Bilirubin Induces Pain Desensitization in Cholestasis by Activating 5-Hydroxytryptamine 3A Receptor in Spinal Cord.

Erliang Kong,Hongqian Wang,Weifeng Yu,Xudong Feng,Yuming Sun,Feixiang Wu,Yan Zhang,Xiaoqiang Wang,Jinmin Zhang

doi:10.3389/fcell.2021.605855

Abstract

BackgroundCholestasis patients often suffer from pain desensitization, resulting in serious complications in perioperative period. This study was aim to investigate the mechanism of bilirubin in cholestasis mediating pain desensitization through 5-hydroxytryptamine 3A (5-HT3A) receptor activation in spinal dorsal horn (SDH).MethodsA cholestasis model was established by bile duct ligation (BDL) in rats. Pain thresholds of rats were measured after BDL or intrathecally injecting bilirubin in the presence or absence of agonist (mCPBG) and antagonists (ondansetron, bicuculline, or CGP55845). Expression of 5-HT3 receptors, and the affinity and binding mode of bilirubin to 5-HT3A receptor were determined. Effects of bilirubin on γ-aminobutyric acid (GABA) pathway and the interactions with 5-HT3A receptor were tested.ResultsBilirubin was elevated significantly in both serum and CSF in BDL rats, accompanied with the up-regulation of pain thresholds. Both of 5-HT3A receptor and GABAA receptor antagonists could reverse the increased pain threshold in BDL rats. Further, 5-HT3A and GABAA receptor expressions were increased in BDL rats or intervention with bilirubin. Molecular docking suggested that bilirubin entered the hydrophobic pocket pre-formed in 5-HT3A receptor with potential hydrogen bonding. Bilirubin also increased GABA concentrations in CSF and GABAergic spontaneous inhibitory postsynaptic current in spinal cord, and directly induced inward currents in HEK293 cells which were overexpressed 5-HT3A receptor by lentivirus.ConclusionIn conclusion, bilirubin induced pain desensitization in cholestasis by activating 5-HT3A receptor in spinal cord. The activation of 5-HT3A receptor might regulate pain threshold by acting on the GABA pathway.

Highlights

MATERIALS AND METHODSCholestasis is characterized by the obstruction of intrahepatic/extrahepatic bile ducts by stones, tumors, or pancreatic diseases (Hegade et al, 2015)
Expressions of 5-HT3 receptors in the spinal cords of bile duct ligation (BDL) rats were determined by western blotting, and the results showed that only the 5-hydroxytryptamine 3A (5-HT3A) receptor expression increased significantly, especially in BDL-3d and BDL-5d, whereas expressions of 5-HT3B, 5-HT3C, 5-HT3D, and 5-HT3E receptors did not change (Figures 2A,B)
Results showed that GABA concentrations in cerebrospinal fluid (CSF) were significantly higher in BDL group (Figure 6G) and in the intrathecally administered bilirubin (1000 μM) group (Figure 6H) than that in control group, and ondansetron could reverse the increase of GABA concentrations

Summary

Introduction

MATERIALS AND METHODSCholestasis is characterized by the obstruction of intrahepatic/extrahepatic bile ducts by stones, tumors, or pancreatic diseases (Hegade et al, 2015). Exploration of the mechanism of pain desensitization in cholestasis can provide theoretical basis and treatment targets for cholestasis. Multiple substances such as 5-hydroxytryptamine (5-HT), endogenous opioids, and cholecystokinin participate in pain transmission in cholestasis (Hasanein and Javanmardi, 2008). Activation of 5-HT3 receptors in SDH produces an anti-nociceptive response, whereas the antagonists of 5-HT3 receptors can reverse this effect. This study was aim to investigate the mechanism of bilirubin in cholestasis mediating pain desensitization through 5-hydroxytryptamine 3A (5-HT3A) receptor activation in spinal dorsal horn (SDH)

Results

Discussion

Conclusion