N-甲基-D-天冬氨酸受体拮抗剂对甲醛诱导的炎症性疼痛中内质网应激的调控作用研究

A preliminary investigation into the effect of the NMDA receptor inhibitor MK801 on endoplasmic reticulum stress in formalin-induced pain

摘要:

目的 探讨鞘内预注射N-甲基-D-天冬氨酸（NMDA）受体拮抗剂地佐环平（MK801）对甲醛诱导的炎症性疼痛中内质网应激反应的影响。方法 大鼠被随机分为对照组、甲醛组、甲醛+溶剂组、甲醛+MK801组。通过记录大鼠舔咬行为时间评估其疼痛行为。甲醛+MK801组在甲醛注射前30 min鞘内注射10 μL（0.1 μg/μL）MK801。采用Western blotting检测同侧脊髓腰膨大部位内质网应激相关蛋白免疫球蛋白重链结合蛋白（BIP）和活化转录因子6（ATF6）的表达。采用免疫组织化学法检测BIP和ATF6在脊髓同侧背角区域的表达及分布。结果 行为学结果显示：与甲醛组、甲醛+溶剂组相比，甲醛+MK801组第一相和第二相伤害性行为显著减少（P 0.05）。Western blotting检测结果显示，与对照组相比，甲醛组和甲醛+溶剂组注射同侧脊髓腰膨大部位BIP和ATF6的表达升高（P 0.05）；而甲醛+MK801组上述蛋白的相对表达量较甲醛组及甲醛+溶剂组均降低（P 0.05）。免疫组织化学染色结果显示，与对照组相比，甲醛组和甲醛+溶剂组注射同侧脊髓腰膨大背角BIP和ATF6免疫染色密度显著增加（P 0.05）；而甲醛+MK801组上述蛋白的免疫染色密度较甲醛组及甲醛+溶剂组均降低（P 0.05）。结论 鞘内注射NMDA受体拮抗剂MK801可显著减弱甲醛诱导的炎症性疼痛中内质网应激反应。

Abstract:

Objective To investigate N-methyl-d-aspartate (NMDA) receptor plays a critical role in the central sensitization of inflammatory pain, and endoplasmic reticulum (ER) stress is prominently involved in formalin-induced inflammatory pain. However, further research is needed to elucidate the relationship between NMDA receptor and ER stress in the context of inflammatory pain. In the present study, we investigated whether the ER stress response contributes to the attenuation of central sensitization following intrathecal pretreatment with the NMDA receptor antagonist MK801 in a formalin-induced inflammatory pain model.Methods Rats were randomly assigned to control, formalin, formalin + vehicle, and formalin + MK801 groups. Nociceptive behavior was assessed by recording licking and biting responses. The formalin + MK801 group received an intrathecal injection of 10 μL (0.1 μg/μL) MK801 30 minutes before formalin administration. The expression levels of ER stress markers immunoglobulin-binding protein (BIP) and activating transcription factor-6 (ATF6), in the lumbar enlargement of the spinal cord were analyzed by Western blotting. Immunohistochemistry was performed to examine the distribution of BIP and ATF6 in the ipsilateral dorsal horn of the spinal cord.Results Behavioral results: Compared with the formalin group and the formalin + vehicle group, the formalin+MK801 group significantly reduced nociceptive behaviors in both phase 1 and phase 2 (P 0.05). Western blotting: Compared with the control group, the expression levels of BIP and ATF6 in the ipsilateral lumbar enlargement of the spinal cord were significantly increased in the formalin group and the formalin + vehicle group (P 0.05). Compared with the formalin group and the formalin + vehicle group, the expression levels of BIP and ATF6 in the ipsilateral lumbar enlargement of the spinal cord were significantly decreased in the formalin+MK801 group (P 0.05). Immunohistochemistry: Compared with the control group, the immunostaining density of BIP and ATF6 in the dorsal horn of the ipsilateral lumbar enlargement were significantly increased in the formalin group and the formalin+vehicle group (P 0.05). Compared with the formalin group and the formalin + vehicle group, the immunostaining density of BIP and ATF6 in the dorsal horn of the ipsilateral lumbar enlargement were significantly decreased in the formalin + MK801 group (P 0.05).Conclusion These findings suggest that the NMDA receptor antagonist MK801 attenuates the ER stress response in formalin-induced inflammatory pain.