Lin Mengjuan, Hu Guiying, Wang Zhengqiang, Yu Baoping, Tan Wei
Department of Gastroenterology, Renmin Hospital of Wuhan University, No. 238 Jiefang Rd, Wuhan, 430060, Hubei, People's Republic of China.
Key Laboratory of Hubei Province for Digestive System Diseases, Wuhan, People's Republic of China.
Dig Dis Sci. 2023 Jun;68(6):2414-2426. doi: 10.1007/s10620-022-07765-5. Epub 2023 Feb 18.
The mechanism underlying irritable bowel syndrome (IBS), a common disease with hyperalgesia, remains elusive. The spinal cholinergic system is involved in pain modulation, but its role in IBS is unknown.
To determine whether high-affinity choline transporter 1 (CHT1, a major determinant of the cholinergic signaling capacity), is implicated in spinal modulation of stress-induced hyperalgesia.
A rat IBS model was established by water avoidance stress (WAS). Visceral sensations were detected by abdominal withdrawal reflex (AWR) and visceromotor response (VMR) to colorectal distension (CRD). Abdominal mechanical sensitivity was determined by von Frey filaments (VFFs) test. RT-PCR, Western blot, and immunostaining were performed for spinal CHT1 expression. Spinal acetylcholine (ACh) was measured by ELISA; the influence of spinal CHT1 on hyperalgesia were evaluated by intrathecal administration of MKC-231 (a choline uptake enhancer) and hemicholinium-3 (HC-3, a specific inhibitor of CHT1). Minocycline treatment was used to explore the role of spinal microglia in hyperalgesia.
After 10 days of WAS, AWR scores and VMR magnitude to CRD, and the number of withdrawal events in VFF test were increased. Double-labeling showed that CHT1 in the dorsal horn was expressed in most of the neurons and almost all the microglia. The CHT1 expression and ACh levels in the spinal cord and the density of CHT1-positive cell in the spinal dorsal horn were enhanced in WAS-exposed rats. HC-3 enhanced pain responses in WAS rats; MKC-231 alleviated pain in WAS rats by upregulating CHT1 expression and increasing ACh production in the spinal cord. Furthermore, microglial activation in the spinal dorsal horn promoted the stress-induced hyperalgesia, and MKC-231 achieved analgesic effects by inhibiting the spinal microglial activation.
CHT1 exerts antinociceptive effects in spinal modulation of chronic stress-induced hyperalgesia by increasing ACh synthesis and suppressing microglial activation. MKC-231 has potential for treating disorders accompanied by hyperalgesia.
肠易激综合征（IBS）是一种伴有痛觉过敏的常见疾病，其潜在机制仍不清楚。脊髓胆碱能系统参与疼痛调节，但其在IBS中的作用尚不清楚。
确定高亲和力胆碱转运体1（CHT1，胆碱能信号传导能力的主要决定因素）是否参与应激诱导的痛觉过敏的脊髓调节。
通过水回避应激（WAS）建立大鼠IBS模型。通过腹部退缩反射（AWR）和对结直肠扩张（CRD）的内脏运动反应（VMR）检测内脏感觉。通过von Frey细丝（VFFs）试验测定腹部机械敏感性。对脊髓CHT1表达进行逆转录聚合酶链反应（RT-PCR）、蛋白质免疫印迹法（Western blot）和免疫染色。通过酶联免疫吸附测定（ELISA）测量脊髓乙酰胆碱（ACh）；通过鞘内注射MKC-231（一种胆碱摄取增强剂）和半胱胺（HC-3，CHT1的特异性抑制剂）评估脊髓CHT1对痛觉过敏的影响。使用米诺环素治疗来探究脊髓小胶质细胞在痛觉过敏中的作用。
WAS处理10天后，对CRD的AWR评分和VMR幅度以及VFF试验中的退缩事件数量增加。双重标记显示，背角中的CHT1在大多数神经元和几乎所有小胶质细胞中表达。WAS处理的大鼠脊髓中CHT1表达、ACh水平以及脊髓背角中CHT1阳性细胞的密度均升高。HC-3增强了WAS大鼠的疼痛反应；MKC-231通过上调CHT1表达和增加脊髓中ACh的产生减轻了WAS大鼠的疼痛。此外，脊髓背角中的小胶质细胞激活促进了应激诱导的痛觉过敏，MKC-231通过抑制脊髓小胶质细胞激活实现镇痛作用。
CHT1通过增加ACh合成和抑制小胶质细胞激活，在慢性应激诱导的痛觉过敏的脊髓调节中发挥镇痛作用。MKC-231具有治疗伴有痛觉过敏疾病的潜力。
