本文選題：突觸后致密蛋白-95 + 長(zhǎng)時(shí)程增強(qiáng)　； 參考：《鄭州大學(xué)》2014年博士論文

【摘要】：每年約150萬(wàn)胎兒或新生兒接受麻醉。研究顯示,幼期(出生后早期或胎兒期)麻醉暴露對(duì)大腦學(xué)習(xí)記憶能力的發(fā)育是一個(gè)危險(xiǎn)因素。異氟醚吸入麻醉新生嚙齒類動(dòng)物可導(dǎo)致長(zhǎng)期的神經(jīng)病理學(xué)改變,這些改變可能會(huì)持續(xù)至成年。但異氟醚對(duì)神經(jīng)元在發(fā)育過(guò)程中急性的影響研究較少,而且幼期經(jīng)歷異氟醚麻醉對(duì)神經(jīng)元發(fā)育和突觸形成的損害機(jī)制尚不清楚。基于這些研究現(xiàn)狀,促使更多研究者重新審視異氟醚在兒科領(lǐng)域應(yīng)用的安全性,并開(kāi)展更多的關(guān)于麻醉藥對(duì)大腦發(fā)育影響的研究。 海馬神經(jīng)元發(fā)育在大腦的學(xué)習(xí)和記憶能力發(fā)育中起著十分關(guān)鍵的作用。海馬神經(jīng)元學(xué)習(xí)和記憶能力在神經(jīng)元發(fā)育和突觸快速形成期極易受到傷害。研究提示,新生大鼠異氟醚麻醉所致的長(zhǎng)期神經(jīng)認(rèn)知功能損害可能與異氟醚導(dǎo)致的海馬區(qū)神經(jīng)元發(fā)育抑制有關(guān)。 N-甲基-D-天冬氨酸(N-methyl-D-aspartate receptors, NMDA)受體在海馬神經(jīng)元學(xué)習(xí)和記憶能力發(fā)育過(guò)程中起關(guān)鍵作用。NMDA受體調(diào)節(jié)海馬神經(jīng)元發(fā)育和突觸的形成。幼期抑制或阻斷NMDA受體將導(dǎo)致發(fā)育期大腦神經(jīng)元發(fā)生退行性病變,并可能持續(xù)至成年。突觸后膜致密蛋白-95(postsynaptic density-95,PSD-95)通過(guò)調(diào)節(jié)谷氨酸釋放、并通過(guò)結(jié)合NMDA受體亞基NR2形成PSD-95/NR2復(fù)合物調(diào)節(jié)NMDA受體活性,參與NMDA受體介導(dǎo)的突觸信號(hào)傳遞和突觸可塑性調(diào)節(jié)等。 我們的預(yù)實(shí)驗(yàn)結(jié)果顯示異氟醚可劑量依賴性地改變PSD-95PDZ的化學(xué)結(jié)合位點(diǎn)結(jié)構(gòu),阻礙PSD-95/NR2復(fù)合物形成,并抑制NMDA受體活性。因此我們推測(cè)PSD-95PDZ介導(dǎo)的蛋白間的相互作用(PSD-95PDZ-mediated proteininteractions, PSD-95PDZ MPI)很可能參與了幼期麻醉暴露對(duì)海馬認(rèn)知功能損害的機(jī)制。 為了驗(yàn)證探究PSD-95PDZ結(jié)構(gòu)域是否參與異氟醚這一臨床常用的吸入麻醉藥物幼期暴露引起的長(zhǎng)期認(rèn)知功能損害機(jī)制,本課題從四個(gè)部分探討PSD-95PDZ參與異氟醚幼期麻醉致長(zhǎng)期認(rèn)知功能損害的機(jī)制：1.異氟醚麻醉對(duì)PSD-95PDZ功能的破壞作用。通過(guò)異氟醚對(duì)出生后7天(PND7)大鼠進(jìn)行的麻醉,應(yīng)用Co-IP觀察異氟醚對(duì)PSD-95/NR2相互作用的干擾和破壞。2. PSD-95PDZ在異氟醚對(duì)海馬神經(jīng)元LTP影響中的作用。通過(guò)構(gòu)建PSD-95PDZ結(jié)構(gòu)域抑制劑(Tat-PSD-95)觀察異氟醚對(duì)海馬神經(jīng)元LTP的影響。3. PSD-95PDZ在幼期異氟醚麻醉對(duì)認(rèn)知功能損害機(jī)制中的作用。通過(guò)上調(diào)表達(dá)PSD-95和應(yīng)用PSD-95PDZ結(jié)構(gòu)域抑制劑探討幼期異氟醚麻醉致長(zhǎng)期認(rèn)知功能損害的機(jī)制。4.PSD-95PDZ在大鼠幼期異氟醚麻醉對(duì)海馬神經(jīng)元發(fā)育和突觸形成影響機(jī)制中的作用。通過(guò)上調(diào)表達(dá)PSD-95和應(yīng)用Tat-PSD-95探討異氟醚麻醉對(duì)大鼠海馬神經(jīng)元發(fā)育和突觸形成的影響。 結(jié)果表明：1Co-IP結(jié)果顯示,異氟醚劑量依賴性的抑制PND7和PND60大鼠的PSD-95與NR2之間的相互作用；2.1.5%異氟醚麻醉、Tat-PSD-95均能夠明顯抑制PND7大鼠海馬腦片的興奮性突觸后場(chǎng)電位(field excitatory postsynaptic potential, fEPSP) LTP。1.5%異氟醚麻醉對(duì)PND60大鼠海馬腦片fEPSP LTP無(wú)明顯影響；3.異氟醚麻醉可導(dǎo)致PND60大鼠短期的海馬依賴的認(rèn)知功能損害(短于7天),但對(duì)PND7大鼠可導(dǎo)致長(zhǎng)期的海馬依賴的認(rèn)知功能損害(長(zhǎng)于7天,持續(xù)至出生后60天以后),Tat-PSD-95可導(dǎo)致與異氟醚相同的PND7大鼠長(zhǎng)期的海馬依賴的認(rèn)知功能損害,上調(diào)海馬神經(jīng)元PSD-95可以減輕異氟醚麻醉對(duì)PND7大鼠長(zhǎng)期的海馬依賴的認(rèn)知功能損害。4.異氟醚麻醉和Tat-PSD-95均抑制海馬神經(jīng)元發(fā)育和突觸形成(抑制PND7大鼠出生后14天海馬神經(jīng)元突觸素SYN-2的表達(dá)、抑制PND7大鼠出生后28天海馬神經(jīng)元神經(jīng)連接蛋白Neuroligin-1的表達(dá)、減少PND7大鼠出生后67天的海馬神經(jīng)突觸致密區(qū)數(shù)目)、促進(jìn)PND7大鼠海馬神經(jīng)元凋亡(上調(diào)PND7大鼠出生后28天海馬神經(jīng)元Caspase-3表達(dá))；上調(diào)海馬神經(jīng)元PSD-95能抑制異氟醚麻醉對(duì)PND7大鼠的這些損害作用。 結(jié)論： 異氟醚幼期麻醉可通過(guò)干擾破壞海馬神經(jīng)元PSD-95PDZ結(jié)構(gòu)域影響PSD-95介導(dǎo)的突觸后蛋白間相互作用,進(jìn)而抑制海馬神經(jīng)元LTP、下調(diào)神經(jīng)發(fā)育和突觸形成蛋白、抑制突觸形成建立、并促進(jìn)神經(jīng)元凋亡等機(jī)制導(dǎo)致長(zhǎng)期的海馬依賴的認(rèn)知功能損害。
[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.

【學(xué)位授予單位】：鄭州大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類號(hào)】：R726.1

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