Mechanistic Study of Pain Inhibition by Activation of Non-nociceptive Afferent Fibers

项目摘要 电刺激低阈值Aβ神经纤维是治疗慢性疼痛的临床策略 包括鸦片类药物在内的药物治疗无效。然而，疼痛的潜在机制 通过Aβ-纤维激活的抑制仍然难以捉摸，限制了进一步的临床和技术改进。我们 主要目的是揭示Aβ-ES抑制疼痛的脊髓神经元和非神经元机制， 明确靶点，合理选择辅助药物，增强镇痛效果，避免副作用。在Aim中 1，通过使用神经病理性疼痛的动物模型，我们将首先描述脊髓神经元机制， Aβ-ES抑制疼痛传递。电生理学研究将揭示不同形式的Aβ-ES 调节浅表背角神经元的功能上不同的子集（例如，抑制性与兴奋性 神经元），并进一步区分潜在的受体机制。我们假设， 神经元优先抑制可能使Aβ-ES与低剂量辅助药物一起用于 回路特异性疼痛抑制增强。我们将测试是否限制内源性腺苷酸 在Aβ-ES过程中，GABA降解或GABA重摄取特异性地增强背角兴奋性抑制， 和投射神经元，并增加疼痛传递的净抑制。在目标2中，我们将推出一个 Aβ-ES激活的非神经痛门控机制。通过进行高通量GCaMP 6成像 和电生理记录，我们将研究Aβ-ES是否激活脊髓星形胶质细胞， 脊髓伤害性传递的抑制，以及该过程是否因神经损伤而减弱。以来 背柱刺激（DCS）的疼痛抑制与Aβ纤维的激活有内在联系，DCS将 可用作体内研究Aβ-ES的原理验证。我们将研究DCS是否会改变星形胶质细胞 反应性标记物，影响促炎介质的水平，并促进小胶质细胞极化， 神经损伤后M1至M2状态。我们将进一步测试胶质细胞衍生的腺苷是否抑制兴奋性或 增强Aβ-ES对投射神经元的抑制作用。我们在目标1和2中的发现 将有助于我们在目标3中开发增强Aβ-ES疼痛抑制的新策略。我们将测试 DCS对神经病理性疼痛相关行为的抑制可以通过限制 内源性腺苷和抑制脊髓中GABA再摄取。由于依赖于使用， 电路特定的功能，我们预计治疗不会引起副作用。我们的发现将揭示 本研究为Aβ-ES的镇痛作用提供了新的机制，并为Aβ-ES的镇痛作用提供了重要的理论基础和药物靶点 未来的转化研究旨在使用低剂量的辅助药物，以提高疗效和特异性 Aβ-ES疗法的疼痛抑制作用。

PROJECT SUMMARY Electrical stimulation (ES) of low-threshold Aβ-nerve fibers is a clinical strategy for treating chronic pain that is refractory to pharmacotherapies, including opiates. However, the mechanisms underlying pain inhibition by Aβ-fiber activation remain elusive, limiting further clinical and technological improvements. Our major goal is to uncover spinal neuronal and non-neuronal mechanisms of pain inhibition by Aβ-ES, and to identify targets for rationally selecting adjuvant drugs to enhance pain inhibition and avoid side effects. In Aim 1, by using animal models of neuropathic pain, we will first delineate spinal neuronal mechanisms by which Aβ-ES inhibits pain transmission. Electrophysiology studies will uncover how different forms of Aβ-ES modulate functionally distinct subsets of superficial dorsal horn neurons (e.g., inhibitory vs. excitatory neurons), and further differentiate the underlying receptor mechanisms. We postulate that the excitatory neuron-preferred inhibition may enable Aβ-ES to be used together with low-dose adjuvant drugs for circuitry-specific enhancement of pain inhibition. We will test whether limiting endogenous adenosine degradation or GABA reuptake during Aβ-ES specifically enhances the inhibition of dorsal horn excitatory and projection neurons and increases the net inhibition of pain transmission. In Aim 2, we will then unveil a non-neuronal pain gating mechanism activated by Aβ-ES. By conducting high-throughput GCaMP6 imaging and electrophysiology recording, we will examine whether Aβ-ES activates spinal astrocytes to induce inhibition of spinal nociceptive transmission, and whether this process is attenuated by nerve injury. Since pain inhibition by dorsal column stimulation (DCS) is intrinsically linked with activation of Aβ-fibers, DCS will be used as a proof-of-principle for studying Aβ-ES in vivo. We will examine whether DCS changes astrocyte reactive markers, affects the levels of pro-inflammatory mediators, and promotes microglial polarization from M1 to M2 state after nerve injury. We will further test whether glia-derived adenosine inhibits excitatory or projection neurons and enhances Aβ-ES–induced inhibition of these neurons. Our findings in Aims 1 and 2 will help us to develop new strategies of enhancing pain inhibition by Aβ-ES in Aim 3. We will test whether inhibition of neuropathic pain-related behavior by DCS can be enhanced by limiting degradation of endogenous adenosine and inhibiting GABA reuptake in the spinal cord. Owing to use-dependent and circuitry-specific features, we expect that the treatment will not induce side effects. Our findings will reveal new mechanisms underlying pain inhibition by Aβ-ES, and will provide important rationales and drug targets for future translational studies aimed at using low-dose adjuvant drugs to improve the efficacy and specificity of pain inhibition by Aβ-ES therapies.