Endothelin Mechanisms in Metastic Prostate Cancer Pain

描述（由申请人提供）：内源性肽内皮素-1（ET-1）对于神经损伤、切口、炎症和癌症后的疼痛和疼痛纤维对非伤害性刺激的敏感性至关重要。在这个建议中，我们试图了解损伤和炎症后皮肤中ET-1的升高与随后的疼痛之间的关系。最近的研究表明，ET-1通过作用于神经末梢周围的组织而不是直接作用于神经引起外周疼痛升高; ET-1及其同源物GPC受体ETA和ETB在痛觉过敏和异常性疼痛的条件下升高。ET受体的拮抗剂能够预防或逆转这些疼痛，证明内源性ET-1在驱动慢性疼痛状态中的重要作用。目前的证据表明，ET-1在皮肤损伤/炎症过程中的作用与其他三种分子/受体密切相关; TRPV 1（辣椒的受体，也被热和H+激活），谷氨酸和CGRP。TRPV 1参与ET-1或CFA注射到大鼠爪子中的皮肤疼痛在非常短的时间内是明显的，并且皮肤神经纤维中TRPV 1的免疫细胞化学染色增加5-10倍。在ET-1或CFA注射后约30分钟内，谷氨酸和CGRP有助于ET-1的疼痛敏化作用;这些物质也以ET-1增强的方式由培养的感觉神经元释放。该提案包含4个具体目标，以解决这些变化的过程：1。目的探讨ET-1和局部炎症后表皮伤害感受器TRPV 1快速增加的机制。2.研究皮肤中ETA受体分布的快速变化（炎症和ET- 1递送后）与对局部ET-1的反应脱敏之间的时间关系。3.确定皮肤细胞中的ET受体是否能够调节角质形成细胞TRPV 1和其他TRP通道（TRPV 3，TRPV 4）。4.探讨ET-1诱导的皮肤谷氨酸、CGRP和ATP的释放在ET-1引起的痛觉和触诱发痛中的作用。实验将整合来自行为、免疫细胞化学、生物化学和细胞生理学技术的结果，用于确定不同途径和物质对ET-1诱导的疼痛的重要性，用于定量ET-1诱导的TRPV受体和皮肤中的ET受体的变化，用于建立角质形成细胞中ET-R调节那些细胞中TRPV受体的能力，以及用于检查ET-1刺激从皮肤释放疼痛诱导物质的能力。

DESCRIPTION (provided by applicant): The endogenous peptide endothelin-1 (ET-1) is essential for pain and the sensitization of pain fibers to non-noxious stimuli after nerve injury, incision, inflammation and in cancer. In this proposal we seek to understand the relationship between the elevation of ET-1 in skin after injury and inflammation and the ensuing pain. Recent work suggests that ET-1 causes elevated pain in the periphery by acting on tissues that surround nerve endings, rather than directly on nerves; ET-1 and its cognate, GPC receptors ETA and ETB are elevated in conditions of hyperalgesia and allodynia. Antagonists of the ET receptors are able to prevent or reverse these pains, testifying to an important role for endogenous ET-1 in driving chronic pain states. Current evidence indicates that ET-1's actions during cutaneous injury/inflammation is closely linked to three other molecules/ receptors; TRPV1 (the receptor for hot peppers that is also activated by heat and by a H+), glutamate, and CGRP. TRPV1's participation in cutaneous pain from ET-1 or CFA injection into the rat's paw is evident within a very short time and immunocytochemical staining of TRPV1 in cutaneous nerve fibers increases 5-10-fold. In ~30 min after ET-1 or CFA injection, glutamate and CGRP contribute to ET-1's pain sensitizing effects; these substances are also released by cultured sensory neurons in a way that is potentiated by ET-1. This proposal contains 4 Specific Aims to address the processes underlying these changes: 1. To determine the mechanism for the rapid increase in TRPV1 in epidermal nociceptors following injection of ET-1 and local inflammation. 2. To investigate the temporal relationship between the rapid changes in ETA receptor distribution in skin (after inflammation and ET- 1 delivery) and the resulting desensitization of responses to local ET-1. 3. To determine if the ET receptors in skin cells are able to modulate keratinocye TRPV1, and other TRP channels (TRPV3, TRPV4). 4. To evaluate the role of ET-1-induced release of cutaneous glutamate, CGRP and ATP in ET-1- induced algesia and allodynia. Experiments will integrate the results from behavioral, immunocytochemical, biochemical and cell physiological techniques, for determining the importance of different pathways and substances for ET-1-induced pain, for quantitating ET-1-induced changes in TRPV receptors and ET receptors in skin, for establishing the ability of ET-Rs in keratinocytes to modulate TRPV receptors in those cells, and for examining ET-1's ability to stimulate release of pain-inducing substances from the skin.