Chronic Inflammatory Pain Models Research Articles

The role of SOCS3 in the hypothalamic paraventricular nucleus in rat model of inflammatory pain

BackgroundInflammatory molecular signals are modulated by a variety of intracellular transduction pathways, the activation of which may induce and amplify the spread of inflammatory response. Suppresser of cytokine signaling 3 (SOCS3) is an established negative feedback regulation transcription factor associated with tumor, diabetes mellitus, inflammation and anaphylaxis. Herein, we investigated whether SOCS3 in the paraventricular nucleus (PVN) can attenuate pro-inflammatory responses, and thereby relieve the inflammatory pain.MethodsAdeno-associated virus (AAV) overexpressing SOCS3 was pre-injected into the PVN. Three weeks later, rat model of chronic inflammatory pain was established via subcutaneous injection of complete Freund’s adjuvant (CFA) into the plantar center of hind paws. The therapeutic effect of SOCS3 was tested by the measurement of thermal and mechanical allodynia. In mechanistic study, the protein level of SOCS3 was evaluated by Western blotting, and the expression of c-fos and Iba-1 were assessed by immunofluorescent staining.ResultsInflammatory pain was associated with upregulated interleukin 6 (IL-6) and SOCS3 in PVN in the acute phase. Thermal hyperalgesia can be relieved by intra-PVN injection of IL-6 neutralizing antibody (NA). Meanwhile, the upregulated c-fos and microglial activation was reversed. Furthermore, SOCS3 expression in PVN was downregulated in the chronic phase. Intra-PVN injection of AAV overexpressing SOCS3 suppressed the activation of neurons and attenuated thermal hyperalgesia and mechanical allodynia.ConclusionInhibition of IL-6 signaling attenuated inflammatory hyperalgesia in the acute phase. SOCS3 overexpression in the PVN attenuated inflammatory pain in the chronic phase via suppression of neuronal activation.

Effect of electroacupuncture on expression of GABAA receptor mRNA in different brain regions in rats with chronic inflammatory pain

To observe the expression of GABAA receptor mRNA in different brain regions of the central nervous system in chronic inflammatory pain rats and the intervention effect of electroacupuncture (EA). A total of 48 SPF male SD rats were randomly divided into a blank control group, a model control group, an EA group and a sham EA group, 12 rats in each group. The model of chronic inflammatory pain was established by injecting Freund's complete adjuvant into the foot. The EA group was treated with EA 28 days after the model establishment. The "Housanli" (ST 36) and "Kunlun" (BL 60) were selected and treated with dilatational wave, 2 Hz/100 Hz in frequency, 0.5-1.5 mA for 30 min; EA was given only once. In the sham EA group, the same acupoints were selected but the needles were only inserted into subcutaneous area; EA was connected for 30 min without electrical stimulation. The behavior changes of mechanical pain threshold and thermal pain threshold before model establishment, 1 day, 3 days, 7 days, 14 days, 21 days and 28 days after the model establishment as well as emotional behavior 29 days after the model establishment were observed; the relative expressions of GABAA receptor mRNA in anterior cingulate cortex, amygdala and hypothalamus were observed. Compared with the blank control group, the change rates of mechanical pain threshold and thermal pain threshold in the model control group were decreased significantly 1 day, 3 days, 7 days, 14 days, 21 days, 28 days after model establishment (P<0.01); 29 days after model establishment, the movement distance and staying time in the central area of open field test in the model control group were decreased significantly (P<0.05). After EA intervention, compared with the model control group and the sham EA group, the change rates of mechanical pain threshold and thermal pain threshold, as well as the movement distance and the staying time of central area were significantly increased in the EA group (P<0.01, P<0.05). Twenty-nine days after model establishment, the expression of GABAA receptor mRNA in anterior cingulate cortex and hypothalamus was not significantly different among all groups (P>0.05). Compared with the blank control group, the expression of GABAA receptor mRNA in the amygdala was decreased significantly in the model control group (P<0.01); compared with the model control group and the sham EA group, the expression of GABAA receptor mRNA in amygdala was increased after intervention in the EA group (P<0.01). Single treatment of EA could significantly increase the mechanical pain threshold and thermal pain threshold, improve abnormal emotional behavior in rats with chronic inflammatory pain, which may be related to the increasing of expression of GABAA receptor mRNA in the amygdala.

Analgesic effect of Fengshi Qutong Capsules on mice with chronic inflammatory pain caused by complete Freund's adjuvant

The aim was to observe the analgesic effect of Fengshi Qutong Capsules(FSQTC) on chronic inflammatory pain in mice, and investigate its effect on p-ERK/COX-2 signal molecular activity. A model of chronic inflammatory pain was induced in mice by complete Freund's adjuvant(CFA). The mice were divided into normal control group, model group, model+FSQTC 0.3, 0.6 and 1.2 g·kg~(-1 )groups, model+positive control drug ibuprofen(IBP, 0.34 mg·kg~(-1)·d~(-1)) group, and normal control+ FSQTC 1.2 g·kg~(-1)group. FSQTC or IBP was given once a day by oral administration. Standard Von Frey fiber was used to evaluate the mechanical pain threshold, and the acetone stimulation was used to induce inflammatory plantar and observe the cold pain reaction scores. The mechanical pain threshold and cold pain reaction scores were observed before administration and 1, 2, 3, 4, 6 h after administration on the first day, as well as 3 h after administration on the 3 rd to 7 th day. The protein levels of PGE_2, COXs-1,2 and p-ERK in the spinal cord of the inflammatory foot and lumbar 4-5 were detected by enzyme-linked immunosorbent assay, Western blot, immunohistochemistry and immunofluorescence. The results showed that the mechanical pain threshold of the model group decreased and the cold pain reaction score increased as compared with the normal group. FSQTC application could dose-dependently increase the mechanical pain threshold and decrease the cold pain reaction score. The effect lasted for 6 h, most significant at 3 h. The effect of ibuprofen was similar to that of the 0.6 g·kg~(-1) dose group. In addition, FSQTC could reduce the abnormally increased protein content of PGE_2, COX-2 and p-ERK in the inflammatory foot and/or spinal cord of the model group, and the effect was most significant in middle and high dose groups. However, it had no effect on COX-1 in the inflammatory foot and spinal cord of mice. The results suggest that FSQTC has ob-vious analgesic effect on chronic inflammatory pain in mice, which may be related to inhibition of p-ERK/COX-2 signaling pathway.

Effect of m-trifluoromethyl-diphenyl diselenide on acute and subchronic animal models of inflammatory pain: Behavioral, biochemical and molecular insights

m-Trifluoromethyl-diphenyl diselenide [(m-CF3-PhSe)2] is an organoselenium molecule that displays multiple pharmacological actions, including the antinociceptive effect. The current study investigated the (m-CF3-PhSe)2 restorative properties in models of acute and chronic inflammatory pain induced by complete Freund's adjuvant (CFA). Male adult Swiss mice received an intraplantar injection of CFA in the hindpaw and 24 h (acute) or 14 days (subchronic) later they were treated with a single or repeated (m-CF3-PhSe)2 schedule via intragastric route, respectively. The mechanical and thermal hypernociceptive behaviors were assessed by von Frey hair and hot plate tests. Samples of injected paw were collected to evaluate the tissue edema and myeloperoxidase (MPO) activity while cerebral contralateral cortex samples were used to determine the inflammatory proteins content (subchronic protocol). The acute (m-CF3-PhSe)2 administration (1 and 10 mg/kg) reduced the hypernociceptive behavior and both paw thickness and MPO activity induced by CFA injection. In the subchronic protocol, the repeated administration with a low effective dosage of (m-CF3-PhSe)2 reduced the mechanical and thermal hypernociception as well as restored the edema and MPO activity in paw samples. In addition, the repeated treatment schedule mitigated the increase in TNF-α, IL-1β and COX-2 content in cerebral contralateral cortex induced by CFA injection. Collectively, these data showed that (m-CF3-PhSe)2 presents anti-inflammatory properties, which could be mediated by an interplay between peripheral and central mechanisms of action, reinforcing the potential biological properties of the compound.

The phthalimide analogues N-3-hydroxypropylphthalimide and N-carboxymethyl-3-nitrophthalimide exhibit activity in experimental models of inflammatory and neuropathic pain.

Phthalimide analogues devoid of the glutarimide moiety exhibit multiple biological activities, thus making them candidates for the treatment of patients with different diseases, including those with inflammatory and painful disorders. In the present study, the activities of five phthalimide analogues devoid of the glutarimide moiety (N-hydroxyphthalimide, N-hydroxymethylphthalimide, N-3-hydroxypropylphthalimide, N-carboxy-3-methylphthalimide, N-carboxymethyl-3-nitrophthalimide) were evaluated in experimental models of acute and chronic inflammatory and neuropathic pain. The phthalimide analogues were administered per os (po) in Swiss mice or Wistar rats. Nociceptive response induced by formaldehyde and mechanical allodynia induced by chronic constriction injury (CCI) of the sciatic nerve or intraplantar (ipl) injection of complete Freund's adjuvant (CFA) were used as experimental models of pain. N-carboxymethyl-3-nitrophthalimide (700 mg/kg, -1 h) inhibited the second phase of the nociceptive response induced by the intraplantar injection of formaldehyde in mice. N-3-hidroxypropylphthalimide (546 mg/kg, -1 h) inhibited both phases of the nociceptive response induced by formaldehyde. Treatment of rats with N-carboxymethyl-3-nitrophthalimide (700 mg/kg) or N-3-hydroxypropylphthalimide (546 mg/kg) inhibited the mechanical allodynia induced by CCI of the sciatic nerve or ipl injection of CFA in rats. Intraperitoneal administration of the opioid antagonist naltrexone (10 mg/kg, -1.5 h) attenuated the antinociceptive activity of N-carboxymethyl-3-nitrophthalimide (700 mg/kg) in the model of nociceptive response induced by formaldehyde. N-3-hydroxypropylphthalimide and N-carboxymethyl-3-nitrophthalimide, two phthalimide analogues devoid of the glutarimide moiety, exhibited activities in different experimental models of pain, including models of chronic inflammatory and neuropathic pain.

P.565 Characterisation of pain-, anxiety- and cognition-related behaviour in the complete Freund's adjuvant model of chronic inflammatory pain in Wistar-Kyoto rats

P.565 Characterisation of pain-, anxiety- and cognition-related behaviour in the complete Freund's adjuvant model of chronic inflammatory pain in Wistar-Kyoto rats

The analgesic effect of refeeding on acute and chronic inflammatory pain

Pain is susceptible to various cognitive factors. Suppression of pain by hunger is well known, but the effect of food intake after fasting (i.e. refeeding) on pain remains unknown. In the present study, we examined whether inflammatory pain behavior is affected by 24 h fasting and 2 h refeeding. In formalin-induced acute inflammatory pain model, fasting suppressed pain behavior only in the second phase and the analgesic effect was also observed after refeeding. Furthermore, in Complete Freund’s adjuvant-induced chronic inflammatory pain model, both fasting and refeeding reduced spontaneous pain response. Refeeding with non-calorie agar produced an analgesic effect. Besides, intraperitoneal (i.p.) administration of glucose after fasting, which mimics calorie recovery following refeeding, induced analgesic effect. Administration of opioid receptor antagonist (naloxone, i.p.) and cannabinoid receptor antagonist (SR 141716, i.p.) reversed fasting-induced analgesia, but did not affect refeeding-induced analgesia in acute inflammatory pain model. Taken together, our results show that refeeding produce analgesia in inflammatory pain condition, which is associated with eating behavior and calorie recovery effect.

Pharmacological mechanisms of alcohol analgesic-like properties in mouse models of acute and chronic pain

Alcohol use and chronic pain are highly comorbid. Acute alcohol use typically produces an analgesic effect. However, chronic use can worsen the progression of chronic pain. In rodent models, acute models of pain have primarily been used to investigate the relationship between alcohol and pain analgesia. Here, we use two models of chronic pain, chronic inflammatory and peripheral neuropathic pain, to investigate acute alcohol's antinociceptive and analgesic properties. We hypothesize that acute ethanol is acting through opioid receptors to create an analgesic-like effect in both reflexive and affective dimensions of pain.Using male and female C57BL/6J mice, oral ethanol administration (0–1.25 g/kg) showed a dose-dependent reversal of mechanical hypersensitivity in both Complete Freund's Adjuvant (CFA) and chronic constriction injury (CCI) models of chronic inflammatory and neuropathic pain. No sex differences were observed. Using the conditioned place preference (CPP) task to assess the subjective responses to ethanol's anti-nociceptive properties, CCI-injured animals showed a preference for the ethanol-paired side, suggesting a reduction in an aversive and pain-like state produced by nerve injury. These effects are likely mediated through the kappa and possibly the mu opioid systems, since ethanol-induced anti-nociception following CCI was fully reversed by pretreatment with the kappa selective antagonist, nor-BNI, or high doses of naltrexone. These data show that ethanol possesses analgesic-like properties in chronic inflammatory and neuropathic pain models in mice and provide new insight into ethanol as it relates to chronic pain.

Brain-derived neurotrophic factor stimulation of T-type Ca2+ channels in sensory neurons contributes to increased peripheral pain sensitivity.

Although brain-derived neurotrophic factor (BDNF) is implicated in the nociceptive signaling of peripheral sensory neurons, the underlying mechanisms remain largely unknown. Here, we elucidated the effects of BDNF on the neuronal excitability of trigeminal ganglion (TG) neurons and the pain sensitivity of rats mediated by T-type Ca2+ channels. BDNF reversibly and dose-dependently enhanced T-type channel currents through the activation of tropomyosin receptor kinase B (TrkB). Antagonism of phosphatidylinositol 3-kinase (PI3K) but not of its downstream target, the kinase AKT, abolished the BDNF-induced T-type channel response. BDNF application activated p38 mitogen-activated protein kinase (MAPK), and this effect was prevented by inhibition of PI3K but not of protein kinase A (PKA). Antagonism of either PI3K or p38 MAPK prevented the BDNF-induced stimulation of PKA activity, whereas PKA inhibition blocked the BDNF-mediated increase in T-type currents. BDNF increased the rate of action potential firing in TG neurons and enhanced the pain sensitivity of rats to mechanical stimuli. Moreover, inhibition of TrkB signaling abolished the increased mechanical sensitivity in a rat model of chronic inflammatory pain, and this effect was attenuated by either T-type channel blockade or knockdown of the channel Cav3.2. Together, our findings indicate that BDNF enhances T-type currents through the stimulation of TrkB coupled to PI3K-p38-PKA signaling, thereby inducing neuronal hyperexcitability of TG neurons and pain hypersensitivity in rats.

RETRACTED ARTICLE: Anti-hyperalgesic properties of a flavanone derivative Poncirin in acute and chronic inflammatory pain models in mice

BackgroundPoncirin is flavanone derivative (isolated from Poncirus trifoliata) with known pharmacological activities such as anti-tumor, anti-osteoporotic, anti-inflammatory and anti-colitic. The present study aimed to explore the anti-allodynic and anti-hyperalgesic potentials of poncirin in murine models of inflammatory pain.MethodsThe analgesic potential of poncirin was evaluated in formalin-, acetic acid-, carrageenan- and Complete Freund’s Adjuvant (CFA)-induced inflammatory pain models in mice. Anti-allodynic and anti-hyperalgesic activities were measured using Von Frey filaments, Randall Selitto, hotplate and cold acetone tests. The serum nitrite levels were determined using Griess reagent. The Quantitative Real-time PCR (qRT-PCR) was performed to assess the effect of poncirin on mRNA expression levels of inflammatory cytokines and anti-oxidant enzymes.ResultsIntraperitoneal administration of poncirin (30 mg/kg) markedly reduced the pain behavior in both acetic acid-induced visceral pain and formalin-induced tonic pain models used as preliminary screening tools. The poncirin (30 mg/kg) treatment considerably inhibited the mechanical hyperalgesia and allodynia as well as thermal hyperalgesia and cold allodynia. The qRT-PCR analysis showed noticeable inhibition of pro-inflammatory cytokines (mRNA expression levels of TNF-α, IL-1β and IL-6) (p < 0.05) in poncirin treated group. Similarly, poncirin treatment also enhanced the mRNA expressions levels of anti-oxidant enzymes such as transcription factor such as nuclear factor (erythroid-derived 2)-like 2 (Nrf2) (p < 0.05), heme oxygenase (HO-1) (p < 0.05) and superoxide dismutase (SOD2) (p < 0.05). Chronic treatment of poncirin for 6 days did not confer any significant hepatic and renal toxicity. Furthermore, poncirin treatment did not altered the motor coordination and muscle strength in CFA-induced chronic inflammatory pain model.ConclusionThe present study demonstrated that poncirin treatment significantly reduced pain behaviors in all experimental models of inflammatory pain, suggesting the promising analgesic potential of poncirin in inflammatory pain conditions.

Anti-hyperalgesic effect of (-)-α-bisabolol and (-)-α-bisabolol/β-Cyclodextrin complex in a chronic inflammatory pain model is associated with reduced reactive gliosis and cytokine modulation

Chronic pain is a continuous or recurring pain which exceeds the normal course of recovery to an injury or disease. According to the origin of the chronic pain, it can be classified as inflammatory or neuropathic. This study aimed to evaluate the antinociceptive and anti-inflammatory effect of (-)-α-bisabolol (BIS) alone and complexed with β-cyclodextrin (βCD) in preclinical models of chronic pain. Chronic pain was induced by Freund's Complete Adjuvant (FCA) or partial lesion of the sciatic nerve (PLSN). Swiss mice were treated with BIS, BIS-βCD (50 mg/kg, p.o) or vehicle (control) and mechanical hyperalgesia, thermal hyperalgesia, muscle strength and motor coordination were evaluated. In addition, levels of TNF-α and IL-10 and expression of the ionized calcium-binding adapter protein (IBA-1) were assessed in the spinal cord of the mice. The complexation efficiency of BIS in βCD was evaluated by High-Performance Liquid Chromatography. BIS and BIS-βCD reduced (p < 0.001) mechanical and thermal hyperalgesia. No alterations were found in force and motor coordination. In addition, BIS and BIS-βCD inhibited (p < 0.05) TNF-α production in the spinal cord and stimulated (p < 0.05) the release of IL-10 in the spinal cord in PLSN-mice. Further, BIS and BIS-βCD reduced IBA-1 immunostaining. Therefore, BIS and BIS-βCD attenuated hyperalgesia, deregulated cytokine release and inhibited IBA-1 expression in the spinal cord in the PLSN model. Moreover, our results show that the complexation of BIS in βCD reduced the therapeutic dose of BIS. We conclude that BIS is a promising molecule for the treatment of chronic pain.

Epigenetic suppression of liver X receptor \u03b2 in anterior cingulate cortex by HDAC5 drives CFA-induced chronic inflammatory pain

BackgroundLiver X receptors (LXRs), including LXRα and LXRβ, are key regulators of transcriptional programs for both cholesterol homeostasis and inflammation in the brain. Here, the modes of action of LXRs and the epigenetic mechanisms regulating LXRβ expression in anterior cingulate cortex (ACC) of chronic inflammatory pain (CIP) are investigated.MethodsThe deficit of LXR isoform and analgesic effect of LXR activation by GW3965 were evaluated using the mouse model of CIP induced by hindpaw injection of complete Freund’s adjuvant (CFA). The mechanisms involved in GW-mediated analgesic effects were analyzed with immunohistochemical methods, ELISA, co-immunoprecipitation (Co-IP), Western blot, and electrophysiological recording. The epigenetic regulation of LXRβ expression was investigated by chromatin immunoprecipitation, quantitative real-time PCR, and sequencing.ResultsWe revealed that CFA insult led to LXRβ reduction in ACC, which was associated with upregulated expression of histone deacetylase 5 (HDAC5), and knockdown of LXRβ by shRNA led to thermal hyperalgesia. Co-IP showed that LXRβ interacted with NF-κB p65 physically. LXRβ activation by GW3965 exerted analgesic effects by inhibiting the nuclear translocation of NF-κB, reducing the phosphorylation of mitogen-activated protein kinases (MAPKs) in ACC, and decreasing the promoted input-output and enhanced mEPSC frequency in ACC neurons after CFA exposure. In vitro experiments confirmed that HDAC5 triggered histone deacetylation on the promoter region of Lxrβ, resulting in downregulation of Lxrβ transcription.ConclusionThese findings highlight an epigenetic mechanism underlying LXRβ deficits linked to CIP, and LXRβ activation may represent a potential novel target for the treatment of CIP with an alteration in inflammation responses and synaptic transmission in ACC.

Protein Kinase C Lambda Mediates Acid-Sensing Ion Channel 1a-Dependent Cortical Synaptic Plasticity and Pain Hypersensitivity.

Chronic pain is a serious debilitating disease for which effective treatment is still lacking. Acid-sensing ion channel 1a (ASIC1a) has been implicated in nociceptive processing at both peripheral and spinal neurons. However, whether ASIC1a also contributes to pain perception at the supraspinal level remains elusive. Here, we report that ASIC1a in ACC is required for thermal and mechanical hypersensitivity associated with chronic pain. ACC-specific genetic deletion or pharmacological blockade of ASIC1a reduced the probability of cortical LTP induction and attenuated inflammatory thermal hyperalgesia and mechanical allodynia in male mice. Using cell type-specific manipulations, we demonstrate that ASIC1a in excitatory neurons of ACC is a major player in cortical LTP and pain behavior. Mechanistically, we show that ASIC1a tuned pain-related cortical plasticity through protein kinase C λ-mediated increase of membrane trafficking of AMPAR subunit GluA1 in ACC. Importantly, postapplication of ASIC1a inhibitors in ACC reversed previously established nociceptive hypersensitivity in both chronic inflammatory pain and neuropathic pain models. These results suggest that ASIC1a critically contributes to a higher level of pain processing through synaptic potentiation in ACC, which may serve as a promising analgesic target for treatment of chronic pain.SIGNIFICANCE STATEMENT Chronic pain is a debilitating disease that still lacks effective therapy. Ion channels are good candidates for developing new analgesics. Here, we provide several lines of evidence to support an important role of cortically located ASIC1a channel in pain hypersensitivity through promoting long-term synaptic potentiation in the ACC. Our results indicate a promising translational potential of targeting ASIC1a to treat chronic pain.

N-Pyrazoloyl and N-thiopheneacetyl hydrazone of isatin exhibited potent anti-inflammatory and anti-nociceptive properties through suppression of NF-κB, MAPK and oxidative stress signaling in animal models of inflammation.

Hydrazide derivatives constitute an important class of compounds for new drug development as they are reported to possess good anti-inflammatory and analgesic activity. The present study was aimed to investigate the role of newly synthesized hydrazide derivatives N-pyrazoloyl hydrazone of isatin (PHI) and N-thiopheneacetyl hydrazone of isatin (THI) in acute and chronic inflammatory pain models induced by carrageenan and complete Freud's adjuvant (CFA). PHI and THI (0.1, 1 and 10mg/kg) pretreatments were provided intraperitoneally to male BALB/c mice prior to inflammatory inducers. Behavioral responses to inflammation and pain were evaluated by assessment of paw edema, mechanical allodynia, mechanical and thermal hyperalgesia. Cytokines production and NF-κB levels were evaluated by ELISA. Western blot analysis was performed for the detection of IκBα, p38, JNK and ERK. Hematoxylin and eosin (H&E) staining and radiographic analysis were performed to evaluate the effect of PHI and THI treatment on bone and soft tissues. Oxidative stress was determined by reduced glutathione, glutathione-S-transferase and catalase assays. Evans blue dye was used to monitor vascular protein leakage. PHI and THI dose dependently (0.1, 1 and 10mg/kg) reduced inflammation and pain in mice, however, the dose of 10mg/kg exhibited significant activity. The anti-inflammatory and analgesic effects were attributed to suppression of pro-inflammatory cytokines (TNF-α, IL-1β, IL-6) productionlevels. PHI and THI significantly blocked CFA-induced activation of NF-κB and MAPK signaling pathways. Oxidative stress and plasma nitrite levels were reduced remarkably. The PHI and THI (10mg/kg) treatment did not exhibit any apparent toxicity on the liver, kidney, muscles strength, and motor co-ordination in mice. Both PHI and THI possess significant anti-inflammatory and analgesic activity via inhibition of inflammatory mediators.

γδ T Cells Modulate Myeloid Cell Recruitment but Not Pain During Peripheral Inflammation.

Circulating immune cells, which are recruited to the site of injury/disease, secrete various inflammatory mediators that are critical to nociception and pain. The role of tissue-resident immune cells, however, remains poorly characterized. One of the first cells to be activated in peripheral tissues following injury are γδT cells, which serve important roles in infection, disease, and wound healing. Using a mouse line lacking these cells, we sought to identify their contribution to inflammatory pain. Three distinct models of peripheral inflammatory pain were used: intraplantar injection of formalin (spontaneous inflammatory pain), incisional wound (acute inflammatory pain), and intraplantar injection of complete Freund's adjuvant (chronic inflammatory pain). Our results show that absence of γδT cells does not alter baseline sensitivity, nor does it result in changes to mechanical or thermal hypersensitivity after tissue injury. Myeloid cell recruitment did show differential changes between models of acute and chronic inflammatory pain. These results were consistent in both male and female mice, suggesting that there are no sex differences in these outcomes. This comprehensive characterization suggests that γδT cells do not contribute to basal sensitivity or the development and maintenance of inflammatory pain.

Convallatoxin enhance the ligand-induced mu-opioid receptor endocytosis and attenuate morphine antinociceptive tolerance in mice

Morphine is a unique opioid analgesic that activates the mu-opioid receptor (MOR) without efficiently promoting its endocytosis that may underlie side effects. Our objective was to discover a novel enhancer of ligand-induced MOR endocytosis and determine its effects on analgesia, tolerance and dependence. We used high-throughput screening to identify convallatoxin as an enhancer of ligand-induced MOR endocytosis with high potency and efficacy. Treatment of cells with convallatoxin enhanced morphine-induced MOR endocytosis through an adaptor protein 2 (AP2)/clathrin-dependent mechanism, attenuated morphine-induced phosphorylation of MOR, and diminished desensitization of membrane hyperpolarization. Furthermore, co-treatment with chronic convallatoxin reduced morphine tolerance in animal models of acute thermal pain and chronic inflammatory pain. Acute convallatoxin administration reversed morphine tolerance and dependence in morphine-tolerant mice. These findings suggest convallatoxin are potentially therapeutic for morphine side effects and open a new avenue to study MOR trafficking.

Global Gene Knockout of Kcnip3 Enhances Pain Sensitivity and Exacerbates Negative Emotions in Rats.

The Ca2+-binding protein Kv channel interacting protein 3 (KChIP3) or downstream regulatory element antagonist modulator (DREAM), a member of the neuronal calcium sensor (NCS) family, shows remarkable multifunctional properties. It acts as a transcriptional repressor in the nucleus and a modulator of ion channels or receptors, such as Kv4, NMDA receptors and TRPV1 channels on the cytomembrane. Previous studies of Kcnip3-/- mice have indicated that KChIP3 facilitates pain hypersensitivity by repressing Pdyn expression in the spinal cord. Conversely, studies from transgenic daDREAM (dominant active DREAM) mice indicated that KChIP3 contributes to analgesia by repressing Bdnf expression and attenuating the development of central sensitization. To further determine the role of KChIP3 in pain transmission and its possible involvement in emotional processing, we assessed the pain sensitivity and negative emotional behaviors of Kcnip3-/- rats. The knockout rats showed higher pain sensitivity compared to the wild-type rats both in the acute nociceptive pain model and in the late phase (i.e., 2, 4 and 6 days post complete Freund’s adjuvant injection) of the chronic inflammatory pain model. Importantly, Kcnip3-/- rats displayed stronger aversion to the pain-associated compartment, higher anxiety level and aggravated depression-like behavior. Furthermore, RNA-Seq transcriptional profiling of the forebrain cortex were compared between wild-type and Kcnip3-/- rats. Among the 68 upregulated genes, 19 genes (including Nr4a2, Ret, Cplx3, Rgs9, and Itgad) are associated with neural development or synaptic transmission, particularly dopamine neurotransmission. Among the 79 downregulated genes, 16 genes (including Col3a1, Itm2a, Pcdhb3, Pcdhb22, Pcdhb20, Ddc, and Sncaip) are associated with neural development or dopaminergic transmission. Transcriptional upregulation of Nr4a2, Ret, Cplx3 and Rgs9, and downregulation of Col3a1, Itm2a, Pcdhb3 and Ddc, were validated by qPCR analysis. In summary, our studies showed that Kcnip3-/- rats displayed higher pain sensitivity and stronger negative emotions, suggesting an involvement of KChIP3 in negative emotions and possible role in central nociceptive processing.

Epigenetic control of hypersensitivity in chronic inflammatory pain by the de novo DNA methyltransferase Dnmt3a2.

Chronic pain is a pathological manifestation of neuronal plasticity supported by altered gene transcription in spinal cord neurons that results in long-lasting hypersensitivity. Recently, the concept that epigenetic regulators might be important in pathological pain has emerged, but a clear understanding of the molecular players involved in the process is still lacking. In this study, we linked Dnmt3a2, a synaptic activity-regulated de novo DNA methyltransferase, to chronic inflammatory pain. We observed that Dnmt3a2 levels are increased in the spinal cord of adult mice following plantar injection of Complete Freund’s Adjuvant, an in vivo model of chronic inflammatory pain. In vivo knockdown of Dnmt3a2 expression in dorsal horn neurons blunted the induction of genes triggered by Complete Freund’s Adjuvant injection. Among the genes whose transcription was found to be influenced by Dnmt3a2 expression in the spinal cord is Ptgs2, encoding for Cox-2, a prime mediator of pain processing. Lowering the levels of Dnmt3a2 prevented the establishment of long-lasting inflammatory hypersensitivity. These results identify Dnmt3a2 as an important epigenetic regulator needed for the establishment of central sensitization. Targeting expression or function of Dnmt3a2 may be suitable for the treatment of chronic pain.

Mansoa alliacea extract presents antinociceptive effect in a chronic inflammatory pain model in mice through opioid mechanisms

In some chronic disorders, as in arthritis, the inflammatory pain persists beyond the inflammation control becoming pathological. Its treatment shows limited efficacy and adverse effects which compromises patients' quality of life. Mansoa alliacea, known as ‘cipo alho’, is popularly used as analgesic and others species of this genus show anti-inflammatory actions. We investigated the anti-inflammatory and antinociceptive potential of M. alliacea extract in an inflammatory pain model which presents inflammatory characteristics similar to those caused by arthritis, through of the intraplantar injection of complete Freund's adjuvant (CFA) in mice. The extract chromatographic analysis revealed the presence of ρ-coumaric, ferulic and chlorogenic acids, luteolin, and apigenin. The treatment with M. alliacea prevented and reversed the CFA-induced mechanical allodynia with maximum inhibition (Imax) of 100% and 90 ± 10%, respectively. The co-administration of M. alliacea extract plus morphine enhanced the anti-allodynic effect with Imax of 100%. The M. alliacea extract also reverted the CFA-induced thermal hyperalgesia with Imax of 3.6 times greater compared to the vehicle and reduced the thermal threshold under physiological conditions. However, M. alliacea extract did not reduce the CFA-induced edema and myeloperoxidase activity. Additionally, non-selective and δ-selective opioid receptor antagonists, but not κ-opioid, prevented extract anti-allodynic effect with Imax of 98 ± 2% and 93 ± 2%, respectively. Moreover, M. alliacea extract did not induce adverse effects commonly caused by opioids and other analgesic drugs, at least in the tested pharmacological doses after the acute treatment. M. alliacea extract presents antinociceptive activity in an inflammatory pain model, which presents inflammatory characteristics similar to those arthritis-induced, without causing adverse effects in tested pharmacological doses. These effects seem to be mediated mainly via δ-opioid receptors.

Concentrations of oxidized linoleic acid derived lipid mediators in the amygdala and periaqueductal grey are reduced in a mouse model of chronic inflammatory pain

Chronic pain is both a global public health concern and a serious source of personal suffering for which current treatments have limited efficacy. Recently, oxylipins derived from linoleic acid (LA), the most abundantly consumed polyunsaturated fatty acid in the modern diet, have been implicated as mediators of pain in the periphery and spinal cord. However, oxidized linoleic acid derived mediators (OXLAMs) remain understudied in the brain, particularly during pain states. In this study, we employed a mouse model of chronic inflammatory pain followed by a targeted lipidomic analysis of the animals’ amygdala and periaqueductal grey (PAG) using LC-MS/MS to investigate the effect of chronic inflammatory pain on oxylipin concentrations in these two brain nuclei known to participate in pain sensation and perception. From punch biopsies of these brain nuclei, we detected twelve OXLAMs in both the PAG and amygdala and one arachidonic acid derived mediator, 15-HETE, in the amygdala only. In the amygdala, we observed an overall decrease in the concentration of the majority of OXLAMs detected, while in the PAG the concentrations of only the epoxide LA derived mediators, 9,10-EpOME and 12,13-EpOME, and one trihydroxy LA derived mediator, 9,10,11-TriHOME, were reduced. This data provides the first evidence that OXLAM concentrations in the brain are affected by chronic pain, suggesting that OXLAMs may be relevant to pain signaling and adaptation to chronic pain in pain circuits in the brain and that the current view of OXLAMs in nociception derived from studies in the periphery is incomplete.