Department of Biomedical Sciences, University of New England, Biddeford, Maine 04005.
Department of Biomedical Sciences, University of New England, Biddeford, Maine 04005
J Neurosci. 2021 Mar 31;41(13):2883-2898. doi: 10.1523/JNEUROSCI.2368-20.2021. Epub 2021 Feb 16.
G-coupled receptors signaling through cAMP provide a key mechanism for the sensitization of nociceptive sensory neurons, and the cAMP effector Epac has been implicated in the transition from acute to chronic pain. Epac exerts its effects through Rap1 and protein kinase C (PKC). To identify targets of Epac-PKC signaling in sensory neurons of the mouse dorsal root ganglion (DRG), we profiled PKC substrate proteins phosphorylated in response to the activation of Epac with the proinflammatory prostaglandin E2 (PGE2). A prominent Epac-dependent phospho-protein band induced by PGE2 was identified by mass spectrometry as the mitochondrial enzyme pyruvate dehydrogenase (Pdha1). In dissociated DRG from both males and females, the recruitment of Pdha1 to phospho-protein fractions was rapidly induced by PGE2 and prevented by selective inhibition of Epac2. Epac activation increased mitochondrial respiration, consistent with an increase in Pdha1 function mediated by Epac2. Hindpaw injection of PGE2 induced heat hyperalgesia in males and females, but Pdha1 phosphorylation occurred only in males. Hyperalgesia was attenuated in males but not in females by systemic inhibition of Epac2, and also by a mitochondrial membrane potential uncoupler, dinitrophenol, supporting a role for mitochondrial regulation in acute hyperalgesia. These findings identify a mechanism for the regulation of mitochondrial function by Epac2 that contributes to acute inflammatory hyperalgesia in male mice. Systemic administration of the cyclooxygenase 2 inhibitor celecoxib suppressed both PGE2-induced heat hyperalgesia and Pdha1 phosphorylation in DRG of males but not females, suggesting that prostaglandin synthesis within the DRG mediates the phosphorylation of Pdha1 in response to hindpaw insult. There has been extensive investigation of mitochondrial dysfunction as a causative factor in neuropathic pain disorders. In contrast, results reported here implicate enhanced mitochondrial function as a contributing factor in the development of acute inflammatory hyperalgesia. We describe a mechanism in which Epac2 activation by prostaglandin receptors leads to phosphorylation of pyruvate dehydrogenase and an increase in mitochondrial respiration in peripheral sensory neurons. Although Epac2 activation leads to Pdha1 (pyruvate dehydrogenase) phosphorylation in dissociated neurons from mice of both sexes, induction of this pathway by hindpaw insult is restricted to males and appears to require intraganglionic prostaglandin synthesis. These findings support a model in which G-coupled receptor modulation of mitochondrial function promotes acute nociceptive signaling and inflammatory hyperalgesia.
G 蛋白偶联受体通过 cAMP 信号转导为伤害感受感觉神经元的敏化提供了一个关键机制，而 cAMP 效应物 Epac 已被牵连到急性痛向慢性痛的转变。Epac 通过 Rap1 和蛋白激酶 C（PKC）发挥作用。为了确定 Epac-PKC 信号在小鼠背根神经节（DRG）感觉神经元中的作用靶点，我们通过用促炎前列腺素 E2（PGE2）激活 Epac，对磷酸化的 PKC 底物蛋白进行了蛋白质谱分析。通过质谱分析，由 PGE2 诱导的 Epac 依赖性磷酸化蛋白条带被鉴定为线粒体酶丙酮酸脱氢酶（Pdha1）。在来自雄性和雌性的分离的 DRG 中，PGE2 迅速诱导 Pdha1 募集到磷酸蛋白级分，并被 Epac2 的选择性抑制所阻止。Epac 激活增加了线粒体呼吸，这与 Epac2 介导的 Pdha1 功能增加一致。在雄性和雌性动物的后爪注射 PGE2 会引起热痛觉过敏，但只有雄性动物发生 Pdha1 磷酸化。在雄性动物中，全身性抑制 Epac2 或使用线粒体膜电位解偶联剂二硝基苯酚均可减轻 Hyperalgesia，而在雌性动物中则没有，这支持了线粒体调节在急性痛觉过敏中的作用。这些发现确定了 Epac2 调节线粒体功能的机制，该机制有助于雄性小鼠的急性炎症性痛觉过敏。全身性给予环氧化酶 2 抑制剂塞来昔布可抑制雄性 DRG 中 PGE2 诱导的热痛觉过敏和 Pdha1 磷酸化，但不能抑制雌性动物，这表明 DRG 内的前列腺素合成介导了后爪损伤时 Pdha1 的磷酸化。已经对线粒体功能障碍作为神经病理性疼痛疾病的致病因素进行了广泛的研究。相比之下，这里报告的结果表明，增强的线粒体功能是急性炎症性痛觉过敏发展的一个促成因素。我们描述了一种机制，即通过前列腺素受体激活 Epac2 导致丙酮酸脱氢酶磷酸化，并增加外周感觉神经元中的线粒体呼吸。尽管 Epac2 激活会导致两性分离神经元中的 Pdha1（丙酮酸脱氢酶）磷酸化，但这种途径的诱导仅限于雄性动物，似乎需要神经节内的前列腺素合成。这些发现支持这样一种模型，即 G 蛋白偶联受体对线粒体功能的调节促进了急性伤害性信号和炎症性痛觉过敏。
