Spared Nerve Injury Model Research Articles

Potassium Channels Kv1.3 and Kir2.1 But Not Kv1.5 Contribute to BV2 Cell Line and Primary Microglial Migration.

(1) Background: As membrane channels contribute to different cell functions, understanding the underlying mechanisms becomes extremely important. A large number of neuronal channels have been investigated, however, less studied are the channels expressed in the glia population, particularly in microglia. In the present study, we focused on the function of the Kv1.3, Kv1.5 and Kir2.1 potassium channels expressed in both BV2 cells and primary microglia cultures, which may impact the cellular migration process. (2) Methods: Using an immunocytochemical approach, we were able to show the presence of the investigated channels in BV2 microglial cells, record their currents using a patch clamp and their role in cell migration using the scratch assay. The migration of the primary microglial cells in culture was assessed using cell culture inserts. (3) Results: By blocking each potassium channel, we showed that Kv1.3 and Kir2.1 but not Kv1.5 are essential for BV2 cell migration. Further, primary microglial cultures were obtained from a line of transgenic CX3CR1-eGFP mice that express fluorescent labeled microglia. The mice were subjected to a spared nerve injury model of pain and we found that microglia motility in an 8 µm insert was reduced 2 days after spared nerve injury (SNI) compared with sham conditions. Additional investigations showed a further impact on cell motility by specifically blocking Kv1.3 and Kir2.1 but not Kv1.5; (4) Conclusions: Our study highlights the importance of the Kv1.3 and Kir2.1 but not Kv1.5 potassium channels on microglia migration both in BV2 and primary cell cultures.

MCRT, a multifunctional ligand of opioid and neuropeptide FF receptors, attenuates neuropathic pain in spared nerve injury model

Chimeric peptide MCRT (YPFPFRTic-NH2 ) was a multifunctional ligand of opioid and neuropeptide FF (NPFF) receptors and reported to be potentially antalgic in acute tail-flick test. Here, we developed spared nerve injury (SNI) model to explore its efficacy in chronic neuropathic pain. Analgesic tolerance, opioid-induced hyperalgesia and gastrointestinal transit were measured for safety evaluation. Intracerebroventricular (i.c.v.) and intraplantar (i.pl.) injections were conducted as central and peripheral routes, respectively. Results demonstrated that MCRT alleviated neuropathic pain effectively and efficiently, with the ED50 values of 4.93nmol/kg at the central level and 3.11nmol/kg at the peripheral level. The antagonist blocking study verified the involvement of mu-, delta-opioid and NPFF receptors in MCRT produced anti-allodynia. Moreover, the separation of analgesia from unwanted effects was preliminarily achieved and that MCRT caused neither analgesic tolerance nor hyperalgesia after chronic i.c.v. administration, nor constipation after i.pl. administration. Notably, the local efficacy of MCRT in SNI mice was about one thousandfold higher than morphine and ten thousandfold higher than pregabalin, indicating a great promise in the future treatment of neuropathic pain.

Activation of Neuronal Voltage-Gated Potassium Kv7/KCNQ/M-Current by a Novel Channel Opener SCR2682 for Alleviation of Chronic Pain.

Treatment of chronic pain remains an unmet medical need. The neuronal voltage-gated potassium Kv7/KCNQ/M channel has been implicated as a therapeutic target for chronic pain. However, whether pharmacological activation of the Kv7 channel can alleviate pain remains elusive. In this study, we show that selective activation of native M-currents by a novel channel opener SCR2682 reduces repetitive firings of dorsal root ganglia (DRG) sensory neurons. Intraperitoneal administration of SCR2682 relieves mechanical allodynia and thermal hyperalgesia in rat models of pain induced by complete Freund's adjuvant (CFA) or spared nerve injury (SNI) in a dose-dependent manner without affecting locomotor activity. The antinociceptive efficacy of SCR2682 can be reversed by the channel-specific blocker XE991. Furthermore, SCR2682 increases Kv7.2/KCNQ2 mRNA and protein expression in DRG neurons from rats in the SNI model of neuropathic pain. Taken together, pharmacological activation of neuronal Kv7 channels by opener SCR2682 can alleviate pain in rats, thus possessing therapeutic potential for chronic pain or hyperexcitability-related neurologic disorders. SIGNIFICANCE STATEMENT: A novel voltage-gated potassium Kv7 channel opener SCR2682 inhibits action potential firings in dorsal root ganglia sensory neurons and exhibits efficacy in antinociception, thus possessing a developmental potential for treatment of chronic pain or epilepsy.

The interaction between P2X3 and TRPV1 in the dorsal root ganglia of adult rats with different pathological pains

Peripheral inflammatory and neuropathic pain are closely related to the activation of purinergic receptor P2X ligand-gated ion channel 3 (P2X3) and transient receptor potential vanilloid 1 (TRPV1), but the interaction between P2X3 and TRPV1 in different types of pathological pain has rarely been reported. In this study, complete Freund’s adjuvant (CFA)-induced inflammatory pain and spared nerve injury (SNI)-induced neuropathic pain models were established in adult rats. The interactions between P2X3 and TRPV1 in the dorsal root ganglion were observed by pharmacological, co-immunoprecipitation, immunofluorescence and whole-cell patch-clamp recording assays. TRPV1 was shown to promote the induction of spontaneous pain caused by P2X3 in the SNI model, but the induction of spontaneous pain behaviour by TRPV1 was not completely dependent on P2X3 in vivo. In both the CFA and SNI models, the activation of peripheral P2X3 enhanced the effect of TRPV1 on spontaneous pain, while the inhibition of peripheral TRPV1 reduced the induction of spontaneous pain by P2X3 in the CFA model. TRPV1 and P2X3 had inhibitory effects on each other in the inflammatory pain model. During neuropathic pain, P2X3 facilitated the function of TRPV1, while TRPV1 had an inhibitory effect on P2X3. These results suggest that the mutual effects of P2X3 and TRPV1 differ in cases of inflammatory and neuropathic pain in rats.

Modulation of microglial activation states by spinal cord stimulation in an animal model of neuropathic pain: Comparing high rate, low rate, and differential target multiplexed programming.

While numerous studies and patient experiences have demonstrated the efficacy of spinal cord stimulation as a treatment for chronic neuropathic pain, the exact mechanism underlying this therapy is still uncertain. Recent studies highlighting the importance of microglial cells in chronic pain and characterizing microglial activation transcriptomes have created a focus on microglia in pain research. Our group has investigated the modulation of gene expression in neurons and glial cells after spinal cord stimulation (SCS), specifically focusing on transcriptomic changes induced by varying SCS stimulation parameters. Previous work showed that, in rodents subjected to the spared nerve injury (SNI) model of neuropathic pain, a differential target multiplexed programming (DTMP) approach provided significantly better relief of pain-like behavior compared to high rate (HRP) and low rate programming (LRP). While these studies demonstrated the importance of transcriptomic changes in SCS mechanism of action, they did not specifically address the role of SCS in microglial activation. The data presented herein utilizes microglia-specific activation transcriptomes to further understand how an SNI model of chronic pain and subsequent continuous SCS treatment with either DTMP, HRP, or LRP affects microglial activation. Genes for each activation transcriptome were identified within our dataset and gene expression levels were compared with that of healthy animals, naïve to injury and interventional procedures. Pearson correlations indicated that DTMP yields the highest significant correlations to expression levels found in the healthy animals across all microglial activation transcriptomes. In contrast, HRP or LRP yielded weak or very weak correlations for these transcriptomes. This work demonstrates that chronic pain and subsequent SCS treatments can modulate microglial activation transcriptomes, supporting previous research on microglia in chronic pain. Furthermore, this study provides evidence that DTMP is more effective than HRP and LRP at modulating microglial transcriptomes, offering potential insight into the therapeutic efficacy of DTMP.

Modulation of SI and ACC response to noxious and non‐noxious electrical stimuli after the spared nerve injury model of neuropathic pain

The current knowledge on the role of SI and ACC in acute pain processing and how these contribute to the development of chronic pain is limited. Our objective was to investigate differences in and modulation of intracortical responses from SI and ACC in response to different intensities of peripheral presumed noxious and non-noxious stimuli in the acute time frame of a peripheral nerve injury in rats. We applied non-noxious and noxious electrical stimulation pulses through a cuff electrode placed around the sciatic nerve and measured the cortical responses (six electrodes in each cortical area) before and after the spared nerve injury model. We found that the peak response correlated with the stimulation intensity and that SI and ACC differed in both amplitude and latency of cortical response. The cortical response to both noxious and non-noxious stimulation showed a trend towards faster processing of non-noxious stimuli in ACC and increased cortical processing of non-noxious stimuli in SI after SNI. We found different responses in SI and ACC to different intensity electrical stimulations based on two features and changes in these features following peripheral nerve injury. We believe that these features may be able to assist to track cortical changes during the chronification of pain in future animal studies. This study showed distinct cortical processing of noxious and non-noxious peripheral stimuli in SI and ACC. The processing latency in ACC and accumulated spiking activity in SI appeared to be modulated by peripheral nerve injury, which elaborated on the function of these two areas in the processing of nociception.

Nerve Injury Decreases Hyperacute Resting-State Connectivity Between the Anterior Cingulate and Primary Somatosensory Cortex in Anesthetized Rats.

A better understanding of neural pain processing and of the development of pain over time, is critical to identify objective measures of pain and to evaluate the effect of pain alleviation therapies. One issue is, that the brain areas known to be related to pain processing are not exclusively responding to painful stimuli, and the neuronal activity is also influenced by other brain areas. Functional connectivity reflects synchrony or covariation of activation between groups of neurons. Previous studies found changes in connectivity days or weeks after pain induction. However, less in known on the temporal development of pain. Our objective was therefore to investigate the interaction between the anterior cingulate cortex (ACC) and primary somatosensory cortex (SI) in the hyperacute (minute) and sustained (hours) response in an animal model of neuropathic pain. Intra-cortical local field potentials (LFP) were recorded in 18 rats. In 10 rats the spared nerve injury model was used as an intervention. The intra-cortical activity was recorded before, immediately after, and three hours after the intervention. The interaction was quantified as the calculated correlation and coherence. The results from the intervention group showed a decrease in correlation between ACC and SI activity, which was most pronounced in the hyperacute phase but a longer time frame may be required for plastic changes to occur. This indicated that both SI and ACC are involved in hyperacute pain processing.

Antinociceptive properties of an isoform-selective inhibitor of Nav1.7 derived from saxitoxin in mouse models of pain.

The voltage-gated sodium channel Nav1.7 is highly expressed in nociceptive afferents and is critically involved in pain signal transmission. Nav1.7 is a genetically validated pain target in humans because loss-of-function mutations cause congenital insensitivity to pain and gain-of-function mutations cause severe pain syndromes. Consequently, pharmacological inhibition has been investigated as an analgesic therapeutic strategy. We describe a small molecule Nav1.7 inhibitor, ST-2530, that is an analog of the naturally occurring sodium channel blocker saxitoxin. When evaluated against human Nav1.7 by patch-clamp electrophysiology using a protocol that favors the resting state, the Kd of ST-2530 was 25 ± 7 nM. ST-2530 exhibited greater than 500-fold selectivity over human voltage-gated sodium channel isoforms Nav1.1-Nav1.6 and Nav1.8. Although ST-2530 had lower affinity against mouse Nav1.7 (Kd = 250 ± 40 nM), potency was sufficient to assess analgesic efficacy in mouse pain models. A 3-mg/kg dose administered subcutaneously was broadly analgesic in acute pain models using noxious thermal, mechanical, and chemical stimuli. ST-2530 also reversed thermal hypersensitivity after a surgical incision on the plantar surface of the hind paw. In the spared nerve injury model of neuropathic pain, ST-2530 transiently reversed mechanical allodynia. These analgesic effects were demonstrated at doses that did not affect locomotion, motor coordination, or olfaction. Collectively, results from this study indicate that pharmacological inhibition of Nav1.7 by a small molecule agent with affinity for the resting state of the channel is sufficient to produce analgesia in a range of preclinical pain models.

AAV-Mediated Combination Gene Therapy for Neuropathic Pain: GAD65, GDNF, and IL-10

Neuropathic pain is a chronic pain state characterized by nerve damage, inflammation, and nociceptive neuron hyperactivity. As the underlying pathophysiology is complex, a more effective therapy for neuropathic pain would be one that targets multiple elements. Here, we generated recombinant adeno-associated viruses (AAVs) encoding three therapeutic genes, namely, glutamate decarboxylase 65, glial cell-derived neurotrophic factor, and interleukin-10, with various combinations. The efficacy for pain relief was evaluated in a rat spared nerve injury model of neuropathic pain. The maximal analgesic effect was achieved when the AAVs expressing all three genes were administered to rats with neuropathic pain. The combination of two virus constructs expressing the three genes was named KLS-2031 and evaluated as a potential novel therapeutic for neuropathic pain. Single transforaminal epidural injections of KLS-2031 into the intervertebral foramen to target the appropriate dorsal root ganglion produced notable long-term analgesic effects in female and male rats. Furthermore, KLS-2031 mitigated the neuroinflammation, neuronal cell death, and dorsal root ganglion hyperexcitability induced by the spared nerve injury. These results suggest that KLS-2031 represents a promising therapeutic option for refractory neuropathic pain.

Upregulation of Nav1.7 by endogenous hydrogen sulfide contributes to maintenance of neuropathic pain.

Nav1.7 is closely associated with neuropathic pain. Hydrogen sulfide (H2S) has recently been reported to be involved in numerous biological functions, and it has been shown that H2S can enhance the sodium current density, and inhibiting the endogenous production of H2S mediated by cystathionine β-synthetase (CBS) using O-(carboxymethyl) hydroxylamine hemihydrochloride (AOAA) can significantly reduce the expression of Nav1.7 and thus the sodium current density in rat dorsal root ganglion (DRG) neurons. In the present study, it was shown that the fluorescence intensity of H2S was increased in a spared nerve injury (SNI) model and AOAA inhibited this increase. Nav1.7 is expressed in DRG neurons, and the expression of CBS and Nav1.7 were increased in DRG neurons 7, 14 and 21 days post-operation. AOAA inhibited the increase in the expression of CBS, phosphorylated (p)-MEK1/2, p-ERK1/2 and Nav1.7 induced by SNI, and U0126 (a MEK blocker) was able to inhibit the increase in p-MEK1/2, p-ERK1/2 and Nav1.7 expression. However, PF-04856264 did not inhibit the increase in CBS, p-MEK1/2, p-ERK1/2 or Nav1.7 expression induced by SNI surgery. The current density of Nav1.7 was significantly increased in the SNI model and administration of AOAA and U0126 both significantly decreased the density. In addition, AOAA, U0126 and PF-04856264 inhibited the decrease in rheobase, and the increase in action potential induced by SNI in DRG neurons. There was no significant difference in thermal withdrawal latency among each group. However, the time the animals spent with their paw lifted increased significantly following SNI, and the time the animals spent with their paw lifted decreased significantly following the administration of AOAA, U0126 and PF-04856264. In conclusion, these data show that Nav1.7 expression in DRG neurons is upregulated by CBS-derived endogenous H2S in an SNI model, contributing to the maintenance of neuropathic pain.

Behavioral, Biochemical and Electrophysiological Changes in Spared Nerve Injury Model of Neuropathic Pain.

Neuropathic pain is a pathological condition induced by a lesion or disease affecting the somatosensory system, with symptoms like allodynia and hyperalgesia. It has a multifaceted pathogenesis as it implicates several molecular signaling pathways involving peripheral and central nervous systems. Affective and cognitive dysfunctions have been reported as comorbidities of neuropathic pain states, supporting the notion that pain and mood disorders share some common pathogenetic mechanisms. The understanding of these pathophysiological mechanisms requires the development of animal models mimicking, as far as possible, clinical neuropathic pain symptoms. Among them, the Spared Nerve Injury (SNI) model has been largely characterized in terms of behavioral and functional alterations. This model is associated with changes in neuronal firing activity at spinal and supraspinal levels, and induces late neuropsychiatric disorders (such as anxious-like and depressive-like behaviors, and cognitive impairments) comparable to an advanced phase of neuropathy. The goal of this review is to summarize current findings in preclinical research, employing the SNI model as a tool for identifying pathophysiological mechanisms of neuropathic pain and testing pharmacological agent.

HDAC6-selective inhibitors decrease nerve-injury and inflammation-associated mechanical hypersensitivity in mice.

HDAC6 is a class IIB histone deacetylase expressed at many levels of the nociceptive pathway. This study tested the ability of novel and selective HDAC6 inhibitors to alleviate sensory hypersensitivity behaviors in mouse models of peripheral nerve injury and peripheral inflammation. We utilized the murine spared nerve injury (SNI) model for peripheral nerve injury and the Complete Freund's Adjuvant (CFA) model of peripheral inflammation. We applied the Von Frey assay to monitor mechanical allodynia. Using the SNI model, we demonstrate that daily administration of the brain-penetrant HDAC6 inhibitor, ACY-738, abolishes mechanical allodynia in male and in female mice. Importantly, there is no tolerance to the antiallodynic actions of these compounds as they produce a consistent increase in Von Frey thresholds for several weeks. We observed a similar antiallodynic effect when utilizing the HDAC6 inhibitor, ACY-257, which shows limited brain expression when administered systemically. We also demonstrate that ACY-738 and ACY-257 attenuate mechanical allodynia in the CFA model of peripheral inflammation. Overall, our findings suggest that inhibition of HDAC6 provides a promising therapeutic avenue for the alleviation of mechanical allodynia associated with peripheral nerve injury and peripheral inflammation.

The gene regulatory network in different brain regions of neuropathic pain mouse models.

Neuropathic pain is directly developed from lesions or somatosensory nervous system diseases that are associated with emotion regulation. In general population, the incidence of neuropathic pain ranges from 7% to 10%, but the underlying mechanism remains largely unknown. Neuropathic pain is often associated with structural and functional abnormalities in multiple brain regions, and its regulation has been shown to correspond with the forebrain, including nucleus accumbens (NAc), medial prefrontal cortex (mPFC) and periaqueductal gray (PAG). To investigate the molecular mechanism of neuropathic pain across different brain regions, we identified the differentially expressed genes (DEGs) between the spared nerve injury model (SNI) mice suffering neuropathic pain and the control Sham mice in NAc, mPFC and PAG three brain regions, and mapped these genes onto a comprehensively functional association network. Thereafter, novel neuropathic pain genes in these three regions were identified using With Random Walk with Restart (RWR) analysis, such as Asic3, Cd200r1 and MT2, besides well-known Capn11 and CYP2E1. Interactions or cross talks among DEGs in NAc, mPFC and PAG three brain regions were discovered. Our results provide novel insights into neuropathic pain and help to explore therapeutic targets in the treatment.

A high‐affinity, bivalent PDZ domain inhibitor complexes PICK1 to alleviate neuropathic pain

Maladaptive plasticity involving increased expression of AMPA‐type glutamate receptors is involved in several pathologies, including neuropathic pain, but direct inhibition of AMPARs is associated with side effects. As an alternative, we developed a cell‐permeable, high‐affinity (~2 nM) peptide inhibitor, Tat‐P4‐(C5)2, of the PDZ domain protein PICK1 to interfere with increased AMPAR expression. The affinity is obtained partly from the Tat peptide and partly from the bivalency of the PDZ motif, engaging PDZ domains from two separate PICK1 dimers to form a tetrameric complex. Bivalent Tat‐P4‐(C5)2 disrupts PICK1 interaction with membrane proteins on supported cell membrane sheets and reduce the interaction of AMPARs with PICK1 and AMPA‐receptor surface expression in vivo. Moreover, Tat‐P4‐(C5)2 administration reduces spinal cord transmission and alleviates mechanical hyperalgesia in the spared nerve injury model of neuropathic pain. Taken together, our data reveal Tat‐P4‐(C5)2 as a novel promising lead for neuropathic pain treatment and expand the therapeutic potential of bivalent inhibitors to non‐tandem protein–protein interaction domains.

The bivalent ligand, MMG22, reduces neuropathic pain after nerve injury without the side effects of traditional opioids.

Functional interactions between the mu opioid receptor (MOR) and the metabotropic glutamate receptor 5 (mGluR5) in pain and analgesia have been well established. MMG22 is a bivalent ligand containing MOR agonist (oxymorphamine) and mGluR5 antagonist (MPEP) pharmacophores tethered by a 22-atom linker. MMG22 has been shown to produce potent analgesia in several models of chronic inflammatory and neuropathic pain (NP). This study assessed the efficacy of systemic administration of MMG22 at reducing pain behavior in the spared nerve injury (SNI) model of NP in mice, as well as its side-effect profile and abuse potential. MMG22 reduced mechanical hyperalgesia and spontaneous ongoing pain after SNI, with greater potency early (10 days) as compared to late (30 days) after injury. Systemic administration of MMG22 did not induce place preference in naive animals, suggesting absence of abuse liability when compared to traditional opioids. MMG22 also lacked the central locomotor, respiratory, and anxiolytic side effects of its monomeric pharmacophores. Evaluation of mRNA expression showed the transcripts for both receptors were colocalized in cells in the dorsal horn of the lumbar spinal cord and dorsal root ganglia. Thus, MMG22 reduces hyperalgesia after injury in the SNI model of NP without the typical centrally mediated side effects associated with traditional opioids.

Lidocaine Alleviates Neuropathic Pain and Neuroinflammation by Inhibiting HMGB1 Expression to Mediate MIP-1α/CCR1 Pathway.

High mobility group box1 (HMGB1) released from sensory nerve tissues can induce neuropathic pain. Whether HMGB1 is implicated in the mechanism underlying the effect of lidocaine in pain management remains to be determined. This study aims to explore the effect of lidocaine in a rat model of spared nerve injury (SNI) and the underlying mechanism. An SNI model was established via nerve ligation. Two weeks after the SNI model was established, rats were intrathecally injected with lidocaine, an HMGB1 antibody (HMG Ab), an MIP-1α antibody (MIP-1α Ab), a CCR1 inhibitor (CCR1-RS) or a CCR5 antagonist (CCR5-Mar). Pain behaviors were assessed before and after model establishment to calculate the number of spontaneous flinches (NSF), paw withdrawal threshold (PWT), paw withdrawal thermal latency (PWL) and sciatic function index (SFI). Cell apoptosis and the inflammatory response in the cerebrospinal fluid (CSF) were detected by TUNEL staining and ELISA. The mRNA and protein expression levels of MIP-1α, CCR1 and CCR5 were determined by RT-PCR and Western blotting. The expression levels of HMGB1, MIP-1α, CCR1 and CCR5 were measured by Western blotting and immunofluorescence. Pain behavior testing in SNI rats showed that SNI rats exhibited an increased NSF and adecreased PWT, PWL and SFI. Cell apoptosis in the spinal dorsal horn and the generation of inflammatory cytokines were enhanced in SNI rats, and the expression levelsof HMGB1, MIP-1α, CCR1 and CCR5 were upregulated. HMGB1 cytoplasmic translocation, the coexpression of MIP-1α with NeuN, and the coexpression of CCR1 and CCR5 with OX42 were also observed in SNI rats. Neuropathic pain and neuroinflammation were suppressed by the intrathecal injection of lidocaine, HMG Ab, MIP-1α Ab, CCR1-RS or CCR5-Mar. Lidocaine inhibited the expression levels of HMGB1, MIP-1α, CCR1 and CCR5, and the HMGB1 antibody suppressed the expression of MIP-1α, CCR1 and CCR5. Lidocaine attenuates neuropathic pain and neuroinflammation by inhibiting HMGB1 to regulate the MIP-1α/CCR1/CCR5 pathway. Graphical Abstract.

Localized sympathectomy reduces peripheral nerve regeneration and pain behaviors in 2 rat neuropathic pain models.

Previous studies have shown that the peripheral nerve regeneration process is linked to pain in several neuropathic pain models. Other studies show that sympathetic blockade may relieve pain in some pain models and clinical conditions. This study examined reduction in peripheral nerve regeneration as one possible mechanism for relief of neuropathic pain by sympathetic blockade. A "microsympathectomy," consisting of cutting the gray rami containing sympathetic postganglionic axons where they enter the L4 and L5 spinal nerves, reduced mechanical hypersensitivity in 2 different rat neuropathic pain models. In the spinal nerve ligation model, in which some functional regeneration and reinnervation of the ligated spinal nerve can be observed, microsympathectomy reduced functional and anatomical measures of regeneration as well as expression of growth-associated protein 43 (GAP43), a regeneration-related protein. In the spared nerve injury model, in which functional reinnervation is not possible and the futile regeneration process results in formation of a neuroma, microsympathectomy reduced neuroma formation and GAP43 expression. In both models, microsympathectomy reduced macrophage density in the sensory ganglia and peripheral nerve. This corroborates previous work showing that sympathetic nerves may locally affect immune function. The results further highlight the challenge of improving pain in neuropathic conditions without inhibiting peripheral nerve regeneration that might otherwise be possible and desired.

Oxycodone‐induced Gene adaptations in the brain reward center in a murine Model of Neuropathic pain

The long‐term use of opioids for the treatment of chronic pain can lead to physical dependence and transition to addiction. Recent studies from our group revealed that chronic neuropathic pain promotes several adaptations in the brain reward regions (Descalzi et al., Sci. Signaling, 2017). Here, we are using Next Generation RNA sequencing and upstream pathway analysis to gain insight on gene expression adaptations triggered by persistent exposure to oxycodone in the presence and in the absence of pain. To this extent, we employed the spared nerve injury model (SNI) of neuropathic pain in adult C57Bl/6 mice and assessed peripheral neuropathy symptoms for 9 weeks. We then applied an oxycodone misuse paradigm, where mice were treated daily with oxycodone for two weeks, and gene expression was monitored three weeks after oxycodone withdrawal. Mice were also assessed for emotional and pain‐related behaviors during the three‐week drug withdrawal period. Oxycodone treatment‐induced thermal hyperalgesia regardless of pain states, while withdrawal produced significant mechanical allodynia and deficits in social interaction. Using RNA Sequencing, we monitored changes in gene expression in the medial prefrontal cortex, the nucleus accumbens, and the ventral tegmental area. Although oxycodone treatment promotes mostly unique transcriptome profiles across brain regions, we observed similar downstream effectors and transcription factors to be affected in pain states when compared with non‐pain states. Furthermore, our findings suggest that chronic pain states exacerbate the behavioral and transcriptomic signatures of oxycodone withdrawal. Overall this work will provide new insights on the influence of long term pain in oxycodone‐induced plasticity in the brain reward center.Support or Funding Information5T32DA007135‐34, NIDA, R01 NS086444, R56 NS111351 NINDS, and P01 DA008227, NIDA

A novel and potent neuronal Kv7 channel opener SCR2682 alleviates chronic pain in rats

Activation of neuronal Kv7/KCNQ/M‐current emerges as an effective intervention for treatment of hyperexcitability‐related neurological diseases including chronic pain that is prevalent and unmet medical need. The aim of this study was to evaluate the anti‐nociceptive effects of a novel and potent Kv7/KCNQ/M channel opener SCR2682 on chronic pain in rats. The pharmacological effects of SCR2682 on native M current and firing of dorsal root ganglia (DRG) neurons were examined using electrophysiological recordings. Two in vivo rat models of complete Freund’s adjuvant (CFA)‐induced inflammatory pain and spared nerve injury (SNI)‐induced neuropathic pain were used to evaluate anti‐nociceptive effects of SCR2682 by measuring the paw withdrawal mechanical threshold (PWMT) and/or thermal latency (PWTL). Locomotor activity in normal rats was tested for safety evaluation. The expression of KCNQ2 in L4 and L5 DRGs of rat SNI model was detected by quantitative RT‐PCR and western blot.We found that SCR2682 (0.1 μM) or an FDA approved anti‐epileptic drug retigabine (RTG) (10 μM) increased M current in rat DRG neurons approximately 30%, which was blocked by Kv7 channel inhibitor XE991 (3 μM). SCR2682 caused the resting membrane potential (RMP) hyperpolarization to −66.4 ± 3.7 mV from −56.7 ± 2.8 mV. Current clamp recordings revealed that perfusion of SCR2682 abolished the firing spikes of DRG neurons. In rat model of CFA‐induced inflammatory pain, intraperitoneal (i.p.) administration of SCR2682 at doses of 0.5, 1 and 2 mg/kg increased the PWMT and PWTL in a dose‐dependent manner, as compared with RTG (7 mg/kg) that also increased the PWMT and PWTL. In the model of SNI‐induced neuropathic pain, SCR2682 (0.5, 1 and 2 mg/kg) or RTG (7 mg/kg) increased the PWMT, and the anti‐nociceptive effect of SCR2682 (2 mg/kg) could be reversed by the channel specific blocker XE991 (3 mg/kg, i.p.). Furthermore, SCR2682 (2 mg/kg.) or RTG (7 mg/kg) significantly increased the expression of KCNQ2 mRNA and protein in DRG of SNI rats, and co‐injection of XE991 (3 mg/kg) with SCR2682 reduced the expression levels of KCNQ2. In the open field test, there was no significant difference in the travel total distance and mean speed of rats treated with RTG (7 mg/kg) or SCR2682 (0.5, 1 and 2 mg/kg). Taken together, our data show that the novel compound SCR2682 reduces the excitability of DRG sensory neurons by activating Kv7 channel, and intraperitoneal administration of SCR2682 alleviates chronic pain in rats.Support or Funding InformationThis project was supported by grants awarded to K.W. from National Natural Sciences Foundation of China (81573410) and the Ministry of Science and Technology of China (2018ZX09711001‐004‐006) and Science and Technology Program of Guangdong (2018B030334001).

IL-27 Counteracts Neuropathic Pain Development Through Induction of IL-10.

Neuroimmune–glia interactions have been implicated in the development of neuropathic pain. Interleukin-27 (IL-27) is a cytokine that presents regulatory activity in inflammatory conditions of the central nervous system. Thus, we hypothesized that IL-27 would participate in the neuropathic pain process. Here, we found that neuropathic pain caused by peripheral nerve injury (spared nerve injury model; SNI), was enhanced in IL-27-deficient(−/−) mice, whereas nociceptive pain is similar to that of wild-type mice. SNI induced an increase in the expression of IL-27 and its receptor subunit (Wsx1) in the sensory ganglia and spinal cord. IL-27 receptor was expressed mainly in resident macrophage, microglia, and astrocytes of the sensory ganglia and spinal cord, respectively. Finally, we identify that the antinociceptive effect of IL-27 was not observed in IL-10−/− mice. These results provided evidence that IL-27 is a cytokine produced after peripheral nerve injury that counteracts neuropathic pain development through induction of the antinociceptive cytokine IL-10. In summary, our study unraveled the role of IL-27 as a regulatory cytokine that counteracts the development of neuropathic pain after peripheral nerve damage. In conclusion, they indicate that immunotherapies based on IL-27 could emerge as possible therapeutic approaches for the prevention of neuropathic pain development after peripheral nerve injury.