Astroglial mechanisms for inflammatory pain

描述（由申请人提供）：组织损伤性炎症性疼痛的特点是机械异常痛和热痛觉过敏，其介导机制不同。特别是，持续性疼痛的特点是机械异常性疼痛。我们对机械性异常痛如何维持的不完全理解阻碍了慢性疼痛的有效治疗。目前的疼痛研究主要集中在神经细胞和神经网络上。关于神经胶质细胞和神经胶质网络的重要性，特别是星形神经胶质网络在慢性疼痛条件下的重要性，我们所知甚少。本应用程序的总体目标是研究脊柱星形胶质细胞和星形胶质网络在持续炎性疼痛期间调节机械性异常痛中的作用。我们的策略是针对组织损伤后在脊髓星形胶质细胞中特异性表达和诱导的信号分子。c-Jun n -末端激酶（INK）是MAP激酶家族的成员。我们的初步研究表明(a) CFA炎症仅在脊髓星形胶质细胞中诱导JNK活化，(b)肽INK抑制剂阻断机械性异常痛觉的维持阶段而不影响热痛觉，(c)星形胶质间隙连接蛋白连接蛋白43 （Cx43）被炎症上调，间隙连接阻滞剂抑制机械性异常痛觉但不影响热痛觉。我们的中心假设是JNK是脊髓星形胶质细胞中的一个关键信号分子，并且炎症持续激活JNK通过星形胶质网络对机械性异常性疼痛的维持做出了重要贡献。这一假设将使用行为测试、免疫组织化学、原位杂交、Western blotting和激酶测定等方法进行检验。信号分子的细胞定位将通过免疫荧光双染色和荧光原位杂交和免疫荧光双染色进行检测。我们将使用CFA炎症模型来实现以下三个具体目标：(1)确定炎症诱导脊髓星形胶质细胞中JNK的持续激活，并且这种激活对于维持机械性异位痛至关重要；(2)确定cx43介导的脊髓星形胶质网络对持续性机械性异位痛的重要性；(3)探索脊髓JNK和星形胶质网络调节机械性异位痛的分子和细胞机制。这些拟议的研究将(a)确定机械性异常性疼痛维持的新分子和细胞机制，(b)记录脊髓星形胶质细胞和神经胶质网络在慢性疼痛中的重要作用，以及(c)揭示脊髓JNK通路作为治疗慢性疼痛的新靶点。

DESCRIPTION (provided by applicant): Tissue injury-induced inflammatory pain is characterized by mechanical allodynia and heat hyperalgesia that are mediated by distinct mechanisms. In particular, persistent pain is characterized by mechanical allodynia. Our incomplete understanding of how mechanical allodynia is maintained hinders the effective treatment of chronic pain. Current pain research focuses primarily on neuronal cells and neural networks. Less is known about the importance of glial cells and the glial network, in particular the astroglial network in chronic pain conditions. The overall goal of this application is to investigate the role of spinal astrocytes and astroglial network in regulating mechanical allodynia during persistent inflammatory pain. Our strategy is to target signaling molecules that are specifically expressed and induced in spinal astrocytes after tissue injury. c-Jun N-terminal kinase (INK) is a member of MAP kinase family. Our pilot studies have shown that (a) CFA inflammation induces JNK activation exclusively in spinal astrocytes, (b) a peptide INK inhibitor blocks maintenance phase of mechanical allodynia without affecting heat hyperalgesia, and (c) the astroglial gap junction protein connexin-43 (Cx43) is upregulated by inflammation and a gap junction blocker suppresses mechanical allodynia but not heat hyperalgesia. Our central hypothesis is that JNK is a critical signaling molecule in spinal astrocytes, and that persistent JNK activation by inflammation makes important contribution to the maintenance of mechanical allodynia via an astroglial network. This hypothesis will be tested using the methods of behavioral testing, immunohistochemistry, in situ hybridization, Western blotting, and a kinase assay. Cellular localization of signaling molecules will be examined by double staining of immunofluorescence and double staining of fluorescence in situ hybridization and immunofluorescence. We will use the CFA inflammation model to accomplish the following three specific aims: (1) to establish that inflammation induces persistent JNK activation in spinal astrocytes and that this activation is essential for the maintenance of mechanical allodynia, (2) to define the importance of Cx43-mediated spinal astroglial network for persistent mechanical allodynia, and (3) to explore molecular and cellular mechanisms by which spinal JNK and the astroglial network regulate mechanical allodynia. These proposed studies will (a) identify novel molecular and cellular mechanisms underlying the maintenance of mechanical allodynia, (b) document important roles of spinal astrocytes and the glial network for chronic pain, and (c) reveal spinal JNK pathway as a new target for the treatment of chronic pain.