Study on the peripheral mechanism of muscular mechanical hyperalgesia using the delayed onset muscle soreness as a model

为了阐明肌肉机械性痛觉过敏的机制，使用由负荷延长收缩（LC）诱导的延迟发作肌肉疼痛的大鼠模型来检查以下三个点。1.通过肌肉压缩或收缩过程中释放的肌肉中致敏物质的变化，以及这些物质对肌肉C纤维受体的致敏作用。2.肌肉C纤维感觉受体神经元中表达的离子通道和接收的变化。3. LC后水肿肌肉中肌肉感觉感受器物理（机械）环境的变化。我们发现：1. ATP的释放是由肌肉的压缩，但其量之间的肌肉LC前和LC后2天没有差异。0.1-100 μM ATP对机械刺激引起的肌C-纤维受体反应有抑制作用。在LC周围不同时间点给予缓激肽受体拮抗剂、考克斯抑制剂和抗神经生长因子（NGF）抗体，提示缓激肽通过B2受体和考克斯-2参与了机械性痛觉过敏的触发过程，而NGF参与了机械性痛觉过敏的维持过程。相应于这些观察结果的表达考克斯-2 mRNA和蛋白增加0 - 12小时后LC，而NGF mRNA增加12小时后2天LC。神经生长因子易化肌肉细纤维受体的机械反应，支持神经生长因子在LC后机械性痛觉过敏中的作用。2.运动后背根节mRNA的抑制性消减杂交分析显示65种物质的RNA上调。其中两种蛋白（疼痛相关膜联蛋白A2和钙结合蛋白1）的RTPCR显示LC后2天显着增加。3. LC后2天的肌肉重约5%，表明I.C.后存在水肿。LC后肌肉水通道蛋白1和4的免疫组化无一致性变化。

To clarify the mechanism for the muscular mechanical hyperalgesia, following three points were examined using a rat model of delayed onset muscle soreness that was induced by loading lengthening contraction (LC). 1. Change of sensitizing substances in the muscle that is released by compression of the muscle or during contraction, and sensitization of muscle C-fiber receptors by these substances. 2. Change in ion channels and reception expressed in muscle C-fiber sensory receptor neurons. 3. Change in physical (mechanical) environment of muscle sensory receptors in edematous muscle after LC. We found: 1. ATP was released by compression of the muscle, but its amount was not different between the muscles before and 2 days after LC. ATP 0.1-100 μM rather suppressed the response of muscle C-fiber receptors to the mechanical stimulation. Administration of bradykinin receptor antagonists, Cox inhibitors and anti-NGF (nerve growth factor) antibody at different time points around LC revealed that bradykinin through B2 receptors and Cox-2 were involved in triggering the process to mechanical hyperalgesia, and that NGF was involved in maintenance of mechanical hyperalgesia. Corresponding to these observations expression of Cox-2 mRNA and protein increased during 0 12 hr after LC whereas NGF mRNA increased during 12 hr 2 days after LC. NGF facilitated the mechanical response of muscle thin-fiber receptors recorded in vitro, supporting the role of NGF in mechanical hyperalgesia after LC. 2. Suppression subtractive hybridization analysis of mRNA in DRGs after exercise revealed upregulation of RNAs of 65 substances. RTPCR of two of them, pain-related annexin A2 and calbindin 1, showed significant increase 2 days after LC. 3. The muscle 2 days after LC was by about 5% heavier, suggesting existence of edema after I.C. Immunohistochemistry of aquaporin 1 and 4 in muscle showed no consistent change after LC.