Withdrawal Latency Time Research Articles

A Buffered Local Anesthetic Without Epinephrine: Development, Characterization, and In Vivo Efficacy and Toxicity Analysis.

Lidocaine hydrochloride (HCl) 2% with 1:100,000 epinephrine (LW/E) is widely used to prevent pain during dental procedures and has been associated with injection sting, jittering effects, slow onset, and a bitter aftertaste. Since LW/E's introduction in 1948, no significant modifications have been proposed. This study aims to design and characterize an improved dental lidocaine HCl injectable formulation without epinephrine (LW/O/E) via buffers, sweeteners, and amino acids. LW/O/E injections were prepared with pH and osmolality values of 6.5-7.0 and 590-610 mOsm/kg. Using the electronic tongue (ETongue), the LW/O/E injectable formulations were characterized for viscosity, injectability, and taste analysis. The results were compared with the LW/E control. In vivo efficacy and anesthetic duration of the samples were measured through radiant heat tail-flick latency (RHTFL) and hot plate (HP) tests and local toxicity was assessed after a single intra-oral injection in Sprague Dawley rats (SDR). The viscosity and injectability values of the LW/O/E samples were found to be comparable to the LW/E injection. ETongue taste analysis showed an improvement in bitterness reduction of the LW/O/E samples compared to the LW/E formulation. Toxicity studies of samples in SDR showed minor and transient signs of erythema/eschar and edema. Anesthetic duration via RHTFL and HP paw withdrawal latency time in SDR were found to be comparable for the LW/O/E Sample 3A and the LW/E injection (p < 0.05). In conclusion, the buffered, higher osmolality and reduced bitterness developed LW/O/E formulation (Sample 3A) could be considered a promising alternative to the LW/E formulation for dental use.

Protective effect of chemically characterized extract of Viola stocksii Boiss. against breast cancer and vincristine induced neuropathic pain by alleviation of oxidative stress and inflammatory markers

Viola stocksii Boiss. is an important medicinal food plant. The powdered form of the species is mixed with butter and used as energy booster in subcontinent. The ethnomedicinal applications of the species include erectile dysfunction, inflammation and fever. In this study, the hydroethanolic extract of V. stocksii (HEEVS) was evaluated for its chemical composition, cytotoxicity, antineuropathic and antioxidant potential. The HPLC-PDA analysis revealed the presence of a total of 13 polyphenols, and 13 compounds were tentatively identified in the GC-MS profiling. The extract showed significant activity against the breast cancer cell lines (MDA-MB-23 and MCF-7) in MTT assay. The in vivo toxicity assessment revealed the biocompatibility of the extract with LD50 > 2000 mg/kg in mice. HEEVS significantly reduced the number of acetic acid induced abdominal writhings at 200 mg/kg. The extract reduced the vincristine induced neuropathic pain (VCINP) by increasing paw withdrawal threshold (g) and tail withdrawal latency time (s) significantly (p < 0.001) at 150 and 200 mg/kg. A marked (p < 0.001) alleviation observed in the levels of inflammatory and oxidative stress markers in sciatic nerve tissues of the animals. The enzyme inhibition and antioxidant assays unveiled the activity of HEEVS against cyclooxygenase COX-I and COX-II and free radical scavenging potential. The in-silico analysis revealed the potential interaction of the selected compounds with COX-I and COX-II. This report suggests that V. stocksii is a biocompatible source with abundant of polyphenols and encourages the use of the species as food and therapeutic alternative for addressing chemotherapy induced neuropathy.

Estrogen receptor β/substance P signaling in spinal cord mediates antinociceptive effect in a mouse model of discogenic low back pain.

Discogenic low back pain (DLBP) is the most commonly described form of back pain. Our previous studies indicated that estrogen-dependent DLBP mechanism was mediated by estrogen receptors (ERs) in the intervertebral disc (IVD) tissue, and the IVD degeneration degree is accompanied by downregulation of ERs, particularly ERβ. However, the neuropathological mechanisms underlying ERs modulation of DLBP are still not well understood. In this study, we investigated the antinociceptive effects of selective ERβ agonists on DLBP-related behavior by regulating substance P in spinal cord and dorsal root ganglia. Two weeks after ovariectomies, 18-week-old female mice were randomly separated into four groups: control group; DLBP sham surgery plus vehicle group; DLBP plus vehicle group; DLBP plus ERβ-specific agonist diarylpropionitrile (DPN) group. Behavioral data was collected including behavioral measures of axial back pain (grip force and tail suspension tests) and radiating hypersensitivity (mechanical sensitivity and cold sensitivity test). Dual label scanning confocal immunofluorescence microscopy was used to observe spatial colocalization of ERβ and substance P in spinal cord. Substance P changes in spinal cord and dorsal root ganglia were measured by immunohistochemistry and real-time PCR. ERβ activation could improve both axial and radiating behavioral disorders of DLBP. DPN facilitated the decrease of the amount of time in immobility 1 week after agonist administration. At the time point of 3 weeks, DPN group spent significantly less time in immobility than the vehicle group. In the grip strength tests, starting from postoperative week 1-week 3, DPN injection DLBP mice showed more resistance to stretch than the vehicle injection DLBP mice. Significant differences of cold withdrawal latency time were observed between the DLBP plus DPN injection and DLBP vehicle injection groups at 2- and 3-week injection time point. DPN significantly reversed the paw withdrawal threshold of DLBP mice at the time point of 1, 2, and 3 weeks. Substance P colocalized with ERβ in spinal dorsal horn, mainly in laminae I and II, a connection site of pain transmission. Substance P levels in dorsal horn and dorsal root ganglia of DLBP group were distinctly increased compared with that of control and DLBP sham group. DPN therapy could decrease substance P content in the dorsal horn and the dorsal root ganglia of DLBP mice compared with that of vehicle-treated DLBP mice. Activation of ERβ is antinociceptive in the DLBP model by controlling substance P in spinal cord and dorsal root ganglia, which might provide a therapeutic target to manage DLBP in the clinic.

Chemical profiling of Justicia vahlii Roth. (Acanthaceae) using UPLC-QTOF-MS and GC-MS analysis and evaluation of acute oral toxicity, antineuropathic and antioxidant activities

Ethnopharmacological relevanceJusticia vahlii Roth. (Acanthaceae), also called as kodasoori and bhekkar is an annual therophyte erect or decumbent herb used traditionally in toothache, skin diseases (itching, topical inflammation) and for the treatment of various respiratory disorders. Aim of the studyThe current study aimed at exploring pain cessation potential of J. vahlii Roth. via murine model of neuropathic pain and its phytochemical, toxicological and antioxidant profiles. Materials and methodsThe hydro-alcoholic extract of J. vahlii (HAEJv) prepared by maceration technique was subjected to preliminary phytochemical screening, total bioactive content determination, UPLC-QTOF-MS and GC-MS analysis. Toxicity assessment was carried out by using brine shrimp lethality assay and acute oral toxicity test. Murine model of neuropathic pain was applied to assess the antineuropathic potential of the species. Furthermore effect of the extract on catalase, superoxide oxide dismutase (SOD), Glutathione (GSH), interleukin-1beta (IL-1β) and total necrosis factor-alpha (TNF-α) was also studied. In vitro antioxidant profile was explored by using four methods; 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2-azinobis(3-ethylbenothiazoline)-6-sulfonic acid (ABTS), CUPric reducing antioxidant capacity (CUPRAC) and Ferric reducing antioxidant power (FRAP) assay. ResultsThe phytochemical screening revealed the presence of phenols, flavonoids, coumarins, alkaloids and lignans as the major classes of secondary metabolites. The extract was found rich in total phenolics content (TPC) and total flavonoids content (TFC) with identification of total 59 bioactives in UPLC-QTOF-MS and 40 compounds in GC-MS analysis. The extract was found nontoxic up to 4000 mg/kg (p.o.) in mice and no mortality observed in brine shrimp lethality assay. The HAEJv significantly reduced number of acetic acid induced abdominal constrictions at 100 mg/kg (p < 0.01) and 200 mg/kg (p < 0.001) and increased paw withdrawal threshold p < 0.05 at 100 mg/kg and p < 0.001 at 200 mg/kg, and an increase in tail withdrawal latency time p < 0.001 at 200 mg/kg was observed. The extract significantly increased levels of catalase, SOD and GSH while decreased IL-1β and TNF-α levels in sciatic nerve tissue of mice. HAEJv showed highest antioxidant activity through CUPRAC method 121.32 ± 1.22 mg trolox equivalent per gram of dry extract (mg TE/g DE) followed by DPPH 81.334 ± 4.35 mg TE/g DE, FRAP 69.89 ± 3.05 mg TE/g DE and ABTS 38.17 ± 2.12 mg TE/g DE. ConclusionThe current study back the traditional use of J. vahlii in pain cessation through antioxidant based antineuropathic pain activity and revealed the extract non-toxic with number of functional phytoconstituents and warrants further research on isolation of the compounds and sub-acute toxicity studies.

Preparation and Analysis of New 1,3,5-Trisubstituted-2-Pyrazolines Derivative for Their Analgesic Potential

The goal of the study was to develop, synthesise, and characterise a novel 1,3,5-trisubstituted-2-pyrazolines derivative, as well as to assess its analgesic potential. The reaction of chalcone derivatives with 4-hydrazinylbenzene sulfonamide hydrochloride and phenyl hydrazine hydrochloride yielded 1,3,5-tri-substituted-2-pyrazolines derivatives. The IR, 1HNMR, and mass spectrum analyses were used to characterise a total of sixteen substances. Analgesic activity of the proposed substances has been tested. The analgesic effect of the produced compounds was tested using two methods: the hot plate test technique and acetic acid induced writhing in mice. To compare the effectiveness, pentazocine and acetyl acetic acid were utilised as reference drugs. The hot plate test technique and acetic acid induced writhing in mice were used to assess the analgesic effect of the 16 produced chemical series A1-A8, and B1-B8. The evaluation's outcomes were viewed using Pentazocine and acetyl acetic acid as the standard drugs. In a 90-minute hot plate test, compounds A2 (10.30 s), A4 (9.45 s), A7 (11.65 s), and A8 (11.26 s) showed a delay in paw withdrawal latency time. Compounds B2 (9.10 s) and B7 (10.42 s) prolong the paw withdrawal latency time after 90 minutes in series B1-B8, reduce the pain feeling, and inhibit pain induced by heat methods. Compounds A2, A5, A6, A7, and A8 from Series A1-A8 showed 83.00, 76.01, 80.34, 86.99, 88.15 percent inhibition, substantially (p0.05 and p0.001, respectively), and decreased the number of wriths caused by 0.6 percent acetic acid at a dosage of 10 mg/kg. Acetylsalicylic acid (10 mg/kg) appears to be more successful in lowering the number of wriths, with a 99.0% reduction in the number of wriths (p0.001). B1, B3, and B4 have the least amount of active activity. These all finding suggest that these synthesized compounds have the potential as analgesic agent.

Exploitation of Thermal Sensitivity and Hyperalgesia in a Mouse Model of Dystonia.

Neuropathic pain is characterized by mechanical allodynia and thermal hyperalgesia to heat, and it affects some 20% of European population. Patients suffering from several neurologic diseases experience neuropathic pain, often finding no relief in therapy. Transgenic mice expressing the gene encoding the human mutant (hMT) or the human wild-type (hWT) torsin A represent a preclinical model of DYT1 dystonia which is the most common form of early-onset inherited dystonia. Baseline thermal sensitivity and hyperalgesia to heat have never been studied in models of dystonia. Therefore, the aim of this research has been to characterize thermal sensitivity in baseline conditions and hyperalgesia to heat after the induction of neuropathic pain through the spinal nerve ligation (SNL) model in mice overexpressing human wild-type and mutated torsin A in comparison to non-transgenic C57BL/6 mice. According to our results, the paw withdrawal latency time to heat in the Hargreaves’ test is significantly lower in the hMT mice (Kruskal–Wallis test = 6.933; p = 0.0312*; hMT vs. hWT p = 0.0317*). On the other hand, no significant differences in SNL-induced thermal hyperalgesia was found among the three strains (Friedman test = 4.933; p = 0.1019). Future studies are needed to better understand the role of torsin A in sensory processing of heat stimuli.

Antinociceptive effect of vapocoolant medium stream spray on hotplate latency in rat pups.

Heel sticks account for most blood tests performed in neonates without analgesia because topical local anesthetics are ineffective on heel glabrous skin. We investigated the antinociceptive effect of an alternative topical analgesic, a vapocoolant spray, on hind paw glabrous skin of rat pups. The spray was applied by two methods: method 1 for 4s at a distance of 8cm and method 2 for 10s at a distance of 18cm. The rat pups were randomized to either method 1 (n=32) or method 2 (n=31). Vapocoolant spray was applied to one hind paw randomly, and saline spray was applied to the contralateral paw. The paws were exposed to a hotplate test to measure withdrawal latency time before and 30s after the spray applications. Additionally, rat pups were tested for tissue toxicity in method 1 (n=20) and method 2 (n=20) after application of the vapocoolant spray before heel sticks three times a day for two consecutive days. Analyses of spray and method effects on hotplate withdrawal latency time were determined by nonparametric Wilcoxon tests to assess paired difference between vapocoolant spray and saline spray and to compare difference in medians between the two methods. Method 1 and method 2 vapocoolant spray applications significantly prolonged the withdrawal latency time compared with saline, a median difference of 0.6s (IQR 0.1-1.2) for method 1 and 9.5s (IQR 5.5-10.7) for method 2 (a 15-fold longer latency time with method 2). Method-2 produced significantly longer withdrawal latency time than method 1 with a difference in median time of 8.9s (CI: 95% 7.3-10.4s, P<.0001). No histopathological changes were detected. Compared with method- 1, the vapocoolant spray in method 2 produced significantly longer withdrawal latency time that is clinically applicable to collecting blood samples after a heel stick.

Mannitol Enhances the Antinociceptive Effects of Diphenhydramine as an Alternative Local Anesthetic.

Mannitol has recently been reported to be effective in enhancing the antinociceptive efficacy of lidocaine. No single study to date, however, has compared diphenhydramine with and without mannitol for nociceptive processing as an alternative local anesthetic. In this study, we examined the antinociceptive efficacy enhancements of diphenhydramine when combined with mannitol. Male Sprague-Dawley rats weighing 230–260 g were used in a hot plate test to evaluate the antinociceptive effects of diphenhydramine. All chemicals were dissolved in isotonic normal saline and administered subcutaneously into the plantar surface of the right hind paw at 10 min before the hot plate test. A subcutaneous injection of 0.5% or 1% diphenhydramine produced significant inhibition of the withdrawal latency time compared with the vehicle treatment. Antinociceptive effects appeared 10 min after the diphenhydramine injections and persisted for over 30 min. The antinociceptive effects of 1% diphenhydramine were not statistically different from those of 1% lidocaine. Although a subcutaneous injection of a 0.5 M mannitol solution alone did not affect the withdrawal latency time, 1% diphenhydramine with 0.5 M mannitol significantly enhanced antinociception. A subcutaneous injection of 1% diphenhydramine with epinephrine (1 : 100,000) solution did not increase the antinociceptive effect of the diphenhydramine. These results suggest that diphenhydramine with mannitol can be used as an alternative local anesthetic.

Anti-nociceptive activity of a few structurally related trimethoxy flavones and possible mechanisms involved.

The present study was designed to investigate the anti-nociceptive activity of a few structurally related trimethoxy flavones (7,2',3'-TMF, 7,2',4'-TMF, 7,3',4'-TMF and 7,5,4'-TMF) and the possible mechanisms involved. Anti-nociceptive activity was evaluated in mice by employing acetic acid-induced writhing, formalin-induced nociception and hot water tail immersion methods. The involvement of opioid, GABAergic, tryptaminergic, adrenergic and dopaminergic mechanisms and K+ATP channels in the anti-nociceptive activity of trimethoxy flavones was investigated using suitable interacting chemicals. Trimethoxy flavones exhibited a significant and dose-dependent inhibition of acetic acid writhing. The paw-licking response time was reduced both in the early and late phases of formalin nociception in a dose-dependent manner by trimethoxy flavones. A significant increase in tail withdrawal latency time was also observed after trimethoxy flavones treatment. These observations revealed the potential anti-nociceptive action of the investigated trimethoxy flavones. Pretreatment with naloxone and bicuculline significantly attenuated the reduction of abdominal constrictions produced by all the tested trimethoxy flavones indicating a definite role of opioid and GABAergic mechanisms in the anti-nociceptive effect of trimethoxy flavones. The anti-nociceptive action elicited by various trimethoxy flavones was differently modulated by glibenclamide, ondansetron, yohimbine and sulpiride. The investigated trimethoxy flavones exhibited promising anti-nociceptive activity in various nociceptive models, and multiple mechanisms are involved in the anti-nociceptive activity of these compounds.

Pathogenesis of spinal cord injury induced edema and neuropathic pain: expression of multiple isoforms of wnk1

Neuropathic pain (NP) is a common occurrence following spinal cord injury (SCI). Identification of specific molecular pathways that are involved in pain syndromes has become a major priority in current SCI research. We have investigated the role of a cation-dependent chloride transporter, Cl-regulatory protein Na(+)-K(+)-Cl(-) 1 (NKCC1), phosphorylation profile of NKCC1 and its specific involvement in neuropathic pain following contusion SCI (cSCI) using a rat model. Administration of the NKCC1 inhibitor bumetanide (BU) increases the mean hindpaw withdrawal latency time (WLT), thermal hyperalgesia (TH) following cSCI. These results demonstrate implication of NKCC1 co-transporter and BUin SCI-induced neuropathic pain. The with-no-lysine (K)-1 (WNK1) kinase has been shown to be an important regulator of NKCC1 phosphorylation in many systems, including nocioception. Mutations in a neuronal-specific exon of WNK1 (HSN2) was identified in patients that have hereditary sensory neuropathy type II (HSANII) also implicates WNK1 in nocioception, such that these patients have loss of perception to pain, touch and heat. In our ongoing research we proposed two studies utilizing our contusion SCI (cSCI) NP model of rat. Study 1 aimed at NKCC1 expression and activity is up-regulated following cSCI in the early edema and chronic neuropathic pain phases. Study 2 aimed at identifying the expression profile of alternatively spliced WNK1 isoforms in animals exhibiting thermal hyperalgesia (TH) following cSCI. Adult male Sprague Dawley rats (275-300 g) following laminectomy received cSCI at T9 with the NYU impactor-device II by dropping 10 g weight from the height of 12.5 mm. Control rats obtained laminectomy but no impaction. Following injury, functional recovery was assessed by BBB locomotor scores on day 1, 7, 14, 21, 35, and 42 and development of thermal hyperalgesia on day 21, 28, 35, and 42 day of injury by monitoring hind paw withdraw latency time (WLT) in seconds compared with the baseline data before injury. Increased NKCC1 may explain observed increase in magnetic resonance imaging (MRI) T2, exhibiting NKCC1 localization in neurons. This data supports NKCC1's role in the pathogenesis of acute and chronic phases of injury, namely spinal cord edema and chronic phase neuropathic pain. NKCC1 dependent chloride influx requires the phosphorylation at specific residues. Probing for the HSN2 exon of WNK1 reveals two key findings: i) the HSN2 exon is found in alternatively spliced neuronal isoforms found at 250 kDa and 230 kDa; ii) the 250 kDa isoform is found only in tissue that is injured. This data implicates the NKCC1/WNK1/WNK1HSN2 involvement in post-injury response that contributes to the development of neuropathic pain. Targeting this system may have therapeutic benefit.

Cannabinoid subtype-2 receptors modulate the antihyperalgesic effect of WIN 55,212-2 in rats with neuropathic spinal cord injury pain

Background context There is increasing evidence for a role of the cannabinoid (CB) system in the development of neuropathic pain (NP) after spinal cord injury (SCI). The nonspecific CB 1 and CB 2 receptor agonists, WIN 55, 212-2 (WIN), have previously been shown to alleviate both mechanical and thermal hyperalgesia (TH) after peripheral nerve injury. Purpose The present study was designed to identify the CB receptors involved in the antihyperalgesic effect of WIN by using selective antagonists for CB 1 and CB 2 receptors. Study design This is an in vivo and behavioral study using a moderate T9 contusion SCI. After injury, TH of the hind paws was measured on postinjury days 21 through 42. Methods Sprague-Dawley rats underwent a contusion SCI using the Multicenter Animal Spinal Cord Injury Study (MASCIS) weight-drop impactor, which induced a moderate T9 SCI. Only animals showing consistent plantar stepping and consistent forelimb and hind limb coordination (Basso, Beattie, and Bresnahan score=15) were tested for TH. Animals exhibiting decreased withdrawal latency time, indicating TH, on or before Day 42, were selected for pharmacological intervention. Animals not exhibiting TH did not receive pharmacological intervention and were sacrificed. Rats underwent hind paw testing before any drug administration (after injury), 45 minutes after selective CB antagonist (AM 251 or AM 630) administration (postantagonist) and again 45 minutes after WIN administration (post-WIN). There were a total of seven treatment groups: saline vehicle control; Dimethyl sulfoxide (DMSO) vehicle control; low-dose WIN (0.2 mg/kg); and high-dose WIN (2.0 mg/kg); AM 251 (3 mg/kg) and AM 630 (1 mg/kg) were given subcutaneously in a total volume of 0.5 mL. Followed by intraperitoneal injection of WIN after each antagonist, sham-operated rats repeated pharmacological intervention used with treatment Groups 5 and 6. Results Thermal hyperalgesia was significantly ameliorated in a dose-dependent manner with systemically administered WIN. Cannabinoid receptor Type 1 antagonist AM 251 pretreatment did not affect the antihyperalgesic effect of WIN. By contrast, pretreatment with the CB 2 receptor antagonist AM 630 significantly attenuated the effect of WIN. Conclusion Taken together, these results suggest a role of the CB 2 receptor in modulating SCI-induced TH. Selective activation of the CB 2 receptor could potentially lead to analgesic effects on NP while avoiding psychotropic side effects in patients with SCI.

The Role of Cation-Dependent Chloride Transporters in Neuropathic Pain Following Spinal Cord Injury

BackgroundAltered Cl- homeostasis and GABAergic function are associated with nociceptive input hypersensitivity. This study investigated the role of two major intracellular Cl- regulatory proteins, Na+-K+-Cl- cotransporter 1 (NKCC1) and K+-Cl- cotransporter 2 (KCC2), in neuropathic pain following spinal cord injury (SCI).ResultsSprague-Dawley rats underwent a contusive SCI at T9 using the MASCIS impactor. The rats developed hyperalgesia between days 21 and 42 post-SCI. Thermal hyperalgesia (TH) was determined by a decrease in hindpaw thermal withdrawal latency time (WLT) between days 21 and 42 post-SCI. Rats with TH were then treated with either vehicle (saline containing 0.25% NaOH) or NKCC1 inhibitor bumetanide (BU, 30 mg/kg, i.p.) in vehicle. TH was then re-measured at 1 h post-injection. Administration of BU significantly increased the mean WLT in rats (p < 0.05). The group administered with the vehicle alone showed no anti-hyperalgesic effects. Moreover, an increase in NKCC1 protein expression occurred in the lesion epicenter of the spinal cord during day 2–14 post-SCI and peaked on day 14 post-SCI (p < 0.05). Concurrently, a down-regulation of KCC2 protein was detected during day 2–14 post-SCI. The rats with TH exhibited a sustained loss of KCC2 protein during post-SCI days 21–42. No significant changes of these proteins were detected in the rostral region of the spinal cord.ConclusionTaken together, expression of NKCC1 and KCC2 proteins was differentially altered following SCI. The anti-hyperalgesic effect of NKCC1 inhibition suggests that normal or elevated NKCC1 function and loss of KCC2 function play a role in the development and maintenance of SCI-induced neuropathic pain.

The Analgesic Interaction between Ketorolac and Morphine in Radiant Thermal Stimulation Rat

Background: Previous studies have suggested synergistic analgesic drug interactions between NSAIDs and opioids in neuropathic and inflammatory pain models. The aim of this study was to investigate the analgesic drug interaction between intraperitoneal (IP) ketorolac and morphine in radiant thermal stimulation rat. Methods: Initially, we assessed the withdrawal latency time of the hindpaw to radiant thermal stimulation every 15 min for 1 hour and every 30 min for next 1 hour after IP normal saline 5 ml (control group). The latency time was changed into percent maximal possible effect (%MPE). Next, IP dose response curves were established for the %MPE of morphine (0.3, 1, 3, 10 mg/kg) and ketorolac (3, 10, 30 mg/kg) to obtain the for each agent. And we confirmed that the IP morphine effect was induced by opioid receptor through IP morphine followed by IP naloxone. At last, we injected three doses of IP ketorolac (3, 10, 30 mg/kg) mixed with one dose of morphine (2 mg/kg) for fixed dose analysis. Results: IP morphine delayed the paw withdrawal latency time dose dependently, but not ketorolac. of IP morphine was 2.1 mg/kg. And the IP morphine effect was reversed to control level by IP naloxone. IP ketorolac morphine combination showed no further additional effects on paw withdrawal latency time over morphine only group. Conclusions: IP ketorolac did not produce antinociceptive effect during radiant thermal stimulation. There was neither additional nor synergistic analgesic interaction between IP morphine and ketorolac in thermal stimulation rat.

Chronic pain and immunity: mononeuropathy alters immune responses in rats

In order to investigate the possible relationship between chronic pain and the immune system, delayed-type hypersensitivity (DTH) and humoral immunity were assessed in Sprague-Dawley rats subjected to unilateral peripheral mononeuropathy induced by sciatic ligation. Paw withdrawal latency (PWL) time was measured twice during the experiment in animals subjected to sciatic nerve ligation or sham surgery. Sciatic nerve-ligated animals showed hyperalgesia in the leg subjected to neural ligation when compared to the contralateral leg. No differences in PWL times existed in sham-operated animals. In order to exclude possible alterations in immune response due to the surgical procedure or to the hyperalgesia testing, a group of control animals, not subjected to surgical procedures or hyperalgesia testing, was also included in the experiment. Three days post-sciatic ligation or sham surgery, both experimental and control animals were sensitized to keyhole limpet hemocyanin (KLH). A secondary sensitization followed 1 week after the initial immunization. Fourteen days after the initial sensitization, KLH was injected into the hind foot pad and vehicle into the contralateral foot pad in order to assess DTH. One group of rats subjected to sciatic nerve ligation was tested for DTH in the hind foot pad ipsilateral to the ligated nerve, while another group was tested in the contralateral foot pad. Twenty-four hours following foot pad injections, the thickness of both paws was measured and animals were bled to test for anti-KLH immunoglobulins. Animals in which mononeuropathy was induced, but not sham-operated or control animals, exhibited an enhanced DTH response to KLH. This enhanced DTH response occurred both ipsilateral and contralateral to the ligated nerve. This increased response was blocked in both cases by the local anesthetic bupivicaine. Two sham-surgery groups and a normal control group were tested similarly. Gamma-immunoglobulin levels against KLH were significantly reduced in the hyperalgesic animals when compared to control animals but were similar when compared to sham-operated animals. This study suggests that chronic nociception causes significant alterations in immune function and strengthens the hypothesis that chronic pain can influence the immune system.

The mouse paw withdrawal assay: A method for determining the effect of calcitonin gene-related peptide on cutaneous heat nociceptive latency time

Previously we have shown that calcitonin gene-related peptide (CGRP) modulates nociception and the effect of opioid analgesics in the central nervous system of mice. Cutaneous primary afferent nerve terminals also contain a high concentration of CGRP, however the lack of a suitable method for assessing cutaneous nociceptive latency changes in the hindpaw skin of the mouse hindered our investigations. We report here on the development of an assay to investigate the effect of CGRP on nociception in the dorsal hindpaw skin. Subcutaneous injection of CGRP produced a modest elevation of withdrawal latency time at doses that were two orders of magnitude greater than the physiologic levels determined in naive animals by radioimmunoassay. This elevation of threshold was minimal when compared to the elevation produced by mepivacaine. These results indicate that subcutaneous injection of CGRP into the dorsal hindpaw skin of the mouse produces a modest increase in paw withdrawal latency times at high, non-physiologic doses.