脊髓GluA2/N-cadherin/CREB信号通路在酒精戒断致术后疼痛慢性化中的作用

Postoperative pain chronicity is closely related to preoperative stress. Our previous studies have shown that social defeat stress increased α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit GluA1 membrane surface expression and GluA2 internalization in the spinal dorsal horn neurons. GluA2 internalization causes it dissociating from N-cadherin and induces N-cadherin redistribution in the synaptic membrane, then activate CREB and downstream gene transcription, finally induces LTP initiation and maintenance and transfers acute pain to chronic pain. In our preliminary studies, we found that alcohol withdrawal increased GluA2 internalization in the membrane of spinal dorsal horn neurons. But we didn’t find significant changes of GluA1 insertion in the cell membrane. This suggests that postoperative pain prolongation in this alcohol withdrawal-induced mouse model is different with that in the stress-induced mouse model. Therefore, we hypothesize that GluA2/N-cadherin/CREB signaling pathway may be involved in the postoperative pain prolongation induced by alcohol withdrawal. To address this central hypothesis, we will use GluA2 knockout mice to explore the effects and the underlying mechanisms of GluA2/N-cadherin/CREB signaling pathway in alcohol withdrawal-induced postoperative pain chronicity by using the methods of pain behavior determination and spinal electrophysiological recording. We hope to further explore the molecular mechanisms of postoperative pain chronicity in different animal models.

术后疼痛慢性化与术前应激有关。我们之前研究发现应激导致的术后疼痛慢性化与脊髓背角AMPA受体亚单位GluA1细胞膜插入和GluA2内吞有关。GluA2内吞导致与其结合的N-cadherin重新分布，激活CREB，促进基因转录，诱导并维持LTP，促进疼痛慢性化。我们的预实验发现酒精戒断可模拟应激导致术后疼痛慢性化，并发现术后脊髓背角神经元上GluA2内吞，但未发现GluA1明显变化，提示酒精戒断模型有别于之前研究的社会挫败应激模型所致的术后疼痛慢性化机制。我们推测GluA2/N-cadherin/CREB信号途径参与了酒精戒断导致的术后疼痛慢性化机制。为此，我们将应用GluA2 Knockout 小鼠，通过疼痛行为测试、脊髓切片电生理记录等技术探讨GluA2/N-cadherin/CREB信号途径在酒精戒断致术后疼痛慢性化中的机制，阐明不同应激模型导致的术后疼痛慢性化的分子机制。

术前饮用酒精是否导致术后疼痛慢性化及其机制尚不清楚。我们的实验发现慢性饮用酒精后撤退（酒精戒断）导致术后急性疼痛向慢性疼痛转变。通过酒精孵育体外培养的脊髓背角神经元，钙成像检测发现酒精导致钙离子内流，钙离子通透型AMPA受体（细胞膜上不含GluA2亚单位）阻断剂NASPM阻断了酒精导致的钙离子内流，进一步的pHluorin融合技术发现，酒精导致AMPA受体亚单位GluA2的内吞，提示酒精通过促进GluA2内吞，使AMPA受体从钙离子不通透型转变为钙离子通透型。进一步研究发现，酒精戒断导致了术后脊髓背角AMPA受体介导的mEPSCs的频率增加，但是不影响幅值，提示酒精戒断导致突触易化。通过检测脊髓背角蛋白发现，AMPA受体亚单位GluA2在细胞膜上表达减少，而细胞内N-Cadherin和P-CREB表达增多，表明GluA2/N-Cadherin/CREB信号通路参与了酒精戒断致术后慢性化过程。该研究提示检测血液酒精浓度可以预测术后疼痛慢性化，同时GluA2也有可能成为酒精成瘾治疗药物的作用靶点。