Spinal CCK1 Receptors Contribute to Somatic Pain Hypersensitivity Induced by Malocclusion via a Reciprocal Neuron-Glial Signaling Cascade.

Recent studies have shown that the incidence of chronic primary pain including temporomandibular disorders (TMD) and fibromyalgia syndrome (FMS) often exhibit comorbidities. We recently reported that central sensitization and descending facilitation system contributed to the development of somatic pain hypersensitivity induced by orofacial inflammation combined with stress. The purpose of this study was to explore whether TMD caused by unilateral anterior crossbite (UAC) can induce somatic pain hypersensitivity, and whether the cholecystokinin (CCK) receptor-mediated descending facilitation system promotes hypersensitivity through neuron-glia cell signaling cascade. UAC evoked thermal and mechanical pain hypersensitivity of the hind paws from day 5 to 70 that peaked at week 4 post UAC. The expression levels of CCK1 receptors, interleukin-18 (IL-18) and IL-18 receptors (IL-18R) were significantly up-regulated in the L4 to L5 spinal dorsal horn at 4 weeks post UAC. Intrathecal injection of CCK1 and IL-18 receptor antagonists blocked somatic pain hypersensitivity. IL-18 mainly co-localized with microglia, while IL-18R mainly co-localized with astrocytes and to a lesser extent with neurons. These findings indicate that the signaling transduction between neurons and glia at the spinal cord level contributes to the descending pain facilitation through CCK1 receptors during the development of the comorbidity of TMD and FMS. PERSPECTIVE: CCK1 receptor-dependent descending facilitation may mediate central mechanisms underlying the development of widespread somatic pain via a reciprocal neuron-glial signaling cascade, providing novel therapeutic targets for the clinical treatment of TMD and FMS comorbidities.

Low-intensity low-frequency pulsed ultrasound ameliorates sciatic nerve dysfunction in a rat model of cisplatin-induced peripheral neuropathy.

Chemotherapy-induced peripheral neuropathy is a neurological complication that frequently occurs during chemotherapeutic intervention, resulting in damaged myelin sheath, motor weakness and/or sensory impairment. This study aims to investigate the therapeutic efficiency of low-intensity pulsed low-frequency ultrasound on cisplatin-induced peripheral neuropathy. Rats were randomly divided into five experimental groups as control, cisplatin administration, 10 mg/kg melatonin treatment after cisplatin administration, 1 MHz frequency 0.5 W/cm2 pulsed ultrasound treatment after cisplatin administration and 1 MHz frequency 1.5 W/cm2 pulsed ultrasound treatment after cisplatin administration. Chemical neuropathy was induced by the injection of 3 mg/kg/week of cisplatin (i.p.) for 5 weeks. Afterwards, melatonin and pulsed ultrasound treatments were applied for 15 consecutive days. Cisplatin administration resulted in a decrease in nociceptive pain perception and nerve conduction velocities together with a decrease in myelin thickness and diameters of axons and myelinated fibers, indicating a dysfunction and degeneration in sciatic nerves. In addition, cisplatin administration led to a decrease, in superoxide dismutase activity, and an increase in malondialdehyde and IL-1ß levels together with an increase in caspase-3 protein expression levels and a decrease in Bcl-2 and Parkin levels. The ultrasound treatments resulted in an increase in nociceptive pain perception and sciatic nerve conduction; led to a decrease in oxidative stress and inflammation, restored nerve degeneration and regulated apoptosis and mitophagy. Taken together, low-intensity pulsed low-frequency ultrasound was efficient in restoring the alterations attributable to cisplatin-induced peripheral neuropathy, and warrants further investigations.

Acid-sensing ion channels modulate bladder nociception.

Sensitization of neuronal pathways and persistent afferent drive are major contributors to somatic and visceral pain. However, the underlying mechanisms that govern whether afferent signaling will give rise to sensitization and pain are not fully understood. In the present report, we investigated the contribution of acid-sensing ion channels (ASICs) to bladder nociception in a model of chemical cystitis induced by cyclophosphamide (CYP). We found that the administration of CYP to mice lacking ASIC3, a subunit primarily expressed in sensory neurons, generates pelvic allodynia at a time point at which only modest changes in pelvic sensitivity are apparent in wild-type mice. The differences in mechanical pelvic sensitivity between wild-type and Asic3 knockout mice treated with CYP were ascribed to sensitized bladder C nociceptors. Deletion of Asic3 from bladder sensory neurons abolished their ability to discharge action potentials in response to extracellular acidification. Collectively, the results of our study support the notion that protons and their cognate ASIC receptors are part of a mechanism that operates at the nerve terminals to control nociceptor excitability and sensitization.NEW & NOTEWORTHY Our study indicates that protons and their cognate acid-sensing ion channel receptors are part of a mechanism that operates at bladder afferent terminals to control their function and that the loss of this regulatory mechanism results in hyperactivation of nociceptive pathways and the development of pain in the setting of chemical-induced cystitis.

IL-23/IL-17A/TRPV1 axis produces mechanical pain via macrophage-sensory neuron crosstalk in female mice.

Although sex dimorphism is increasingly recognized as an important factor in pain, female-specific pain signaling is not well studied. Here we report that administration of IL-23 produces mechanical pain (mechanical allodynia) in female but not male mice, and chemotherapy-induced mechanical pain is selectively impaired in female mice lacking Il23 or Il23r. IL-23-induced pain is promoted by estrogen but suppressed by androgen, suggesting an involvement of sex hormones. IL-23 requires C-fiber nociceptors and TRPV1 to produce pain but does not directly activate nociceptor neurons. Notably, IL-23 requires IL-17A release from macrophages to evoke mechanical pain in females. Low-dose IL-17A directly activates nociceptors and induces mechanical pain only in females. Finally, deletion of estrogen receptor subunit α (ERα) in TRPV1+ nociceptors abolishes IL-23- and IL-17-induced pain in females. These findings demonstrate that the IL-23/IL-17A/TRPV1 axis regulates female-specific mechanical pain via neuro-immune interactions. Our study also reveals sex dimorphism at both immune and neuronal levels.

TREK-1 potassium channels participate in acute and long-lasting nociceptive hypersensitivity induced by formalin in rats.

TREK-1 channels are expressed in small nociceptive dorsal root ganglion (DRG) neurons where they participate in acute inflammatory and neuropathic pain. However, the role of TREK-1 in persistent pain is not well understood. The aim of this study was to investigate the local peripheral and spinal participation of TREK-1 in formalin-induced acute and long-lasting nociceptive hypersensitivity. Local peripheral or intrathecal pre-treatment with spadin, selective blocker of TREK-1, increased acute flinching behavior and secondary mechanical allodynia and hyperalgesia behavior observed 6 days after formalin injection. Local peripheral or intrathecal pre-treatment with BL-1249, selective opener of TREK-1, decreased long-lasting secondary mechanical allodynia and hyperalgesia induced by formalin. Pre-treatment with BL-1249 prevented the pro-nociceptive effect of spadin on acute nociception and long-lasting mechanical allodynia and hyperalgesia in rats. Pre-treatment with two recombinant channels that produce a high TREK-1 current, S300A and S333A (non-phosphorylated state of TREK-1), reduced formalin-induced acute pain and long-lasting mechanical allodynia and hyperalgesia. Besides, post-treatment with S300A, S333A or BL-1249 reversed long-lasting mechanical allodynia and hyperalgesia induced by formalin. Formalin increased TREK-1 expression at 1 and 6 days in DRG and dorsal spinal cord in rats, whereas that it increased c-fos expression at the DRG. Intrathecal repeated transfection of rats with S300A and S333A or injection with BL-1249 reduced formalin-induced enhanced c-fos expression. Data suggest that TREK-1 activity at peripheral and spinal sites reduces neuronal excitability in the process of acute and long-lasting nociception induced by formalin in rats.

Selective blockade of transient receptor potential vanilloid 4 reduces cyclophosphamide-induced bladder pain in mice.

Transient receptor potential vanilloid 4 (TRPV4) is a non-selective cation channel activated by various physical stimuli such as cell swelling and shear stress. TRPV4 is expressed in bladder sensory nerves and epithelium, and its activation produces urinary dysfunction in rodents. However, there have been few reports regarding its involvement in bladder pain. Therefore, we investigated whether TRPV4 is involved in bladder pain in mouse cystitis model. Intraperitoneal injection of cyclophosphamide (CYP; 300 mg/kg) produced mechanical hypersensitivity in the lower abdomen associated with a severe inflammatory bladder in mice. The mechanical threshold was reversed significantly in Trpv4-knockout (KO) mice. Repeated injections of CYP (150 mg/kg) daily for 4 days provoked mild bladder inflammation and persistent mechanical hypersensitivity in mice. Trpv4-KO mice prevented a reduction of the mechanical threshold without an alteration in bladder inflammation. A selective TRPV4 antagonist also reversed the mechanical threshold in chronic cystitis mice. Although expression of Trpv4 was unchanged in the bladders of chronic cystitis mice, the level of phosphorylated TRPV4 was increased significantly. These results suggest involvement of TRPV4 in bladder pain of cystitis mice. A TRPV4 antagonist might be useful for patients with irritable bladder pain such as those with interstitial cystitis/painful bladder syndrome.

Involvement of TCF7L2 in generation of morphine-induced antinociceptive tolerance and hyperalgesia by modulating TLR4/ NF-κB/NLRP3 in microglia.

Morphine is an opioid agonist and a nonselective mu, kappa and delta receptor agonist. It is a commonly used analgesic drug for the treatment of acute and chronic pain as well as cancer pain. Morphine is particularly important to address the problem of morphine tolerance. Tcf7l2, known as a risk gene for schizophrenia and autism, encodes a member of the LEF1/TCF transcription factor family. TCF7L2 is an important transcription factor that is upregulated in neuropathic pain models. However, the relationship between TCF7L2 and morphine tolerance has not been reported. In this study, we found that morphine tolerance led to the upregulation of TCF7L2 in the spinal cord, and also led to the upregulation of TCF7L2 expression in glial cells, which promoted inflammation related signal, and activated TLR4 / NF-κB/NLRP3 pathway. In addition, TCF7L2 regulated microglial cell activation induced by chronic morphine treatment. Mechanically, we found that TCF7L2 transcriptionally regulated TLR4 expression, and the depletion of TCF7L2 alleviated morphine tolerance induced by chronic morphine treatment, and further alleviated pain hypersensitivity induced by chronic morphine treatment. We therefore suggested that TCF7L2 regulates the activation of TLR4/ NF-κB/NLRP3 pathway in microglia, and is involved in the formation of morphine tolerance. Our results provide a new idea for the regulation mechanism of morphine tolerance.

TNF-α, CXCL-1 and IL-1 ß as activators of the opioid system involved in peripheral analgesic control in mice.

Tissue injury results in the release of inflammatory mediators, including a cascade of nociceptive substances, which contribute to development of hyperalgesia. In addition, during this process endogenous analgesic substances are also peripherally released with the aim of controlling the hyperalgesia. Thus, the present study aimed to investigate whether inflammatory mediators TNF-α, IL-1ß, CXCL1, norepinephrine (NE) and prostaglandin E2 (PGE2) may be involved in the deflagration of peripheral endogenous modulation of inflammatory pain by activation of the opioid system. Thus, male Swiss mice and the paw withdrawal test were used. All substances were injected by the intraplantar route. Carrageenan, TNF-α, CXCL-1, IL1-ß, NE and PGE2 induced hyperalgesia. Selectives µ (clocinamox), δ (naltrindole) and κ (norbinaltorphimine, nor-BNI) and non-selective (naloxone) opioid receptor antagonists potentiated the hyperalgesia induced by carrageenan, TNF-α, CXCL-1 and IL1-ß. In contrast, when the enzyme N-aminopeptidase involved in the degradation of endogenous opioid peptides was inhibited by bestatin, the hyperalgesia was significantly reduced. In addition, the western blotting assay indicated that the expression of the opioid δ receptor was increased after intraplantar injection of carrageenan. The data obtained in this work corroborate the hypothesis that TNF-α, CXCL-1 and IL-ß cause, in addition to hyperalgesia, the release of endogenous substances such as opioid peptides, which in turn exert endogenous control over peripheral inflammatory pain.

Novel selective agonist of GPR18, PSB-KK-1415 exerts potent anti-inflammatory and anti-nociceptive activities in animal models of intestinal inflammation and inflammatory pain.

BACKGROUND: GPR18 is a recently deorphanized receptor which was reported to act with several endogenous cannabinoid ligands. Here, we aimed to describe the role of GPR18 in intestinal inflammation and inflammatory pain. METHODS: The anti-inflammatory activity of selective GPR18 agonist, PSB-KK-1415, and antagonist, PSB-CB5, was characterized in semi-chronic and chronic mouse models of colitis induced by 2,4,6-trinitrobenzenesulfonic acid (TNBS). The extent of inflammation was evaluated based on the macroscopic and microscopic scores, quantification of myeloperoxidase (MPO) activity, and Western blot analyses of tumor necrosis factor-α (TNF-α) and interleukin-6 in colonic tissue. The expression of GPR18 in colonic samples from patients with Crohn's disease (CD) was quantified using real-time PCR. The anti-nociceptive potential of the agonist in intestinal inflammation was evaluated in the mouse model of inflammatory pain. KEY RESULTS: In semi-chronic colitis, PSB-KK-1415 reduced macroscopic score (1.79 ± 0.22 vs. 2.61 ± 0.48), expression of TNF-α (1.89 ± 0.36 vs. 2.83 ± 0.64), and microscopic score (5.00 ± 0.33 vs. 6.45 ± 0.40), all compared to mice with colitis. In chronic colitis, PSB-KK-1415 decreased macroscopic score (3.33 ± 1.26 vs. 4.00 ± 1.32) and MPO activity (32.23 ± 8.51 vs. 41.33 ± 11.64) compared to inflamed mice. In the mouse model of inflammatory pain, PSB-KK-1415 decreased the number of pain-induced behaviors in both, controls (32.60 ± 2.54 vs. 58.00 ± 6.24) and inflamed mice (60.83 ± 2.85 vs. 85.00 ± 5.77) compared to animals without treatment with PSB-KK-1415 (P < 0.005 for both). Lastly, we showed an increased expression of GPR18 in CD patients compared to healthy controls (3.77 ± 1.46 vs. 2.38 ± 0.66, p = 0.87). CONCLUSIONS & INFERENCES: We showed that GPR18 is worth considering as a potential treatment target in intestinal inflammation and inflammatory pain.

Synergistic interaction between haloperidol and gabapentin in a model of neuropathic nociception in rat.

Preclinical studies have reported that sigma-1 receptor antagonists may have efficacy in neuropathic pain states. The sigma-1 receptor is a unique ligand-operated chaperone present in crucial areas for pain control, in both the peripheral and central nervous system. This study assesses the synergistic antihyperalgesic and antiallodynic effect of haloperidol, a sigma-1 antagonist, combined with gabapentin in rats with peripheral neuropathy. Wistar rats male were subjected to chronic constriction injury (CCI) of the sciatic nerve. The effects of systemic administration of gabapentin and the sigma-1 receptor antagonist, haloperidol, were examined at 11 days post-CCI surgery. An analysis of Surface of Synergistic Interaction was used to determine whether the combination's effects were synergistic. Twelve combinations showed various degrees of interaction in the antihyperalgesic and antiallodynic effects. In hyperalgesia, three combinations showed additive effects, four combinations showed supra-additive effects, and three combinations produced an effect limited by the maximum effect. In allodynia, five combinations showed additive effects, two combinations showed supra-additive effects, and five combinations produced antihyperalgesic effects limited by the maximum effect. These findings indicate that the administration of some specific combination of gabapentin and haloperidol can synergistically reduce nerve injury-induced allodynia and hyperalgesia. This suggests that the haloperidol-gabapentin combination can improve the antiallodynic and antihyperalgesic effects in a neuropathic pain model.

Effects of Transcutaneous Auricular Vagus Nerve Stimulation on Peripheral and Central Tumor Necrosis Factor Alpha in Rats with Depression-Chronic Somatic Pain Comorbidity.

Depression and pain disorders share a high degree of comorbidity. Inflammatory mechanisms play an important role in the pathogenesis of depression-chronic somatic pain comorbidity. In this study, we investigated the effects of acupuncture on blood and brain regional tumor necrosis factor alpha (TNF-α) in rats with depression and chronic somatic pain comorbidity. Forty Sprague-Dawley rats were randomly divided into the following 4 groups with 10 each: control, model, model treated with transcutaneous auricular vagus nerve stimulation (taVNS), and model treated with electroacupuncture (EA). Chronic unpredictable mild stress (CUMS) with chronic constriction injury of the sciatic nerve (CCI) was used to produce depression and chronic somatic pain comorbidity in the latter 3 groups. The rats of the taVNS and EA groups received, respectively, taVNS and EA at ST 36 for 28 days. Pain intensity was measured using a mechanical withdrawal threshold and thermal stimulation latency once biweekly. Depressive behavior was examined using a sucrose preference test at baseline and the end of modeling and intervention. The level of plasma TNF-α and the expression of TNF-α in the prefrontal cortex (PFC), hippocampus, amygdala, and hypothalamus were measured. While CUMS plus CCI produced remarkable depression-like behavior and pain disorders, EA and taVNS significantly improved depression and reduced pain intensity. CUMS plus CCI also resulted in a significant increase in plasma TNF-α level and the expression in all brain regions examined compared to the intact controls. Both EA and taVNS interventions, however, suppressed the elevated level of TNF-α. These results suggest that EA and taVNS have antidepressant and analgesic effects. Such effects may be associated with the suppression of TNF-α-related neuroinflammation.

Sialorphin Potentiates Effects of [Met5]Enkephalin without Toxicity by Action other than Peptidase Inhibition.

This dose-response study investigated the effects of sialorphin on [Met5]enkephalin (ME)-induced inhibition of contractions in mouse vas deferens and antinociception in male rats. Differences were compared among combinations of three chemical peptidase inhibitors: amastatin, captopril, and phosphoramidon. The ratio of potencies of ME in mouse vas deferens pretreated with both sialorphin (100 µM) and a mixture of the three peptidase inhibitors (1 µM each) was higher than that with the mixture of peptidase inhibitors alone at any dose. Intrathecal administration of sialorphin (100-400 nmol) significantly and dose dependently increased ME (3 nmol)-induced antinociception with the mixture of three peptidase inhibitors (10 nmol each). The degree of antinociception with a combination of any two of the peptidase inhibitors (10 nmol each) in the absence of sialorphin was less than that in the presence of sialorphin (200 nmol). Pretreatment with both sialorphin (200 nmol) and the mixture of three peptidase inhibitors (10 nmol each) produced an approximately 100-fold augmentation in ME (10 nmol)-induced antinociception, but without signs of toxicity such as motor dysfunction in rats. Radioligand receptor binding assay revealed that sialorphin did not affect either binding affinity or maximal binding capacity of [d-Ala2,N-MePhe4,Gly-ol5]enkephalin. These results indicate that sialorphin potentiates the effects of ME without toxicity by a mechanism other than peptidase inhibition and with no effect on its affinity to µ-opioid receptors. SIGNIFICANCE STATEMENT: Sialorphin is regarded as an endogenous peptidase inhibitor that interacts with enkephalin-degrading enzymes. The results of these in vitro and in vivo studies confirm that sialorphin potentiates the effects of [Met5]enkephalin without toxicity by an action other than peptidase inhibition. This suggests that sialorphin offers the advantage of reducing or negating the side effects of opioid drugs and endogenous opioid peptides.

The leaves of Bougainvillea spectabilis suppressed inflammation and nociception in vivo through the modulation of glutamatergic, cGMP, and ATP-sensitive K+ channel pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Bougainvillea spectabilis is an ornamental shrub from Nyctaginaceae family, widely used in the traditional medicine in the treatment of pain, inflammation, and ulcer. Some research investigated the analgesic potential of this plant, however, the in-depth analysis of its antinociceptive properties and molecular mechanism(s) are yet to be revealed. PURPOSE OF THE STUDY: This study, therefore, investigated the antinociceptive potential of methanol extract of the leaves of B. spectabilis (MEBS) with possible molecular mechanism(s) of action using several pre-clinical models of acute and chronic pain in mice. MATERIALS AND METHODS: The dry leaf powder of B. spectabilis was macerated with 100% methanol, and then dried crude extract was used for in vivo experiments. Following the acute toxicity test with 500, 1000, and 2000 mg/kg b.w. doses of MEBS, the central antinociceptive activities of the extract (50, 100, and 200 mg/kg b.w.) were evaluated using hot plate and tail immersion tests, whereas the peripheral activities were investigated using acetic acid-induced writhing, formalin-induced licking and oedema, and glutamate-induced licking tests. Moreover, the possible involvements of cGMP and ATP-sensitive K+ channel pathways in the observed antinociceptive activities were also investigated using methylene blue (20 mg/kg b.w.) and glibenclamide (10 mg/kg b.w.), respectively. We also performed GC/MS-MS analysis of MEBS to identify the phyto-constituents and in silico modelling of the major compounds for potential molecular targets. RESULTS: Our results demonstrated that MEBS at 50, 100, and 200 mg/kg b.w. doses were not effective enough to suppress centrally mediated pain in the hot plate and tail immersion models. However, the extract was potent (at 100 and 200 mg/kg b.w. doses) in reducing peripheral nociception in the acetic acid-induced writhing and inflammatory phase of the formalin tests. Further analyses revealed that MEBS could interfere with glutamatergic system, cGMP and ATP-sensitive K+ channel pathways to show its antinociceptive properties. GC/MS-MS analysis revealed 35 different phytochemicals with potent anti-inflammatory and antinociceptive properties including phytol, neophytadiene, 2,4-Di-tert-butylphenol, fucoxanthin, and Vit-E. Prediction analysis showed high intestinal absorptivity and low toxicity profiles of these compounds with capability to interact with glutamatergic system, inhibit JAK/STAT pathway, scavenge nitric oxide and oxygen radicals, and inhibit expression of COX3, tumor necrosis factor, and histamine. CONCLUSION: Taken together, these results suggested the antinociceptive potentials of MEBS which were mediated through the modulation of glutamatergic, cGMP, and ATP-sensitive K+ channel pathways. These also suggested that MEBS could be beneficial in the treatment of complications associated with nociceptive pain.

Changes in opioid receptors, opioid peptides and morphine antinociception in mice subjected to early life stress.

Recent studies have shown that the endogenous opioid system is considerably affected by early life stress such as child abuse. Here, we investigated whether early life stress changes the endogenous opioid receptors and their peptides, and if such stress impacts morphine antinociception. We used mice affected by maternal separation and social isolation (MSSI) as an early life stress model. In the tail-flick test, 10-week-old MSSI mice showed a significant decrease in morphine antinociception compared to age-matched control mice. The number of c-Fos-positive cells increased in the periaqueductal gray (PAG), nucleus accumbens, and thalamus of control mice after the morphine injections, whereas hardly any positive cells were detected in the same areas of MSSI mice. The expression of µ- and κ-opioid receptor (MOR and KOR, respectively) messenger RNA (mRNA) was significantly decreased in the PAG of MSSI mice, whereas KOR expression was significantly increased in the amygdala of MSSI mice. The expression of δ-opioid receptor (DOR) mRNA was significantly reduced in the PAG and rostral ventromedial medulla of MSSI mice compared to control mice. Moreover, the lack of morphine antinociception was observed in 18-week-old MSSI mice. Our findings suggest that the supraspinal opioid system may be affected by early life stress exposure, and that this exposure may impact morphine antinociception.

Antinociceptive activity of Schinus terebinthifolia leaf lectin (SteLL) in sarcoma 180-bearing mice.

ETHNOPHARMACOLOGY RELEVANCE: Schinus terebinthifolia Raddi leaves have been used in folk medicine due to several properties, including antitumor and analgesic effects. The variable efficacy and adverse effects of analgesic drugs have motivated the search for novel antinociceptive agents. It has been reported that the S. terebinthifolia leaf lectin (SteLL) has antitumor activity against sarcoma 180 in mice. AIM OF THE STUDY: This work aimed to evaluate whether SteLL would reduce cancer pain using an orthotopic tumor model. MATERIALS AND METHODS: A sarcoma 180 cell suspension was inoculated into the right hind paws of mice, and the treatments (150 mM NaCl, negative control; 10 mg/kg morphine, positive control; or SteLL at 1 and 2 mg/kg) were administered intraperitoneally 24 h after cell inoculation up to 14 days. Spontaneous nociception, mechanical hyperalgesia, and hot-plate tests were performed. Further, the volume and weight of the tumor-bearing paws were measured. RESULTS: SteLL (2 mg/kg) improved limb use during ambulation. The lectin (1 and 2 mg/kg) also inhibited mechanical hyperalgesia and increased the latency time during the hot-plate test. Naloxone was found to reverse this effect, indicating the involvement of opioid receptors. The tumor-bearing paws of mice treated with SteLL exhibited lower volume and weight. CONCLUSION: SteLL reduced hyperalgesia due to sarcoma 180 in the paws of mice, and this effect can be related to its antitumor action.

Anti-nociceptive and antioxidant activity of betaine on formalin- and writhing tests induced pain in mice.

Pain is a physiological response which is mediated via the central and peripheral nervous system. Betaine, is a methyl glycine derivative and a commonly used nutrient supplement. The main purpose of the current paper is to determine the possible anti-nociceptive and antioxidant activity and sedative effect of betaine in mice. Adult male albino mice were divided into two categories, formalin and writhing tests. In the formalin test, mice were injected with betaine (10, 20 and 30 mg/kg) or morphine (5 mg/kg). For co-injections mice received betaine (30 mg/kg) + naloxone (2 mg/kg) or atropine (1 mg/kg), chlorpheniramine (20 mg/kg), flumazenil (5 mg/kg), cimetidine (12.5 mg/kg) and cyproheptadine (4 mg/kg). Then the formalin test was done and paw licking time was determined. In the writhing test, injections were the same but the animals were injected with acetic acid (0.6 %) and the percentage of writhing inhibition was recorded. At the end of the study, blood antioxidant levels were determined. According to the results, betaine reduced the pain response in a dose-dependent manner. Co-administration of the naloxone + betaine or flumazenil + betaine significantly decreased the anti-nociceptive effect of betaine on the licking and biting time of the injected paw and inhibited the number of writhing movements. Betaine decreased malondialdehyde (MDA) and improved superoxide dismutase (SOD) and glutathione peroxidase (GPx) levels in formalin receiving mice. No adverse locomotion and sedation effect were observed in betaine-treated mice. These findings suggest that betaine has anti-nociceptive and antioxidant activity in mice, and its anti-nociceptive role interacts with opioidergic and GABA receptors.

Antinociceptive, antiedematous, and antiallodynic activity of 1H-pyrrolo[3,4-c]pyridine-1,3(2H)-dione derivatives in experimental models of pain.

The aim of the presented study was to examine the potential antinociceptive, antiedematous (anti-inflammatory), and antiallodynic activities of two 1H-pyrrolo[3,4-c]pyridine-1,3(2H)-dione derivatives (DSZ 1 and DSZ 3) in various experimental models of pain. For this purpose, the hot plate test, the capsaicin test, the formalin test, the carrageenan model, and oxaliplatin-induced allodynia tests were performed. In the hot plate test, only DSZ 1 in the highest dose (20 mg/kg) was active but its effects appear to be due to sedatation rather than antinociceptiveness. In capsaicin-induced neurogenic pain model, both compounds displayed a significant antinociceptive activity. In the formalin test, DSZ 1 and DSZ 3 (5-20 mg/kg) revealed antinociceptive activity in both phases but it was more pronounced in the second phase of the test. In this test, pretreatment with caffeine, DPCPX reversed the antinociceptive effect of DSZ 3. On the other hand, pretreatment with L-NAME diminished the antinociceptive effect of DSZ 1. Pretreatment with naloxone did not affect antinociceptive activity of both compounds. Similar to ketoprofen, DSZ 1 and DSZ 3 showed antiedematous (antiinflammatory) and antihyperalgesic activity, and similar to lidocaine local anesthetic activity. Furthermore, both compounds (5 and 10 mg/kg) reduced tactile allodynia in acute and chronic phases of neuropathic pain. In the in vitro studies, DSZ 1 and DSZ 3 reduced the COX-2 level in LPS-activated RAW 264.7 cells, which suggests their anti-inflammatory activity. In conclusion, both DSZ 1 and DSZ 3 displayed broad spectrum of activity in several pain models, including neurogenic, tonic, inflammatory, and chemotherapy-induced peripheral neuropathic pain.

Spinal Serum- and Glucocorticoid-Regulated Kinase 1 (SGK1) Signaling Contributes to Morphine-Induced Analgesic Tolerance in Rats.

Accumulating evidence indicates that phosphorylated serum- and glucocorticoid-regulated kinase 1 (SGK1) is associated with spinal nociceptive sensitization by modulating glutamatergic N-methyl-D-aspartate receptors (NMDARs). In this study, we determined whether spinal SGK1 signaling contributes to the development of morphine analgesic tolerance. Chronic morphine administration markedly induced phosphorylation of SGK1 in the spinal dorsal horn neurons. Intrathecal injection of SGK1 inhibitor GSK-650394 reduced the development of morphine tolerance with a significant leftward shift in morphine dose-effect curve. Furthermore, spinal inhibition of SGK1 suppressed morphine-induced phosphorylation of nuclear factor kappa B (NF-κB) p65 and upregulation of NMDAR NR1 and NR2B expression in the spinal dorsal horn. In contrast, intrathecal administration of NMDAR antagonist MK-801 had no effect on the phosphorylation of SGK1 in morphine-treated rats. In addition, morphine-induced upregulation of NR2B, but not NR1, was significantly abolished by intrathecal pretreatment with PDTC, a specific NF-κB activation inhibitor. Finally, spinal delivery of SGK1 small interfering RNA exhibited similar inhibitory effects on morphine-induced tolerance, phosphorylation of NF-κB p65, as well as upregulation of NR1 and NR2B expression. Our findings demonstrate that spinal SGK1 contributes to the development of morphine tolerance by enhancing NF-κB p65/NMDAR signaling. Interfering spinal SGK1 signaling pathway could be a potential strategy for prevention of morphine tolerance in chronic pain management.

Cystitis-induced bladder pain is Toll-like receptor 4 dependent in a transgenic autoimmune cystitis murine model: a MAPP Research Network animal study.

Altered Toll-like receptor (TLR)4 activation has been identified in several chronic pain conditions but has not been well studied in interstitial cystitis/bladder pain syndrome (IC/BPS). Our previously published human studies indicated that patients with IC/BPS present altered systemic TLR4-mediated inflammatory responses, which were significantly correlated with reported pain severity. In the present study, we sought to determine whether altered TLR4 activation plays a role in pelvic/bladder pain seen in patients with IC/BPS using our validated IC/BPS-like transgenic autoimmune cystitis model (URO-OVA). URO-OVA mice developed responses consistent with pelvic and bladder pain after cystitis induction, which was associated with increased splenocyte production of TLR4-mediated proinflammatory cytokines IL-1ß, IL-6, and TNF-α. Increased spinal expression of mRNAs for proinflammatory cytokines IL-6 and TNF-α, glial activation markers CD11b and glial fibrillary acidic protein, and endogenous TLR4 ligand high mobility group box 1 was also observed after cystitis induction. Compared with URO-OVA mice, TLR4-deficient URO-OVA mice developed significantly reduced nociceptive responses, although similar bladder inflammation and voiding dysfunction, after cystitis induction. Intravenous administration of TAK-242 (a TLR4-selective antagonist) significantly attenuated nociceptive responses in cystitis-induced URO-OVA mice, which was associated with reduced splenocyte production of TLR4-mediated IL-1ß, IL-6, and TNF-α as well as reduced spinal expression of mRNAs for IL-6, TNF-α, CD11b, glial fibrillary acidic protein, and high mobility group box 1. Our results indicate that altered TLR4 activation plays a critical role in bladder nociception independent of inflammation and voiding dysfunction in the URO-OVA model, providing a potential mechanistic insight and therapeutic target for IC/BPS pain.

Rapamycin alleviates proinflammatory cytokines and nociceptive behavior induced by chemotherapeutic paclitaxel.

Background/Aims: Paclitaxel is largely used as a chemotherapeutic agent for the treatment of several types of cancers. However, one of the significant limiting complications of paclitaxel is painful peripheral neuropathy during its therapy. The purposes of this study were to examine (1) the effects of blocking mammalian target of rapamycin (mTOR) on mechanical and thermal hypersensitivity evoked by paclitaxel; and (2) the underlying mechanisms responsible for the role of mTOR in regulating paclitaxel-induced neuropathic pain. Methods: Behavioral test was performed to determine mechanical and thermal sensitivity in rats. ELISA was used to examine the levels of proinflammatory cytokines (PICs including IL-1ß, IL-6, and TNF-α) and substance P and calcitonin gene-related peptide (CGRP) in the dorsal root ganglion (DGR); and Western blot analysis was used to examine expression of mTOR signal pathway. Results: Paclitaxel increased mechanical and thermal sensitivity as compared with vehicle control animals (P < 0.05 vs. controls). Paclitaxel also amplified the expression of p-mTOR, mTOR-mediated phosphorylation of p70 ribosomal S6 protein kinase 1 (p-S6K1), 4E-binding protein 1 (p-4E-BP1) in the DRG. Blocking mTOR using rapamycin attenuated peripheral painful neuropathy observed in paclitaxel rats (P < 0.05 vs. without rapamycin). This inhibitory effect was accompanied with decreases of IL-1ß, IL-6, and TNF-α as well as substance P and CGRP. In addition, inhibition of phosphatidylinositide 3-kinase (p-PI3K) attenuated expression of p-mTOR and PICs/substance P/CGRP in paclitaxel rats and this further attenuated mechanical and thermal hypersensitivity. Conclusions: The data revealed specific signaling pathways leading to paclitaxel-induced peripheral neuropathic pain, including the activation of PI3K-mTOR, PIC signal, and substance P and CGRP. Inhibition of these pathways alleviates neuropathic pain. Targeting one or more of these molecular mediators may present new opportunities for treatment and management of neuropathic pain observed during chemotherapeutic application of paclitaxel.