脊髓κ-阿片受体在瑞芬太尼诱导痛觉过敏中的作用及相关机制研究

Opioid-induced hyperalgesia (OIH) is the new side effect during and after the administration of opioids. The mechanisms of OIH in detail have not been clarified. It is widely believed they are related with mu-opioid receptor. Remifentanil (Rem) is the representative of OIH. Our previous study showed that the central sensitization played a key role in remifentanil-induced hyperalgesia and the pretreatment of buprenorphine with the effect of kappa-opioid receptor (KOR) antagonist could prevent remifentanil-induced hyperalgesia. However, the role of kappa-opioid receptor (KOR) in spinal sensitization involved in the postoperative hyperalgesia induced by remifentanil and its mechanisms have not been reported at home and abroad. In this study, based on a rat of remifentanil-induced postoperative hyperalgesia model, we want to investigate followed by intrathecal injection of KOR agonist or antagonist: (1) the changes of remifentanil-induced mechanical and thermal pain thresholds; (2) the expression changes of KOR in the spinal dorsal horn by immunofluorescence-histochemical method，Western-blot and RT-PCR techniques in order to evaluate the role of spinal KOR in remifentanil-induced postoperative hyperalgesia and clarify its cellular signal transduction pathway; (3) the changes of postsynaptic density by activating KOR in spinal cord during remifentanil-induced postoperative; (4) the changes of long-term potential(LTP) in spinal dorsal neurons underlying remifentanil-induced hyperalgesia in vivo rat model of recording spinal LTP. To sum up, in the present study, we hope to elucidate that the activation of spinal KOR may be involved in spinal central sensitization mechanisms of remifentanil-induced postoperative hyperalgesia and to provide the experimental evidence and the new target to treat remifentanil-induced postoperative hyperalgesia in future.

阿片诱导痛觉过敏(OIH)是阿片药物新近发现的副作用。OIH确切机制尚未阐明，多认为与μ-阿片受体有关。瑞芬太尼（Rem）是诱发OIH的典型代表，我们已证实中枢敏化在OIH中起关键作用，并发现有κ-阿片受体（KOR）拮抗作用的丁丙诺啡预处理可以防止OIH，但脊髓KOR在脊髓中枢敏化的具体作用机制，迄今国内外尚无报道。本课题基于已建立的Rem诱导术后痛觉过敏的大鼠模型,通过鞘内注射KOR拮抗剂或激动剂：⑴观察Rem诱导大鼠术后疼痛行为学变化；⑵通过免疫荧光组织化学法、Werstern-blot和RT-PCR技术测定脊髓KOR的表达及其信号转导通路；⑶通过电镜观察脊髓神经元突触后致密物的密度；⑷建立脊髓长时程增强电位（LTP）的整体动物模型，记录脊髓背角LTP。通过上述研究，力图阐明脊髓KOR在Rem诱导术后痛觉过敏的作用及其相关机制，为临床防治瑞芬太尼诱导术后痛觉过敏提供实验依据和新的靶点。

阿片诱导痛觉过敏(OIH)的确切机制尚未阐明，多认为与μ-阿片受体有关。我们已证实中枢敏化在OIH中起关键作用，瑞芬太尼（Rem）是诱发OIH的典型代表，而脊髓KOR在瑞芬太尼诱导脊髓中枢敏化的具体作用机制，目前尚不清楚。本课题基于已建立的Rem诱导术后痛觉过敏的大鼠模型，选择鞘内置管成功的大鼠，随机分为6组（n = 6）：假手术组（C组），异氟烷组（I组），瑞芬太尼组（RF-组），瑞芬太尼手术组（RF+组），瑞芬太尼+nor-BNI组（nor-BNI：κ-阿片受体拮抗剂)，瑞芬太尼+U50488H组（U50488H：κ-阿片受体激动剂）。观察各组大鼠双侧跖底术后1 h，2 h，4 h，1 d，2 d, 3 d的机械性触痛阈（PWT）、机械性痛阈（PPW）和热刺激痛阈（PWL）的变化。通过疼痛行为学观察证实持续泵注瑞芬太尼后可以诱导大鼠术后痛觉过敏，明确鞘内注射κ-阿片受体激动剂可以防止瑞芬太尼诱导的大鼠术后痛觉过敏，而鞘内注射κ-阿片受体拮抗剂又恢复了瑞芬太尼诱导的大鼠术后痛觉过敏。通过Werstern-blot和RT-PCR技术测定脊髓KOR及mRNA的表达水平，发现瑞芬太尼诱发术后痛觉过敏与下调脊髓κ-阿片受体有关。透射电镜观察发现瑞芬太尼诱导脊髓中枢敏化的分子学基础与脊髓背角神经元突触后致密物的长度、厚度和面积增加、突触小泡数量增多有关。通过在体电生理学记录脊髓长时程增强电位（LTP），发现LTP的形成是瑞芬太尼诱导脊髓中枢敏化的直接证据，鞘内给予脊髓κ-阿片受体激动剂则明显抑制脊髓LTP现象。上述结果证实：瑞芬太尼诱导大鼠术后痛觉过敏与抑制脊髓κ-阿片受体有关。