Thermal Nociceptive Tests Research Articles

Selective knockdown of NMDA receptors in primary afferent neurons decreases pain during phase 2 of the formalin test

The role of NMDA receptors (NMDARs) expressed by primary afferent neurons in nociception remains controversial. The aim of this study was to develop mice with a tissue selective knockdown of NMDARs in these neurons and to evaluate their behavioral responses to different types of painful stimuli. Mice with floxed NMDAR NR1 subunit gene (fNR1) were crossed with mice expressing Cre recombinase under the control of the peripherin promotor (Prph-Cre). Male Prph-Cre+ floxed NR1 mice were compared to Cre− littermates. Both quantitative RT/PCR and Western blotting indicated a ∼75% reduction in NR1 expression in dorsal root ganglia (DRG) extracts with no effect on NR1 expression in spinal cord, brain or the enteric nervous system. Immunocytochemistry with antibodies to NR1 revealed decreased staining in all size classes of DRG neurons. NMDA produced a detectable increase in [Ca 2+] i in 60% of DRG neurons cultured from Cre− mice, but only 15% of those from Cre+ mice. Furthermore, the peak [Ca 2+] i responses were 64% lower in neurons from Cre+ mice. There was no significant difference between Cre+ and Cre− mice in response latencies to the hotplate or tail withdrawal tests of thermal nociception, nor was there a difference in withdrawal thresholds to mechanical stimuli of the tail or paw. However, compared to Cre− littermates, Cre+ knockdown mice had a 50% decrease in the phase 2 response to formalin injection ( P<0.001). There was no effect on phase 1 responses. These results suggest that NMDA receptors expressed by primary afferent nerves play an important role in the development of sensitized pain states.

H3 Receptors and Pain Modulation: Peripheral, Spinal, and Brain Interactions

Histamine H(3) receptors (H(3)Rs), distributed within the brain, the spinal cord, and on specific types of primary sensory neurons, can modulate pain transmission by several mechanisms. In the skin, H(3)Rs are found on certain Aβ fibers, and on keratinocytes and Merkel cells, as well as on deep dermal, peptidergic Aδ fibers terminating on deep dermal blood vessels. Activation of H(3)Rs on the latter in the skin, heart, lung, and dura mater reduces calcitonin gene-related peptide and substance P release, leading to anti-inflammatory (but not antinociceptive) actions. However, activation of H(3)Rs on the spinal terminals of these sensory fibers reduces nociceptive responding to low-intensity mechanical stimuli and inflammatory stimuli such as formalin. These findings suggest that H(3)R agonists might be useful analgesics, but these drugs have not been tested in clinically relevant pain models. Paradoxically, H(3) antagonists/inverse agonists have also been reported to attenuate several types of pain responses, including phase II responses to formalin. In the periaqueductal gray (an important pain regulatory center), the H(3) inverse agonist thioperamide releases neuronal histamine and mimics histamine's biphasic modulatory effects in thermal nociceptive tests. Newer H(3) inverse agonists with potent, selective, and brain-penetrating properties show efficacy in several neuropathic and arthritis pain models, but the sites and mechanisms for these actions remain poorly understood.

Combination of dexmedetomidine with buprenorphine enhances the antinociceptive effect to a thermal stimulus in the cat compared with either agent alone

ObjectiveTo evaluate the sedative and antinociceptive effects of combinations of dexmedetomidine and buprenorphine in cats. Study designExperimental randomized study. AnimalsTwelve purpose-bred neutered domestic short-hair cats (4 male and 8 female) weighing 4.6 kg (range 3.7–5.5 kg) aged from 2 to 5 years. MethodsSix cats per group were administered buprenorphine (B) at 10 (B10) or 20 μg kg−1 (B20) or dexmedetomidine (D) at 20 (D20) or 40 μg kg−1 (D40) or a combination of B10/D20. A feline thermal nociceptive threshold testing device was used to evaluate the antinociceptive effects of the drugs before and up to 24 hours after drug treatment. Sedation was scored using a 100 mm visual analogue scale (VAS). ResultsThermal thresholds increased significantly after administration of all but D20. Area under the curve (AUC, hours °C) for the first 6 hours (mean ± SD) for B20 (281 ± 17.8) was significantly greater than B10 (260 ± 11.4), D20 (250 ± 7.9) and D40 (255 ± 11.4). The AUC for B10/D20 (273 ± 12.2) was significantly greater than D20 but not the other treatments. No sedation was seen after administration of B10 or B20 and maximal sedation was seen for all animals in the D40 and B10/D20 groups and most animals in the D20 group. ConclusionsD20 alone had the smallest analgesic effect; B10 alone provided no sedation but their combination gave good sedation with analgesia comparable with B20. Clinical relevanceThis combination could be a useful multimodal sedative/analgesic regimen in cats.

Involvement of the nitric oxide/ CGMP/K ATP pathway in antinociception induced by exercise in rats

Aims Physical exercise is responsible for increasing the nociceptive threshold. The present study aimed to investigate the involvement of the nitric oxide/ CGMP/K ATP pathway in antinociception induced by acute aerobic exercise (AAc) in rats. Main methods Wistar rats performed exercise in a rodent treadmill, according to an AAc protocol. The nociceptive threshold was measured by mechanical and thermal nociceptive tests (paw-withdrawal, tail-flick and face-flick). To investigate the involvement of the NO/ CGMP/K ATP pathway the following nitric oxide synthase (NOS) unspecific and specific inhibitors were used: N-nitro- l-arginine (NOArg), Aminoguanidine, N 5-(1-Iminoethyl)- l-ornithine dihydrocloride (L-NIO), N ω-Propyl- l-arginine (L-NPA); guanylyl cyclase inhibitor, 1H-[1,2,4]oxidiazolo[4,3-a]quinoxalin-1-one (ODQ); and K ATP channel blocker, Glybenclamide; all administered subcutaneously at a dose of 2 mg/kg 10 min before exercise started. Plasma and cerebrospinal fluid (CSF) nitrite levels were determined by spectrophotometry. Key findings In the paw-withdrawal, tail-flick and face-flick tests, the AAc protocol produced antinociception, which lasted for more than 15 min. This effect was significantly reversed ( P < 0.05) by NOS specific and unspecific inhibitors, guanylyl cyclase inhibitor (ODQ) and K ATP channel blocker (Glybenclamide). Acute exercise was also responsible for increasing nitrite levels in both plasma and cerebrospinal fluid. Significance Taken together, these results suggest that the NO/ CGMP/K ATP pathway participates in antinociception induced by exercise.

Protein Kinase C Mediates the Synergistic Interaction Between Agonists Acting at α2-Adrenergic and Delta-Opioid Receptors in Spinal Cord

Coactivation of spinal alpha(2)-adrenergic receptors (ARs) and opioid receptors produces antinociceptive synergy. Antinociceptive synergy between intrathecally administered alpha(2)AR and opioid agonists is well documented, but the mechanism underlying this synergy remains unclear. The delta-opioid receptor (DOP) and the alpha(2A)ARs are coexpressed on the terminals of primary afferent fibers in the spinal cord where they may mediate this phenomenon. We evaluated the ability of the DOP-selective agonist deltorphin II (DELT), the alpha(2)AR agonist clonidine (CLON) or their combination to inhibit calcitonin gene-related peptide (CGRP) release from spinal cord slices. We then examined the possible underlying signaling mechanisms involved through coadministration of inhibitors of phospholipase C (PLC), protein kinase C (PKC) or protein kinase A (PKA). Potassium-evoked depolarization of spinal cord slices caused concentration-dependent release of CGRP. Coadministration of DELT and CLON inhibited the release of CGRP in a synergistic manner as confirmed statistically by isobolograpic analysis. Synergy was dependent on the activation of PLC and PKC, but not PKA, whereas the effect of agonist administration alone was only dependent on PLC. The importance of these findings was confirmed in vivo, using a thermal nociceptive test, demonstrating the PKC dependence of CLON-DELT antinociceptive synergy in mice. That inhibition of CGRP release by the combination was maintained in the presence of tetrodotoxin in spinal cord slices suggests that synergy does not rely on interneuronal signaling and may occur within single subcellular compartments. The present study reveals a novel signaling pathway underlying the synergistic analgesic interaction between DOP and alpha(2)AR agonists in the spinal cord.

Functional Interaction Between TRPV1 and μ-Opioid Receptors in the Descending Antinociceptive Pathway Activates Glutamate Transmission and Induces Analgesia

The transient receptor potential vanilloid-1 (TRPV1) receptor is involved in peripheral and spinal nociceptive processing and is a therapeutic target for pain. We have shown previously that TRPV1 in the ventrolateral periaqueductal gray (VL-PAG) tonically contributes to brain stem descending antinociception by stimulating glutamate release into the rostral ventromedial medulla and off neuron activity. Because both opioid and vanilloid systems integrate and transduce pain sensation in these pathways, we studied the potential interaction between TRPV1 and mu-opioid receptors in the VL-PAG-rostral ventromedial medulla (RVM) system. We found that the TRPV1 agonist, capsaicin, and the mu-receptor agonist [D-Ala(2),N-Me-Phe(4),Gly(5)-ol]enkephalin, when coadministered into the ventrolateral-PAG at doses nonanalgesic per se, produce 1) antinociception in tests of thermal nociception; 2) stimulation of glutamate release into the RVM; and 3) inhibition of on neuron activity in the RVM. These effects were all antagonized by the TRPV1 and opioid receptor antagonists 5'-iodo-resiniferatoxin and naloxone, respectively, thus suggesting the existence of a TRPV1-mu-opioid interaction in the VL-PAG-RVM system. By using double immunofluorescence techniques, we found that TRPV1 and mu-opioid receptors are coexpressed in several neurons of the VL-PAG. These findings suggest that mu-receptor activation not only acts on inhibitory neurons to disinhibit PAG output neurons but also interacts with TRPV1 activation at increasing glutamate release into the RVM, possibly by acting directly on PAG output neurons projecting to the RVM.

Allopurinol for pain relief: more than just crystal clearance?

Gout and pain are synonymous, and a study in this issue of the BJP reports a novel anti-nociceptive effect of allopurinol, the drug most commonly used to treat gout. Allopurinol works by inhibiting xanthine oxidase (XO), the enzyme responsible for converting hypoxanthine to uric acid which is deposited as crystals in the joints of gout sufferers. Hypoxanthine is a metabolite of, and a possible precursor to, adenosine. Schmidt et al., find that acute inhibition of XO with allopurinol produces a modest adenosine A(1) receptor-mediated anti-nociceptive effect in common tests of chemical and thermal nociception in mice. A concomitant increase in cerebrospinal fluid levels of adenosine supports their hypothesis that inhibiting XO increases adenosine levels via salvage from hypoxanthine. Elevating endogenous adenosine levels by inhibiting metabolism is a well-established strategy for producing anti-nociception in many preclinical models, but inhibiting XO is likely to be particularly beneficial in some chronic pain states because of the pro-nociceptive reactive oxygen species that are produced by XO activity. Thus, allopurinol may have unexpected benefits in pain associated with chronic inflammation, diabetes and vascular dysfunction.

Behavioral characterization of dysbindin-1 deficient sandy mice

Dysbindin-1 (dystrobrevin binding protein-1) has been reported as a candidate gene associated with schizophrenia. Dysbindin-1 mRNA and protein levels are significantly reduced in the prefrontal cortex and hippocampus of schizophrenia subjects. To understand the in-vivo functions of dysbindin-1, we studied schizophrenia relevant behaviors in adult male Sandy homozygous (sdy/sdy) and heterozygous (sdy/+) mice that have a natural mutation in dysbindin-1 gene (on a DBA/2J background) resulting in loss of protein expression. Spontaneous locomotor activity of sdy/sdy and sdy/+ mice in novel environment was not significantly different from DBA/2J controls. However, on repeated testing in the same environment for 7 days, sdy/sdy mice, in contrast to DBA/2J controls showed a lack of locomotor habituation. Locomotor activating effect of a low dose of d-amphetamine (2.5 mg/kg ip), a behavioral measure of mesolimbic dopamine activity, was significantly reduced in the mutant mice. Interestingly, sdy/sdy mice showed enhanced locomotor sensitization to repeated five daily injection of amphetamine. Possible cognitive impairment in Sandy mutants was revealed in novel object recognition test as sdy/sdy and sdy/+ mice spent significantly less time exploring novel objects compared to DBA/2J. Sdy/sdy mice also showed deficits in emotionally motivated learning and memory showing greater freezing response to auditory conditioned stimulus (CS) in fear conditioning paradigm. In thermal nociceptive test, the latency of paw withdrawal in sdy/sdy and sdy/+ animals was significantly higher compared to DBA/2J indicating hypoalgesia in the mutants. Taken together, these data suggest that dysbindin-1 gene deficiency leads to significant changes in cognition and altered responses to psychostimulants.

The comparison of effects of processed Aconiti tuber, U50488H and MK-801 on the antinociceptive tolerance to morphine

In the previous studies, we demonstrated that an oriental herbal medicine, processed Aconiti tuber (PAT), at subanalgesic doses could inhibit or reverse the antinociceptive tolerance to morphine. In the present study, we compared the effect of PAT, trans-(±)-3,4-dichloro- N-methyl- N-(2-(1-pyrrolidin)cyclohexyl)-benzeneacetamide methane sulfonate hydrate (U50488H), a selective kappa opioid receptor (KOR) agonist, and (−)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine-maleate (MK-801), a N-methyl- d-aspartate (NMDA) receptor antagonist, on the antinociceptive tolerance to morphine in the same experimental condition. Mice received subcutaneous morphine (10 mg/kg), and oral PAT at a subanalgesic dose (0.3 g/kg for mechanical or 1.0 g/kg for thermal test), or intraperitoneal U50488H at a subanalgesic dose (3 mg/kg), or MK-801 at a subanalgesic dose (0.1 mg/kg) once daily for 14 days. The mechanical nociceptive threshold was measured before, and at 60 min by tail pressure testing, and thermal nociceptive latency was measured before, and at 30 min by hot plate testing, after daily morphine injections. PAT and U50488H could not only inhibit the development of morphine tolerance but also reverse the already-developed morphine tolerance, while MK-801 could only inhibit the development of morphine tolerance but not reverse the already-developed morphine tolerance, in both mechanical and thermal nociceptive tests. These data suggested that PAT, an indirect-acting KOR agonist, share the common pharmacological property of KOR agonists on morphine tolerance, and that PAT may be superior to some NMDA receptor antagonists which do not reverse already-developed morphine tolerance.

Decreased spinal cord opioid receptor mRNA expression and antinociception in a Theiler's murine encephalomyelitis virus model of multiple sclerosis

Multiple sclerosis patients typically experience increased pain that is relatively insensitive to opiate treatment. The mechanistic basis for this increased nociception is currently poorly understood. In the present study, we utilized the Theiler's murine encephalomyelitis virus (TMEV) model of MS to examine possible changes in spinal cord opioid receptor mRNA over the course of disease progression. TMEV infection led to significantly decreased mu, delta and kappa opioid receptor mRNA expression as analyzed by quantitative real-time PCR in both male and female mice at days 90, 150 and 180 post-infection (PI). Since opioid receptor mRNA expression decreased in TMEV mice, we examined whether opiate analgesia is also altered. TMEV infected female mice had significantly decreased opiate analgesia in thermal nociceptive tests beginning at day 90 PI, while TMEV-infected male mice did not display significantly decreased opiate analgesia until day 120 PI. The novel finding that opioid receptor expression is significantly decreased in the spinal cord of TMEV mice could explain the increased nociception and loss of opiate analgesia observed in both TMEV mice and multiple sclerosis patients.

Antinociceptive Synergistic Interaction Between Clonidine and Ouabain on Thermal Nociceptive Tests in the Rat

The antinociceptive effect produced by spinal injection of clonidine (an alpha(2)-adrenergic agonist) is mediated by a cholinergic mechanism. We aimed in the current study to evaluate the antinociceptive interaction between intrathecally administered ouabain, an inhibitor of Na(+), K(+)-ATPase, and clonidine. We used rats chronically implanted with lumbar intrathecal catheters to examine the ability of intrathecal clonidine and ouabain and the mixtures of clonidine-ouabain to alter tail-flick latency. To characterize the interaction, isobolographic analysis was performed. Intrathecal clonidine (0.5-10 microg) and ouabain (0.1-5 microg) produced significant dose- and time-dependent antinociception in the tail-flick tests. The median effective dose (ED(50)) values for intrathecally administered ouabain and clonidine were 2.3 microg and 4.7 microg, respectively. The experimental point for the ouabain-clonidine combination decreased significantly (P < .05) below the lines of additivity. Isobolographic analysis exhibited a synergistic interaction after the coadministration of ouabain and clonidine. No motor impairment was observed in the animals after intrathecal administration of the combination of ouabain and clonidine or clonidine alone. Intrathecal pretreatment with atropine but not yohimbine blocked the antinociceptive effect of ouabain and attenuated its interaction with spinal clonidine. These results suggest that the synergistic interaction of ouabain and clonidine were probably mediated, at least in part, via an enhancement of cholinergic transmission in the spinal nociceptive processing system. Although intrathecal clonidine produces pronounced analgesia, antinociceptive doses of intrathecal clonidine produce several side effects, including hypotension, bradycardia, and sedation. This article presents antinociceptive synergistic interaction between clonidine and ouabain on thermal nociceptive tests in the rat.

Augmentation of spinal morphine analgesia and inhibition of tolerance by low doses of μ‐ and δ‐opioid receptor antagonists

Ultralow doses of naltrexone, a non-selective opioid antagonist, have previously been found to augment acute morphine analgesia and block the development of tolerance to this effect. Since morphine tolerance is dependent on the activity of micro and delta receptors, the present study investigated the effects of ultralow doses of antagonists selective for these receptor types on morphine analgesia and tolerance in tests of thermal and mechanical nociception. Effects of intrathecal administration of mu-receptor antagonists, CTOP (0.01 ng) or CTAP (0.001 ng), or a delta-receptor antagonist, naltrindole (0.01 ng), on spinal morphine analgesia and tolerance were evaluated using the tail-flick and paw-pressure tests in rats. Both micro and delta antagonists augmented analgesia produced by a sub-maximal (5 microg) or maximal (15 microg) dose of morphine. Administration of the antagonists with morphine (15 microg) for 5 days inhibited the progressive decline of analgesia and prevented the loss of morphine potency. In animals exhibiting tolerance to morphine, administration of the antagonists with morphine produced a recovery of the analgesic response and restored morphine potency. Combining ultralow doses of micro- or delta-receptor antagonists with spinal morphine augmented the acute analgesic effects, inhibited the induction of chronic tolerance and reversed established tolerance. The remarkably similar effects of micro- and delta-opioid receptor antagonists on morphine analgesia and tolerance are interpreted in terms of blockade of the latent excitatory effects of the agonist that limit expression of its full activity.

(666): Anabolic-androgenic steroid effects on pain and opioid analgesia

It has been suggested that anabolic-androgenic steroids such as testosterone promote body building in part because they reduce pain sensitivity. The present study was designed to determine whether male rats’ nociceptive and opioid antinociceptive sensitivity changed after chronic exposure to high-dose testosterone. Adult, gonadally intact Sprague-Dawley male rats were given once-daily injections of testosterone propionate (TP, 5 mg/kg s.c.) or safflower oil vehicle for 28 days. Beginning 3 hr after the vehicle or TP injection on day 28, latency to respond on two tests of acute thermal nociception (hotplate and tail withdrawal) and a test of acute mechanical nociception (paw pressure) was recorded, or behavior on a test of inflammatory nociception (formalin test) was recorded; in the latter group of rats, the development of thermal hyperalgesia and mechanical allodynia was also evaluated periodically for a month after formalin injection, during which daily vehicle or TP injections were continued.

Activation of α 2-adrenoceptors in the trigeminal region produces sex-specific modulation of nociception in the rat

Sex-related differences in the sensitivity to pain and in the response to analgesics have been reported including higher perceptual responses to experimentally induced pain and the higher prevalence of many pain syndromes in women compared with men. This study examines whether α 2-adrenoceptor-mediated antinociceptive effects are reduced by estrogen which could account for the sex-related differences in pain perception and modulation. Clonidine, an α 2-adrenoceptor agonist, has been shown to inhibit noxious stimulus-evoked nociceptive behavior as well as the responses of nociceptive neurons in the medullary dorsal horn. Intracisternal microinjection of clonidine (7 μg/5 μl) through the implanted PE-10 cannulae dorsal to the trigeminal region in male, ovariectomized (OVX), and diestrous (DiE) Sprague–Dawley rats produced a strong antinociceptive effect on N-methyl- d-aspartic acid (NMDA)-induced nociceptive scratching behavior and heat-induced face withdrawal nociceptive tests. However, it failed to produce any inhibition in the estradiol-treated ovariectomized (OVX+E) group regardless of the dose of estradiol (1, 10 or 100 μg/100 μl sesame oil) or in the proestrous (ProE) group. Further, clonidine produced dose-dependent effects in male and OVX groups but not in the OVX+E group on the NMDA-induced nociceptive behavior. Finally, the effect of clonidine was reversed by yohimbine, an α 2-adrenoceptor antagonist, in male and OVX groups on thermal nociceptive test. These results lead us to conclude that activation of α 2-adrenoceptors produces sex-specific, estrogen dependent modulation of nociception in the trigeminal region of the rat. A decreased α 2-adrenoceptor-mediated inhibition could be one of the factors responsible for the higher prevalence of pain syndromes in females.

Morphine Antinociceptive Potency on Chemical, Mechanical, and Thermal Nociceptive Tests in the Rat

Many tests are used to assess nociception in laboratory animals. The objective of this study was to compare morphine potency across tests. Rats were injected with saline or morphine (1-20 mg/kg SC), and nociception was assessed 15-20 min later. A consistent definition of antinociception—a change in response greater than 4 times the standard deviation above the mean for the saline-treated controls—was used to compare morphine potency on different tests. These data revealed 4 things. 1) Morphine potency was greatest on the paw pressure, hot plate, and tail withdrawal tests and lowest on the formalin test. 2) Stimulus intensity had no effect on morphine potency on the hot plate (ED 50 = 4.5, 2.8, and 2.6 mg/kg for 49°C, 52°C, and 55°C tests, respectively) or tail withdrawal tests (ED 50 = 2.9 and 2.6 for 48°C and 52°C water, respectively). 3) Assessment of morphine potency using a within-subjects cumulative dosing procedure resulted in lower ED 50 values compared to data collected using a between-subjects design (hot plate: 2.6 vs 4.9; tail withdrawal: 2.6-2.9 vs 5.7 mg/kg). 4) Adjusting the cutoff value from 4 to 5, 6, 7, and 8 standard deviations greater than the mean resulted in a progressive increase in ED 50 values. These data demonstrate that morphine potency is dependent, in part, on the nociceptive test even when all other factors (eg, species, strain, age, gender, and cutoff value) are held constant. Perspective The ability of morphine to block nociception is influenced by many factors. The present study shows that the test used to assess nociception, but not the stimulus intensity, can have a dramatic effect on morphine potency. This finding shows that morphine potency varies depending on how pain is assessed.

Facilitation of spinal morphine analgesia in normal and morphine tolerant animals by neuropeptide SF and related peptides

Neuropeptide FF and related synthetic amidated peptides have been shown to elicit sustained anti-nociceptive responses and potently augment spinal anti-nociceptive actions of spinal morphine in tests of thermal and mechanical nociception. Recent studies have described the occurrence of another octapeptide, neuropeptide SF (NPSF) in the spinal cord and the cerebrospinal fluid and demonstrated its affinity for the NPFF receptors. This study examined the effects of NPSF and two putative precursor peptides, EFW-NPSF and NPAF, on the spinal actions of morphine in normal and opioid tolerant rats using the tailflick and pawpressure tests. In normal rats, NPSF demonstrated weak intrinsic activity but sub-effective doses of the peptide significantly increased the magnitude and duration of spinal morphine anti-nociception in both tests. A low-dose of NPSF also augmented the spinal actions of a delta receptor agonist, deltorphin. The morphine-potentiating effect of NPSF was shared by EFW-NPSF and the octadecapeptide NPAF. In animal rendered tolerant by continuous intrathecal infusion of morphine for 6 days, low dose NPSF itself elicited a significant anti-nociceptive response and potently increased morphine-induced response in both tests. In animals made tolerant by repeated injections of intrathecal morphine, administration of NPSF, EFW-NPSF, and NPAF with morphine reversed the loss of the anti-nociceptive effect and restored the agonist potency. The results demonstrate that in normal animals NPSF and related peptides exert strong potentiating effect on morphine anti-nociception at the spinal level and in tolerant animals these agents can reverse the loss of morphine potency.

Inflammation-Susceptible Lewis Rats Show Less Sensitivity Than Resistant Fischer Rats in the Formalin Inflammatory Pain Test and With Repeated Thermal Testing

Comparisons between Lewis and Fischer inbred strains of rats are used frequently to study the effect of inherent differences in function of the hypothalamic-pituitary-adrenal axis on pain-relevant traits, including differential susceptibility to chronic inflammatory disease and differential responsiveness to analgesic drugs. Increasing use of genetic models including transgenic knockout mice and inbred strains of rodents has raised our awareness of, and the importance of, thorough characterization (or phenotyping) of the strains of rodents being compared. Furthermore, genetic variability in analgesic sensitivity is correlated with, and may be caused by, genetically determined baseline sensitivity. Thus in this study, baseline inflammatory and thermal nociceptive sensitivities were measured in awake male and female Lewis and Fischer rats to examine whether the results could explain relevant strain differences reported in the literature. The effect of maternal separation was also examined and no effect was found on nociceptive sensitivity, corticosterone responses, or the development of adjuvant-induced arthritis, a model of rheumatoid arthritis. Lewis rats and female rats were more sensitive to thermal nociception in the tail withdrawal test (mean of 3 trials) than Fischer rats and male rats, respectively. Unexpectedly, the more inflammation-susceptible Lewis rats were less sensitive in the formalin inflammatory nociception test, and showed a significant decrease in sensitivity with repeated thermal nociceptive testing, whereas Fischer rats did not. These results affect the interpretation of previously observed results. Further study of the underlying mechanisms and the relevance to differential susceptibility to chronic inflammation is warranted.

Anti-hyperalgesic effects of intrathecally administered neuropeptide W-23, and neuropeptide B, in tests of inflammatory pain in rats

Neuropeptide W-23 (NPW23) is an endogenous ligand of both GPR7 and GPR8, and neuropeptide B (NPB) is an endogenous ligand of GPR7. GPR7 mRNA has been detected in regions of the cortex, the hippocampus, the hypothalamus, and the spinal cord in the rat, but GPR8 has not been found in rodents. GPR7 and GPR8 receptors have structural features in common with both opioid and somatostatin receptors. The effects of intrathecal (i.t.) application of NPW23 and NPB were tested in two inflammatory pain models (plantar injection of formalin or carrageenan) and two thermal nociceptive tests (52.5 °C and 50.5 °C hot plates) and one mechanical nociceptive test in the rat. I.t. injection of either NPW23 or NPB decreased the number of agitation behaviors induced by paw formalin injection and attenuated the level of mechanical allodynia, but not the level of thermal hyperalgesia, induced by paw carrageenan injection in a dose-dependent manner at a dose between 0.1 and 10 μg, significantly. The effects of either 10 μg of NPW23 or 10 μg of NPB were not antagonized by 10 μg of naloxone. I.t. injection of either NPW23 or NPB had no effect in both the 52.5 °C hot plate test or in the 50.5 °C hot plate tests at a dose between 1 and 100 μg. I.t. injection of either 10 μg of NPW23 or 10 μg of NPB had no effect in the mechanical nociceptive test. I.t. injection of either 10 μg of NPW23 or 10 μg of NPB significantly suppressed the expression of Fos-like immunoreactivity of the L4–5 spinal dorsal horn induced by paw formalin injection. These data suggest that both spinally-applied NPW23 and NPB suppressed the input of nociceptive information to the spinal dorsal horn, produced an analgesic effect in the formalin test, and attenuated the level of mechanical allodynia in the carrageenan test, and that either spinally applied NPW23 or spinally applied NPB had no effect in the physiological condition.

Absence of diabetic hyperalgesia in bradykinin B 1 receptor-knockout mice

Experimental evidence has shown that the inducible bradykinin (BK) B 1 receptor (BKB 1-R) subtype is involved in the development of hyperalgesia associated with type 1 diabetes. Selective BKB 1-R antagonists inhibited, whereas selective agonists increased the hyperalgesic activity in diabetic mice in thermal nociceptive tests. Here we evaluate the development of diabetic hyperalgesia in a BKB 1-R-knockout (KO) genotype compared to wild-type (WT) mice. The BKB 1-R-KO mice were backcrossed for 10 generations to C57BL/6 genetic background before use in the experiments. Diabetes was induced by streptozotocin (STZ) and thermal nociception was assessed by the hot plate and tail immersion tests. The hyperalgesia observed in wild type mice was totally absent in the BKB 1-R-KO mice. Furthermore, the selective BKB 1-R agonist, desArg 9BK, significantly increased the hyperalgesic activity in diabetic WT mice but had no effect on nociceptive responses in diabetic BKB 1-R-KO mice. Taken together, the results confirm the crucial role of the BKB 1-R, upregulated alongside inflammatory diabetes, in the development of diabetes-induced hyperalgesia.

Pathways for the bradykinin B1 receptor-mediated diabetic hyperalgesia in mice

Experimental evidence has shown that the bradykinin B1 receptor (BKB1-R) is involved in the development of hyperalgesia associated with diabetes since specific BKB1-R antagonists significantly inhibited the hyperalgesic activity observed in streptozotocin (STZ)-mice in thermal nociceptive tests. The involvement of the nitric oxide (NO), the substance P (SP) and the calcitonin gene-related peptide (CGRP) pathways in mediating BKB1-R-induced hyperalgesia was evaluated. Diabetes was induced in male CD-1 mice by injecting STZ (200 mg/kg; i.p.). Nociception was assessed using the hot plate and tail immersion tests, one week following the injection of STZ. The nitric oxide synthase (NOS) inhibitors (L-NNA, 20 mg/kg; L-NMMA, 30 mg/kg and AGUA, 50 mg/kg; i.p.), the SP antagonists (sendide and L-732,138, 100 microg/kg; i.v.) and the CGRP antagonist (hCGRP8-37, 100 microg/kg; i.v.) significantly attenuated the hyperalgesic activity and also reversed the potentiating effect of the BKB1- R agonist, DBK on diabetic hyperalgesia in STZ-mice. These results support the involvement of BKB1-R in the development of diabetic hyperalgesia in STZ-mice through activation of the NO, SP and CGRP pathways.