人们对三叉神经系统内的疼痛，尤其是牙痛知之甚少。本研究旨在确定单个或多个牙髓损伤是否会引起持续性疼痛、其与三叉神经中枢伤害性通路的关系以及电针 (EA) 是否在持续性牙痛模型中提供持久的镇痛和神经保护作用。使用单牙髓损伤（SDPI）和多发牙髓损伤（MDPI）模型来诱导三叉神经病理性疼痛。使用机械退头阈值（HWT）评估牙痛相关行为的迹象。免疫荧光和蛋白质印迹方案用于监测星形胶质细胞活化、凋亡相关蛋白的变化以及 GABA 能中间神经元可塑性。 SDPI 小鼠的 HWT 从第 1 天到第 14 天表现出最初的显着下降，随后从第 21 天到第 42 天逐步恢复。从第 49 天到第 70 天，HWT 增加并恢复到对照值。相比之下，MDPI 小鼠的 HWT 从第 1 天到第 70 天持续下降。MDPI 增加了 Vi/Vc 过渡区的胶质纤维酸性蛋白 (GFAP) 并降低了谷氨酰胺合成酶 (GS) 和谷氨酸转运蛋白 1 (GLT1) 的表达。在 SDPI 后第 70 天脑干中未观察到星形胶质细胞标记物的变化，而在 SDPI 后第 70 天未观察到星形胶质细胞标记物的变化。术后第 70 天，观察到裂解的半胱氨酸天冬氨酸蛋白酶 3（裂解的 caspase-3）和 Bcl-2 相关 X 蛋白 (Bax) 的表达增加，同时 B 细胞淋巴瘤/白血病 2 (Bcl-2) 的表达减少。 MDPI 但不在 SDPI 之后。仅在 MDPI 后第 70 天观察到谷氨酸脱羧酶 (GAD65) 表达下调。 MDPI 引起的从第 1 天到第 70 天的 HWT 降低的影响通过 12 个疗程的 EA 治疗（MDPI 后第 1 到 21 天）而减弱。 MDPI 70 天后观察到的星形胶质细胞 GFAP、GS 和 GLT-1 以及裂解的 caspase-3、Bax、Bcl-2 和 GAD65 表达的变化通过 EA 治疗得以逆转。结果表明，小鼠持续性牙痛是由 MDPI 引起的，而不是由 SDPI 引起的。这种效应与三叉神经 GABA 能中间神经元可塑性以及星形胶质细胞的形态和功能变化有关。 EA 发挥持久的镇痛和神经保护作用，可能与神经元-胶质细胞串扰机制的调节有关。版权所有 © 2023 Ballon Romero、Fuh、Hung、Lee、Huang、Chien 和 Chen。

Pain within the trigeminal system, particularly dental pain, is poorly understood. This study aimed to determine whether single or multiple dental pulp injuries induce persistent pain, its association with trigeminal central nociceptive pathways and whether electroacupuncture (EA) provides prolonged analgesic and neuroprotective effects in a persistent dental pain model. Models of single dental pulp injury (SDPI) and multiple dental pulp injuries (MDPI) were used to induce trigeminal neuropathic pain. The signs of dental pain-related behavior were assessed using the mechanical head withdrawal threshold (HWT). Immunofluorescence and western blot protocols were used to monitor astrocyte activation, changes in apoptosis-related proteins, and GABAergic interneuron plasticity. SDPI mice exhibited an initial marked decrease in HWT from days one to 14, followed by progressive recovery from days 21 to 42. From days 49 to 70, the HWT increased and returned to the control values. In contrast, MDPI mice showed a persistent decrease in HWT from days one to 70. MDPI increased glial fibrillary acidic protein (GFAP) and decreased glutamine synthetase (GS) and glutamate transporter-1 (GLT1) expression in the Vi/Vc transition zone of the brainstem on day 70, whereas no changes in astrocytic markers were observed on day 70 after SDPI. Increased expression of cleaved cysteine-aspartic protease-3 (cleaved caspase-3) and Bcl-2-associated X protein (Bax), along with decreased B-cell lymphoma/leukemia 2 (Bcl-2), were observed at day 70 after MDPI but not after SDPI. The downregulation of glutamic acid decarboxylase (GAD65) expression was observed on day 70 only after MDPI. The effects of MDPI-induced lower HWT from days one to 70 were attenuated by 12 sessions of EA treatment (days one to 21 after MDPI). Changes in astrocytic GFAP, GS, and GLT-1, along with cleaved caspase-3, Bax, Bcl-2, and GAD65 expression observed 70 days after MDPI, were reversed by EA treatment. The results suggest that persistent dental pain in mice was induced by MDPI but not by SDPI. This effect was associated with trigeminal GABAergic interneuron plasticity along with morphological and functional changes in astrocytes. EA exerts prolonged analgesic and neuroprotective effects that might be associated with the modulation of neuron-glia crosstalk mechanisms.Copyright © 2023 Ballon Romero, Fuh, Hung, Lee, Huang, Chien and Chen.