Monosodium urate crystal interleukin-1ß release is dependent on Toll-like receptor 4 and transient receptor potential V1 activation.

OBJECTIVE: The present study aimed to elucidate the mechanisms involved in MSU-induced IL-1ß release in a rodent animal model of acute gout arthritis. METHODS: Painful (mechanical and thermal hypersensitivity, ongoing pain and arthritis score) and inflammatory (oedema, plasma extravasation, cell infiltration and IL-1ß release) parameters were assessed several hours after intra-articular injection of MSU (100 µg/articulation) in wild-type or knockout mice for Toll-like receptor 4 (TLR4), inducible nitric oxide synthase (iNOS), transient receptor potential (TRP) V1 and the IL-1 receptor (IL-1R). Also, wild-type animals were treated with clodronate, lipopolysaccharide from Rhodobacter sphaeroides (LPS-RS) (TLR4 antagonist), spleen tyrosine kinase (SYK) inhibitor (iSYK), aminoguanidine (AMG, an iNOS inhibitor) or SB366791 (TRPV1 antagonist). Nitrite/nitrate and IL-1ß levels were measured on the synovial fluid of wild-type mice, 2 h after intra-articular MSU injections, or medium from macrophages stimulated for MSU (1000 µg) for 2 h. RESULTS: Intra-articular MSU injection caused robust nociception and severe inflammation from 2 up to 6 h after injection, which were prevented by the pre-treatment with clodronate, LPS-RS, iSYK, AMG and SB366791, or the genetic ablation of TLR4, iNOS, TRPV1 or IL-1R. MSU also increased nitrite/nitrate and IL-1ß levels in the synovial fluid, which was prevented by clodronate, LPS-RS, iSYK and AMG, but not by SB366791. Similarly, MSU-stimulated peritoneal macrophages released nitric oxide, which was prevented by LPS-RS, iSYK and AMG, but not by SB366791, and released IL-1ß, which was prevented by LPS-RS, iSYK, AMG and SB366791. CONCLUSION: Our data indicate that MSU may activate TLR4, SYK, iNOS and TRPV1 to induce the release of IL-1ß by macrophages, triggering nociception and inflammation during acute gout attack.

Neuroimmune-Glia Interactions in the Sensory Ganglia Account for the Development of Acute Herpetic Neuralgia.

Herpetic neuralgia is the most important symptom of herpes zoster disease, which is caused by Varicella zoster Nevertheless, the pathophysiological mechanisms involved in herpetic neuralgia are not totally elucidated. Here, we examined the neuroimmune interactions at the sensory ganglia that account for the genesis of herpetic neuralgia using a murine model of Herpes Simplex Virus Type-1 (HSV-1) infection. The cutaneous HSV-1 infection of mice results in the development of a zosteriform-like skin lesion followed by a time-dependent increase in pain-like responses (mechanical allodynia). Leukocytes composed mainly of macrophages and neutrophils infiltrate infected DRGs and account for the development of herpetic neuralgia. Infiltrating leukocytes are responsible for driving the production of TNF, which in turn mediates the development of herpetic neuralgia through downregulation of the inwardly rectifying K+ channel Kir4.1 in satellite glial cells. These results revealed that neuroimmune-glia interactions at the sensory ganglia play a critical role in the genesis of herpetic neuralgia. In conclusion, the present study elucidates novel mechanisms involved in the genesis of acute herpetic pain and open new avenues for its control.SIGNIFICANCE STATEMENT Acute herpetic neuralgia is the most important symptom of herpes zoster disease and it is very difficult to treat. Using a model of peripheral infection of mice with HSV-1, we have characterized for the first time the neuroimmune-glia interactions in the sensory ganglia that account for the development of acute herpetic neuralgia. Among these mechanisms, leukocytes composed mainly of macrophages and neutrophils infiltrate infected sensory ganglia and are responsible for driving the production of TNF. TNF, via TNFR1, mediates herpetic neuralgia development through downregulation of the inwardly rectifying K+ channel Kir4.1 in satellite glial cells. This study elucidates novel mechanisms involved in the genesis of acute herpetic neuralgia and open new avenues for its control.

The role of kinin B1 receptor and the effect of angiotensin I-converting enzyme inhibition on acute gout attacks in rodents.

OBJECTIVE: Verify the role of the kinin B1 receptors (B1R) and the effect of ACE inhibitors (ACEi) on acute gout induced by monosodium urate (MSU) crystals in rodents. METHODS: Painful (overt pain and allodynia) and inflammatory parameters (joint oedema, leukocyte trafficking, interleukin-1ß levels) of acute gout attacks were assessed several hours after an intra-articular injection of MSU (1.25 or 0.5 mg/articulation) into the ankle of rats or mice, respectively. The role of B1R was investigated using pharmacological antagonism or gene deletion. Additionally, B1R immunoreactivity in ankle tissue and sensory neurons, kininase I activity and des-Arg(9)-bradykinin synovial levels were also measured. Similar tools were used to investigate the effects of ACEi on a low dose of MSU (0.0125 mg/articulation)-induced inflammation. RESULTS: Kinin B1R antagonism or gene deletion largely reduced all painful and inflammatory signs of gout. Furthermore, MSU increased B1R expression in articular tissues, the content of the B1 agonist des-Arg(9)-bradykinin and the activity of the B1 agonist-forming enzyme kininase I. A low dose of MSU crystals, which did not induce inflammation in control animals, caused signs of acute gout attacks in ACEi-treated animals that were B1R-dependent. CONCLUSIONS: Kinin B1R contributes to acute gouty attacks, including the ones facilitated by ACEi. Therefore, B1R is a potential therapeutic target for the treatment and prophylaxis of gout, especially in patients taking ACEi.

Effect of Uncaria tomentosa extract on purinergic enzyme activities in lymphocytes of rats submitted to experimental adjuvant arthritis model.

BACKGROUND: Considering that adjuvant arthritis is an experimental model of arthritis widely used for preclinical testing of numerous anti-arthritic agents, which were taken by a large number of patients worldwide, it is of great interest to investigate the therapeutic action of compounds with anti-inflammatory properties, such as Uncaria tomentosa extract. Moreover, there are no studies demonstrating the effect of U. tomentosa on the metabolism of adenine nucleotides published so far. Thus, the purpose of the present study is to investigate the effects of U. tomentosa extract on E-NTPDase and E-ADA activities in lymphocytes of Complete Freund's Adjuvant (CFA) arthritis induced rats. METHODS: To evaluate the effect of U. tomentosa extract on the activity of E-NTPDase and ADA in lymphocytes, the rats were submitted to an experimental adjuvant arthritis model. Peripheral lymphocytes were isolated and E-NTPDase and E-ADA activities were determined. Data were analyzed by a one- or two-way ANOVA. Post hoc analyses were carried out by the Student-Newman-Keuls (SNK) Multiple Comparison Test. RESULTS: E-NTPDase activity was increased in arthritic untreated. Arthritic rats which received U. tomentosa extract, presented similar results to the control group. However, results obtained for adenosine hydrolysis by E-ADA were not altered in arthritic rats. U. tomentosa extract did not alter E-NTPDase and E-ADA activity in healthy animals. CONCLUSIONS: The present investigation supports the hypothesis that the increased E-NTPDase activity verified in arthritic rats might be an attempt to maintain basal levels of ATP and ADP in the extracellular medium, since the arthritis induction causes tissue damage and, consequently, large amounts of ATP are released into this milieu. Also, it highlights the possibility to use U. tomentosa extract as an adjuvant to treat arthritis.

Transient receptor potential ankyrin 1 receptor stimulation by hydrogen peroxide is critical to trigger pain during monosodium urate-induced inflammation in rodents.

OBJECTIVE: Gout is a common cause of inflammatory arthritis and is provoked by the accumulation of monosodium urate (MSU) crystals. However, the underlying mechanisms of the pain associated with acute attacks of gout are poorly understood. The aim of this study was to evaluate the role of transient receptor potential ankyrin 1 (TRPA-1) and TRPA-1 stimulants, such as H2 O2 , in a rodent model of MSU-induced inflammation. METHODS: MSU or H2 O2 was injected into the hind paws of rodents or applied in cultured sensory neurons, and the intracellular calcium response was measured in vitro. Inflammatory or nociceptive responses in vivo were evaluated using pharmacologic, genetic, or biochemical tools and methods. RESULTS: TRPA-1 antagonism, TRPA-1 gene deletion, or pretreatment of peptidergic TRP-expressing primary sensory neurons with capsaicin markedly decreased MSU-induced nociception and edema. In addition to these neurogenic effects, MSU increased H2 O2 levels in the injected tissue, an effect that was abolished by the H2 O2 -detoxifying enzyme catalase. H2 O2 , but not MSU, directly stimulated sensory neurons through the activation of TRPA-1. The nociceptive responses evoked by MSU or H2 O2 injection were attenuated by the reducing agent dithiothreitol. In addition, MSU injection increased the expression of TRPA-1 and TRP vanilloid channel 1 (TRPV-1) and also enhanced cellular infiltration and interleukin-1ß levels, and these effects were blocked by TRPA-1 antagonism. CONCLUSION: Our results suggest that MSU injection increases tissue H2 O2 , thereby stimulating TRPA-1 on sensory nerve endings to produce inflammation and nociception. TRPV-1, by a previously unknown mechanism, also contributes to these responses.

Antinociceptive effect of novel trihalomethyl-substituted pyrazoline methyl esters in formalin and hot-plate tests in mice.

The aim of the present study was to evaluate the antinociceptive potential of four novel pyrazoline methyl ester compounds on chemical and thermal models of pain in mice. The following 5-trihalomethylated-4,5-dihydro-1H-pyrazole methyl ester compounds were tested: 3-methyl-5-trifluoromethyl-(MPF3), 4-methyl-5-trifluoromethyl-(MPF4), 3-methyl-5-trichloromethyl-(MPCl3) and 4-methyl-5-trichloromethyl-(MPCl4). MPF3, MPF4, MPCl3 and MPCl4 (0.03-1.0 mmol/kg) given intraperitoneally decreased neurogenic and inflammatory phases of nociception in the formalin test. Moreover, MPF3, MPF4, MPCl3, MPCl4 (0.1-1.0 mmol/kg) and dipyrone (1.5 mmol/kg) also produced a dose-dependent antinociceptive effect in the hot-plate test. However, MPF3, MPF4, MPCl3 and MPCl4 did not impair motor coordination in the rotarod test or spontaneous locomotion in the open field test. The antinociceptive effect of MPF4 (1.0 mmol/kg, i.p.) was reversed by the opioid receptor antagonist naloxone (2 mg/kg, i.p.), but not by the alpha(2)-adrenergic receptor antagonist yohimbine (0.15 mg/kg, i.p.) or by p-chlorophenylalanine ethyl ester (PCPA, 300 mg/kg, i.p.) treatment. In contrast to morphine (5 mg/kg, i.p.), MPF4 given daily for up to 8 days did not generate a tolerance to its antinociceptive effect. However, similar to morphine (11 mg/kg, i.p.), MPF4 reduced gastrointestinal transit in mice. Taken together these results demonstrate that these novel pyrazoline methyl esters tested may be promising prototypes of additional mild analgesics.

Antinociceptive action of 4-methyl-5-trifluoromethyl-5-hydroxy-4, 5-dihydro-1H-pyrazole methyl ester in models of inflammatory pain in mice.

AIMS: The aim of the present study was to evaluate the antinociceptive effect of the novel pyrazoline methyl ester: 4-methyl-5-trifluoromethyl-5-hydroxy-4,5-dihydro-1H-pyrazole methyl ester (MPF4). MAIN METHODS: The effect of MPF4 was assessed in two models of pain: arthritic pain caused by Complete Freund's Adjuvant (CFA) and postoperative pain caused by surgical incision in mice. KEY FINDINGS: MPF4 given intraperitoneally (1.0 mmol/kg, i.p.) produced marked antinociception in inflammatory allodynia caused by CFA. The antinociceptive effect produced by MPF4 was reversed with the pre-treatment of animals with naloxone or naltrindole. Oral administration of MPF4 (1.0 mmol/kg, p.o), dipyrone (1.0 mmol/kg, p.o.) and morphine (0.026 mmol/kg, p.o.) also produced an anti-allodynic effect. However, none of the compounds evaluated reversed the paw edema produced by CFA. Moreover, MPF4, dipyrone and morphine also produced an anti-allodynic effect in the surgical incisional pain model. The maximal inhibitions obtained with preemptive drug treatment were 66+/-7%, 73+/-9% and 88+/-8% for MPF4 (1.0 mmol/kg, p.o.), dipyrone (1.0 mmol/kg, p.o.) and morphine (0.026 mmol/kg, p.o.), respectively. The maximal inhibitions obtained with curative drug treatment were 53+/-9%, 83+/-7% and 84+/-7%, for MPF4, dipyrone and morphine, respectively. Unlike indomethacin, MPF4 did not induce gastric lesions at the dose that caused the highest antinociception (1.0 mmol/kg, p.o). The anti-allodynic action of MPF4, dipyrone and morphine was not associated with impairment of motor activity. SIGNIFICANCE: The results of the present study suggest that MPF4 represents a potential target for the development of new drugs to treat persistent inflammatory pain.

Antinociceptive activity of Mirabilis jalapa in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: The infusion or decoction of Mirabilis jalapa leaves is used in traditional medicine in Brazil to treat inflammatory and painful diseases. AIM OF THE STUDY: The present study examined the antinociceptive effect of Mirabilis jalapa extracts from leaves and stems in models of pain in mice. MATERIALS, METHODS AND RESULTS: The crude hydroethanolic extract from leaves (CrdL) was more potent than the crude extract from stems (CrdS) to inhibit abdominal constrictions induced by acetic acid, with ID(50) values of 5.5 (2.3-13.1) and 18.0 (11.3-28.5) mg/kg, respectively. Among the fractions tested, the Eta fraction from leaves (Eta) was more effective (maximal inhibition of 83+/-8%) and potent (ID(50) of 1.1 (0.6-2.1) mg/kg) to induce antinociception. Eta and CrdL also possessed an antinociceptive effect in the tail-flick test. Pre-treatment with naloxone did not modify the antinociceptive effect of Eta, but co-administration with atropine completely prevented it. This suggests that the antinociceptive effect might depend on the cholinergic system. Instead, Eta was not able to alter the acetylcholinesterase activity in blood or spinal cord. Concerning side effects, Eta did not alter locomotor activity, body temperature, gastrointestinal transit and did not produce gastric lesions. CONCLUSION: Our results demonstrate that Mirabilis jalapa presents antinociceptive activity in mice, which supports its folkloric use as an analgesic.

Synthetic and Biological Studies of Sesquiterpene Polygodial: Activity of 9-Epipolygodial against Drug-Resistant Cancer Cells.

Polygodial, a terpenoid dialdehyde isolated from Polygonum hydropiper L., is a known agonist of the transient receptor potential vanilloid 1 (TRPV1). In this investigation a series of polygodial analogues were prepared and investigated for TRPV1-agonist and anticancer activities. These experiments led to the identification of 9-epipolygodial, which has antiproliferative potency significantly exceeding that of polygodial. 9-Epipolygodial was found to maintain potency against apoptosis-resistant cancer cells as well as those displaying the multidrug-resistant (MDR) phenotype. In addition, the chemical feasibility for the previously proposed mechanism of action of polygodial, involving the formation of a Paal-Knorr pyrrole with a lysine residue on the target protein, was demonstrated by the synthesis of a stable polygodial pyrrole derivative. These studies reveal rich chemical and biological properties associated with polygodial and its direct derivatives. These compounds should inspire further work in this area aimed at the development of new pharmacological agents, or the exploration of novel mechanisms of covalent modification of biological molecules with natural products.

Wittig derivatization of sesquiterpenoid polygodial leads to cytostatic agents with activity against drug resistant cancer cells and capable of pyrrolylation of primary amines.

Many types of cancer, including glioma, melanoma, non-small cell lung cancer (NSCLC), among others, are resistant to proapoptotic stimuli and thus poorly responsive to current therapies based on the induction of apoptosis in cancer cells. The current investigation describes the synthesis and anticancer evaluation of unique C12-Wittig derivatives of polygodial, a sesquiterpenoid dialdehyde isolated from Persicaria hydropiper (L.) Delabre. These compounds were found to undergo an unprecedented pyrrole formation with primary amines in a chemical model system, a reaction that could be relevant in the biological environment and lead to the pyrrolation of lysine residues in the target proteins. The anticancer evaluation of these compounds revealed their promising activity against cancer cells displaying various forms of drug resistance, including resistance to proapoptotic agents. Mechanistic studies indicated that compared to the parent polygodial, which displays fixative general cytotoxic action against human cells, the C12-Wittig derivatives exerted their antiproliferative action mainly through cytostatic effects explaining their activity against apoptosis-resistant cancer cells. The possibility for an intriguing covalent modification of proteins through a novel pyrrole formation reaction, as well as useful activities against drug resistant cancer cells, make the described polygodial-derived chemical scaffold an interesting new chemotype warranting thorough investigation.

Gallic acid functions as a TRPA1 antagonist with relevant antinociceptive and antiedematogenic effects in mice.

The transient receptor potential ankyrin 1 (TRPA1) has been identified as a relevant target for the development of novel analgesics. Gallic acid (GA) is a polyphenolic compound commonly found in green tea and various berries and possesses a wide range of biological activities. The goal of this study was to identify GA as a TRPA1 antagonist and observe its antinociceptive effects in different pain models. First, we evaluated the ability of GA to affect cinnamaldehyde-induced calcium influx. Then, we observed the antinociceptive and antiedematogenic effects of GA (3-100 mg/kg) oral administration after the intraplantar (i.pl.) injection of TRPA1 agonists (allyl isothiocyanate, cinnamaldehyde, or hydrogen peroxide-H2O2) in either an inflammatory pain model (carrageenan i.pl. injection) or a neuropathic pain model (chronic constriction injury) in male Swiss mice (25-35 g). GA reduced the calcium influx mediated by TRPA1 activation. Moreover, the oral administration of GA decreased the spontaneous nociception triggered by allyl isothiocyanate, cinnamaldehyde, and H2O2. Carrageenan-induced allodynia and edema were largely reduced by the pretreatment with GA. Moreover, the administration of GA was also capable of decreasing cold and mechanical allodynia in a neuropathic pain model. Finally, GA was absorbed after oral administration and did not produce any detectable side effects. In conclusion, we found that GA is a TRPA1 antagonist with antinociceptive properties in relevant models of clinical pain without detectable side effects, which makes it a good candidate for the treatment of painful conditions.

Antinociceptive and antiedematogenic effect of pecan (Carya illinoensis) nut shell extract in mice: a possible beneficial use for a by-product of the nut industry.

Abstract Background: Interest in pecan (Carya illinoensis) nut shells, a by-product of the nut industry, has increased due to its anti-inflammatory and antioxidant activities. The goal of this study was to evaluate the antinociceptive and antiedematogenic activity and the mechanisms of the pecan shell aqueous extract (AE). Methods: First, we performed fingerprinting of C. illinoensis AE. The antinociceptive and antiedematogenic effects of AE intragastric (i.g.) administration in mice (male Swiss mice 20-30 g) were evaluated using the acetic acid test or after subcutaneous (s.c.) paw injection of diverse transient receptor potential ankyrin 1 (TRPA1) agonists, including hydrogen peroxide (H2O2), allyl isothiocyanate, or cinnamaldehyde. We also observed AE antinociceptive and antiedematogenic effects after carrageenan s.c. paw injection and measured H2O2 production. Moreover, we observed the development of adverse effects after AE i.g. treatment. Results: The high-performance liquid chromatography fingerprinting of AE showed the presence of rutin. AE or rutin i.g. treatment produced antinociception in the acetic acid test and reduced the nociception and edema mediated by H2O2 s.c. hind paw injection or nociception induced by other TRPA1 agonists. Moreover, AE or rutin reduced the hyperalgesia, edema, and H2O2 production induced by carrageenan s.c. paw injection. No motor, gastric, or toxicological alterations were observed after AE administration. Conclusions: Collectively, the present results show that AE and its constituent rutin produced antinociceptive and antiedematogenic action in models of acute and persistent inflammatory nociception and it seems to be related to the inhibition of TRPA1 receptor activation.

The antinociceptive and anti-inflammatory effects of the crude extract of Jatropha isabellei in a rat gout model.

ETHNOPHARMACOLOGICAL RELEVANCE: Jatropha isabellei Müll Arg. (Euphorbiaceae) is a medicinal plant that has been used in South American folk medicine for the treatment of arthritic diseases, particularly gout. AIM OF THE STUDY: This study was designed to verify the antinociceptive, anti-inflammatory and hypouricemic potential of Jatropha isabellei. MATERIALS AND METHODS: Rats were orally administered with the crude extract (100-300 mg/kg) or a fraction that is rich in alkaloids (0.15 mg/kg) of Jatropha isabellei. An intra-articular (i.a.) injection of 50 µl of monosodium urate (MSU) crystals (1.25mg/site) was used to generate the gout model to assess the effect of the treatment on nociception (thermal and mechanical hyperalgesia) and inflammation (oedema and neutrophil infiltration). The effect of Jatropha isabellei on the serum levels of uric acid was evaluated in a model of hyperuricaemia induced by the intraperitoneal injection of potassium oxonate (250 mg/kg). The side effects were analysed using an open-field test, gastric lesion assessment and by measuring the levels of the ALT and AST enzymes. RESULTS: Our study demonstrated that the crude extract of Jatropha isabellei and a fraction rich in alkaloids were able to prevent the thermal hyperalgesia, mechanical allodynia, oedema and neutrophil infiltration induced by intra-articular MSU injection in rats. On the other hand, treatment with Jatropha isabellei did not alter the uric acid levels increased by potassium oxonate in the hyperuricaemia model. In addition, Jatropha isabellei did not induce gastric lesions or liver damage and did not alter spontaneous locomotor activity. CONCLUSION: The crude extract of Jatropha isabellei and its fraction rich in alkaloid presents antinociceptive and anti-inflammatory effects in a rat gout model, similar to that observed after treatment with colchicine, supporting the traditional use of this plant in gouty patients.

Mechanisms involved in abdominal nociception induced by either TRPV1 or TRPA1 stimulation of rat peritoneum.

Abdominal pain is a frequent symptom of peritoneal cavity irritation, but little is known about the role of the receptors for irritant substances, transient receptor potential vanilloid 1 (TRPV1) and ankyrin 1 (TRPA1), in this painful condition. Thus, we investigated the abdominal nociception caused by peritoneal stimulation with TRPV1 (capsaicin) and TRPA1 (allyl isothiocyanate, AITC) agonists and their mechanisms in rats. The intraperitoneal (i.p.) injection of either capsaicin or AITC (0.03-10 mg/kg) induced short-term (up to 20 min) and dose-dependent abdominal nociception, and also produced c-fos expression in spinal afferents of the dorsal horn. TRPV1 antagonism prevented (94 ± 4% inhibition) nociception induced by capsaicin but not by AITC. In contrast, the TRPA1 antagonism almost abolished AITC-induced nociception (95 ± 2% inhibition) without altering the capsaicin response. Moreover, nociception induced by either capsaicin or AITC was reduced by the desensitisation of TRPV1-positive sensory fibres with resiniferatoxin (73 ± 18 and 76 ± 15% inhibitions, respectively) and by the NK1 receptor antagonist aprepitant (56 ± 5 and 53 ± 8% inhibitions, respectively). Likewise, the i.p. injections of capsaicin or AITC increased the content of substance P in the peritoneal fluid. Nevertheless, neither the mast cell membrane stabiliser cromoglycate, nor the H1 antagonist promethazine, nor depletion of peritoneal macrophages affected abdominal nociception induced either by capsaicin or AITC. Accordingly, neither capsaicin nor AITC increased the histamine content in the peritoneal fluid or provoked peritoneal mast cell degranulation in vitro. Collectively, our findings suggest that TRPV1 and TRPA1 stimulation in the peritoneum produces abdominal nociception that is mediated by sensory fibres activation.

Effect of ω-conotoxin MVIIA and Phα1ß on paclitaxel-induced acute and chronic pain.

The treatment with the chemotherapeutic agent paclitaxel produces a painful peripheral neuropathy, and is associated with an acute pain syndrome in a clinically significant number of patients. However, no standard therapy has been established to manage the acute pain or the chronic neuropathic pain related to paclitaxel. In the present study, we evaluated the analgesic potential of two N-type voltage-gated calcium channel (VGCC) blockers, ω-conotoxin MVIIA and Phα1ß, on acute and chronic pain induced by paclitaxel. Adult male rats were treated with four intraperitoneal injections of paclitaxel (1+1+1+1mg/kg, in alternate days) and the development of mechanical hyperalgesia was evaluated 24h (acute painful stage) or 15days (chronic painful stage) after the first paclitaxel injection. Not all animals showed mechanical hyperalgesia 24h after the first paclitaxel injection, but those that showed developed a more intense mechanical hyperalgesia at the chronic painful stage. Intrathecal administration (i.t.) of ω-conotoxin MVIIA (3-300pmol/site) or Phα1ß (10-300pmol/site) reduced the mechanical hyperalgesia either at the acute or at the chronic painful stage induced by paclitaxel. When administered at the acute painful stage, ω-conotoxin MVIIA (300pmol/site, i.t.) and Phα1ß (300pmol/site, i.t.) prevented the worsening of chronic mechanical hyperalgesia. Furthermore, Phα1ß (30-300pmol/site, i.t.) elicited less adverse effects than ω-conotoxin MVIIA (10-300 pmol/site, i.t.). Taken together, our data evidence the involvement of N-type VGCC in pain sensitization induced by paclitaxel and point out the potential of Phα1ß as a safer alternative than ω-conotoxin MVIIA to treat the pain related to paclitaxel.

Antinociceptive effect of Mirabilis jalapa on acute and chronic pain models in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: The infusion or decoction of Mirabilis jalapa leaves is used in traditional medicine in Brazil to treat inflammatory and painful diseases. Thus, the present study was designed to investigate whether the leaf ethyl acetate (Eta) fraction from Mirabilis jalapa exhibits antinociceptive effect in clinically relevant pain models in mice. Furthermore, we have investigated the role of cholinergic system in the antinociceptive action produced by Eta in mice. MATERIALS AND METHODS: The effect of Eta administered orally (10mg/kg, p.o.) in mice was verified on the painful hypersensitivity (mechanical allodynia) in models of chronic inflammation (subcutaneous injection of complete Freund's Adjuvant-CFA in the plantar surface of the right hind paw), postoperative (paw surgical incision) and neuropathic (partial sciatic nerve ligation) pain. In the chronic inflammation model, we further verified the effect of Eta treatment on paw edema and interleukin-1ß (IL-1ß) levels. We also investigated the role of muscarinic and nicotinic receptors in the antiallodynic action produced by Eta as well as the possible action of Eta on in vitro and ex vivo acetylcholinesterase activity in CFA treated animals. Furthermore, we verified the effect of Eta treatment on the parameters of liver and kidney lesion (level of urea, and activity of aspartate aminotransferase and alanine aminotransferase). RESULTS: Eta produced marked reduction in the allodynia caused by CFA, surgical incision and partial sciatic nerve ligation. However, Eta did not alter the paw edema or the increase of IL-1ß levels produced by CFA. The antinociceptive effect of Eta was reversed by the pre-treatment of animals with the antagonists of muscarinic (atropine, 5mg/kg, s.c) or nicotinic (mecamylamine, 0.001mg/kg, s.c.) receptors. Eta did not alter in vitro acetylcholinesterase activity in blood or spinal cord samples, but it reversed the increase in the acetylcholinesterase activity observed in the spinal cord samples from mice injected with CFA. Moreover, Eta did not alter the indicators of liver or kidney lesion. CONCLUSIONS: Based on its use in traditional medicine, the results of the present study confirmed the antinociceptive properties of Eta in clinically relevant pain models. Also its effect on the CFA-induced chronic inflammation seems to be related to acetylcholinesterase inhibition and cholinergic system.

A novel, potent, oral active and safe antinociceptive pyrazole targeting kappa opioid receptors.

Pyrazole compounds are an intriguing class of compounds with potential analgesic activity; however, their mechanism of action remains unknown. Thus, the goal of this study was to explore the antinociceptive potential, safety and mechanism of action of novel 1-pyrazole methyl ester derivatives, which were designed by molecular simplification, using in vivo and in vitro methods in mice. First, tree 1-pyrazole methyl ester derivatives (DMPE, MPFE, and MPCIE) were tested in the capsaicin test and all presented antinociceptive effect; however the MPClE (methyl 5-trichloromethyl-3-methyl-1H-pyrazole-1-carboxylate) was the most effective. Thus, we selected this compound to assess the effects and mechanisms in subsequent pain models. MPCIE produced antinociception when administered by oral, intraperitoneal, intrathecal and intraplantar routes and was effective in the capsaicin and the acetic acid-induced nociception tests. Moreover, this compound reduced the hyperalgesia in diverse clinically-relevant pain models, including postoperative, inflammatory, and neuropathic nociception in mice. The antinociception produced by orally administered MPClE was mediated by κ-opioid receptors, since these effects were prevented by systemically pre-treatment with naloxone and the κ-opioid receptor antagonist nor-binaltorphimine. Moreover, MPCIE prevented binding of the κ-opioid ligand [(3)H]-CI-977 in vitro (IC50 of 0.68 (0.32-1.4) µM), but not the TRPV1 ([(3)H]-resiniferatoxin) or the α2-adrenoreceptor ([(3)H]-idazoxan) binding. Regarding the drug-induced side effects, oral administration of MPClE did not produce sedation, constipation or motor impairment at its active dose. In addition, MPCIE was readily absorbed after oral administration. Taken together, these results demonstrate that MPClE is a novel, potent, orally active and safe analgesic drug that targets κ-opioid receptors.

Role of peripheral polyamines in the development of inflammatory pain.

Polyamines (putrescine, spermidine and spermine) are aliphatic amines that are produced by the action of ornithine decarboxylase (ODC) in a rate-limiting and protein kinase C (PKC)-regulated step. Because high levels of polyamines are found in the synovial fluid of arthritic patients, the aim of the present study was to identify the role of peripherally produced polyamines in a model of inflammatory pain induced by adjuvant. The subcutaneous injection of Complete Freund's adjuvant (CFA, 50 µL/paw) caused the development of mechanical allodynia and edema. Moreover, it increased ODC expression and activity and PKC activation. Administration of the selective ODC inhibitor DFMO (10 µmol/paw) attenuated the development of allodynia and edema and decreased ODC activity in both control and CFA-treated animals. Furthermore, administration of the PKC inhibitor GF109203X (1 nmol/paw) reduced allodynia and ODC activity in animals injected with CFA. A subcutaneous injection of putrescine (10 µmol/paw), spermidine (3-10 µmol/paw) or spermine (0.3-3 µmol/paw) into the rat paw also caused mechanical allodynia and edema. The present results suggest that endogenously synthesized polyamines are involved in the development of nociception and edema caused by an adjuvant. Moreover, polyamine production in inflammatory sites seems to be related to an increase in ODC activity stimulated by PKC activation. Thus, controlling polyamine synthesis and action could be a method of controlling inflammatory pain.