DREADDed microglia in pain: Implications for spinal inflammatory signaling in male rats
DREADDed microglia in pain: Implications for spinal inflammatory signaling in male rats
复制标题
疼痛中可怕的小胶质细胞：对雄性大鼠脊髓炎症信号的影响。
DOI：
10.1016/j.expneurol.2018.03.005
复制
发表时间：
2018-06-01
影响因子：
5.3
通讯作者：
Watkins, Linda R.
中科院分区：
文献类型：
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作者：
Grace, Peter M.;Wang, Xiaohui;Watkins, Linda R.
选择性药理学工具的缺乏是小胶质细胞体内研究的主要障碍。为了解决这个问题，我们开发了G（q）-和G（i）-偶联的设计者受体，其由设计者药物（DREADD）独家激活，以分别选择性刺激或抑制小胶质细胞。通过AAV 9载体鞘内转染CD 68（小胶质细胞/巨噬细胞）启动子下的DREADD。鞘内转染Gq（刺激性）DREADD的幼稚雄性大鼠在鞘内给予DREADD选择性配体氯氮平-N-氧化物（CNO）后表现出显著的异常性疼痛，鞘内给予白细胞介素-1受体拮抗剂可消除这种疼痛。在鞘内转染G（i）（抑制性）DREADDs的雄性大鼠中，鞘内注射CNO可减轻慢性压迫性损伤诱导的异常性疼痛。为了探索机制，在体外用G（q）或G（i）DREADD稳定转染BV 2细胞。CNO处理诱导表达G（q）-DREADD的细胞产生促炎介质本身，并抑制表达G（i）DREADD的细胞产生脂多糖和CCL 2诱导的炎症信号。这些研究是第一个使用DREADD操纵小胶质细胞功能的研究，它可以最终证明胶质细胞在疼痛中的作用，不受所有其他抑制胶质细胞的药物存在的神经元脱靶效应的混淆。因此，这些研究是第一个最终证明完整脊髓中驻留的脊髓小胶质细胞的体内刺激a）对于异常性疼痛是足够的，并且B）对于由外周神经损伤诱导的异常性疼痛是必需的。DREADDs是一种独特的工具，选择性地探索小胶质细胞在体内的生理和病理作用。
The absence of selective pharmacological tools is a major barrier to the in vivo study of microglia. To address this issue, we developed a G(q)-and G(i)-coupled Designer Receptor Exclusively Activated by a Designer Drug (DREADD) to enable selective stimulation or inhibition of microglia, respectively. DREADDs under a CD68 (microglia/ macrophage) promoter were intrathecally transfected via an AAV9 vector. Naive male rats intrathecally transfected with Gq (stimulatory) DREADDs exhibited significant allodynia following intrathecal administration of the DREADD-selective ligand clozapine-N-oxide (CNO), which was abolished by intrathecal interleukin-1 receptor antagonist. Chronic constriction injury-induced allodynia was attenuated by intrathecal CNO in male rats intrathecally transfected with G(i) (inhibitory) DREADDs. To explore mechanisms, BV2 cells were stably transfected with G(q) or G(i) DREADDs in vitro. CNO treatment induced pro-inflammatory mediator production per se from cells expressing G(q)-DREADDs, and inhibited lipopolysaccharide-and CCL2-induced inflammatory signaling from cells expressing G(i) DREADDs. These studies are the first to manipulate microglia function using DREADDs, which allow the role of glia in pain to be conclusively demonstrated, unconfounded by neuronal off-target effects that exist for all other drugs that also inhibit glia. Hence, these studies are the first to conclusively demonstrate that in vivo stimulation of resident spinal microglia in intact spinal cord is a) sufficient for allodynia, and b) necessary for allodynia induced by peripheral nerve injury. DREADDs are a unique tool to selectively explore the physiological and pathological role of microglia in vivo.