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TRPV1 介导甲醛激活初级传入伤害性感受器细胞的作用

田丽娟, 杜意如, 萧勇, 吕卓敏, 于耀清, 崔秀玉, 陈军

首都医科大学疼痛生物医学研究所，北京 100069； 第四军医大学唐都医院疼痛生物医学研究所暨功能性脑疾病研究所，西安 710038

摘要

福尔马林致痛模型是化学伤害性刺激引起急性和持续性痛常用的动物模型。本研究旨在细胞和整体水平上证明TRPV1受体是否参与甲醛溶液(甲醛是福尔马林溶液的主要成分)对初级伤害性感受器细胞的激活过程。运用电流钳记录的92 个背根神经节细胞中，大约有34% 的中、小细胞对甲醛溶液敏感，产生动作电位的发放，其产生动作电位的潜伏期是(367.34±32.96)s，并且单个动作电位复极化相具有伤害性感受器细胞的特征性，如动作电位间期较长、复极化相有“驼峰”以及较长的后超极化相。在同时给予TRPV1 受体抑制剂Capsazepine (CPZ)时，甲醛溶液诱发背根神经节细胞产生的放电可以被阻断，但对细胞膜去极化不能完全阻断。在激光共聚焦钙成像中记录的160 个背根神经节细胞中，有32% 的细胞对甲醛溶液敏感，可以产生胞内钙离子浓度的升高，同时加入CPZ 后，67% 的这些甲醛敏感性细胞的胞内钙浓度升高可以被抑制。在电压钳状态下，甲醛溶液可以诱发背根神经节细胞产生内向电流，大约有41% 的细胞对甲醛溶液敏感，实验证明甲醛溶液诱发产生内向电流的幅度呈剂量依赖。当同时给予CPZ 时比TRPA1 受体选择性抑制剂HC-030031 抑制内向电流的幅度明显。利用行为学技术方法证明足底注射CPZ 可以明显抑制福尔马林溶液所致的第一相全程和第二相内仅第25~55 min 的自发缩足反射。以上结果提示，甲醛溶液选择性地激活一个亚群的初级传入伤害性感受器细胞，甲醛激活的动作电位的发放主要参与福尔马林诱发的第一相伤害性感受的产生。本实验进一步证明甲醛激活初级伤害性感受器细胞可能是通过TRPV1 受体和/ 或TRPA1受体参与介导的。

关键词： 福尔马林试验; 伤害性感受; TRPV1 受体; 背根神经节; 膜片钳; 钙成像

分类号：R441.1

Mediating roles of the vanilloid receptor TRPV1 in activation of rat primary afferent nociceptive neurons by formaldehyde

TIAN Li-Juan, DU Yi-Ru, XIAO Yong, LV Zhuo-Min, YU Yao-Qing, CUI Xiu-Yu, CHEN Jun

Institute for Biomedical Sciences of Pain, Capital Medical University, Beijing 100069, China； Institute for Biomedical Sciences of Pain and Institute for Functional Brain Disorders, Tangdu Hospital, the Fourth Military Medical University, Xi’an 710038, China

Abstract

The formalin test is a commonly used animal model of acute and tonic pain. However, the molecular targets of formaldehyde
(FA, the main ingredient of the formalin solution) on primary nociceptor cells remain controversial. In this report, the effects of FA on
electrophysiologically-identified primary nociceptor cells were evaluated in vitro and the roles of the vanilloid receptor TRPV1 in FAproduced
activation of primary nociceptors were also examined at both cellular and behavioral levels. Of 92 acutely dissociated dorsal
root ganglion (DRG) cells recorded by current patch-clamp technique, 34% were discharged by FA application with the mean onset
latencies of the first action potential (AP) being (367.34±32.96) s. All the FA-sensitive cells were identified as nociceptor cells by their
distinguishable features of AP including longer duration, existence of a hump (a shoulder or inflection) on the repolarizing phase, and
longer after-hyperpolarization of APs. Co-application of capsazepine (CPZ), a competitive antagonist of TRPV1 receptors, could
block FA-evoked firing with partial inhibition on the membrane depolarization of all cells tested. Of another 160 cells examined by
confocal calcium imaging, 32% were shown to respond to FA with an intracellular Ca2+ rise. Of 51 FA-sensitive cells, 67% were
suppressed by CPZ, suggesting partial involvement of TRPV1 in mediation of the FA-evoked intracellular Ca2+ rise. Under voltageclamp
mode, 41% of DRG cells were evoked to give rise to inward current with the remaining 59% being unchanged. In separate
experiments on the other 56 FA-sensitive cells, concentration-dependent increase in the FA-evoked current amplitude was demonstrated.
In comparison with controls, the FA-evoked inward current could be significantly suppressed by CPZ that was further enhanced by
HC-030031, a TRPA1 selective antagonist. Finally, local effects of CPZ were confirmed in the formalin test and it was shown that the
formalin-induced paw flinches were strongly suppressed by CPZ in phase 1 but with phase 2 being significantly suppressed only
during 25-55 min. It is therefore concluded that FA can directly activate a subpopulation of primary nociceptor cells and the FA-induced
AP discharges are likely to contribute mainly to phase 1, but not phase 2 of the formalin-induced nociception. The activation of primary
nociceptor cells by FA is likely to be mediated, at least in part, through TRPV1 and/or TRPA1 receptors.

Key words: Formalin test; nociceptor; TRPV1 receptor; dorsal root ganglion; patch clamp; calcium imaging

收稿日期：2009-09-23　　录用日期：2009-09-28

通讯作者：陈军　　E-mail: junchen@fmmu.edu.cn

引用本文：

田丽娟, 杜意如, 萧勇, 吕卓敏, 于耀清, 崔秀玉, 陈军. TRPV1 介导甲醛激活初级传入伤害性感受器细胞的作用[J]. 生理学报 2009; 61 (5): 404-416.

TIAN Li-Juan, DU Yi-Ru, XIAO Yong, LV Zhuo-Min, YU Yao-Qing, CUI Xiu-Yu, CHEN Jun. Mediating roles of the vanilloid receptor TRPV1 in activation of rat primary afferent nociceptive neurons by formaldehyde. Acta Physiol Sin 2009; 61 (5): 404-416 (in Chinese with English abstract).