Yishan Lei

Spinal activation of alpha7-nicotinic acetylcholine receptor attenuates posttraumatic stress disorder-related chronic pain via suppression of glial activation

The high prevalence of chronic pain in posttraumatic stress disorder (PTSD) individuals has been widely reported by clinical studies, which emphasized an urgent need to uncover the underlying mechanisms and identify potential therapeutic targets. Recent studies suggested that targeting activated glia and their pro-inflammatory products may provide a novel and effective therapy for the stress-related pain. In this study, we investigated whether activation of alpha-7 nicotinic acetylcholine receptor (α7 nAChR), a novel anti-inflammatory target, could attenuate PTSD-related chronic pain. The experiments were conducted in a rat model of single prolonged stress (SPS), an established model of PTSD-pain comorbidity. We found that SPS exposure produced persistent mechanical allodynia. Immunohistochemical and enzyme-linked immuno sorbent assay analysis showed that SPS also induced elevated activation of glia cells (including microglia and astrocytes) and accumulation of pro-inflammatory cytokines in spinal cord. In another experiment, we found that intrathecal injection of PHA-543613, a selective α7 nAchR agonist, attenuated the SPS-evoked allodynia in a dose dependent manner. However, this anti-hyperalgesic effect was blocked by pretreatment with methyllycaconitine (MLA), a selective α7 nAchR antagonist. Further analyses showed that PHA-543613 suppressed SPS-induced spinal glial activation and SPS-elevated spinal pro-inflammatory cytokines, and these were abolished by MLA. Taken together, the present study showed that spinal activation of α7 nAChR by PHA-543613 attenuated mechanical allodynia induced by PTSD-like stress, and the suppression of spinal glial activation may underlie this anti-hyperalgesic effect. Our study demonstrated the therapeutic potential of targeting α7 nAChR in the treatment of PTSD-related chronic pain.

The effects of dexmedetomidine pretreatment on the pro- and anti-inflammation systems after spinal cord injury in rats

Excessive inflammatory responses play important roles in the aggravation of secondary damage to an injured spinal cord. Dexmedetomidine (DEX), a selective α2-adrenoceptor agonist, has recently been implied to be neuroprotective in clinical anesthesia, but the underlying mechanism is elusive. As signaling through Toll-like receptor 4 (TLR4) and nicotinic receptors (nAChRs, notably α7nAChR) play important roles in the pro- and anti-inflammation systems in the central nervous system, respectively, this study investigated whether and how they were modulated by DEX pretreatment in a rat model of spinal cord compression. The model was used to mimic perioperative compressive spinal cord injury (SCI) during spinal correction. DEX preconditioning improved locomotor scores after SCI, which was accompanied by increased α7nAChR and acetylcholine (Ach, an endogenous ligand of α7nAChR) expression as well as PI3K/Akt activation. However, there was a decrease in Ly6h (a negative regulator for α7nAChR trafficking), TLR4, PU.1 (a critical transcriptional regulator of TLR4), HMGB1 (an endogenous ligand of TLR4), and caspase 3-positive cells, which was prevented by intrathecal preconditioning with antagonists of either α2R, α7nAChR or PI3K/Akt. In addition, application of an α7nAChR agonist produced effects similar to those of DEX after SCI, while application of an α7nAChR antagonist reversed these effects. Furthermore, both α7nAChR and TLR4 were mainly co-expressed in NeuN-positive cells of the spinal ventral horn, but not in microglia or astrocytes after SCI. These findings imply that the α2R/PI3K/Akt/Ly6h and α7nAChR/PI3K/Akt/PU.1 cascades are required for upregulated α7nAChR and downregulated TLR4 expression by DEX pretreatment, respectively, which provided a unique insight into understanding DEX-mediated neuroprotection.

Activated Glia Increased the Level of Proinflammatory Cytokines in a Resiniferatoxin-Induced Neuropathic Pain Rat Model.

Background and Objectives Administration of resiniferatoxin (RTX) can mimic the clinical symptoms of postherpetic neuralgia. However, it is unclear whether activated glia contribute to the pathogenesis of RTX-induced neuropathic pain; furthermore, the relationship between p38, N-methyl-D-aspartate receptor type 2B (NR2B) as well as proinflammatory cytokines and activated glia remains unknown. Methods Intraperitoneal injection of RTX was performed to induce neuropathic pain in rats. Mechanical allodynia and thermal hyperalgesia were assessed by von Frey filaments and a radiant heat stimulus, respectively. Western blot and immunofluorescence labeling examined the expression of NR2B, activated glia markers, p38, and proinflammatory cytokines in the spinal cord. We further investigated the effect of the glial inhibitors, fluorocitrate and minocycline, on nociceptive behaviors and expression of p38, NR2B, and proinflammatory cytokines. Results Resiniferatoxin leads to an increase of paw withdrawal latency to a heat stimulus and caused a mechanical allodynia within 2 weeks. The expression of tumor necrosis factor &agr;, IL-1&bgr;, p38, and NR2B was up-regulated in RTX-induced neuropathic pain rat model and lasted for at least 49 days. Microglia were activated at the early phase of the disease, whereas activated astrocytes were detected in the sustainment phase. Both minocycline and fluorocitrate attenuated the nociceptive behaviors and expression of related proteins. Conclusions Activated glia participate in the pathogenesis of RTX-induced neuropathic pain and are likely to be the source of proinflammatory cytokines. Inhibition of glia contributes to an analgesic effect. These findings provide a novel strategy for the treatment of postherpetic neuralgia.

Mas-Related Gene (Mrg) C Activation Attenuates Bone Cancer Pain via Modulating Gi and NR2B.

Objective This study is to investigate the role of Mas-related gene (Mrg) C in the pathogenesis and treatment of bone cancer pain (BCP). Methods BCP mouse model was established by osteosarcoma cell inoculation. Pain-related behaviors were assessed with the spontaneous lifting behavior test and mechanical allodynia test. Expression levels of MrgC, Gi, and NR2B in the spinal cord were detected with Western blot analysis and immunohistochemistry. Results Pain-related behavior tests showed significantly increased spontaneous flinches (NSF) and decreased paw withdrawal mechanical threshold (PWMT) in mouse models of BCP. Western blot analysis showed that, compared with the control group and before modeling, all the expression levels of MrgC, Gi, and NR2B in the spinal cord of BCP mice were dramatically elevated, which were especially increased at day 7 after operation and thereafter, in a time-dependent manner. Moreover, the treatment of MrgC agonist BAM8-22 significantly up-regulated Gi and down-regulated NR2B expression levels, in the spinal cord of BCP mice, in a time-dependent manner. On the other hand, anti-MrgC significantly down-regulated Gi expression, while dramatically up-regulated NR2B expression, in the BCP mice. Similar results were obtained from the immunohistochemical detection. Importantly, BAM8-22 significantly attenuated the nociceptive behaviors in the BCP mice. Conclusion Our results indicated the MrgC-mediated Gi and NR2B expression alterations in the BCP mice, which might contribute to the pain hypersensitivity. These findings may provide a novel strategy for the treatment of BCP in clinic.

Hippocampal activation of microglia may underlie the shared neurobiology of comorbid posttraumatic stress disorder and chronic pain.

The high comorbidity rates of posttraumatic stress disorder and chronic pain have been widely reported, but the underlying mechanisms remain unclear. Emerging evidence suggested that an excess of inflammatory immune activities in the hippocampus involved in the progression of both posttraumatic stress disorder and chronic pain. Considering that microglia are substrates underlying the initiation and propagation of the neuroimmune response, we hypothesized that stress-induced activation of hippocampal microglia may contribute to the pathogenesis of posttraumatic stress disorder-pain comorbidity. We showed that rats exposed to single prolonged stress, an established posttraumatic stress disorder model, exhibited persistent mechanical allodynia and anxiety-like behavior, which were accompanied by increased activation of microglia and secretion of pro-inflammatory cytokines in the hippocampus. Correlation analyses showed that hippocampal activation of microglia was significantly correlated with mechanical allodynia and anxiety-like behavior. Our data also showed that both intraperitoneal and intra-hippocampal injection of minocycline suppressed single prolonged stress-induced microglia activation and inflammatory cytokines accumulation in the hippocampus, and attenuated both single prolonged stress-induced mechanical allodynia and anxiety-like behavior. Taken together, the present study suggests that stress-induced microglia activation in the hippocampus may serve as a critical mechanistic link in the comorbid relationship between posttraumatic stress disorder and chronic pain. The novel concept introduces the possibility of cotreating chronic pain and posttraumatic stress disorder.

Intraoperative electroacupuncture relieves remifentanil-induced postoperative hyperalgesia via inhibiting spinal glial activation in rats

Background Accumulating studies have suggested that remifentanil, the widely-used opioid analgesic in clinical anesthesia, can activate the pronociceptive systems and enhance postoperative pain. Glial cells are thought to be implicated in remifentanil-induced hyperalgesia. Electroacupuncture is a complementary therapy to relieve various pain conditions with few side effects, and glial cells may be involved in its antinociceptive effect. In this study, we investigated whether intraoperative electroacupuncture could relieve remifentanil-induced postoperative hyperalgesia by inhibiting the activation of spinal glial cells, the production of spinal proinflammatory cytokines, and the activation of spinal mitogen-activated protein kinases. Methods A rat model of remifentanil-induced postoperative hyperalgesia was used in this study. Electroacupuncture during surgery was conducted at bilateral Zusanli (ST36) acupoints. Behavior tests, including mechanical allodynia and thermal hyperalgesia, were performed at different time points. Astrocytic marker glial fibrillary acidic protein, microglial marker Iba1, proinflammatory cytokines, and phosphorylated mitogen-activated protein kinases in the spinal cord were detected by Western blot and/or immunofluorescence. Results Mechanical allodynia and thermal hyperalgesia were induced by both surgical incision and remifentanil infusion, and remifentanil infusion significantly exaggerated and prolonged incision-induced pronociceptive effects. Glial fibrillary acidic protein, Iba1, proinflammatory cytokines (interleukin-1β and tumor necrosis factor-α), and phosphorylated mitogen-activated protein kinases (p-p38, p-JNK, and p-ERK1/2) were upregulated after surgical incision, remifentanil infusion, and especially after their combination. Intraoperative electroacupuncture significantly attenuated incision- and/or remifentanil-induced pronociceptive effects, spinal glial activation, proinflammatory cytokine upregulation, and phosphorylated mitogen-activated protein kinase upregulation. Conclusions Our study suggests that remifentanil-induced postoperative hyperalgesia can be relieved by intraoperative electroacupuncture via inhibiting the activation of spinal glial cells, the upregulation of spinal proinflammatory cytokines, and the activation of spinal mitogen-activated protein kinases.

Activation of spinal alpha-7 nicotinic acetylcholine receptor shortens the duration of remifentanil-induced postoperative hyperalgesia by upregulating KCC2 in the spinal dorsal horn in rats

Background Accumulating evidence has shown that the signal from spinal brain-derived neurotrophic factor/tyrosine receptor kinase B-K+-Cl− cotransporter-2 plays a critical role in the process of pain hypersensitivity. The activation of alpha-7 nicotinic acetylcholine receptors could have an analgesic effect on remifentanil-induced postoperative hyperalgesia. Nevertheless, whether intrathecal administration of PNU-120596, an alpha-7 nicotinic acetylcholine receptors selective type II positive allosteric modulator, before surgery could affect the duration of remifentanil-induced postoperative hyperalgesia remains unknown, and the effects of alpha-7 nicotinic acetylcholine receptors activation on the brain-derived neurotrophic factor/tyrosine receptor kinase B-K+-Cl− cotransporter-2 signal in the spinal dorsal horn of rats with remifentanil-induced postoperative hyperalgesia is still enigmatic. Results We demonstrated that the brain-derived neurotrophic factor/tyrosine receptor kinase B-K+-Cl− cotransporter-2 signal played a critical role in the development of remifentanil-induced postoperative hyperalgesia. Intrathecal administration of PNU-120596 (8 µg/kg, 15 min before surgery) was associated with earlier signs of recovery from remifentanil-induced postoperative hyperalgesia. Simultaneously, remifentanil-induced postoperative hyperalgesia-induced K+-Cl− cotransporter-2 downregulation was partly reversed and coincided with a decreased expression of brain-derived neurotrophic factor/tyrosine receptor kinase B in the spinal dorsal horn, approximately correlating with the time course of the nociceptive behavior. Moreover, intrathecal administration of the K+-Cl− cotransporter-2 inhibitor VU0240551 significantly reduced the analgesic effect of PNU-120596 on remifentanil-induced postoperative hyperalgesia. Conclusions The activation of alpha-7 nicotinic acetylcholine receptors induced a shorter duration of remifentanil-induced postoperative hyperalgesia by restoring the brain-derived neurotrophic factor/tyrosine receptor kinase B-K+-Cl− cotransporter-2 signal in the spinal dorsal horn of rats, which provides new insight into treatment in clinical postoperative pain management.

The effects of an intraperitoneal single low dose of ketamine in attenuating the postoperative skin/muscle incision and retraction-induced pain related to the inhibition of N-methyl-d-aspartate receptors in the spinal cord

BACKGROUND Chronic postoperative pain (CPOP) is a common clinical problem which might be related to central sensitization. It has been widely accepted that NMDA (N-methyl-D-aspartate) receptors are among the triggers of central sensitization. Ketamine is a non-competitive NMDA receptor antagonist that is widely used in alleviating postoperative pain, but its effect on CPOP has been rarely reported. In the present study, the skin/muscle incision and retraction (SMIR) model was used to investigate the role of NMDARs in chronic postoperative pain and the effect of an intraperitoneal single low dose ketamine (10mg/kg) of attenuating SMIR-induced CPOP. METHODS We assessed pain behaviours after a SMIR operation by paw withdrawal threshold (PWMT) and paw withdrawal latency (PWMTL). Western blotting were performed to examine the role of NMDARs in SMIR-induced CPOP and the effect of ketamine on the expression and phosphorylation of NMDARs. RESULTS The SMIR operation induced long-lasting mechanical hyperalgesia, and the up-regulation of phosphorylated NMDARs and total NMDARs at the spinal level. A single intraperitoneal administration of low dose ketamine (10mg/kg) during surgery alleviated pain behaviors and inhibited the up-regulation of phosphorylated NMDARs and total NMDARs. CONCLUSIONS Our datas suggested that NMDARs play important roles in SMIR-induced CPOP. A single intraperitoneal low dose of ketamine could attenuate SMIR-induced CPOP, which might be associated with the inhibition of NMDARs. Our finding might provide a new, simple method of addressing CPOP.

Dehydrocorydaline attenuates bone cancer pain by shifting microglial M1/M2 polarization toward the M2 phenotype:

Bone cancer pain remains a major challenge in patients with primary or metastatic bone cancer due to a lack of understanding the mechanisms. Previous studies have revealed the two distinct functional polarization states of microglia (classically activated M1 and alternatively activated M2) in the spinal cord in nerve injury–induced neuropathic pain. However, whether microglia in the spinal cord polarize to M1 and M2 phenotypes and contribute to the development of bone cancer pain remains unclear. In this study, we used a mouse model with bone cancer to characterize the M1/M2 polarization of microglia in the spinal cord during the development of bone cancer pain, and investigated the antinociceptive effects of dehydrocorydaline, an alkaloidal component isolated from Rhizoma corydalis on bone cancer pain. Our results show that microglia in the spinal cord presented increased M1 polarization and decreased M2 polarization, while overproduction of IL-1β and inhibited expression of IL-10 was detected during bone cancer pain development. Intraperitoneal administration of dehydrocorydaline (10 mg/kg) had significant antinociceptive effects on day 14 after osteosarcoma cell implantation, accompanied by suppressed M1 phenotype and upregulated M2 phenotype of microglia in the spinal cord, while alleviated inflammatory response was observed then. These results suggest that the imbalanced polarization of microglia toward the M1 phenotype in the spinal cord may contribute to the development of bone cancer pain, while dehydrocorydaline helps to attenuate bone cancer pain, with microglial polarization shifting toward the M2 phenotype in the spinal cord.