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，我们采用负荷延长收缩（LC）诱导的延迟性肌肉酸痛大鼠模型，对以下三点进行了研究。1. 肌肉受压或收缩时释放的致敏物质在肌肉中的变化，以及这些物质对肌肉c -纤维受体的致敏作用2. 肌c -纤维感觉受体神经元离子通道和受体表达的变化。3. LC后水肿肌肉感觉受体物理（机械）环境的变化。我们发现：1；ATP是通过压迫肌肉释放的，但在LC前和LC后2天的肌肉中，ATP的释放量没有差异。ATP在0.1 ~ 100 μM范围内明显抑制肌肉c -纤维受体对机械刺激的反应。在LC周围的不同时间点给予缓激素受体拮抗剂、Cox抑制剂和抗NGF（神经生长因子）抗体，表明缓激素通过B2受体和Cox-2参与触发机械性痛觉过敏的过程，NGF参与机械性痛觉过敏的维持。相应的，在LC后0 - 12小时Cox-2 mRNA和蛋白的表达增加，而NGF mRNA在LC后12小时2天的表达增加。NGF促进了体外记录的肌肉薄纤维受体的机械反应，支持NGF在LC后机械性痛觉过敏中的作用。2. 运动后DRGs中mRNA的抑制减法杂交分析显示65种物质的rna上调。其中疼痛相关膜联蛋白A2和钙结合蛋白1的RTPCR在LC后2天显著升高。3. LC后2天肌肉重约5%，提示LC后存在水肿。LC后肌肉中水通道蛋白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.