Neohesperidin Alleviates the Neuropathic Pain Behavior of Rats by Downregulating the P2X4 Receptor.

Neuropathic pain (NP) is a type of chronic pain affecting 6-8% of human health as no effective drug exists. The purinergic 2X4 receptor (P2X4R) is involved in NP. Neohesperidin (NH) is a dihydroflavonoside compound, which has anti-inflammatory and antioxidative properties. This study aimed to investigate whether NH has an effect on P2X4R-mediated NP induced by chronic constriction injury (CCI) of the sciatic nerve in rats. In this study, the CCI rat model was established to observe the changes of pain behaviors, P2X4R, and satellite glial cells (SGCs) activation in dorsal root ganglion (DRG) after NH treatment by using RT-PCR, immunofluorescence double labeling and Western blotting. Our results showed CCI rats had mechanical and thermal hyperalgesia with an increased level of P2X4R. Furthermore, SGCs were activated as indicated by increased expression of glial fibrillary acidic protein and increased tumor necrosis factor-alpha receptor 1and interleukin-1ß. In addition, phosphorylated extracellular regulated protein kinases and interferon regulatory factor 5 in CCI rats increased. After NH treatment in CCI rats, the levels of above protein decreased, and the pain reduced. Overall, NH can markedly alleviate NP by reducing P2X4R expression and SGCs activation in DRG.

Astragalin Alleviates Neuropathic Pain by Suppressing P2X4-Mediated Signaling in the Dorsal Root Ganglia of Rats.

Neurologic damage often leads to neuropathic pain, for which there are no effective treatments owing to its complex pathogenesis. The purinergic receptor P2X4 is closely associated with neuropathic pain. Astragalin (AST), a compound that is used in traditional Chinese medicine, has protective effects against allergic dermatitis and neuronal injury, but its mechanism of action is not well understood. The present study investigated whether AST can alleviate neuropathic pain in a rat model established by chronic constriction injury (CCI) to the sciatic nerve. The model rats exhibited pain behavior and showed increased expression of P2X4 and the activated satellite glial cell (SGC) marker glial fibrillary acidic protein in dorsal root ganglia (DRG). AST treatment partly abrogated the upregulation of P2X4, inhibited SGC activation, and alleviated pain behavior in CCI rats; it also suppressed ATP-activated currents in HEK293 cells overexpressing P2X4. These data demonstrate that AST relieves neuropathic pain by inhibiting P2X4 and SGC activation in DRG, highlighting its therapeutic potential for clinical pain management.

Exploring the molecular mechanism of the effect of puerarin on P2X3.

P2X3 is a ligand-gated nonselective cation channel and permeable to Na+, K+ and Ca2+. Adenosine triphosphate (ATP) activation of the P2X3 on primary sensory ganglion neurons is involved in nociceptive transmission. Puerarin is a major active ingredient extracted from the traditional Chinese medicine Ge-gen. Puerarin inhibits nociceptive signal transmission by inhibiting the P2X3 in the dorsal root ganglia (DRG) and sympathetic ganglia, but its molecular mechanism is unclear. The aim of this study was to explore the molecular mechanism of puerarin on the P2X3. Here, molecular docking results revealed that puerarin binds well to the human P2X3 protein in the vicinity of the ATP binding pocket. Protein-ligand docking showed that the V64A mutation reduced the effect of puerarin but had little effect on ATP. V64A site-directed mutagenesis of P2X3 was performed using an overlap extension PCR technique. The wild-type and V64A mutant pEGFP-C1-P2X3 recombinant plasmids were transfected into HEK 293 cells. The electrophysiology results demonstrated that puerarin exerted an obvious inhibitory effect on ATP-activated currents in HEK 293 cells transfected with the wild-type P2X3, while little inhibition was observed in HEK 293 cells transfected with the mutant P2X3. These studies suggest that puerarin inhibits the P2X3 by binding to V64A.