Formalin-induced Hyperalgesia Research Articles

Effects of riboflavin on hyperalgesia and serum glutamine-to-glutamate ratio in rats with painful diabetic neuropathy

Previous studies have explored the antinociceptive effects of riboflavin (vitamin B2) across various experimental models. However, there remains a gap in the literature regarding its potential to alleviate neuropathic pain in diabetes. This study aims to investigate the effects of riboflavin on hyperalgesia and serum glutamine-to-glutamate ratio in rats with painful diabetic neuropathy. In fasted rats, a model of painful diabetic neuropathy was induced through intraperitoneal injection of streptozotocin. In the fifth week post-injection, diabetic rats experiencing neuropathic pain were administered daily doses of riboflavin (25 or 50 mg), dissolved in their drinking water, for a duration of two weeks. Results demonstrate that riboflavin significantly reduced mechanical and cold-induced hyperalgesia in diabetic rats compared to controls. Formalin-induced hyperalgesia was alleviated by riboflavin in the second phase. Additionally, riboflavin supplementation increased the serum glutamine-to-glutamate ratio in these animals. These findings highlight the therapeutic potential of riboflavin in managing neuropathic pain associated with diabetes.

Orofacial anti-hypernociceptive effect of citral in acute and persistent inflammatory models in rats

Orofacial pain has significant psychological and physiological effects. Citral (3,7-dimethyl-2,6-octadienal) is the main component of Cymbopogon citratus (DC) Stapf, an herb with analgesic properties. Although citral has been considered a potent analgesic, its putative effects on orofacial pain are still unknown. ObjectiveThe objective of this study is to test the hypothesis that citral modulates orofacial pain using two experimental models: formalin-induced hyperalgesia in the vibrissae area and during persistent temporomandibular hypernociception using Complete Freund's Adjuvant - CFA test. MethodsFor the formalin test, citral (100 and 300 mg/kg, oral gavage) or its vehicle (Tween 80, 1 %) were given 1 h before the formalin injection subcutaneously (sc) into the vibrissae area. For the CFA model, we analyzed the prophylactic (100 mg/kg of citral by oral gavage, 1 h before CFA injection) and the chronic therapeutic (citral treatment 1-hour post-CFA injection and daily post-CFA injection) effect of citral or its vehicle in animals treated with CFA for 8 days. ResultsCitral caused a decrease in formalin-induced local inflammation and the time spent performing nociceptive behavior in a dose-dependent fashion. Similarly, prophylactic and therapeutic citral treatment decreased the CFA-induced persistent mechanical hypernociception in the temporomandibular area. ConclusionOur data strengthen the notion that citral plays a powerful antinociceptive role by decreasing orofacial hypernociception in formalin and CFA models.

Investigation of the anti-inflammatory and analgesic activities of promising pyrazole derivative

The development of new COX-2 inhibitors with analgesic and anti-inflammatory efficacy as well as minimal gastrointestinal, renal and cardiovascular toxicity, is of vital importance to patients suffering from chronic course pain and inflammatory conditions. This study aims at evaluating the therapeutic activity and adverse drug reactions associated with the use of the newly synthesized pyrazole derivative, compound AD732, E-4-[3-(4-methylphenyl)-5-hydroxyliminomethyl-1H-pyrazol-1-yl]benzenesulfonamide, as compared to indomethacin and celecoxib as standard agents. Anti-inflammatory activity was assessed using carrageenan-induced rat paw edema and cotton pellet granuloma tests; formalin-induced hyperalgesia and hot plate tests were done to study analgesic activity. In vitro tests to determine COX-1/COX-2 selectivity and assessment of renal and gastric toxicity upon acute exposure to AD732 were also conducted. Compound AD732 exhibited promising results; higher anti-inflammatory and analgesic effects compared to standard agents, coupled with the absence of ulcerogenic effects and minimal detrimental effects on renal function. Additionally, compound AD732 was a less potent inhibitor of COX-2 in vitro than celecoxib, which may indicate lower potential cardiovascular toxicity. It may be concluded that compound AD732 appears to be a safer and more effective molecule with promising potential for the management of pain and inflammation.

(256) Peripherally-Restricted Dual-Acting Kappa/Delta Opioid Agonist (CA1001) Prevents Formalin-Induced Hyperalgesia

Peripheral mu-opioid receptors (MOR) are responsible for opioid-induced constipation, but in the uninjured state do not participate significantly in the pain pathway. On the other hand, kappa- (KOR) and delta-opioid (DOR) receptors are constitutively present in the periphery. However, the DOR only becomes active following the induction of the inflammatory response. The relative contribution of DOR agonism to KOR agonism was examined in the formalin model. Subcutaneous plantar injection of formalin causes a bi-phasic nocifensive behavioral response in rodents. The early phase (phase 1) lasts for about 5-10 minutes, following which an interphase occurs without any discernible nociceptive reactions, after which the late phase (phase 2) nociceptive reaction ensues continuing from about 20-60 minutes. Phase 2 of the formalin model is a model of continuously present persistent pain. The late phase (phase 2), in particular, is considered as a pharmacodynamic surrogate of central sensitization. A novel dual-acting, peripherally-restricted kappa-delta opioid agonist (CA1001) was compared to a peripherally-restricted kappa agonist (ICI204448) in the formalin model in the mouse. Following IACUC approval, mice were randomly pretreated with inert vehicle, ICI204448 1mg/kg, ICI204448 10mg/kg, CA1001 1mg/kg, or CA1001 10 mg/kg. Spontaneous nocifensive behaviors were blindly assessed (video recording). Neither agent was effective in reducing the acute (0-5 minutes) response to formalin injection (phase 1). CA1001 1 mg/kg was as effective as ICI204448 10 mg/kg in reducing formalin-induced responses at 20-35 minutes; CA1001 10 mg/kg was significantly more effective than ICI204448 10 mg/kg (phase 2; p

Osthole ameliorates neurogenic and inflammatory hyperalgesia by modulation of iNOS, COX-2, and inflammatory cytokines in mice.

Osthole is a bioactive component reported in medicinal plants such as Angelica pubescens and Cnidium monnieri, known for analgesic activity. However, the toxicity, median effective dose (ED50), and dual modulation of nitric oxide and cyclooxygenase pathways along with inflammatory cytokines of osthole are yet to be determined. The animals (mice) were assessed for general behaviour and mortality in varying doses (50, 300, and 2000mgkg-1) of osthole for acute toxicity over 14days. The analgesic activity was investigated using acetic acid and formalin-induced hyperalgesia, and anti-inflammatory activity was explored in carrageenan-induced paw oedema. ED50 of osthole was calculated using Design Expert software. Involvement of nitric oxide and cyclooxygenase pathways was investigated by agonist challenges with L-arginine and substance P, respectively. The expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) was determined in spinal sections by immunohistochemical analysis. Lipopolysaccharide (LPS) challenge was used to assess in vivo effect on inflammatory cytokines (TNFα and IL-6). Acute toxicity studies revealed no behavioural abnormality or mortality on osthole treatment and unremarkable histological findings. Osthole was found to significantly decrease acetic acid and formalin-induced hyperalgesia (ED50 = 5.43mgkg-1) and carrageenan-induced paw oedema with no toxicity symptoms. Osthole produced a marked decrease in iNOS and COX-2 expression as well as TNFα and IL-6. The findings corroborate to modulation of iNOS and COX-2 and inflammatory cytokines by osthole. This study provides promising insights and prospects for application of osthole in pain management.

Analgesic Effects of Duloxetine on Formalin-Induced Hyperalgesia and Its Underlying Mechanisms in the CeA.

In rodents, the amygdala has been proposed to serve as a key center for the nociceptive perception. Previous studies have shown that extracellular signal-regulated kinase (ERK) signaling cascade in the central nucleus of amygdala (CeA) played a functional role in inflammation-induced peripheral hypersensitivity. Duloxetine (DUL), a serotonin and noradrenaline reuptake inhibitor, produced analgesia on formalin-induced spontaneous pain behaviors. However, it is still unclear whether single DUL pretreatment influences formalin-induced hypersensitivity and what is the underlying mechanism. In the current study, we revealed that systemic pretreatment with DUL not only dose-dependently suppressed the spontaneous pain behaviors, but also relieved mechanical and thermal hypersensitivity induced by formalin hindpaw injection. Consistent with the analgesic effects of DUL on the pain behaviors, the expressions of Fos and pERK that were used to check the neuronal activities in the spinal cord and CeA were also dose-dependently reduced following DUL pretreatment. Meanwhile, no emotional aversive behaviors were observed at 24 h after formalin injection. The concentration of 5-HT in the CeA was correlated with the dose of DUL in a positive manner at 24 h after formalin injection. Direct injecting 5-HT into the CeA suppressed both the spontaneous pain behaviors and hyperalgesia induced by formalin injection. However, DUL did not have protective effects on the formalin-induced edema of hindpaw. In sum, the activation of CeA neurons may account for the transition from acute pain to long-term hyperalgesia after formalin injection. DUL may produce potent analgesic effects on the hyperalgesia and decrease the expressions of p-ERK through increasing the concentration of serotonin in the CeA.

Inhibition of monoacylglycerol lipase: Another signalling pathway for potential therapeutic targets in migraine?

Background Drugs that modulate endocannabinoid signalling are effective in reducing nociception in animal models of pain and may be of value in the treatment of migraine. Methods We investigated the anti-nociceptive effects of inhibition of monoacylglycerol lipase (MGL), a key enzyme in the hydrolysis of the 2-arachidonoylglycerol, in a rat model of migraine based on nitroglycerin (NTG) administration. We evaluated c-fos expression in specific brain areas and nociceptive behavior in trigeminal and extra-trigeminal body areas. Results URB602, a reversible MGL inhibitor, did not show any analgesic effect in the tail flick test, but it inhibited NTG-induced hyperalgesia in both the tail flick test and the formalin test applied to the hind paw or to the orofacial area. Quite unexpectedly, URB602 potentiated formalin-induced hyperalgesia in the trigeminal area when used alone. The latter result was also confirmed using a structurally distinct, irreversible MGL inhibitor, JZL184. URB602 did not induce neuronal activation in the area of interest, but significantly reduced the NTG-induced neuronal activation in the ventrolateral column of the periaqueductal grey and the nucleus trigeminalis caudalis. Conclusions These findings support the hypothesis that modulation of the endocannabinoid system may be a valuable approach for the treatment of migraine. The topographically segregated effect of MGL inhibition in trigeminal/extra-trigeminal areas calls for further mechanistic research.

The Effect of Carvacrol on Inflammatory Pain and Motor Coordination in Rats

Abstract Carvacrol is a monoterpenic phenol and an active ingredient of the plant essential oils of the family Lamiaceae. We have investigated the analgesic effect of carvacrol, the possible dependence of the effect in relation to animal sex, and the impact of carvacrol on motor coordination in rats. Hyperalgesia was induced by formalin (1.5%), which was administered SC in the upper lip of rat. Hyperalgesia and effects of carvacrol and indomethacin were measured by using the orofacial formalin test. The influence on motor coordination in animals treated with carvacrol was investigated by using the rota-rod test. Carvacrol administered PO in pre-treatment (45 min. prior to formalin) at a single dose of 50, 75 and 100 mg /kg BW, in the male, 50 and 100 mg /kg BW, in female rats caused a dose-dependent antinociceptive effect. This effect of carvacrol was significantly higher (P<0.01, P<0.001) in male rats. Compared with indomethacin administered during pre-treatment (2 mg/kg, PO), carvacrol (100 mg/kg) exhibits significantly higher (P <0.05 and P <0.001) antinociceptive effect on formalininduced hyperalgesia in male rats. In the rota-rod test carvacrol did not disturb the motor coordination in male rats, nor the dose of carvacrol with clear antinociceptive properties exhibited depressive effect on the CNS of treated rats. Keeping in mind that the monoterpene carvacrol is of plant origin, with potentially less side effects and without residues, it is realistic to expect the possibility of its therapeutic use in the treatment of inflammatory pain in animals.

Antihyperalgesic Activity of Rhodiola rosea in a Diabetic Rat Model.

Preclinical Research Rhodiola rosea L. (Crassulaceae) is used for enhancing physical and mental performance. Recent studies demonstrated that R. rosea had anti-inflammatory activity in animal models, for example, carrageenan- and nystatin-induced edema in rats, possibly by inhibiting phospholipase A2 and cyclooxygenases-1 and -2. In addition, R. rosea had antinociceptive activity in thermal and chemical pain tests as well as mechanical hyperalgesia. The purpose of the present study was to assess the antihyperalgesic effect of an ethanol extract of Rhodiola rosea (R. rosea) in a diabetic rat model. Rats were administered a single dose of streptozotocin (STZ; 50 mg/kg, i.p.) and hyperalgesia was evaluated four weeks later. Formalin-evoked (0.5%) flinching was increased in diabetic rats compared with nondiabetic controls Systemic (1-100 mg/kg, i.p.) and local (0.1-10 mg/paw into the dorsal surface of the right hind paw) administration of R. rosea ethanol extract dose-dependently reduced formalin-induced hyperalgesia in diabetic rats. The antihyperalgesic effect of R. rosea was compared with gabapentin. These results suggest that R. rosea ethanol extract may have potential as a treatment for diabetic hyperalgesia.

Inferior Alveolar Nerve Transection Enhanced Formalin-Induced Nocifensive Responses in the Upper Lip: Systemic Buprenorphine Had More Antinociceptive Efficacy over Morphine

This study was designed to investigate the efficacy of a partial μ-opioid agonist, buprenorphine, against the formalin-induced hyperalgesia in the upper lip in chronically inferior alveolar nerve (IAN)-transected rats. Subcutaneous injection of diluted formalin into the upper lip in the IAN-transected rats showed an increased number of pain-related behavior (PRB; face-rubbing behavior) in every phase up to 45 min (p < 0.01) compared with that in the nontransected sham control rats. The numbers of c-Fos-immunoreactive (IR) cells in the superficial layers of the trigeminal nucleus caudalis (VcI/II) at the rostral (0-0.7 mm caudal to the obex) and middle levels (1.4-2.2 mm caudal to the obex) 2 h after the formalin injection in the IAN-transected rats were significantly increased compared with those in the control rats. The PRB in phases 1 and 2 (0-15 and 15-30 min after formalin injection) in rats with preadministration of morphine (3 mg/kg i.p.) or buprenorphine (100 µg/kg i.p.) was significantly (p < 0.05) smaller than those in the control rats. There was no significant difference in the efficacy between morphine and buprenorphine at these doses. The antinociceptive efficacy in phase 2 of buprenorphine (100 µg/kg) was higher (p < 0.05) than that of morphine (3 mg/kg) in the IAN-transected rats. The number of c-Fos-IR cells in the VcI/II at every level (0-3.6 mm caudal to the obex) after formalin injection was significantly decreased (p < 0.01) with preadministration of morphine (3 mg/kg) or buprenorphine (100 µg/kg) in the control rats. In the IAN-transected rats, the number of c-Fos-IR cells in the caudal VcI/II (2.2-3.6 mm caudal to the obex) after formalin injection was significantly decreased (p < 0.01) with preadministration of buprenorphine (100 µg/kg) but not so much (2.2-2.9 mm caudal to the obex, p < 0.05; 2.9-3.6 mm caudal to the obex, p > 0.05) with preadministration of morphine (3 mg/kg). These results indicate that IAN transection enhanced formalin-induced nocifensive responses in the upper lip, the dermatome of the intact nerve neighboring the IAN. Systemic preadministration of buprenorphine had more antinociceptive effects on the formalin-induced nocifensive behavior in the upper lip compared with morphine in the IAN-transected rats.

Critical Role of Protease-activated Receptor 2 Activation by Mast Cell Tryptase in the Development of Postoperative Pain

Studies have indicated that nearly half of all surgical patients still have inadequate pain relief. Thus, it is crucial to understand the mechanisms involved in postoperative pain in order to better treat it. Thus, the aim of this study was to investigate the involvement of mast cell degranulation, tryptase and its substrate, the protease-activated receptor 2, in a model of postoperative pain in mice. We evaluated the effect of the compound 48/80 (to cause mast cell mediator depletion), cromoglycate or ketotifen (mast cell stabilizers), gabexate (tryptase inhibitor) or N3-methylbutyryl-N-6-aminohexanoyl-piperazine (protease-activated receptor 2 antagonist) in a postoperative pain model in mice (n = 5-10). Mast cell degranulation and tryptase activity were also evaluated in the operated tissue (n= 5-8). The pre-treatment with compound 48/80 or ketotifen was able to prevent nociception throughout the postoperative hyperalgesia course (until 5 days after surgery), whereas cromoglycate presented a shorter effect (until 1 day). Gabexate or N3-methylbutyryl-N-6-aminohexanoyl-piperazine also produced a short-lasting effect in preventing postoperative nociception. However, neither gabexate, N3-methylbutyryl-N-6-aminohexanoyl-piperazine nor cromoglycate was capable of reversing nociception when administered after incision. Surgery led to early mast cell degranulation on the incised tissue and increased tryptase activity in tissue perfusates. Cromoglycate fully prevented the tryptase release in the perfusate and the compound 48/80 substantially reduced tryptase activity in the incised tissue. Thus, the mast cell degranulation with the subsequent release of tryptase and protease-activated receptor 2 activation are potential targets for the development of novel therapies to prevent, but not reverse, postoperative pain.

Antinociceptive Effects of Intraperitoneal and Intrathecal Vitamin E in the Rat Formalin Test

BackgroundVitamin E is widely known to be one of the reactive oxygen species (ROS) scavengers and a drug that can easily be obtained, and it has been shown to attenuate the pain responses induced by various causes in animal pain models. Thus, this experiment was conducted to assess the antinociceptive effects of vitamin E by comparing intraperitoneal and intrathecal injections in rats subjected to the formalin test.MethodsAfter the intraperitoneal and intrathecal injections of vitamin E were carried out, respectively (IP: 500 mg/kg, 1 g/kg, and 2 g/kg, IT: 3 mg/kg, 10 mg/kg, and 30 mg/kg), the formalin test was perfumed. As soon as 5% formalin was injected into left hind paw, the number of flinches induced by pain was measured at 5-minute intervals for 1 hour.ResultsFormalin injected into the left hind paw induced biphasic nociceptive behavior in all animals. Intraperitoneal injection of vitamin E diminished the nociceptive behavior in a dose-dependent manner during the early and late phase. Intrathecal vitamin E diminished nociceptive behavior dose dependently during the late phase but showed no significant difference in the early phase.ConclusionsVitamin E attenuated acute nociception when it was injected systemically, while both systemic and intrathecal injection produced analgesia in a rat model of formalin-induced hyperalgesia.

Regulation of neuronal functions by the E3-ubiquitinligase protein associated with MYC (MYCBP2).

The E3-ubiquitinligase MYCBP2 regulates neuronal growth, synaptogenesis and synaptic plasticity by modulating several signaling pathways including the p38 MAPK signaling cascade. We found that loss of MYCBP2 in peripheral sensory neurons inhibits the internalization of transient receptor potential vanilloid receptor 1 (TRPV1) in a p38 MAPK-dependent manner. This prevented desensitization of activity-induced calcium increases and prolongs formalin-induced thermal hyperalgesia in mice. Besides its function in pain perception TRPV1 is also involved in the regulation of neuronal growth. Therefore, the observed effect of MYCBP2 on TRPV1 internalization could be part of the mechanisms underlying its well documented regulatory role in neuronal growth. The clarification of the mechanism is important for the understanding of the different MYCBP2-functions in diverse neuronal subpopulations and species.

Activation of metabotropic glutamate receptor 7 in spinal cord inhibits pain and hyperalgesia in a novel formalin model in sheep

This study set out to characterize the contribution of group III metabotropic glutamate receptor 7 activation to nociceptive behaviour and mechanical hypersensitivity in a novel formalin test in sheep. The mGlu receptor 7 allosteric agonist, N,N'-dibenzhydrylethane-1,2-diamine dihydrochloride (AMN082; 2-20 mM), the nonselective group III mGlu receptor agonist L-(+)-2-amino-4-phosphonobutyric acid (0.2-20 mM) and drug vehicle were injected intrathecally into naive subjects (n=7 per group), or 5 min preformalin (3%; 0.2 ml)/saline injection (intradermal), into the lower forelimb of adult female sheep (n=5-7 per group). Forelimb withdrawal thresholds to noxious mechanical stimulation and pain behaviours (time spent nonweight bearing or flinching) were assessed for up to 180 min. Formalin induced a characteristic biphasic pain-behaviour response and mechanical hyperalgesia between 1-5 and 30-120 min postinjection. Treatment with AMN082, but not L-(+)-2-amino-4-phosphonobutyric acid significantly inhibited both early and late phase formalin-induced hyperalgesia and pain behaviours. AMN082 also induced a rapid but short lasting analgesia in naive subjects. These data suggest that enhancing endogenous metabotropic glutamate receptor 7 activity in spinal cord, using the novel allosteric modulator, AMN082, blocks pain and hyperalgesia, and may be of therapeutic benefit for the treatment of inflammatory pain.

Stress-induced hyperalgesia is associated with a reduced and delayed GABA inhibitory control that enhances post-synaptic NMDA receptor activation in the spinal cord

GABA and glutamate are both affected by stress and are involved in nociception. Thus, we determined whether stress-induced enhancement of inflammatory hyperalgesia is mediated by an imbalance between glutamate and GABA neurotransmission. Male rats were subjected daily to 10 to 20minutes per day of either forced swimming (FS) or sham swimming for 3 consecutive days; nonconditioned rats served as controls. Some rats were treated i.p. with ketamine (5mg/kg), diazepam (2mg/kg), flumazenil (0.1mg/kg), or vehicle (0.9% NaCl), 30 to 60minutes before each conditioning session or nociception assessment. Pain behavior, spinal nociceptive neuronal activation and GABA and glutamate release were respectively evaluated by the formalin test, the expression of c-Fos and in vivo microdialysis of superficial laminae of the lumbar spinal cord, 48hours after the last conditioning session. Nitric oxide metabolites (NOx) were determined as markers of post-synaptic NMDA receptor activation. FS stress enhanced formalin-induced hyperalgesia, increased pain-elicited c-Fos expression, decreased basal and delayed pain-induced GABA release, and increased basal and induced glutamate release. Hyperalgesia and c-Fos overexpression were blocked only by prestress treatment with diazepam and post-stress treatment with ketamine, whereas changes in GABA and glutamate release were reversed by prestress treatment with diazepam. Diazepam effects were blocked by flumazenil. NOx increased in lumbar spinal cord of FS rats by a mechanism antagonized by ketamine. Thus, stress-induced hyperalgesia is initiated by a decreased and delayed GABA release and GABA-A receptor activation, whereas it is maintained by increased glutamate release and NMDA glutamate receptor activation at the spinal level.

A nitric oxide (NO)-donating gabapentin, NCX 1236, exerts marked anti-allodynic/anti-hyperalgesic effects in rodent models of neuropathic pain

Nitric oxide (NO) plays a role in the development and maintenance of pain following nerve injury. This study was conducted to address the anti-allodynic/anti-hyperalgesic effects of NCX 1236, a novel NO-donating gabapentin, using three well defined rodent models, namely: the formalin-induced hyperalgesia in mice, the chronic constriction injury (CCI) in mice and the spinal nerve ligation (SNL) in rats. Plasma gabapentin exposure after treatment with NCX 1236 and gabapentin was also monitored. NCX 1236 resulted in similar gabapentin plasma exposure as for gabapentin when administered orally at equimolar doses. More specifically, gabapentin plasma levels increased rapidly after oral administration of either gabapentin or NCX 1236 (Tmax=30 min) and decayed during the following hours to reach basal levels 6h post-dosing. Like gabapentin (30 mg/kg, po), equimolar doses of NCX 1236 did not significantly modify the 1st phase of formalin-induced hyperalgesia (199±20; 252±44, 188±32 sec, respectively for vehicle, gabapentin and NCX 1236). However, NCX 1236 diminished the licking/flinching time more efficiently than gabapentin during the 2nd phase of this test (417±46; 301±37*; 195±30 sec*#, respectively for vehicle, gabapentin and NCX 1236, *p<0.05 vs. vehicle, #p<0.05 vs. gabapentin). In the CCI mice model, a single oral administration of NCX 1236 inhibited thermal hyperalgesia more effectively than gabapentin at similar exposure level (thermal threshold: 4.6±0.3, 5.8±0.5* and 7.7±0.5 sec*# for vehicle, gabapentin and NCX 1236, *p<0.05 vs. vehicle, #p<0.05 vs. gabapentin). Finally, NCX 1236 demonstrated superior activity compared to gabapentin on mechanical allodynia in SNL rats (mechanical threshold: 4.4±1.8, 3.8±1.2 and 10.7±2.2g*#, respectively for vehicle, gabapentin and NCX 1236, *p<0.05 vs. vehicle, #p<0.05 vs. gabapentin). These data suggest that, by virtue of its NO-donating properties, NCX 1236 reduces thermal and mechanical hyperalgesia/allodynia more efficiently than gabapentin at similar exposure in well defined rodent models of neuropathic pain. Nitric oxide (NO) plays a role in the development and maintenance of pain following nerve injury. This study was conducted to address the anti-allodynic/anti-hyperalgesic effects of NCX 1236, a novel NO-donating gabapentin, using three well defined rodent models, namely: the formalin-induced hyperalgesia in mice, the chronic constriction injury (CCI) in mice and the spinal nerve ligation (SNL) in rats. Plasma gabapentin exposure after treatment with NCX 1236 and gabapentin was also monitored. NCX 1236 resulted in similar gabapentin plasma exposure as for gabapentin when administered orally at equimolar doses. More specifically, gabapentin plasma levels increased rapidly after oral administration of either gabapentin or NCX 1236 (Tmax=30 min) and decayed during the following hours to reach basal levels 6h post-dosing. Like gabapentin (30 mg/kg, po), equimolar doses of NCX 1236 did not significantly modify the 1st phase of formalin-induced hyperalgesia (199±20; 252±44, 188±32 sec, respectively for vehicle, gabapentin and NCX 1236). However, NCX 1236 diminished the licking/flinching time more efficiently than gabapentin during the 2nd phase of this test (417±46; 301±37*; 195±30 sec*#, respectively for vehicle, gabapentin and NCX 1236, *p<0.05 vs. vehicle, #p<0.05 vs. gabapentin). In the CCI mice model, a single oral administration of NCX 1236 inhibited thermal hyperalgesia more effectively than gabapentin at similar exposure level (thermal threshold: 4.6±0.3, 5.8±0.5* and 7.7±0.5 sec*# for vehicle, gabapentin and NCX 1236, *p<0.05 vs. vehicle, #p<0.05 vs. gabapentin). Finally, NCX 1236 demonstrated superior activity compared to gabapentin on mechanical allodynia in SNL rats (mechanical threshold: 4.4±1.8, 3.8±1.2 and 10.7±2.2g*#, respectively for vehicle, gabapentin and NCX 1236, *p<0.05 vs. vehicle, #p<0.05 vs. gabapentin). These data suggest that, by virtue of its NO-donating properties, NCX 1236 reduces thermal and mechanical hyperalgesia/allodynia more efficiently than gabapentin at similar exposure in well defined rodent models of neuropathic pain.

A Novel COX‐2 Inhibitor Pyrazole Derivative Proven Effective as an Anti‐Inflammatory and Analgesic Drug

The introduction of new COX-2 inhibitors with high efficacy and enhanced safety profile would be a great achievement in the development of anti-inflammatory drugs. This study was designed to screen and assess the anti-inflammatory and analgesic activities as well as some of the expected side effects of some pyrazole derivatives, newly synthesized as potential COX-2 inhibitors at the Faculty of Pharmacy, Alexandria University and compared to indomethacin and celecoxib. Twelve compounds were screened for their anti-inflammatory activity using carrageenan-induced paw oedema and cotton pellet granuloma tests. On the basis of their apparent anti-inflammatory activity, four compounds with different substitutions were selected for the evaluation of their analgesic activity using the formalin-induced hyperalgesia and hot-plate tests. Compound AD 532, ((4-(3-(4-Methylphenyl)-4-cyano-1H-pyrazol-1-yl)benzenesulfonamide)), showed very promising results. In the single-dose and subchronic toxicity studies, compound AD 532 showed no ulcerogenic effect and produced minimal effects on renal function. Furthermore, compound AD 532 was a less potent inhibitor of COX-2 in vitro than celecoxib, which may indicate lower potential cardiovascular toxicity. It is concluded that compound AD 532 appears to be a promising and safe option for the management of chronic inflammatory conditions. This study recommends more in-depth investigation into the therapeutic effects and toxicity profile of this compound including its cardiovascular toxicity.

Spinal d-Amino Acid Oxidase Contributes to Neuropathic Pain in Rats

D-amino acid oxidase (DAO) is an enzyme catalyzing oxidative deamination of neutral and polar d-amino acids and is expressed in the kidneys, liver, and central nervous system (CNS) including the spinal cord. We have previously demonstrated that DAO gene deletion/mutation by using mutant ddY/DAO(-/-) mice and systemic administration of the DAO inhibitor sodium benzoate blocked formalin-induced hyperalgesia in mice. In this study, we further investigated the potential role of DAO in neuropathic pain in a rat model of tight L(5)/L(6) spinal nerve ligation. After L(5)/L(6) spinal nerve ligation, the mRNA expression (measured by real-time quantitative polymerase chain reaction) and enzyme activity (measured by a colorimetric method) of DAO in the lumbar spinal cord were markedly increased, in agreement with the development of neuropathic pain (mechanical allodynia). Intraperitoneal injection of sodium benzoate (400 mg/kg) specifically blocked mechanical allodynia in neuropathic rats and formalin-induced hyperalgesia but did not suppress acute pain responses in the tail-flick test or formalin test. Systemic injection of sodium benzoate also inhibited DAO activity in the lumbar spinal cord of rats. Furthermore, direct intrathecal (spinal cord) injection of benzoate (30 mug/rat) specifically blocked spinal nerve ligation-induced mechanical allodynia in neuropathic rats and formalin-induced hyperalgesia (but not acute pain) in the formalin test. Based on the above results, we conclude that spinal DAO plays a pronociceptive (rather than an antinociceptive) role and might be a target molecule for the treatment of chronic pain of neuropathic origin.

Diabetic Painful and Insensate Neuropathy: Pathogenesis and Potential Treatments

Advanced peripheral diabetic neuropathy (PDN) is associated with elevated vibration and thermal perception thresholds that progress to sensory loss and degeneration of all fiber types in peripheral nerve. A considerable proportion of diabetic patients also describe abnormal sensations such as paresthesias, allodynia, hyperalgesia, and spontaneous pain. One or several manifestations of abnormal sensation and pain are described in all the diabetic rat and mouse models studied so far (i.e., streptozotocin-diabetic rats and mice, type 1 insulinopenic BB/Wor and type 2 hyperinsulinemic diabetic BBZDR/Wor rats, Zucker diabetic fatty rats, and nonobese diabetic, Akita, leptin- and leptin-receptor-deficient, and high-fat diet-fed mice). Such manifestations are 1) thermal hyperalgesia, an equivalent of a clinical phenomenon described in early PDN; 2) thermal hypoalgesia, typically present in advanced PDN; 3) mechanical hyperalgesia, an equivalent of pain on pressure in early PDN; 4) mechanical hypoalgesia, an equivalent to the loss of sensitivity to mechanical noxious stimuli in advanced PDN; 5) tactile allodynia, a painful perception of a light touch; and 5) formalin-induced hyperalgesia. Rats with short-term diabetes develop painful neuropathy, whereas those with longer-term diabetes and diabetic mice typically display manifestations of both painful and insensate neuropathy, or insensate neuropathy only. Animal studies using pharmacological and genetic approaches revealed important roles of increased aldose reductase, protein kinase C, and poly(ADP-ribose) polymerase activities, advanced glycation end-products and their receptors, oxidative-nitrosative stress, growth factor imbalances, and C-peptide deficiency in both painful and insensate neuropathy. This review describes recent achievements in studying the pathogenesis of diabetic neuropathic pain and sensory disorders in diabetic animal models and developing potential pathogenetic treatments.

A comparative study on anti-hyperalgesia effect of MTA and Ketoprofen in inflammatory pain.

Mineral trioxide aggregate (MTA) is an endodontic material with different clinical applications e.g. root-end filling, pulp capping and perforation repair. It has been reported to possess antimicrobial and antifungal activities. The aim of this study was to examine the effect of White MTA on formalin-induced hyperalgesia in a rat with inflammatory pain. Inflammatory pain was induced by subcutaneous (SC) injection of formalin (40 µL, 2.5%) into the rat upper lip. The nociceptive behavioral responses i.e. shaking of the lower jaw and face rubbing were quantified. 40 µL of eugenol (50 mg/kg), WMTA (20 mg/0.2 mL) or ketoprofen were injected solely or in combination with formalin 2.5% and the behavioral responses were compared with those observed after formalin treatment alone. One-way ANOVA, Tukey were used for analysis of data. Formalin 2.5% provoked a biphasic nociceptive response, with an early and short lasting first tonic phase followed by a second phase. Solely SC injection of either WMTA or ketoprofen (a non steroidal anti-inflammatory drug) did not stimulate any significant nociceptive behaviour. However, injection of eugenol (a pain relieving agent) induced the early phase not the tonic phase of nociceptive response. WMTA, eugenol or ketoprofen injection 20 min before formalin injection attenuated the first phase but somehow prevented the induction of the second phase of nociceptive responses which were produced by formalin. Behavioural nociceptive responses including shaking of the lower jaw and face rubbing were significantly reduced when the subject was pretreated with either WMTA or ketoprofen (P<0.001). In this study, WMTA induced pain reduction by suppression of the formalininduced nociceptive response.