Postoperative Pain Model Research Articles

Changes in median eminence of fatty acid–binding protein 3 in a mouse model of pain

AimsFatty acid–binding protein (FABP) regulates polyunsaturated fatty acid (PUFA) intracellular trafficking and signal transduction. Our previous studies demonstrated that the alteration of PUFA in the hypothalamus is involved in pain process. However, how FABP subtypes change during pain remain unclear. Here, we examined the expression changes and localization in the hypothalamic FABP subtype in postoperative pain model mice.MethodsPaw incision–induced postoperative methods were adopted as a pain model in male ddY mice. Mechanical allodynia was examined using the von Frey test. The analysis of several FABPs mRNA was measured by real‐time PCR, and cellular localization of its protein level was measured by immunofluorescent study.ResultsPostoperative pain mouse elicited mechanical allodynia on Day 2 after paw incision, and mRNA expression of FABP3 increased significantly in the hypothalamus in the postoperative pain mouse model compared to that in control mice. FABP3 protein expressed in the median eminence and the arcuate nucleus, and colocalized with Iba‐1, which is a microglial cell marker. Its protein level significantly increased in the median eminence on Day 2 after incision and returned to the control level on Day 4 after incision.ConclusionsOur findings indicate that FABP3 in the median eminence may change in pain stimuli and may represent a molecular link controlling pain.

Local Effects of Natural Alkylamides from Acmella Oleracea and Synthetic Isobutylalkyl Amide on Neuropathic and Postoperative Pain Models in Mice

Local Effects of Natural Alkylamides from Acmella Oleracea and Synthetic Isobutylalkyl Amide on Neuropathic and Postoperative Pain Models in Mice

Antinociceptive and anxiolytic-like effects of Lavandula angustifolia essential oil on rat models of orofacial pain.

Nociceptive and inflammatory orofacial pain is highly prevalent in the population, which justifies the search for safer analgesics. There is increasing evidence of the analgesic and anxiolytic potential of Lavandula angustifolia essential oil (LAV EO), which may represent, when administered through inhalation, may represent a safer alternative for pain treatment. to evaluate whether LAV EO has antinociceptive effect in the formalin test, and anti-hyperalgesic and anxiolytic-like effects in rats subjected to a model of orofacial postoperative pain. Female Wistar rats were exposed to LAV EO (5%) by inhalation for 30 minutes. After exposure, animals were injected with formalin (2.5%, 50 μL) or saline into the hind paw or upper lip and the number of flinches or facial grooming time, respectively, were evaluated. Likewise, on day 3 after intraoral mucosa incision, the animals were exposed to LAV EO and facial mechanical, and heat hyperalgesia were assessed. The influence of LAV EO inhalation on anxiety-like behavior was assessed in operated rats by testing them on the open field (OF) and elevated plus maze (EPM). LAV EO reduced the phase II of the paw formalin test and both phases of the orofacial formalin test. On day three post-incision, LAV EO reduced heat and mechanical hyperalgesia, from 30 minutes up to three hours, and reduced the anxiety-like behavior in operated rats without causing locomotor deficit. LAV EO inhalation results in antinociceptive and anxiolytic-like effects in orofacial pain models, which encourages further studies on LAV EO indications and effectiveness on orofacial pain conditions.

The PPARγ agonist pioglitazone produces a female-predominant inhibition of hyperalgesia associated with surgical incision, peripheral nerve injury, and painful diabetic neuropathy

Pioglitazone, an agonist at peroxisome proliferator-activated receptor gamma, is FDA-approved for the treatment of insulin resistance in type 2 diabetes. Numerous studies in male rodents suggest that pioglitazone inhibits inflammatory and neuropathic pain, but few included female subjects. To address this gap, we compared the effects of pioglitazone in both sexes in the intraplantar methylglyoxal model (MG) model of chemical pain and painful diabetic neuropathy (PDN), the plantar incision model (PIM) of postoperative pain, the spared nerve injury (SNI) model of traumatic nerve injury, and the ZDF rat and db/db mouse models of PDN. We administered pioglitazone by one-time intrathecal or intraperitoneal injection or by adding it to chow for 6 weeks, followed by measurement of hypersensitivity to non-noxious mechanical, noxious mechanical, heat, and/or cold stimuli. In all mouse models, injection of pioglitazone decreased pain-like behaviors with greater potency and/or efficacy in females as compared to males: heat and mechanical hypersensitivity in the MG model (0.1–10 mg/kg); mechanical hypersensitivity in the PIM model (10 μg); mechanical and cold hypersensitivity in the SNI model (100 mg/kg); and heat hypersensitivity in the db/db model (100 mg/kg). Furthermore, co-administration of low doses of morphine (1 mg/kg) and pioglitazone (10 mg/kg) decreased SNI-induced mechanical and cold hypersensitivity in female but not male mice. In the ZDF rat, pioglitazone (100 mg/kg) decreased heat and mechanical hypersensitivity with no sex difference. In the db/db model, pioglitazone had no effect when given into chow for 6 weeks at 0.3, 3 or 30 mg/kg doses. We conclude that females exhibit greater anti-hyperalgesic responses to pioglitazone in mouse models of chemical-induced nociception, postsurgical pain, neuropathic pain, and PDN. These findings set the stage for clinical trials to determine whether pioglitazone has analgesic properties across a broad spectrum of chronic pain conditions, particularly in women.This article is part of the Special Issue on ‘Advances in mechanisms and therapeutic targets relevant to pain’.

Comparative analysis of two laser wavelengths in the stimulation of acupuncture points for analgesic effects in an animal model.

This study compares the effectiveness of two laser wavelengths for stimulating acupoints in an experimental model of acute postoperative pain. Forty-five Wistar rats were randomly assigned to receive treatment on their left hind paw, contralateral to a surgical procedure. Laser treatments were performed with Green Laser-GL (532 nm, 70 mW and 7J/cm2 of energy), Red Laser-RL (660 nm, 100 mW and 7J/cm2 of energy), or with Laser Off-LO. After each application, the animals were evaluated with a Von Frey analgesiometer to check for painful sensitivity on their right (with surgery) and left (without surgery) hind paws. Neuropeptides and cytokine levels in the incision site tissue of the right paw were measured by ELISA after 1, 6 and 24 hours. It was possible to observe that, in this pain model, both lasers promoted analgesia and that the GL altered the levels of TNF-α and IL-1β.

The analgesic effects of ulinastatin either as a single agent or in combination with sufentanil: A novel therapeutic potential for postoperative pain

Ulinastatin is a broad-spectrum protease inhibitor widely used for the treatment of various inflammation-related diseases owing to its recognized excellent anti-inflammatory and cytoprotective properties. However, whether ulinastatin can relieve postoperative pain remains unclear. In this study, we evaluated the analgesic effects of ulinastatin administered either as a single agent or in combination with sufentanil in a validated preclinical rat model of postoperative pain induced by plantar incision. We found that incisional surgery on the hind paw of these rats induced sustained ipsilateral mechanical pain hypersensitivity that lasted for at least 10 days. A single intraperitoneal (i.p.) injection of ulinastatin prevented the development and reversed the maintenance of incision-induced mechanical pain hypersensitivity in a dose-dependent manner. However, ulinastatin had no effect on the baseline nociceptive threshold. Moreover, repeated i.p. injections of ulinastatin persistently attenuated incision-induced mechanical pain hypersensitivity and promoted recovery from the surgery. The rats did not develop any analgesic tolerance over the course of repeated injections of ulinastatin. A single i.p. injection of ulinastatin was also sufficient to inhibit the initiation and maintenance of incision-induced hyperalgesic priming when the rats were subsequently challenged with an ipsilateral intraplantar prostaglandin E2 injection. Furthermore, the combined administration of ulinastatin and sufentanil significantly enhanced the analgesic effect of sufentanil on postoperative pain, which involved mechanisms other than a direct influence on opioid receptors. These findings demonstrated that ulinastatin had a significant analgesic effect on postoperative pain and might be a novel pharmacotherapeutic agent for managing postoperative pain either alone or as an adjuvant.

Synergism between oral paracetamol and nefopam in a murine model of postoperative pain.

The use of paracetamol or nefopam for postoperative pain control is limited by the need of high doses associated with unwanted effects. Previous works suggest positive interactions between both compounds that may be exploited to obtain potentiation of antinociception. Mechanical and heat antinociception induced by oral doses of paracetamol, nefopam or their combination was studied by isobolographic analysis in a murine model of postsurgical pain. The effective doses that produced 50% antinociception (ED50 ) were calculated from the log dose-response curves for each compound. Subsequently, the effects of ED8.7 s, ED12.5 s, ED17.5 s and ED35 s of nefopam and paracetamol combined were assessed. Oral paracetamol induced dose-dependent relief of postoperative sensitivity and showed higher efficacy reducing mechanical hypersensitivity (ED50 177.3±15.4mg/kg) than heat hyperalgesia (ED50 278.6±43mg/kg). Oral nefopam induced dose-dependent antinociception with similar efficacy for mechanical and heat hypersensitivity (ED50 s 5.42±0.81 vs. 5.83±0.72). Combinations of increasing isoeffective doses revealed that combined ED17.5 s (85.76mg/kg paracetamol and 1.9mg/kg nefopam) and ED35 s (132.67mg/kg and 3.73mg/kg) showed synergistic effects leading to 75% and 90% mechanical antinociception, respectively. These mixtures were defined by interaction indexes of 0.43 and 0.41 and ratios 45:1 and 35:1 paracetamol:nefopam, respectively. The same combinations showed additive effects for the inhibition of incisional thermal hyperalgesia. This work describes a synergistic antinociceptive interaction between low doses of nefopam and paracetamol for the treatment of postoperative hypersensitivity to peripheral stimuli. The promising results obtained on reflexive nociceptive responses of young male mice subjected to plantar surgery highlight the interest of further research evaluating the effects of this mixture on the affective-motivational component of pain and in females and additional age groups. Confirmation of pain-relieving efficacy and safety of this oral combination clinically available in European and Asian countries could provide a useful tool for postsurgical pain management. Early postoperative pain is currently undertreated and has been recognized as a relevant source of chronic postsurgical pain. Oral efficient treatments could facilitate fast-track surgeries and patient recovery at home. Here, we identify in a mouse model of postoperative pain a potent synergistic oral combination consisting of low paracetamol and nefopam doses that provides relief of postsurgical hypersensitivity to mechanical and thermal stimuli. Oral multimodal paracetamol-nefopam mixtures represent a potential clinically available pharmacological strategy for the relief of incisional sensitivity and the promotion of patient recovery.

Effect of pregabalin on nociceptive thresholds and immune responses in a mouse model of incisional pain.

BackgroundIt is known that some analgesics as well as pain can affect the immune system. The aim of this study was to investigate the analgesic effect and immunomodulation of pregabalin (PGB) in a mouse incisional pain model.MethodsA postoperative pain model was induced by hind paw plantar incision in male BALB/c mice. Mice were randomly divided into four groups (n = 8) a saline-treated incision (incision), PGB-treated incision (PGB-incision), sham controls without incision or drug treatment (control), and a PGB-treated control (PGB-control). In the PGB treated groups, PGB was administered intraperitoneally (IP) 30 minutes before and 1 hour after the plantar incision. Changes of the mechanical nociceptive thresholds following incision were investigated. Mice were euthanized for spleen harvesting 12 hours after the plantar incision, and natural killer (NK) cytotoxicity to YAC 1 cells and lymphocyte proliferation responses to phytohemagglutinin were compared among these four groups.ResultsMechanical nociceptive thresholds were decreased after plantar incision and IP PGB administration recovered these decreased mechanical nociceptive thresholds (P < 0.001). NK activity was increased by foot incision, but NK activity in the PGB-incision group was significantly lower than that in the Incision group (P < 0.001). Incisional pain increased splenic lymphocyte proliferation, but PGB did not alter this response.ConclusionsIncisional pain alters cell immunity of the spleen in BALB/c mice. PGB showed antinocieptive effect on mouse incisional pain and attenuates the activation of NK cells in this painful condition. These results suggest that PGB treatment prevents increases in pain induced NK cell activity.

Promising potential of articaine-loaded poly(epsilon-caprolactone) nanocapules for intraoral topical anesthesia.

To determine whether the permeation capacity and analgesic efficacy of articaine (ATC) could be increased and cytotoxicity decreased by encapsulation in poly(ɛ-caprolactone) nanocapsules (ATCnano), aiming at local or topical anesthesia in dentistry. Cellular viability was evaluated (using the MTT test and fluorescence microscopy) after 1 h and 24 h exposure of HaCaT cells to ATC, ATCnano, ATC with epinephrine (ATCepi), and ATC in nanocapsules with epinephrine (ATCnanoepi). The profiles of permeation of 2% ATC and 2% ATCnano across swine esophageal epithelium were determined using Franz-type vertical diffusion cells. Analgesic efficacy was evaluated with a von Frey anesthesiometer in a postoperative pain model in rats, comparing the 2% ATC, 2% ATCnano, 2% ATCepi, and 2% ATCnanoepi formulations to 4% ATCepi (a commercially available formulation). We show that use of the nanocapsules decreased the toxicity of articaine (P<0.0001) and increased its flux (P = 0.0007). The 2% ATCepi and 4% ATCepi formulations provided higher analgesia success and duration (P<0.05), compared to 2% ATC, 2% ATCnano, and 2% ATCnanoepi. Articaine-loaded poly(ɛ-caprolactone) nanocapsules constitute a promising formulation for intraoral topical anesthesia (prior to local anesthetic injection), although it is not effective when injected in inflamed tissues for pain control, such as irreversible pulpitis.

Antinociceptive Effects of Sinomenine Combined With Ligustrazine or Paracetamol in Animal Models of Incisional and Inflammatory Pain.

The management of postoperative and inflammatory pain has been a pressing challenge in clinical settings. Sinomenine (SN) is a morphinan derived alkaloid with remarkable analgesic properties in various kinds of pain models. The aim of the current study is to investigate if SN can enhance the effect of ligustrazine hydrochloride (LGZ) or paracetamol (PCM) in animal models of postoperative and inflammatory pain. And to determine if the combined therapeutic efficacies can be explained by pharmacokinetics changes. Pharmacological studies were performed using a rat model of incisional pain, and a mouse model of carrageenan induced inflammatory pain. Pharmacokinetic studies were performed using a microdialysis sampling and HPLC-MS/MS assay method to quantify SN, LGZ, and PCM levels in blood and extracellular fluid in brain. We found that SN plus LGZ or SN plus PCM produced marked synergistic analgesic effects. However, such synergy was subjected to pain modalities, and differed among pain models. Pharmacological discoveries could be partially linked to pharmacokinetic alterations in SN combinations. Though further evaluation is needed, our findings advocate the potential benefits of SN plus LGZ for postoperative pain management, and SN plus PCM for controlling inflammatory pain.

Phospholipase Cβ3 Expressed in Mouse DRGs is Involved in Inflammatory and Postoperative Pain.

BackgroundPrevious studies suggested that phospholipase Cβ3 (PLCβ3), which is a common downstream component in the signaling cascade, plays an important role in peripheral mechanisms of perception including nociception. However, detailed profiles of PLCβ3-expressing dorsal root ganglion (DRG) neurons and involvement of PLCβ3 in inflammatory and postoperative pain have not been fully investigated.PurposeWe evaluated neurochemical char0acteristics of PLCβ3-expressing DRG neurons in mice and then we examined the effects of selective knockdown of PLCβ3 expression in DRGs on inflammatory and postoperative pain.MethodsMale C57BL/6-strain mice were used. For the inflammatory model, each mouse received subcutaneous injection of complete Freund’s adjuvant (CFA) in the left hindpaw. For the postoperative pain model, a plantar incision was made in the left hindpaw. PLCβ3 antisense oligodeoxynucleotide or PLCβ3 mismatch oligodeoxynucleotide was intrathecally administered once a day for three consecutive days in each model. The time courses of thermal hyperalgesia and mechanical hyperalgesia were investigated. Changes in PLCβ3 protein levels in DRGs were evaluated by Western blotting.ResultsImmunohistochemical analysis showed that high proportion of the PLCβ3-positive profiles were biotinylated isolectin B4-positive or transient receptor potential vanilloid subfamily 1-positive. PLCβ3 protein level in DRGs during CFA-induced inflammation was comparable to that at baseline. Intrathecal administration of PLCβ3 antisense oligodeoxynucleotide, which significantly suppressed PLCβ3 expression in DRGs, did not affect pain thresholds in normal conditions but inhibited CFA-induced thermal and mechanical hyperalgesia both at the early and late phases compared to that in mismatch oligodeoxynucleotide-treated mice. Intrathecal administration of PLCβ3 antisense oligodeoxynucleotide also inhibited surgical incision-induced thermal and mechanical hyperalgesia.ConclusionOur results uncover a unique role of PLCβ3 in the development and maintenance of inflammatory pain induced by CFA application and in those of surgical incision-induced pain, although PLCβ3 does not play a major role in thermal nociception or mechanical nociception in normal conditions.

TRPV1-Targeted Drugs in Development for Human Pain Conditions.

The transient receptor potential vanilloid-1 (TRPV1) is a non-specific cation channel known for its sensitivity to pungent vanilloid compound (i.e. capsaicin) and noxious stimuli, including heat, low pH or inflammatory mediators. TRPV1 is found in the somatosensory system, particularly primary afferent neurons that respond to damaging or potentially damaging stimuli (nociceptors). Stimulation of TRPV1 evokes a burning sensation, reflecting a central role of the channel in pain. Pharmacological and genetic studies have validated TRPV1 as a therapeutic target in several preclinical models of chronic pain, including cancer, neuropathic, postoperative and musculoskeletal pain. While antagonists of TRPV1 were found to be a valuable addition to the pain therapeutic toolbox, their clinical use has been limited by detrimental side effects, such as hyperthermia. In contrast, capsaicin induces a prolonged defunctionalisation of nociceptors and thus opened the door to the development of a new class of therapeutics with long-lasting pain-relieving effects. Here we review the list of TRPV1 agonists undergoing clinical trials for chronic pain management, and discuss new indications, formulations or combination therapies being explored for capsaicin. While the analgesic pharmacopeia for chronic pain patients is ancient and poorly effective, modern TRPV1-targeted drugs could rapidly become available as the next generation of analgesics for a broad spectrum of pain conditions.

Phα1β, a dual blocker of TRPA1 and Cav2.2, as an adjuvant drug in opioid therapy for postoperative pain

Opioids are the “gold standard” treatment for postoperative pain, but these drugs also have limiting adverse effects. Thus, adjuvant drugs might be useful in opioid therapy for postoperative pain. The aim of the present study was to evaluate the effect of Phα1β, a dual blocker of Cav2 and TRPA1 channels, on antinociceptive and adverse actions of morphine in a model of postoperative pain. Phα1β (100–300 pmol/site) or morphine (3–10 mg/kg), alone, largely reduced postoperative nociception. However, Phα1β (100 pmol/site) or morphine (10 mg/kg) also produced motor impairment. Lower doses of Phα1β (30 pmol/site) or morphine (1 mg/kg), that did not have an effect alone, showed antinociceptive effect when concomitantly administrated. Moreover, co-administration of Phα1β (30 pmol/site) with morphine (1 or 10 mg/kg) was unable to cause motor impairment. Preoperative repeated treatment with morphine increased the expression of Cav2 and TRPA1 channels in spinal cord, and caused tolerance and withdrawal syndrome, which were reversed with a single injection of Phα1β (30 pmol/site). When injected postoperatively, escalating doses of morphine worsened postoperative hyperalgesia, induced tolerance, and withdrawal syndrome. Similarly, Phα1β (30 pmol/site) reversed these adverse effects. Single or repeated morphine caused constipation, which was not altered by Phα1β. Thus, a low dose of Phα1β potentiated the analgesia, and reversed some adverse effects of morphine on operated mice, indicating the potential use of this agent as an adjuvant drug in opioid therapy for postoperative pain.

Effect of intraperitoneally administered propentofylline in a rat model of postoperative pain.

BackgroundIn this study, we sought to evaluate whether systemic propentofylline (PPF) has antiallodynic effects in a rat model of postoperative pain, and to assess the mechanism involved.MethodsAfter plantar incision, rats were intraperitoneally injected with various doses of PPF to evaluate its antiallodynic effect. To investigate the involved mechanism, rats were intraperitoneally injected with yohimbine, dexmedetomidine, prazosin, naloxone, atropine or mecamylamine, following the incision of the rat hind paws, and then PPF was administered intraperitoneally. The mechanical withdrawal threshold (MWT) was evaluated using von Frey filaments at various time points and serum levels of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 were measured to determine the inflammatory response level.ResultsMWT was significantly increased after intraperitoneal injection of 30 mg/kg of PPF when compared with the control group. Injection of PPF and yohimbine, atropine or mecamylamine showed significant decreases in the MWT, while injection of PPF and dexmedetomidine showed a significant increase. Systemic administration of PPF inhibited the post-incisional increase in serum level of TNF-α and IL-1β.ConclusionsSystemic administration of PPF following surgery presented antiallodynic effects in a rat model of postoperative pain. The antiallodynic effects against mechanical allodynia could be mediated by α-adrenergic and cholinergic receptors.

Antinociception induced by artemisinin nanocapsule in a model of postoperative pain via spinal TLR4 inhibition.

Artemisinin (ART) was initially described for the control of inflammation and pain. However, the mechanisms involved with its antinociceptive effect are still poorly understood. Thus, this present study aimed to investigate the effect of ART in both free and nanocapsulated form on postoperative pain, as well as the participation of the spinal Toll-like receptor 4 (TLR4) in this process. Postoperative pain was induced using the skin/muscle incision retraction (SMIR) model in male Swiss mice. After 3 and 28days of SMIR, the animals received an intrathecal injection of free or nanocapsulated ART, and the nociceptive threshold was evaluated by von Frey filament test. To evaluate the involvement of the microglia, astrocytes, and TLR4, minocycline (a microglia inhibitor), fluorocitrate (an astrocyte inhibitor), and Lipopolysaccharide Rhodobacter sphaeroides (LPS-RS), a TLR4 antagonist, were intrathecally injected on the third day of SMIR. The levels of spinal TLR4 protein and proinflammatory cytokines tumor necrosis factor-alpha (TNF-α), and interleukin-1-beta (IL-1β) were quantified by western blot and enzyme-linked immunosorbent assay, respectively. The results showed that free ART reduced postoperative pain (P < 0.001, F5,30 = 7.49, 16.66% for 1000ng dose; and P < 0.01, F5,30 = 7.49, 14.58% for 500ng dose) on the 3rd day of SMIR; while the ART nanocapsule had this effect on both the third (P < 0.001; F5,30 = 4.94; 43.75, 39.58 and 72.91% for the 250, 500 and 1000ng doses, respectively) and 28th (P < 0.05; F5,30 = 7.71; 29.16 and 33.33% for the 500 and 1000ng doses, respectively) day. The ART nanocapsule had a more potent and longer antinociceptive effect than free ART or morphine. Postoperative pain was also reduced by minocycline and LPS-RS. The ART nanocapsule also reduced the increased levels of TLR4, TNF-α, and IL-1β induced by SMIR. These data suggest that the ART nanocapsule has a potent analgesic effect on postoperative pain at the spinal level, and this response involves the inhibition of TLR4 and the proinflammatory cytokines TNF-α and IL-1β.

Intrathecal Oxytocin Improves Spontaneous Behavior and Reduces Mechanical Hypersensitivity in a Rat Model of Postoperative Pain.

The first few days post-surgery, patients experience intense pain, hypersensitivity and consequently tend to have minor locomotor activity to avoid pain. Certainly, injury to peripheral tissues produces pain and increases sensitivity to painful (hyperalgesia) and non-painful (allodynia) stimuli. In this regard, preemptive pharmacological treatments to avoid or diminish pain after surgery are relevant. Recent data suggest that the neuropeptide oxytocin when given at spinal cord level could be a molecule with potential preemptive analgesic effects, but this hypothesis has not been properly tested. Using a validated postoperative pain model (i.e. plantar incision), we evaluated in male Wistar rats the potential preemptive antinociceptive effects of intrathecal oxytocin administration measuring tactile hypersensitivity (across 8 days) and spontaneous motor activity (across 3 days). Hypersensitivity was evaluated using von Frey filaments, whereas spontaneous activity (total distance, vertical activity episodes, and time spent in the center of the box) was assessed in real time using a semiautomated open-field system. Under these conditions, we found that animals pretreated with spinal oxytocin before plantar incision showed a diminution of hypersensitivity and an improvement of spontaneous behavior (particularly total distance and vertical activity episodes). This report provides a basis for addressing the therapeutic relevance of oxytocin as a potential preemptive analgesic molecule.

Toll-like receptor 4 (TLR4) signaling in the trigeminal ganglion mediates facial mechanical and thermal hyperalgesia in rats

There is increasing evidence that the toll-like receptor 4 (TLR4) signaling pathway contribute to development of hyperalgesia in the trigeminal system. The aim of the present study was to investigate the role of TLR4 in the trigeminal ganglion (TG) in facial hyperalgesia induced by injection of Lipopolysaccharide (LPS) or intraoral mucosal incision, which is an orofacial postoperative pain model, in male Wistar rats. The TLR4 antagonist (LPS-RS, 20 µg/10 µL) was administrated 30 min before LPS injection into the TG (10 µg/10 µL) or oral mucosa (10 µg/50 µL). In the postoperative pain model, rats were treated with LPS-RS (20 µg/10 µL) into the TG for three consecutive days after the incision. Facial heat and mechanical hyperalgesia were assessed hourly after LPS injection or intraoral incision. In addition, expression of NFκB was assessed in the TG on day 3 after intraoral incision. Our results showed that blockade of TLR4 in the TG attenuated facial heat and mechanical hyperalgesia induced by LPS or by mucosal incision, and that both conditions are associated to increase of phosphorylated NFκB in the TG. In conclusion, the present study suggests that activation of TLR4-NFκB signaling pathway in the TG contributes to the development of facial heat and mechanical hyperalgesia and may contribute to pain in inflammatory oral conditions.

Targeting spinal neuropeptide Y1 receptor-expressing interneurons to alleviate chronic pain and itch.

An accelerating basic science literature is providing key insights into the mechanisms by which spinal neuropeptide Y (NPY) inhibits chronic pain. A key target of pain inhibition is the Gi-coupled neuropeptide Y1 receptor (Y1). Y1 is located in key sites of pain transmission, including the peptidergic subpopulation of primary afferent neurons and a dense subpopulation of small, excitatory, glutamatergic/somatostatinergic interneurons (Y1-INs) that are densely expressed in the dorsal horn, particularly in superficial lamina I-II. Selective ablation of spinal Y1-INs with an NPY-conjugated saporin neurotoxin attenuates the development of peripheral nerve injury-induced mechanical and cold hypersensitivity. Conversely, conditional knockdown of NPY expression or intrathecal administration of Y1 antagonists reinstates hypersensitivity in models of chronic latent pain sensitization. These and other results indicate that spinal NPY release and the consequent inhibition of pain facilitatory Y1-INs represent an important mechanism of endogenous analgesia. This mechanism can be mimicked with exogenous pharmacological approaches (e.g. intrathecal administration of Y1 agonists) to inhibit mechanical and thermal hypersensitivity and spinal neuron activity in rodent models of neuropathic, inflammatory, and postoperative pain. Pharmacological activation of Y1 also inhibits mechanical- and histamine-induced itch. These immunohistochemical, pharmacological, and cell type-directed lesioning data, in combination with recent transcriptomic findings, point to Y1-INs as a promising therapeutic target for the development of spinally directed NPY-Y1 agonists to treat both chronic pain and itch.

Pre-clinical evaluation of new dibucaine formulations for preventive analgesia

We have previously developed ammonium sulphate gradient loaded liposomes to encapsulate dibucaine. Thus, the purpose of this study was to evaluate the pre-clinical safety and effectiveness of this novel ionic liposomal formulation of dibucaine (DBC), as described in previous work. Effectiveness was evaluated in vivo on Wistar rats (n = 8) that received plain DBC or liposomal DBC (DBCLUV). Control empty liposomes (without DBC) or saline were also used as control. Sciatic nerve block was performed using the formulations or controls (0.4 mL). A hindpaw incision-based postoperative pain model was used to evaluate mechanical hypersensitivity with von Frey filaments. To verify antiinflamatory activity protein levels of TNF-α, IL-1β, substance P and CGRP were measured by ELISA in the hindpaw tissue after 1 and 6 hours of the incision. To corroborate drug safety, sciatic nerve Schwann cell cultures were treated with the aforementioned formulations and assessed for cell viability (MTT assay) and death (flow cytometry assay). Histopathology of the tissues surrounding the sciatic nerve region was also assessed 2 and 7 days after treatment. All animals presented post incisional hypersensitivity and DBCLUV showed longer analgesic effect (p < 0.001). DBCLUV reduced TNF-α and CGRP levels (p < 0.05). Histopathological evaluation showed greater inflammatory reaction after the administration of control liposomes when compared to DBC (p < 0.05). There was no difference in Schwann cell viability and death between plain and encapsulated DBC. DBCLUV was safe and enhanced anaesthesia duration due to slow release of dibucaine from ammonium sulphate gradient loaded liposomes.

Spinal administration of the multi-functional opioid/neuropeptide FF agonist BN-9 produced potent antinociception without development of tolerance and opioid-induced hyperalgesia

Chronic opioids treatment is impeded by the development of analgesic tolerance and opioid-induced hyperalgesia. Recent studies have shown that multi-functional opioid compounds produce analgesic activities with limited side effects. We developed a novel multi-functional peptide targeting opioid and neuropeptide FF receptors named BN-9, which produced potent and non-tolerance forming antinociceptive effect after supraspinal and systemic administrations. In the present study, the analgesic properties and potential side effects of intrathecal BN-9 were investigated in a range of preclinical rodent models. In complete Freund's adjuvant-induced inflammatory pain model, intrathecal BN-9 dose-dependently produced analgesic effect via opioid receptors, and the spinal antinociceptive effect was augmented by the neuropeptide FF receptor antagonist RF9. In contrast, in plantar incision-induced postoperative pain model, BN-9 exhibited potent anti-allodynic effect via opioid receptors and, at least partially, neuropeptide FF receptors. In mouse models of acetic acid-induced visceral pain and formalin pain, BN-9-induced spinal antinociception was mainly mediated by opioid receptors, independent of neuropeptide FF receptors. Furthermore, at the spinal level, chronic treatments with BN-9 did not lead to analgesic tolerance and cross-tolerance to morphine. Moreover, opioid-induced hyperalgesia was observed after repeated administration of morphine, but not BN-9. Taken together, our present study suggests that intrathecal BN-9 produces potent and non-tolerance forming antinociception, and does not cause opioid-induced hyperalgesia. Thus, BN-9 might serve as a promising lead compound in the development of multi-functional opioid analgesics with minimized side effects.