Abstract:
Drive from peripheral neurons is essential in almost all pain states, but pharmacological silencing of these neurons to effect analgesia has proved problematic. Reversible gene therapy using long-lived chemogenetic approaches is an appealing option. We used the genetically activated chloride channel PSAM4-GlyR to examine pain pathways in mice. Using recombinant AAV9-based delivery to sensory neurons, we found a reversal of acute pain behavior and diminished neuronal activity using in vitro and in vivo GCaMP imaging on activation of PSAM4-GlyR with varenicline. A significant reduction in inflammatory heat hyperalgesia and oxaliplatin-induced cold allodynia was also observed. Importantly, there was no impairment of motor coordination, but innocuous von Frey sensation was inhibited. We generated a transgenic mouse that expresses a CAG-driven FLExed PSAM4-GlyR downstream of the Rosa26 locus that requires Cre recombinase to enable the expression of PSAM4-GlyR and tdTomato. We used NaV1.8 Cre to examine the role of predominantly nociceptive NaV1.8+ neurons in cancer-induced bone pain (CIBP) and neuropathic pain caused by chronic constriction injury (CCI). Varenicline activation of PSAM4-GlyR in NaV1.8-positive neurons reversed CCI-driven mechanical, thermal, and cold sensitivity. Additionally, varenicline treatment of mice with CIBP expressing PSAM4-GlyR in NaV1.8+ sensory neurons reversed cancer pain as assessed by weight-bearing. Moreover, when these mice were subjected to acute pain assays, an elevation in withdrawal thresholds to noxious mechanical and thermal stimuli was detected, but innocuous mechanical sensations remained unaffected. These studies confirm the utility of PSAM4-GlyR chemogenetic silencing in chronic pain states for mechanistic analysis and potential future therapeutic use.
摘要:
来自外周神经元的驱动在几乎所有疼痛状态下都是必不可少的,但是这些神经元的药理学沉默以实现镇痛已被证明是有问题的。使用长寿命化学遗传学方法的可逆基因治疗是一个吸引人的选择。我们使用遗传激活的氯通道PSAM4-GlyR来检查小鼠的疼痛途径。使用基于重组AAV9的递送至感觉神经元,我们发现,在varenicline激活PSAM4-GlyR后,使用体外和体内GCaMP成像,急性疼痛行为逆转,神经元活性降低.还观察到炎性热痛觉过敏和奥沙利铂诱导的冷异常性疼痛的显著降低。重要的是,运动协调没有损害,但无害的冯·弗雷的感觉被抑制了.我们产生了转基因小鼠,其在Rosa26基因座下游表达CAG驱动的FLExedPSAM4-GlyR,其需要Cre重组酶以使得能够表达PSAM4-GlyR和tdTomato。我们使用NaV1.8Cre检查了主要的伤害性NaV1.8神经元在癌症诱导的骨痛(CIBP)和慢性收缩损伤(CCI)引起的神经性疼痛中的作用。NaV1.8阳性神经元中PSAM4-GlyR的伐尼克兰激活逆转了CCI驱动的机械,热,冷敏感性。此外,通过负重评估,用NaV1.8感觉神经元中表达PSAM4-GlyR的CIBP治疗小鼠可逆转癌症疼痛。此外,当这些小鼠接受急性疼痛检测时,检测到对有害机械和热刺激的戒断阈值升高,但无害的机械感觉没有受到影响。这些研究证实了PSAM4-GlyR化学遗传沉默在慢性疼痛状态中用于机理分析和潜在的未来治疗用途的实用性。重要性陈述慢性疼痛是一个巨大的问题。周围神经阻滞在许多慢性疼痛情况下是有效的,证明外周驱动在慢性疼痛中的重要性。我们使用基于修饰的配体门控氯通道PSAM4-GlyR的化学遗传学工具在体外和体内沉默背根神经节神经元。这种方法减少了急性和慢性疼痛模型中的疼痛样行为,包括抗性疼痛,如神经性疼痛或癌症引起的骨痛。我们产生了一个以Cre依赖性方式表达PSAM4-GlyR的小鼠系,提供了一种有用的研究工具,不仅可以解决疼痛状态中伤害性感觉神经元的作用,还可以解决在正常和病理条件下整个神经系统中遗传定义的神经元集的功能。