腹外侧眶皮层内5-HT6受体参与神经病理性痛与抑郁共病调控的机制研究

The comorbidity of neuropathic pain and depression is a high clinical disease. Since the mechanism of their coexistence is unclear, it is still lack of effective drugs. The ventrolateral orbital cortex (VLO) is involved in the modulation of chronic pain and depression, and 5-HT6 receptors are a new target for the treatment of depression. Therefore, the project is designed to examine whether 5-HT6 receptors in the VLO are involved in modulating nociception and depression in the spared nerve injury-induced neuropathic pain model (SNI) in rats and mice. Using behavioral test, in vivo electrophysiological methods and neuropharmacological methods, we will observe the effect of VLO microinjection of 5-HT6 receptor agonist on SNI-induced paw withdraw threshold and depression behavior, as well as, the effect of 5-HT6 receptor antagonist or glutamate receptor antagonist. By using morphological method, we will examine the synapse connection of 5-HTergic terminals, 5-HT6 receptors, glutamatergic neurons and projecting neurons from the VLO to the periaquaeductal gray (PAG). The molecular biological and patch-clamp techniques will be used to observe the signal transduction mechanisms induced by activation of 5-HT6 receptors. These studies will provide new evidence for that, under the neuropathic pain state, activation of 5-HT6 receptors in the VLO may produce anti-neuropathic pain by directly activating the PAG-brainstem descending inhibitory system or by a presynaptic facilitation from excited the VLO neurons projecting to the PAG, leading to the activation of the PAG-brainstem descending inhibitory system and producing antinociceptive effect at the spinal cord level. Moreover, activation of 5-HT6 receptors in the VLO may produce anti-depression via activating the phosphorylation of MAPK pathway and increasing the levels of BNDF. This study will elucidate more mechanisms for the role of VLO in modulating the comorbidity of chronic pain and depression. Moreover, it provide active targets for developing the new drugs for reatment neuropathic pain and depression.

神经病理性痛与抑郁共病是临床高发病，机制不明且缺乏有效的治疗药物。腹外侧眶皮层（VLO）参与慢性痛与抑郁的调控且5-HT6受体为治疗抑郁的新靶点。因此，本项目以神经病理性痛大鼠和小鼠为对象，用行为学、在体电生理学和神经药理学技术观察激活VLO内5-HT6受体对神经病理性痛与抑郁共病的效应；用形态学方法研究VLO内5-HT能神经末梢、5-HT6受体与谷氨酸能神经元和投射神经元之间的联系；用分子生物学和膜片钳技术观察激活VLO内5-HT6受体产生镇痛和抗抑郁效应的胞内信号机制。以阐明激活VLO内5-HT6受体可能通过直接激活或突触前易化谷氨酸能突触传递间接激活PAG-脑干下行抑制系统实现镇痛效应，并通过激活胞内MAPK通路磷酸化水平进而上调BDNF表达产生抗神经病理性痛伴发的抑郁行为。为大脑皮层参与慢性痛与抑郁共病调制机制提供新资料，为开发新型抗神经病理性痛和抑郁的药物及作用靶点开拓思路。

神经病理性痛与抑郁共病是临床高发病，机制不明且缺乏有效的治疗药物。腹外侧眶皮层（VLO）参与慢性痛与抑郁的调控且5-HT6受体为治疗抑郁的新靶点。因此，本项目以神经病理性痛大鼠为对象，研究激活VLO内5-HT6受体对神经病理性痛与抑郁共病的效应及其作用机制。我们发现激活VLO的5-HT6受体可以抑制SNI神经病理性疼痛大鼠的触诱发痛。这一作用可被选择性5-HT6受体拮抗剂、AC、PKA抑制剂和非选择性谷氨酸拮抗剂翻转。免疫荧光双标染色结果显示：VLO内5-HT6受体在谷氨酸能神经元上大量表达。该结果证实激活VLO内5-HT6受体产生抗神经病理性疼痛的效应，这一效应是通过AC-cAMP-PKA信号通路调节VLO内谷氨酸能神经元的活动，进而激活PAG下行抑制系统实现镇痛效应。而且激活VLO内的5-HT6 受体也可以通过AC- cAMP-PKA通路参与下行易化炎性痛效应。进一步研究发现，SNI 手术4 周后，大鼠表现出显著的焦虑抑郁样行为，且VLO内5-HT6 受体、p-ERK/ERK 和p-CREB/CREB 表达均下降。激活VLO内 5-HT6 受体可减轻SNI大鼠的焦虑抑郁样行为，这一作用可被选择性5-HT6 受体拮抗剂、AC和PKA 抑制剂和 MEK1/2 抑制剂翻转，表明激活 VLO内5-HT6受体产生抗神经病理性痛诱发焦虑抑郁的效应，这一效应通过上调AC-cAMP- PKA信号通路磷酸化水平进而促进BDNF的表达水平的上调实现的。我们发现激活VLO内的谷氨酸受体可以产生抗神经病性痛效应，该效应主要由 non-NMDA 受体所介导，也可通过部分 NMDA 和代谢性谷氨酸受体介导。进步一我们证实大鼠 SNI 术后VLO 中AMPA受体表达上调，是主要参与VLO下行镇痛环路的谷氨酸受体。激活VLO 中的CPAR可以显著的产生镇痛作用，并且也能够显著的改善与慢性痛共病的抑郁焦虑样行为。以上研究结果说明，激活VLO内5-HT6受体可能通过直接激活或突触前易化谷氨酸能突触传递间接激活PAG-脑干下行抑制系统实现镇痛效应，并通过激活胞内MAPK通路磷酸化水平进而上调BDNF表达产生抗神经病理性痛伴发的抑郁行为。为大脑皮层参与慢性痛与抑郁共病调制机制提供新资料，为开发新型抗神经病理性痛和抑郁的药物及作用靶点开拓思路。