赖的认知功能损害。4.异氟醚麻醉和Tat-PSD-95均抑制海马神经元发育和突触形成(抑制PND7大鼠出生后14天海马神经元突触素SYN-2的表达、抑制PND7大鼠出生后28天海马神经元神经连接蛋白Neuroligin-1的表达、减少PND7大鼠出生后67天的海马神经突触致密区数目)、促进PND7大鼠海马神经元凋亡(上调PND7大鼠出生后28天海马神经元Caspase-3表达)；上调海马神经元PSD-95能抑制异氟醚麻醉对PND7大鼠的这些损害作用。结论：异氟醚幼期麻醉可通过干扰破坏海马神经元PSD-95PDZ结构域影响PSD-95介导的突触后蛋白间相互作用,进而抑制海马神经元LTP、下调神经发育和突触形成蛋白、抑制突触形成建立、并促进神经元凋亡等机制导致长期的海马依赖的认知功能损害。
[Abstract]:About 1 million 500 thousand babies or newborns are anesthetized every year. Studies have shown that exposure to anaesthesia in the young (early or fetal stages) is a risk factor for the development of brain learning and memory. There are few studies on the acute effects of neurons in the development process, and the mechanism of the damage to the development of neurons and the formation of synapses by isoflurane is unclear. Based on these studies, more researchers have reexamined the safety of isoflurane in the field of Pediatrics, and more about the development of anesthetics on brain development. The study of influence.
The development of hippocampal neurons plays a crucial role in the learning and memory development of the brain. The learning and memory ability of hippocampal neurons is extremely vulnerable to the development of neurons and the rapid formation of synapses. The growth inhibition of the neurons in the horse area is related.
The N- methyl -D- aspartic acid (N-methyl-D-aspartate receptors, NMDA) receptor plays a key role in the learning and memory development of hippocampal neurons. The.NMDA receptor regulates the development of hippocampal neurons and the formation of synapses. The inhibition or blocking of the NMDA receptor at the young stage may lead to degenerative changes in the developmental brain neurons and may continue to occur. Adult. The postsynaptic membrane densification protein -95 (postsynaptic density-95, PSD-95) regulates the release of glutamate and regulates the activity of NMDA receptor by combining the NMDA receptor subunit NR2 to form the PSD-95/NR2 complex, and participates in the synaptic transmission and synaptic plasticity regulation mediated by the NMDA receptor.
Our experimental results show that isoflurane can change the chemical binding site structure of PSD-95PDZ in a dose-dependent manner, inhibit the formation of PSD-95/NR2 complex and inhibit the activity of NMDA receptor. Therefore, we speculate that the interaction of PSD-95PDZ mediated proteins (PSD-95PDZ-mediated proteininteractions, PSD-95PDZ MPI) is likely to be involved in the young. The mechanism of stage anaesthesia exposure on hippocampal cognitive impairment.
In order to verify whether the PSD-95PDZ domain is involved in the long-term cognitive impairment induced by isoflurane, a clinically common inhalation anesthetic, this topic explores the mechanism of PSD-95PDZ participation in the long-term cognitive impairment induced by isoflurane anesthesia from four parts: 1. the damage to PSD-95PDZ function by isoflurane anesthesia Effect. The effects of isoflurane on the interaction of PSD-95/NR2 and the effect of.2. PSD-95PDZ on the LTP effect of isoflurane on hippocampal neurons were observed by isoflurane on the 7 day (PND7) rats. The effect of isoflurane on the LTP of hippocampal neurons was observed by.2. PSD-95PDZ. The effect of isoflurane on the LTP of hippocampal neurons was observed by the construction of PSD-95PDZ domain inhibitor (Tat-PSD-95).3.. The role of PSD-95PDZ in the mechanism of cognitive impairment in juvenile isoflurane anesthesia. Through up regulation of the expression of PSD-95 and the application of PSD-95PDZ domain inhibitors to explore the mechanism of long-term cognitive impairment induced by isoflurane anesthesia at young stage.4.PSD-95PDZ in the mechanism of the effects of isoflurane on the development of neurogenesis and synapse formation in the hippocampus of the young rats The effects of isoflurane anesthesia on the development and synapse formation of hippocampal neurons in rats were investigated by up-regulated expression of PSD-95 and Tat-PSD-95.
The results showed that 1Co-IP results showed that isoflurane dose-dependent inhibition of the interaction between PSD-95 and NR2 in PND7 and PND60 rats, and 2.1.5% isoflurane anesthesia, Tat-PSD-95 could significantly inhibit the excitatory postsynaptic potential of hippocampal slices in PND7 rats (field excitatory postsynaptic potential,) isoflurane Intoxication had no obvious effect on fEPSP LTP in hippocampal slices of PND60 rats; 3. isoflurane anesthesia could lead to short-term hippocampal dependence of cognitive impairment in PND60 rats (shorter than 7 days), but PND7 rats could lead to long-term hippocampal Dependence Cognitive Impairment (longer than 7 days, until 60 days after birth), and Tat-PSD-95 could lead to the same P as isoflurane. ND7 rat's long-term hippocampal dependence cognitive impairment, up regulation of hippocampal neuron PSD-95 can reduce the cognitive impairment of the long-term hippocampal dependence of isoflurane anesthesia in PND7 rats.4. isoflurane anesthesia and Tat-PSD-95 inhibit the development of hippocampal neurons and synapse formation (inhibition of hippocampal synaptophysin SYN-2 14 days after the birth of PND7 rats SYN-2 To inhibit the expression of neural connexin Neuroligin-1 in hippocampal neurons of PND7 rats at 28 days after birth, reduce the number of hippocampal synapses in PND7 rats at 67 days after birth and promote the apoptosis of hippocampal neurons in PND7 rats (up regulation of Caspase-3 expression in hippocampus 28 days after birth of PND7 rats); up regulation of PSD-95 in hippocampal neurons. The effects of isoflurane anesthesia on PND7 rats were observed.
Conclusion:
Isoflurane anesthesia can affect the PSD-95PDZ domain of hippocampal neurons by interfering with the interaction of postsynaptic proteins mediated by PSD-95, thus inhibiting the LTP of hippocampal neurons, downregulating neurodevelopment and synaptic formation protein, inhibiting the formation of synapse formation, and promoting neuronal apoptosis and other mechanisms leading to long-term hippocampal dependence cognitive function. Damage.

【学位授予单位】：郑州大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R726.1

【参考文献】