ERK在臂旁核與杏仁核中央被核之突觸傳導與可塑性功能的角色研究

Role of extracellular signal-regulated kinase in synaptic transmission and plasticity of a nociceptive input on capsular central amygdaloid neurons

鄭信忠(Sin-Jhong Cheng)

指導教授 ： 閔明源

國立臺灣大學/生命科學院/動物學研究所/博士(2011年)

https://doi.org/10.6342/NTU.2011.10866

摘要

中文摘要
在過去的研究中發現許多慢性疼痛模式動物，杏仁核的中央壁核細胞(central capsular amygdaloid neurons，CeAC)都有大量的ERK磷酸化蛋白表現。並且證實ERK蛋白磷酸化會增加臂旁核(parabrachial nucleus，PB)將疼痛的訊息傳送至杏仁核CeAC的突觸傳導效能。在這個研究裡，我們直接探討ERK蛋白活化後對PBA-CeAC突觸傳導功能的影響。以全細胞鑲嵌記錄方式對杏仁核CeAC的主要細胞進行記錄。利用電流刺激引發興奮性突觸後電流(EPSCs)以及在有鍶的條件下誘發非同步性的興奮性突觸後電流(aEPSCs)，並分析EPSCs以及aEPSCs的結果。施予3μM phorbol 12,13-diacetate(PDA)激活ERK蛋白，可使PBA-CeAC的突觸傳導效能增益，並增加aEPSCs的大小與產生頻率。倘若在切片後先施予ERK蛋白的抑制劑(50μM U0126)則PDA促使PBA-CeAC突觸傳導效能的長期增益效用就會被遏止住，而且aEPSCs的大小也不再受PDA的影響。然而，EPSCs還是會在給予PDA期間，造成短暫的小幅度增益，而且aEPSCs的頻率依然受到PDA的影響而增加。但是若給予PKC的抑制劑Chelerythrine， 則會對PDA造成的短暫小幅度增益有進一步的抑制效果。由此結果可知PDA造成短暫的突觸傳導效能增益，是來自於突觸前PKC蛋白的活化使突觸小泡的釋放機率增加所產生的影響；另外PDA也會活化突觸後ERK蛋白而促使PBA-CeAC突觸形成長期的突觸傳導效能增益。高頻率刺激(100 Hz 刺激三回合，每回合之間的間距為60秒)可以在PBA-CeAC突觸活化NMDA接受器而產生長期的突觸傳導效能增益(LTP)，然而其分子機制也是仰賴PKC-ERK蛋白的活化途徑。在腓腸肌施打pH4酸性溶液所建立出來的肌肉酸痛動物(AIMP)或是NRG-1變異的小鼠，在其大腦切片可觀查到CeAC核區的神經細胞ERK磷酸化的表現量有明顯的增加，並且發現AIMP老鼠或是NRG-1變異小鼠，PBA-CeAC的突觸傳導效能也有顯著的被提高。增強AIMP或是NRG-1變異小鼠PBA-CeAC突觸傳導效能的分子機制可能與正常老鼠使用PDA增進突觸後傳導效能以及高頻率刺激所誘發出NMDAR-dependent LTP的分子機制共通，都是需要使ERK蛋白被活化。由於CeAC在疼痛感的管理上扮演著重要的角色，所以可以將活化ERK的機轉---使PBA與CeAC核區的突觸傳導效能提升---解釋為是中樞系統對疼痛敏感化的部份分子機制。

關鍵字

中樞敏感化 ； 長期的突觸傳導效能增益 ； 酸誘發肌肉酸痛 ； PDA ； PKC蛋白.

並列摘要

Abstract
It has been recently shown that, in the chronic pain animal models, there is an increased level of pohsphorylated extracellular signal-regulated kinase (ERK) in the central capsular amygdaloid neurons (CeAC) and an enhancement of synaptic transmission of the parabrachio-amygdaloid (PBA) pathway that transfer nociceptive signal from parabrachial nucleus onto CeAC neurons. In this study we directly test the effect of ERK activation on the PBA-CeAC synapse. Whole-cell patch-clamp recordings were obtained from CeAC neurons. Electrical stimulus- evoked excitatory postsynaptic currents (EPSCs) and asynchronous EPSCs (aEPSCs) in the presence of strontium (Sr) were analysised. Bath application of 3μM Phorbol 12,13-diacetate (PDA) , an ERK activator, caused a noticeable enhancement of synaptic transmission of PBA-CeAC synapse and significantly increased the amplitude and frequency of aEPSCs. Slices pretreated with an ERK inhibitor (50μM U0126) had block the long-last synaptic plasticity and amplitude of aEPSCs, but showed a significant enhancement of EPSCs during application of PDA and the frequency of aEPSCs had no effect. This immediate enhancement of EPSC was blocked by 10μM Chelerythrine, a PKC inhibitor. These results showed that the immediate enhancement of EPSC was dependent on pre-synaptic PKC activation that enhanced the vesicle release. The long-last synaptic potentiation induced by PDA was dependent on postsynaptic PKC-ERK activation in PBA-CeAC synapses. NMDAR-dependent LTP (Long-term potentiation) of PBA-CeAC EPSC was induced by high frequency tetanus stimulation (100 Hz × 3 trains / 60s，HFS), which also depended on PKC-ERK signaling pathway. In slices from acid-induced muscle pain (AIMP) and NRG-1 mutant mice, the p-ERK level in CeAC was significantly increased and the PBA-CeAC synaptic transmission was postsynaptically enhanced. The enhanced PBA-CeAC synaptic transmission in AIMPand NRG-1 mutan mice might share common mechanisms with the postsynaptic potentiation effect of PDA and HFS induced NMDAR-dependent LTP in normal slices, both requiring activation of ERK. Since CeAC plays an important role in affective component of pain, enhanced synaptic function of nociceptive input from PBA onto CeAC neurons might partially account for supraspinal mechanisms underlying central sensitization.

並列關鍵字

Central sensitization ； Long-term potentiation (LTP) ； acid-induced muscle pain ； PDA ； PKC

參考文獻

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