RVM CCK and Neuropathic Pain

描述（由申请人提供）：最近的证据表明，引发神经性疼痛的过程可能与维持这种疼痛的过程不同。持续性神经性疼痛状态需要由延髓头端腹内侧（RVM）引起的下行易化，这一发现表明，在损伤后第6天（及以后），但在损伤后第3天，注射到RVM中的利多卡因有效地阻断了实验性神经性疼痛。这种下行促进的开始可能是由表达μ阿片受体的RVM细胞的活性引起的，因为用皂草素-μ阿片缀合物选择性损伤这些细胞或手术损伤背外侧索（DLF）（一种通向背角的下行纤维通路）预防和逆转神经性疼痛的表达。这些发现暗示了损伤后RVM下行易化过程的时间依赖性激活，以维持而不是引发异常疼痛。驱动这种疼痛的RVM神经可塑性的性质尚不清楚。这一机制的一个线索是我们观察到RVM微量注射CCK 2受体拮抗剂产生可逆性阻断已建立的神经病理性疼痛。此外，将CCK-8（S）微量注射到未受伤大鼠的RVM中产生触觉和热超敏反应，使人联想到神经损伤诱导的疼痛状态。这些作用被DLF或表达μ阿片受体的RVM细胞的损伤所阻止，这表明RVM CCK可能通过激活RVM神经元来促进损伤诱导的疼痛，所述RVM神经元由μ阿片受体的表达表型定义。基于这些原因，我们假设神经损伤导致RVM CCK活性的时间相关性增加，以驱动下行易化并维持神经损伤诱导的疼痛。这一假设将通过以下具体目标进行检验。目的1将表征（a）RVM中的基础CCK活性，以及幼稚大鼠中CCK活性对机械和热皮肤输入的响应变化，以及（B）神经损伤后RVM中CCK活性的时间依赖性。具体而言，将在脊神经结扎损伤后的早期和晚期时间点测量RVM中响应于机械和热皮肤输入的基础CCK释放和CCK释放。目的2将确定RVM CCK可能起作用以驱动下行易化的CCK受体。CCK和CCK 2受体类型在RVM损伤前和损伤后的功能作用将通过显微注射选择性CCK受体拮抗剂来评估。CCK受体的空间分布将通过半定量原位杂交和受体放射自显影进行分析。目的2还将研究CCK受体可能与μ阿片受体在RVM神经元中共定位的可能性。拟议的实验将揭示RVM可塑性的一些功能，可能是维持神经病理性疼痛的关键。由于患有神经性疼痛的患者可能在发生诱发性损伤后很长时间内寻求治疗，因此了解维持神经性状态的机制对于合理治疗干预的发展至关重要。在这方面，拟议的实验可能揭示了CCK受体拮抗剂作为治疗神经性疼痛的重要作用。

DESCRIPTION (provided by applicant): Recent evidence suggests that processes that initiate neuropathic pain may differ from those that maintain such pain. That persistent neuropathic pain states require descending facilitation arising from the rostral ventromedial medulla (RVM) is suggested by the finding that lidocaine injected into the RVM effectively blocks experimental neuropathic pain at post-in jury day 6 (and later), but not at post-injury day 3. Onset of this descending facilitation may arise from activity of RVM cells that express mu opioid receptors, since either a selective lesion of these cells with a saporin-mu opioid conjugate or surgical lesion of the dorsolateral funiculus (DLF), a descending fiber pathway to the dorsal horn, prevents and reverses the expression of neuropathic pain. These findings implicate a time-dependent activation of descending facilitatory processes in the RVM following injury to maintain, rather than initiate, abnormal pain. The nature of the RVM neuroplasticity that drives such pain is unknown. One clue to this mechanism is our observation that RVM microinjection of a CCK 2 receptor antagonist produces a reversible blockade of established neuropathic pain. In addition, microinjection of CCK-8(S) into the RVM of uninjured rats produces tactile and thermal hypersensitivity reminiscent of states of nerve-injury induced pain. These effects are prevented by lesions of the DLF, or of RVM cells expressing mu opioid receptors, suggesting the possibility that RVM CCK may promote injury- induced pain by activating RVM neurons that are phenotypically defined by the expression of mu opioid receptors. For these reasons, we hypothesize that nerve-injury results in a time-related increase in RVM CCK activity to drive descending facilitation and to maintain nerve-injury induced pain. This hypothesis will be tested by the following specific aims. Aim 1 will characterize (a) the basal CCK activity in the RVM, and changes in CCK activity in response to mechanical and thermal cutaneous inputs in naive rats, and (b) the time-dependent nature of CCK activity in the RVM following nerve injury. Specifically, basal CCK release, and CCK release in response to mechanical and thermal cutaneous inputs in the RVM will be measured at early and late time points after spinal nerve ligation injury. Aim 2 will identify the CCK receptor(s) at which RVM CCK may act to drive descending facilitation. The functional roles of CCK and CCK2 receptor types in the RVM pre- and post-injury will be evaluated pharmacologically by microinjection of selective CCK receptor antagonists. The spatial distribution of the CCK receptors will be analyzed by semi-quantitative in situ hybridization and receptor autoradiography. Aim 2 will also examine the possibility that CCK receptors may co-localize with mu opioid receptors in RVM neurons. The proposed experiments will reveal some features of the RVM plasticity that may be critical in maintaining neuropathic pain. As patients who suffer from neuropathic pain are likely to seek treatment long after the precipitating injury has occurred, understanding the mechanisms that maintain the neuropathic state will be essential for the development of rational therapeutic interventions. In this regard, the proposed experiments may reveal an important role for CCK receptor antagonists as therapy for neuropathic pain.