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Background: Migraine research has highlighted the pivotal role of nitric oxide (NO) in migraine pathophysiology. Nitric oxide donors such as glyceryl trinitrate (GTN) induce migraine attacks in humans, whereas spontaneous migraine attacks can be aborted by inhibiting NO production. The present study aimed to investigate how GTN triggers migraine through its three nitric oxide synthase (NOS) isoforms (neuronal NOS (nNOS), endothelial NOS (eNOS) and inducible NOS (iNOS)) via a suspected feed-forward phenomenon. Methods: Migraine-relevant hypersensitivity was induced by repeated injection of GTN in an in vivo mouse model. Cutaneous tactile sensitivity was assessed using von Frey filaments. Signaling pathways involved in this model were dissected using non-selective and selective NOS inhibitors, knockout mice lacking eNOS or nNOS and their wild-type control mice. Also, we tested a soluble guanylate cyclase inhibitor and a peroxynitrite decomposition catalyst (Ntotal = 312). Results: Non-selective NOS inhibition blocked GTN-induced hypersensitivity. This response was partially associated with iNOS, and potentially nNOS and eNOS conjointly. Furthermore, we found that the GTN response was largely dependent on the generation of peroxynitrite and partly soluble guanylate cyclase. Conclusions: Migraine-relevant hypersensitivity induced by GTN is mediated by a possible feed-forward phenomenon of NO driven mainly by iNOS but with contributions from other isoforms. The involvement of peroxynitrite adds to the notion that oxidative stress reactions are also involved. [ABSTRACT FROM AUTHOR]

Background: Pituitary adenylate cyclase-activating peptide (PACAP) is a neuropeptide pivotal in migraine pathophysiology and is considered a promising new migraine drug target. Although intravenous PACAP triggers migraine attacks and a recent phase II trial with a PACAP-inhibiting antibody showed efficacy in migraine prevention, targeting the PACAP receptor PAC1 alone has been unsuccessful. The present study investigated the role of three PACAP receptors (PAC1, VPAC1 and VPAC2) in inducing migraine-relevant hypersensitivity in mice. Methods: Hindpaw hypersensitivity was induced by repeated PACAP38 injections. Tactile sensitivity responses were quantified using von Frey filaments in three knockout (KO) mouse strains, each lacking one of the PACAP-receptors (Ntotal = 160). Additionally, ex vivo wire myography was used to assess vasoactivity of the carotid artery, and gene expression of PACAP receptors was examined by qPCR. Results: PACAP38 induced hypersensitivity in WT controls (p < 0.01) that was diminished in VPAC1 and VPAC2 KO mice (p < 0.05). In contrast, PAC1 KO mice showed similar responses to WT controls (p > 0.05). Myograph experiments supported these findings showing diminished vasoactivity in VPAC1 and VPAC2 KO mice. We found no upregulation of the non-modified PACAP receptors in KO mice. Conclusions: This study assessed all three PACAP receptors in a migraine mouse model and suggests a significant role of VPAC receptors in migraine pathophysiology. The lack of hypersensitivity reduction in PAC1 KO mice suggests the involvement of other PACAP receptors or compensatory mechanisms. The results indicate that targeting only individual PACAP receptors may not be an effective migraine treatment. [ABSTRACT FROM AUTHOR]

Supplemental Digital Content is Available in the Text. High-speed videography of hind paw withdrawal produced variable results across experimenters, did not consistently reflect intensity-dependent pain scores, and did not differentiate inflamed and naive animals. Few analgesics identified using preclinical models have successfully translated to clinical use. These translational limitations may be due to the unidimensional nature of behavioral response measures used to assess rodent nociception. Advances in high-speed videography for pain behavior allow for objective quantification of nuanced aspects of evoked paw withdrawal responses. However, whether videography-based assessments of mechanical hypersensitivity outperform traditional measurement reproducibility is unknown. First, we determined whether high-speed videography of paw withdrawal was reproducible across experimenters. Second, we examined whether this method distinguishes behavioral responses exhibited by naive mice and mice with complete Freund's adjuvant (CFA)-induced inflammation. Twelve experimenters stimulated naive C57BL/6 mice with varying mechanical stimuli. Paw withdrawal responses were recorded with high-speed videography and scored offline by one individual. Our group was unable to replicate the original findings produced by high-speed videography analysis. Surprisingly, ∼80% of variation was not accounted for by variables previously reported to distinguish between responses to innocuous and noxious stimuli (paw height, paw velocity, and pain score), or by additional variables (experimenter, time-of-day, and animal), but rather by unidentified factors. Similar high-speed videography assessments were performed in CFA- and vehicle-treated animals, and the cumulative data failed to reveal an effect of CFA injection on withdrawal as measured by high-speed videography. This study does not support using paw height, velocity, or pain score measurements from high-speed recordings to delineate behavioral responses to innocuous and noxious stimuli. Our group encourages the continued use of traditional mechanical withdrawal assessments until additional high-speed withdrawal measures are validated in established pain models. [ABSTRACT FROM AUTHOR]

Objective: To characterize the circadian features of the trigeminal ganglion in a mouse model of headache. Background: Several headache disorders, such as migraine and cluster headache, are known to exhibit distinct circadian rhythms of attacks. The circadian basis for these rhythmic pain responses, however, remains poorly understood. Methods: We examined trigeminal ganglion ex vivo and single‐cell cultures from Per2::LucSV reporter mice and performed immunohistochemistry. Circadian behavior and transcriptomics were investigated using a novel combination of trigeminovascular and circadian models: a nitroglycerin mouse headache model with mechanical thresholds measured every 6 h, and trigeminal ganglion RNA sequencing measured every 4 h for 24 h. Finally, we performed pharmacogenomic analysis of gene targets for migraine, cluster headache, and trigeminal neuralgia treatments as well as trigeminal ganglion neuropeptides; this information was cross‐referenced with our cycling genes from RNA sequencing data to identify potential targets for chronotherapy. Results: The trigeminal ganglion demonstrates strong circadian rhythms in both ex vivo and single‐cell cultures, with core circadian proteins found in both neuronal and non‐neuronal cells. Using our novel behavioral model, we showed that nitroglycerin‐treated mice display circadian rhythms of pain sensitivity which were abolished in arrhythmic Per1/2 double knockout mice. Furthermore, RNA‐sequencing analysis of the trigeminal ganglion revealed 466 genes that displayed circadian oscillations in the control group, including core clock genes and clock‐regulated pain neurotransmitters. In the nitroglycerin group, we observed a profound circadian reprogramming of gene expression, as 331 of circadian genes in the control group lost rhythm and another 584 genes gained rhythm. Finally, pharmacogenetics analysis identified 10 genes in our trigeminal ganglion circadian transcriptome that encode target proteins of current medications used to treat migraine, cluster headache, or trigeminal neuralgia. Conclusion: Our study unveiled robust circadian rhythms in the trigeminal ganglion at the behavioral, transcriptomic, and pharmacogenetic levels. These results support a fundamental role of the clock in pain pathophysiology. Plain Language Summary: Several headache diseases, such as migraine and cluster headache, have headaches that occur at the same time each day. We learned that the trigeminal ganglion, an important pain structure in several headache diseases, has a 24‐hour cycle that might be related to this daily cycle of headaches. Our genetic analysis suggests that some medications may be more effective in treating migraine and cluster headache when taken at specific times of the day. Plain Language Summary: Several headache diseases, such as migraine and cluster headache, have headaches that occur at the same time each day. We learned that the trigeminal ganglion, an important pain structure in several headache diseases, has a 24‐hour cycle that might be related to this daily cycle of headaches. Our genetic analysis suggests that some medications may be more effective in treating migraine and cluster headache when taken at specific times of the day. [ABSTRACT FROM AUTHOR]

Introduction/Aims: We have recently isolated and expanded skin‐derived Schwann cells (Sk‐SCs) from human skin and showed that they are largely similar to nerve‐derived Schwann cells (N‐SCs). Here, we extend our investigation into functional assessments of the nude rats that received human Sk‐SCs and N‐SCs after intraneural delivery into crushed and decellularized tibial nerve in adult nude rats. Methods: Sk‐SCs, N‐SCs, dermal fibroblasts, or control culture medium was injected into the crushed and decellularized tibial nerve using in situ repeated freeze‐thaw cycles. Animals were then subjected to a ladder rung walking test, nociceptive von Frey testing, and walking gait analysis weekly. Animals were euthanized 6 weeks after surgery, gastrocnemius and soleus muscles were weighed, distal nerves were harvested, and whole semithin cross‐sections were analyzed using segmentation software. Results: N‐SC–injected and dermal fibroblast–injected animals improved significantly at 4 to 6 weeks postinjury in nociceptive assessment compared with medium‐injected controls. Sk‐SCs recovered more rapidly in tibial functional index at 2 weeks postinjury compared with medium‐injected controls. No significant difference was observed for the ladder rung walking test or muscle weight ratio. Histologically, the number of myelinated axons was significantly higher in all cell injection groups compared with medium‐injected controls. No significant difference was observed in g ratio, axon diameter, or myelin thickness. Discussion: Cell injection significantly improved axon regeneration across an in situ decellularized nerve segment. However, a more human cell–permissive animal model is required to delineate functional differences between cell types for preclinical transplantation studies. [ABSTRACT FROM AUTHOR]

The objective of this study was to determine to what extent the oculocardiac reflex (OCR) progressively rises with increasing external pressures over the superior eyelid. Controlled external weights were applied to the left eyelid of ten conscious healthy adult beagle dogs, using an electronic von Frey aesthesiometer with a modified probe. Four different weights (200 g, 300 g, 400 g and 800 g) were applied on four different non-consecutive days (with one-week intervals). Pressure was applied by pushing the device against the superior eyelid. The baseline heart rate was primarily recorded using an electrocardiograph (ECG). Then, indices of heart rate variability such as R-R intervals and vasovagal tonus index (VVTI) were calculated from the ECG tracings using twenty consecutive QRS complexes from each dog, before and after a selected set of graded weights were applied to the eye. Median R-R interval significantly increased when an external weight of 200 g was applied and peaked at 300 g. VVTI values peaked at 400 g. With higher pressures, the proportional increase was smaller in both R-R intervals and VVTI. We concluded that external eye pressure using a weight of 200 g or more is sufficient to elicit an OCR response in conscious dogs. Pressures above 400 g started to activate compensatory mechanisms that counteracted with the OCR, probably linked to an adrenergic response induced by the animal's discomfort. Changes in R-R intervals might be used as an early OCR marker, while VVTI apparently was less influenced by potential sympathetic responses. [ABSTRACT FROM AUTHOR]

Background: Calcitonin gene-related peptide (CGRP) antagonizing drugs represents the most important advance in migraine therapy for decades. However, these new drugs are only effective in 50–60% of patients. Recent studies have shown that the pituitary adenylate cyclase-activating peptide (PACAP38) pathway is independent from the CGRP signaling pathway. Here, we investigate PACAP38 signaling pathways in relation to glyceryl trinitrate (GTN), levcromakalim and sumatriptan. Methods: In vivo mouse models of PACAP38-, GTN-, and levcromakalim-induced migraine were applied using tactile sensitivity to von Frey filaments as measuring readout. Signaling pathways involved in the three models were dissected using PACAP-inhibiting antibodies (mAbs) and sumatriptan. Results: We showed that PACAP mAbs block PACAP38 induced hypersensitivity, but not via signaling pathways involved in GTN and levcromakalim. Also, sumatriptan has no effect on PACAP38-induced hypersensitivity relevant to migraine. This is the first study testing the effect of a PACAP-inhibiting drug on GTN- and levcromakalim-induced hypersensitivity. Conclusions: Based on the findings in our mouse model of migraine using migraine-inducing compounds and anti-migraine drugs, we suggest that PACAP acts via a distinct pathway. Using PACAP38 antagonism may be a novel therapeutic target of interest in a subgroup of migraine patients who do not respond to existing therapies. [ABSTRACT FROM AUTHOR]

Abstract: Identifying the genetic determinants of pain is a scientific imperative given the magnitude of the global health burden that pain causes. Here, we report a genetic screen for nociception, performed under the auspices of the International Mouse Phenotyping Consortium. A biased set of 110 single-gene knockout mouse strains was screened for 1 or more nociception and hypersensitivity assays, including chemical nociception (formalin) and mechanical and thermal nociception (von Frey filaments and Hargreaves tests, respectively), with or without an inflammatory agent (complete Freund's adjuvant). We identified 13 single-gene knockout strains with altered nocifensive behavior in 1 or more assays. All these novel mouse models are openly available to the scientific community to study gene function. Two of the 13 genes (Gria1 and Htr3a) have been previously reported with nociception-related phenotypes in genetically engineered mouse strains and represent useful benchmarking standards. One of the 13 genes (Cnrip1) is known from human studies to play a role in pain modulation and the knockout mouse reported herein can be used to explore this function further. The remaining 10 genes (Abhd13, Alg6, BC048562, Cgnl1, Cp, Mmp16, Oxa1l, Tecpr2, Trim14, and Trim2) reveal novel pathways involved in nociception and may provide new knowledge to better understand genetic mechanisms of inflammatory pain and to serve as models for therapeutic target validation and drug development. [ABSTRACT FROM AUTHOR]

Thermal nociception involves the transmission of temperature-related noxious information from the periphery to the CNS and is a heritable trait that could predict transition to persistent pain. Rodent forward genetics complement human studies by controlling genetic complexity and environmental factors, analysis of end point tissue, and validation of variants on appropriate genetic backgrounds. Reduced complexity crosses between nearly identical inbred substrains with robust trait differences can greatly facilitate unbiased discovery of novel genes and variants. We found BALB/cByJ mice showed enhanced sensitivity on the 53.5°C hot plate and mechanical stimulation in the von Frey test compared to BALB/cJ mice and replicated decreased gross brain weight in BALB/cByJ versus BALB/cJ. We then identified a quantitative trait locus (QTL) on chromosome 13 for hot plate sensitivity (LOD = 10.7; p < 0.001; peak = 56 Mb) and a QTL for brain weight on chromosome 5 (LOD = 8.7; p < 0.001). Expression QTL mapping of brain tissues identified H2afy (56.07 Mb) as the top transcript with the strongest association at the hot plate locus (FDR = 0.0002) and spliceome analysis identified differential exon usage within H2afy associated with the same locus. Whole brain proteomics further supported decreased H2AFY expression could underlie enhanced hot plate sensitivity, and identified ACADS as a candidate for reduced brain weight. To summarize, a BALB/c reduced complexity cross combined with multiple-omics approaches facilitated identification of candidate genes underlying thermal nociception and brain weight. These substrains provide a powerful, reciprocal platform for future validation of candidate variants. [ABSTRACT FROM AUTHOR]

Abstract: Glycinergic neurons and glycine receptors (GlyRs) exert a critical control over spinal nociception. Prostaglandin E2 (PGE2), a key inflammatory mediator produced in the spinal cord in response to peripheral inflammation, inhibits a certain subtype of GlyRs (α3GlyR) that is defined by the inclusion of α3 subunits and distinctly expressed in the lamina II of the spinal dorsal horn, ie, at the site where most nociceptive nerve fibers terminate. Previous work has shown that the hyperalgesic effect of spinal PGE2 is lost in mice lacking α3GlyRs and suggested that this phenotype results from the prevention of PGE2-evoked protein kinase A (PKA)-dependent phosphorylation and inhibition of α3GlyRs. However, direct proof for a contribution of this phosphorylation event to inflammatory hyperalgesia was still lacking. To address this knowledge gap, a phospho-deficient mouse line was generated that carries a serine to alanine point mutation at a strong consensus site for PKA-dependent phosphorylation in the long intracellular loop of the GlyR α3 subunit. These mice showed unaltered spinal expression of GlyR α3 subunits. In behavioral experiments, they showed no alterations in baseline nociception, but were protected from the hyperalgesic effects of intrathecally injected PGE2 and exhibited markedly reduced inflammatory hyperalgesia. These behavioral phenotypes closely recapitulate those found previously in GlyR α3-deficient mice. Our results thus firmly establish the crucial role of PKA-dependent phosphorylation of α3GlyRs in inflammatory hyperalgesia. [ABSTRACT FROM AUTHOR]

Background and aims: Preclinical animal models are crucial to study pain mechanisms and assess antinociceptive effects of medications. One major problem with current animal behavioral models is their lack of face validity with human nociception and the vulnerability for false-positive results. Here, we evaluated the usefulness of rotarod as a new way to assess inflammatory nociception in rodents. Methods: Adult male mice were injected with saline or Complete Freund's Adjuvant (CFA) in the left hindpaws. Mechanical allodynia and rotarod performance were evaluated before and after the administration of CFA. Mechanical allodynia was measured using von Frey filaments. Long-term effect of CFA on rotarod performance was also assessed for 2 weeks. Results: Our results showed that CFA administration decreased pain threshold and increased sensitivity to von Frey filaments compared to control group. In rotarod experiments, the starting speed of the rod rotation started at four RPM, and accelerated until it reached 40 RPM in 5 min. Rotarod performance was enhanced from day to day in the control group. However, rotarod performance in CFA group was attenuated after CFA administration, which was significant after 24 h compared to vehicle. This attenuation was blocked by ibuprofen. Haloperidol administration (positive control) produced similar results to CFA administration. CFA did not produce significant attenuation of rotarod performance after 1 week post-injection. Conclusions: Collectively, our findings could encourage the use of rotarod assay to measure acute (but not chronic) inflammatory nociception as a useful tool in rodents. [ABSTRACT FROM AUTHOR]

Abstract Pain is one of the cardinal signs of inflammation and is present in many inflammatory conditions. Therefore, anti-inflammatory drugs such as NSAIDs also have analgesic properties. We previously showed that prostaglandin D 2 -glycerol ester (PGD 2 -G), endogenously produced by cyclooxygenase-2 from the endocannabinoid 2-arachidonoylglycerol, has anti-inflammatory effects in vitro and in vivo that are partly mediated by DP1 receptor activation. In this work, we investigated its effect in a model of carrageenan-induced inflammatory pain. PGD 2 -G decreased hyperalgesia and edema, leading to a faster recovery. Moreover, PGD 2 -G decreased carrageenan-induced inflammatory markers in the paw as well as inflammatory cell recruitment. The effects of PGD 2 -G were independent from metabolite formation (PGD 2 and 15d-PGJ 2 -G) or DP1 receptor activation in this model. Indeed PGD 2 delayed recovery from hyperalgesia while 15d-PGJ 2 -G worsened the edema. However, while PGD 2 -G decreased hyperalgesia in this model of inflammatory pain, it had no effect when tested in the capsaicin-induced pain model. While the targets mediating the effects of this bioactive lipid in inflammatory pain remain to be elucidated, our findings further support the interest of anti-inflammatory lipid mediators in the management of inflammatory pain. Highlights • PGD 2 -G decreases inflammation and pain in carrageenan-induced hyperalgesia in mice. • These effects are not due to hydrolysis of PGD 2 -G into PGD 2. • 15d-PGJ 2 -G, a PGD 2 -G metabolite, does not mediate its effects on inflammatory pain. • PGD 2 -G decreases macrophage activation and neutrophil infiltration in the paw. • The targets mediating the effects of PGD 2 -G remain to be elucidated. [ABSTRACT FROM AUTHOR]

Background: Flavonoids, a group of polyphenols responsible for protective role against many diseased conditions, provide antioxidant activity which is the reason for their medicinal properties. Tactile allodynia is a behavioral biomarker of neuropathy that is well estimated by von Frey filaments and Randall–Selitto test. Objective: Ficus carica Lam. leaves were studied for the conformation of flavonoids in ethyl acetate fraction of methanolic extract (FCEA) using GC-HRMS for the identification of flavonoids. It was analyzed for antioxidant activity by in vitro free radical scavenging activity, performed using 2,2-diphenyl-1-picrylhydrazyl (DPPH) followed by blood glucose-level estimation, evaluation of neuropathic pain, and kidney and liver function tests in diabetic rats. Materials and Methods: The shade-dried leaves of F. carica Lam. were extracted with methanol and after that fractionated using ethyl acetate (FCEA). The characterization of FCEA was established using GC-HRMS. In vitro free radical scavenging activity was performed using DPPH assay. Diabetes was induced using streptozotocin (40 mg/kg/intraperitoneally), and effects of FCEA were studied on blood glucose level, neuropathy markers, and liver and kidney functions of diabetic rats. Results: GC-HRMS results highlighted the presence of quercetin, kaempferol, and chrysin in FCEA with free radical scavenging activity of 78.35% and IC50 value of 5.508 μM. FCEA reduces glucose levels and also shows protective effects in case of diabetic neuropathy as it increases the threshold of withdrawal latency in tactile allodynia and also decreases the serum glutamic-oxaloacetic transaminase, serum glutamic-pyruvic transaminase, blood urea nitrogen, and creatinine levels. Conclusion: The protective effects of FCEA against diabetic neuropathy, hepatoprotective and nephroprotective effects might be due to strong antioxidant property of important flavonoids present which is confirmed in the study. [ABSTRACT FROM AUTHOR]

Abstract Gulf war illness (GWI) is a chronic multi-symptom disease that afflicts 25–33% of troops that were deployed in the 1990–1991 Gulf War. GWI symptoms include cognitive, behavioral and emotional deficits, as well as migraines and pain. It is possible that exposure to Gulf War agents and prophylactics contributed to the reported symptomology. Pyridostigmine bromide (PB) and permethrin (PER) were given to protect from nerve gas attacks and insect vector born disease, respectively. Previous studies have demonstrated that 10 days of exposure to these chemicals can cause symptoms analogous to those observed in GWI, including impairment of long-term memory in mice. Other studies using this model have shown chronic neuroinflammation, and chronic neuroinflammation can lead to altered nociceptive sensitivity. At 10-weeks after the 10-day PB and PER exposure paradigm, we observed lowered nociceptive threshold on the Von Frey test that was no longer evident at 28 weeks and 38 weeks post-exposure. We further determined that vagus nerve stimulation, initiated at 38 weeks after exposure, restores the lowered nociceptive sensitivity. Therefore, stimulating the vagus nerve appears to influence nociception. Future studies are need to elucidate possible mechanisms of this effect. [ABSTRACT FROM AUTHOR]

Highlights • Sumatriptan increased mechanical thresholds in the periorbital region for four days. • Pulsed FUS increased mechanical thresholds post therapy in the periorbital region. • Ablative HIFU did not increase mechanical thresholds post therapy. • FUS may provide a therapeutic option for patients suffering from chronic migraines. Abstract Chronic migraines (CM) are the third most common disease and are refractory to medical treatment in 15% of patients. Currently, temporary relief is achieved with steroid blocks or pulsed radiofrequency ablation, which have short-term benefits. Our project aims to develop a non-invasive treatment for medically refractory chronic migraine, which does not require a permanent implant. This project investigates the safety and effectiveness of pulsed focused ultrasound (FUS) in a validated rodent headache model of cutaneous allodynia associated with chronic migraine (CM) as compared to sumatriptan and ablative lesioning. We demonstrate a significant reduction in mechanical thresholds as measured through Von Frey filaments in CM in the forepaw and periorbital region (p < 0.001). Sumatriptan and pulsed FUS both significantly improve thresholds at day 3 after treatment in the periorbital region. Ablative lesioning has no effect. This study provides initial evidence that FUS may provide an important therapeutic option for patients suffering from CM. [ABSTRACT FROM AUTHOR]

Objective: Two well‐known spinal cord stimulation (SCS) paradigms, conventional (Con) and burst SCS, are hypothesized to exert their antinociceptive effects through different stimulation‐induced mechanisms. We studied the course of the behavioral antinociceptive effect during 60 minutes of SCS and 30 minutes post‐SCS in a rat model of chronic neuropathic pain. Methods: Animals underwent a unilateral partial sciatic nerve ligation, after which quadripolar electrodes were implanted into the epidural space at vertebral level T13 (n = 43 rats). While receiving either Con SCS or biphasic burst SCS, the pain behavior of the rats was assessed by means of paw withdrawal thresholds (WTs) in response to the application of von Frey monofilaments. Results: After 15 minutes of Con SCS (n = 21), WTs significantly differed from baseline (P = 0.04), whereas WTs of the burst SCS group (n = 22) did not. After 30 minutes of SCS, WTs of the Con SCS and burst SCS groups reached similar levels, both significantly different from baseline, indicating a comparable antinociceptive effect for these SCS paradigms. Yet, the WTs of the burst SCS group were still significantly increased compared with baseline at 30 minutes post‐stimulation, whereas the WTs of the Con SCS group were not. Conclusions: To conclude, biphasic burst SCS results in a delayed antinociceptive effect after onset of the stimulation, as compared with Con SCS, in a chronic neuropathic pain model. Furthermore, biphasic burst SCS seems to exhibit a delayed wash‐out of analgesia after stimulation is turned off. [ABSTRACT FROM AUTHOR]

Anxiety disorders are the most prevalent psychiatric disorders, and they are highly comorbid with chronic pain conditions. The central nucleus of the amygdala (CeA) is known not only for its role in the regulation of anxiety but also as an important site for the negative affective dimension of pain. Pituitary adenylate cyclase activating polypeptide (PACAP), a neuropeptide whose terminals are abundant in the CeA, is strongly implicated in the stress response as well as in pain processing. Here, using Cre-dependent viral vectors, we explored in greater detail the role of the PACAP projection to the CeA that originates in the lateral parabrachial nucleus (LPB). We first performed a circuit mapping experiment by injecting an anterograde Cre-dependent virus expressing a fluorescent reporter in the LPB of PACAP-Cre mice and observing their projections. Then, we used a chemogenetic approach (a Cre-dependent Designer Receptors Activated by Designer Drugs, DREADDs) to assess the effects of the direct stimulation of the PACAP LPB to CeA projection on general locomotor activity, anxiety-like behavior (using a defensive withdrawal test), and mechanical pain sensitivity (using the von Frey test). We found that the CeA, together with other areas, is one of the major downstream projection targets of PACAP neurons originating in the lateral parabrachial nucleus (LPB). In the DREADD experiment, we then found that the selective activation of this neuronal pathway is sufficient to increase both anxiety-like behavior and mechanical pain sensitivity in mice, without affecting general locomotor activity. In conclusion, our data suggest that the dysregulation of this circuit may contribute to a variety of anxiety disorders and chronic pain states, and that PACAP may represent an important therapeutic target for the treatment of these conditions. • We used a circuit mapping approach to assess downstream targets of LPB PACAP neurons. • Considerable PACAP terminals originating in LPB were observed in CeA, BNST, and VMH. • We used chemogenetics to assess the effect of the activation of LPB-CeA PACAP neurons. • Pathway activation increased anxiety-like behavior and mechanical pain sensitivity. • Dysregulation of this circuit may contribute to anxiety disorders and chronic pain. [ABSTRACT FROM AUTHOR]

Background: Systemic inflammation is known to induce sickness behaviors, including decreased social interaction and pain. We have reported increased serum inflammatory cytokines in a rat model of repetitive strain injury (rats perform an upper extremity reaching task for prolonged periods). Here, we sought to determine if sickness behaviors are induced in this model and the effectiveness of conservative treatments.Methods: Experimental rats underwent initial training to learn a high force reaching task (10 min/day, 5 days/week for 6 weeks), with or without ibuprofen treatment (TRHF vs. TRHF + IBU rats). Subsets of trained animals went on to perform a high repetition high force (HRHF) task for 6 or 12 weeks (2 h/day, 3 days/week) without treatment, or received two secondary interventions: ibuprofen (HRHF + IBU) or a move to a lower demand low repetition low force task (HRHF-to-LRLF), beginning in task week 5. Mixed-effects models with repeated measures assays were used to assay duration of social interaction, aggression, forepaw withdrawal thresholds and reach performance abilities. One-way and two-way ANOVAs were used to assay tissue responses. Corrections for multiple comparisons were made.Results: TRHF + IBU rats did not develop behavioral declines or systemic increases in IL-1beta and IL-6, observed in untreated TRHF rats. Untreated HRHF rats showed social interaction declines, difficulties performing the operant task and forepaw mechanical allodynia. Untreated HRHF rats also had increased serum levels of several inflammatory cytokines and chemokines, neuroinflammatory responses (e.g., increased TNFalpha) in the brain, median nerve and spinal cord, and Substance P and neurokinin 1 immunoexpression in the spinal cord. HRHF + IBU and HRHF-to-LRLF rats showed improved social interaction and reduced inflammatory serum, nerve and brain changes. However, neither secondary treatment rescued HRHF-task induced forepaw allodynia, or completely attenuated task performance declines or spinal cord responses.Conclusions: These results suggest that inflammatory mechanisms induced by prolonged performance of high physical demand tasks mediate the development of social interaction declines and aggression. However, persistent spinal cord sensitization was associated with persistent behavioral indices of discomfort, despite use of conservative secondary interventions indicating the need for prevention or more effective interventions. [ABSTRACT FROM AUTHOR]

Neuropathic pain is a debilitating, chronic condition with a significant unmet need for effective treatment options. Recent studies have demonstrated that in addition to neurons, non-neuronal cells such as microglia contribute to the initiation and maintenance of allodynia in rodent models of neuropathic pain. The Ca 2+ - activated K + channel, K Ca 3.1 is critical for the activation of immune cells, including the CNS-resident microglia. In order to evaluate the role of K Ca 3.1 in the maintenance of mechanical allodynia following peripheral nerve injury, we used senicapoc, a stable and highly potent K Ca 3.1 inhibitor. In primary cultured microglia, senicapoc inhibited microglial nitric oxide and IL-1β release. In vivo , senicapoc showed high CNS penetrance and when administered to rats with peripheral nerve injury, it significantly reversed tactile allodynia similar to the standard of care, gabapentin. In contrast to gabapentin, senicapoc achieved efficacy without any overt impact on locomotor activity. Together, the data demonstrate that the K Ca 3.1 inhibitor senicapoc is effective at reducing mechanical hypersensitivity in a rodent model of peripheral nerve injury. [ABSTRACT FROM AUTHOR]

The upregulation of the tetrodotoxin-resistant voltage-gated sodium channel NaV1.9 has previously been associated with inflammatory hyperalgesia. Na1.9 knockout (KO) mice, however, did not seem insensitive in conventional tests of acute nociception. Using electrophysiological, neurochemical, and behavioral techniques, we now show NaV1.9-null mice exhibit impaired mechanical and thermal sensory capacities and reduced electrical excitability of nociceptors. In single-fiber recordings from isolated skin, the electrical threshold of NaV1.9 KO C fibers was elevated by 55% and the median von Frey threshold was 32 mN in contrast to 8 mN in wild types (WTs). The prevalence of C mechano-heat-sensitive (CMH) fibers was only 25.6% in NaV1.9 KO animals compared to 75.8% in the WT group, and the heat threshold of these CMH fibers was 40.4°C in the control vs 44°C in the KO group. Compound action potential recordings from isolated sciatic nerve segments of NaV1.9 KO mice revealed lower activity-induced slowing of conduction velocity upon noxious heat stimulation: 8% vs 30% in WTs. Heat-induced calcitonin gene-related peptide release from the skin was less in the KO than in the WT group. The reduced noxious heat sensitivity was finally confirmed with the Hargreaves test using 2 rates of radiant heating of the plantar hind paws. In conclusion, NaV1.9 presumably contributes to acute thermal and mechanical nociception in mice, most likely through increasing the excitability but probably also by amplifying receptor potentials irrespective of the stimulus modality. [ABSTRACT FROM AUTHOR]

Friedreich’s Ataxia (FA) is a pediatric neurodegenerative disease whose clinical presentation includes ataxia, muscle weakness, and peripheral sensory neuropathy. The KIKO mouse is an animal model of FA with frataxin deficiency first described in 2002, but neurobehavioral deficits have never been described in this model. The identification of robust neurobehavioral deficits in KIKO mice could support the testing of drugs for FA, which currently has no approved therapy. We tested 13 neurobehavioral tasks to identify a robust KIKO phenotype: Open Field, Grip Strength Test(s), Cylinder, Skilled Forelimb Grasp Task(s), Treadmill Endurance, Locotronic Motor Coordination, Inverted Screen, Treadscan, and Von Frey. Of these, Inverted Screen, Treadscan and Von Frey produced significant neurobehavioral deficits at >8 months of age, and relate to the clinically relevant endpoints of muscle strength and endurance, gait ataxia, and peripheral insensitivity. Thus we identify robust phenotypic measures related to Friedreich's ataxia clinical endpoints which could be used to test effectiveness of potential drug therapy. [ABSTRACT FROM AUTHOR]

The anterior cingulate cortex (ACC) has been extensively cited as a key area for processing pain affect. While local field potential (LFP) studies in other fields have yielded a great deal of information about neural oscillations, there is a poverty in the pain literature about the neural LFP profile related to pain, particularly in freely moving animals. In this study, we revealed the LFP profile in the ACC in freely moving rats during carrageenan inflammation. Mechanical allodynia was recorded before and after unilateral injection of carrageenan/saline in the left hindpaw. LFP activity in the ACC was recorded at baseline, after injection, and after injection with mechanical stimulation to the paw using a von Frey filament. This study uniquely reveals that carrageenan injection significantly recruited ACC LFP activity in delta, theta, and alpha bands (0–13 Hz). Application of von Frey mechanical stimulation to the carrageenan-injected paw resulted in a significant increase in delta, theta, and alpha bands over and above what was recruited by carrageenan alone and further expanded the LFP range to additionally include beta activity (13–30 Hz). Taken together, these data reveal significant changes in the lowest-frequency activities in the LFP range during painful inflammation, which merit attention. LFP is a powerful window to reveal wide-range, integrated synaptic processing by low-frequency cellular events during behavior. Information about LFP during pain broadens the scope of our understanding of pain mechanisms, our greatest resource for designing management approaches. [ABSTRACT FROM AUTHOR]

Mechanical sensitivity is commonly affected in chronic pain and other neurological disorders. To discover mechanisms of individual differences in punctate mechanosensation, we performed quantitative trait locus (QTL) mapping of the response to von Frey monofilament stimulation in BXD recombinant inbred (BXD) mice. Significant loci were detected on mouse chromosome (Chr) 5 and 15, indicating the location of underlying polymorphisms that cause heritable variation in von Frey response. Convergent evidence from public gene expression data implicates candidate genes within the loci: von Frey thresholds were strongly correlated with baseline expression of Cacna2d1, Ift27 and Csnk1e in multiple brain regions of BXD strains. Systemic gabapentin and PF-670462, which target the protein products of Cacna2d1 and Csnk1e, respectively, significantly increased von Frey thresholds in a genotype-dependent manner in progenitors and BXD strains. Real-time polymerase chain reaction confirmed differential expression of Cacna2d1 and Csnk1e in multiple brain regions in progenitors and showed differential expression of Cacna2d1 and Csnk1e in the dorsal root ganglia of the progenitors and BXD strains grouped by QTL genotype. Thus, linkage mapping, transcript covariance and pharmacological testing suggest that genetic variation affecting Cacna2d1 and Csnk1e may contribute to individual differences in von Frey filament response. This study implicates Cacna2d1 and Ift27 in basal mechanosensation in line with their previously suspected role in mechanical hypersensitivity. Csnk1e is implicated for von Frey response for the first time. Further investigation is warranted to identify the specific polymorphisms involved and assess the relevance of these findings to clinical conditions of disturbed mechanosensation. [ABSTRACT FROM AUTHOR]

Recent studies have suggested that in humans and animals with significant skeletal pain, changes in the mechanical hypersensitivity of the skin can be detected. However, whether measuring changes in skin hypersensitivity can be a reliable surrogate for measuring skeletal pain itself remains unclear. To explore this question, we generated skeletal pain by injecting and confining GFP-transfected NCTC 2472 osteosarcoma cells unilaterally to the femur of C3H male mice. Beginning at day 7 post-tumor injection, animals were administered vehicle, an antibody to the P2X3 receptor (anti-P2X3) or anti-NGF antibody. Pain and analgesic efficacy were then measured on days 21, 28, and 35 post-tumor injection using a battery of skeletal pain-related behaviors and von Frey assessment of mechanical hypersensitivity on the plantar surface of the hind paw. Animals with bone cancer pain treated with anti-P2X3 showed a reduction in skin hypersensitivity but no attenuation of skeletal pain behaviors, whereas animals with bone cancer pain treated with anti-NGF showed a reduction in both skin hypersensitivity and skeletal pain behaviors. These results suggest that although bone cancer can induce significant skeletal pain-related behaviors and hypersensitivity of the skin, relief of hypersensitivity of the skin is not always accompanied by attenuation of skeletal pain. Understanding the relationship between skeletal and skin pain may provide insight into how pain is processed and integrated and help define the preclinical measures of skeletal pain that are predictive end points for clinical trials. [ABSTRACT FROM AUTHOR]

Despite much evidence that combination of morphine and gabapentin can be beneficial for managing postoperative pain, the nature of the pharmacological interaction of the two drugs remains unclear. The aim of this study was to assess the interaction of morphine and gabapentin in range of different dose combinations and investigate whether co-administration leads to synergistic effects in a preclinical model of postoperative pain. The pharmacodynamic effects of morphine (1, 3 and 7 mg/kg), gabapentin (10, 30 and 100 mg/kg) or their combination (9 combinations in total) were evaluated in the rat plantar incision model using an electronic von Frey device. The percentage of maximum possible effect (%MPE) and the area under the response curve (AUC) were used for evaluation of the antihyperalgesic effects of the drugs. Identification of synergistic interactions was based on Loewe additivity response surface analyses. The combination of morphine and gabapentin resulted in synergistic antihyperalgesic effects in a preclinical model of postoperative pain. The synergistic interactions were found to be dose dependent and the increase in observed response compared to the theoretical additive response ranged between 26 and 58% for the synergistic doses. The finding of dose-dependent synergistic effects highlights that choosing the right dose–dose combination is of importance in postoperative pain therapy. Our results indicate benefit of high doses of gabapentin as adjuvant to morphine. If these findings translate to humans, they might have important implications for the treatment of pain in postoperative patients. [ABSTRACT FROM AUTHOR]

Aims This study analyzed the sensorial, structural and functional response of rats subjected to paw immobilization. Main methods Animal pelvis, hip, knee and ankle were immobilized using waterproof tape during two weeks for assessment of sensorial response to thermal (hot plate test) and mechanical stimuli (Von Frey test), motor system structure (histology and radiography) and muscle function (soleus contractility). Key findings Disuse animals became more responsive to thermal stimuli (49%), although less responsive to mechanical challenge (58%). Disuse animals showed local injuries such as reduction in muscle fiber diameter (16.7% in gastrocnemius, 5.7% in soleus), contractile activity (55% of the control maximal tonic contraction) and tibia cortical thickness (9.3%), besides increased nitrite:protein ratio, suggestive of protein degradation. Disuse also evoked systemic adaptations that include increase in serum lactate dehydrogenase (36.1%) and alkaline phosphatase (400%), but reduction in calcium (8.4%) and total serum protein (5.5%), especially albumin (34.2%). Significance Two weeks of functional paw disuse leads to local and systemic harmful adaptive changes in sensorial and structural systems. This study brings new insights into nervous and motor system mechanism associated with therapeutic limb immobilization in muscle and skeletal pathological conditions. [ABSTRACT FROM AUTHOR]

Spinal nerve L5/L6 ligation (SNL) in rats has become the standard for mechanistic studies of peripheral neuropathy and screening for novel analgesics. Conventional SNL in our hybrid mice resulted in a wide range of allodynia. Anatomical evaluation indicated that a variable number of lumbar vertebrae existed, resulting in L4/L5 or L5/L6 being ligated. Surprisingly, L4/L5 ligation did not result in ipsilateral hind limb paralysis and produced robust allodynia. Following a recent report that the mouse L4 neural segment is homologous with rat L5 we generated L4, L5 or both L4 and L5 (L4/L5) ligations in C57 mice after establishing a modified set of surgical landmarks. In contrast to rats, L4 ligation in these mice did not result in hind limb paralysis. Robust allodynia was observed in all three ligation groups. Nerve degeneration confirmed that L4 and L5, respectively, are primary contributors to the tibial and sural branches of the sciatic nerve in mice. A larger von Frey sensitive area reflected the wider distribution of Wallerian degeneration in the hindlimb of L4- compared to L5-ligated mice. Ligation of mouse L4 and L5 spinal nerves produces consistent, robust neuropathic pain behaviors and is suitable as a model for investigating mechanisms of neuropathic pain and for testing of novel analgesics. Gabapentin, used as a validation drug in neuropathic pain models and as a reference compound for novel analgesics, significantly reduced allodynia in the mice tested (L4/L5 ligations). Given the ease of surgery, robust allodynia, and larger von Frey sensitive area, we conclude that combined ligation of spinal nerves L4 and L5 optimizes the SNL model in mice. [ABSTRACT FROM AUTHOR]

There is a paucity of data on the role of microglia and neuroinflammatory processes in the association between chronic pain and depression. The current study examined the effect of the microglial inhibitor minocycline on depressive-like behaviour, spinal nerve ligation (SNL)-induced mechanical and cold allodynia and associated changes in the expression of genes encoding microglial markers (M1 vs. M2 polarisation) and inflammatory mediators in the prefrontal cortex in the olfactory bulbectomised (OB) rat model of depression. Acute minocycline administration did not alter OB-induced depressive-like behaviour but prevented SNL-induced mechanical allodynia in both OB and sham rats. In comparison, chronic minocycline attenuated OB-induced depressive-like behaviour and prevented the development of SNL-induced mechanical allodynia in OB, but not sham, rats. Further analysis revealed that SNL-induced mechanical allodynia in OB rats was attenuated by chronic minocycline at almost all time-points over a 2 week testing period, an effect observed only from day 10 post-SNL in sham rats. Chronic administration of minocycline reduced the expression of CD11b, a marker of microglial activation, and the M1 pro-inflammatory cytokine IL-1β, in the prefrontal cortex of sham-SNL animals. In comparison, the expression of the M2 microglia marker (MRC2) and anti-inflammatory cytokine IL-10 was increased, as were IL-1β, IL-6 and SOCS3, in the prefrontal cortex of OB-SNL animals following chronic minocycline. Thus, chronic minocycline attenuates neuropathic pain behaviour and modulates microglial activation and the central expression of inflammatory mediators in a manner dependent on the presence or absence of a depressive-like phenotype. [ABSTRACT FROM AUTHOR]

This study assessed the clinical effects and the mechanical antinociceptive potential of intravenous (IV) tramadol in horses.A blinded and randomized study was designed with 7 horses treated with 1 (Tr1), 2 (Tr2) or 3 (Tr3) mg kg-1 of tramadol IV. The heart rate, respiratory rate (fR), arterial pressure, degree of sedation, gastrointestinal motility (GI), behavior changes and the mechanical nociceptive threshold (MNT) were evaluated. The MNT was determined with von Frey device method.Tr3 had a significant increase in their fR and more pronounced behavioral changes than other treatments. The Tr1 showed a significant increase in arterial pressure. The GI reduced significantly, mainly in Tr2. The tramadol did not change the MNT of the horses. The clinical alterations observed with the different treatments were considered mild and transitory, being most evident in Tr2. However the tramadol did not have any analgesic effect with any of the doses evaluated. [ABSTRACT FROM AUTHOR]

Abstract: Although spinal cord stimulation (SCS) of the dorsal columns is an established method for treating chronic neuropathic pain, patients still suffer from a substantial level of pain. From a clinical perspective it is known that the location of the SCS is of pivotal importance, thereby suggesting a segmental spinal mode of action. However, experimental studies suggest that SCS acts also through the modulation of supraspinal mechanisms, which might suggest that the location is unimportant. Here we investigated the effect of the rostrocaudal location of SCS stimulation and the effectiveness of pain relief in a rat model of chronic neuropathic pain. Adult male rats (n=45) were submitted to a partial ligation of the sciatic nerve. The majority of animals developed tactile hypersensitivity in the nerve lesioned paw. All allodynic rats were submitted to SCS (n=33) for 30minutes (f=50Hz; pulse width 0.2ms). In one group (n=16) the electrodes were located at the level where the injured sciatic nerve afferents enter the spinal cord (T13), and in a second group (n=17) the electrodes were positioned at more rostral levels (T11) as verified by X-ray. A repositioning experiment of electrodes from T12 to T13 was performed in 2 animals. Our data demonstrate that SCS of the dorsal columns at the level where the injured fibers enter the spinal cord dorsal horn result in a much better pain-relieving effect than SCS at more rostral levels. From this we conclude that SCS in treatment of neuropathic pain acts through a segmental spinal site of action. [Copyright &y& Elsevier]

Abstract: Maternal methamphetamine (MA) abuse during pregnancy has been proved to induce various impacts on the development of infant and child. In this study, we examined whether prenatal exposure to MA would affect the development of nociceptive system by measuring the responses to noxious stimulation in the developing rat. Adult female Sprague–Dawley rats received bi-daily subcutaneous injection of methamphetamine (5mg/kg) or isovolumetric normal saline since the day of mating till the day of delivery. Birth profiles of the offspring including birth length, weight, and body temperature were recorded during the first postnatal month. Mechanical withdrawal thresholds were measured by von Frey filaments on postnatal day (PND) 30 and 60, and hyperalgesic behaviors following plantar formalin injection (2%, 50μl) were evaluated on PND 60. The birth body weight and length of rats born to MA-injected dam rats (MA group) were significantly lower than those of the control rats during the first postnatal month; however, their body temperature was significantly higher than those of the control rats during the first 3 days after birth. The MA group rats had significantly lower tactile withdrawal values in von Frey test and higher pain scores in the late phase of pain in the formalin test than those of the control rats. There is a gender difference of nociceptive hypersensitivity manifested as that the female MA group rats had significantly lower withdrawal thresholds and higher pain scores in response to formalin injection than the male MA group rats. These results suggest that prenatal MA exposure could predispose an alteration in the development of nociceptive neuronal network, which leads to a long-lasting status of hypersensitivity to pain stimulations in the offspring. [Copyright &y& Elsevier]

Abstract: Following spinal cord injury (SCI), individuals lose normal sensation and often develop debilitating neuropathic pain. Basic research has helped to elucidate many of the underlying mechanisms, but unanswered questions remain concerning how sensation changes after SCI and potential negative consequences of regenerative therapies. Mouse models provide an opportunity to explore these questions using genetic markers and manipulations. However, despite the increasing use of mice in pain and sensory research, the responses to sensory stimuli after SCI are poorly characterized in this species. This study evaluated behavioral responses to mechanical and nociceptive stimuli applied to the hindlimbs and the dorsal trunk in C57BL/6 mice after mid-thoracic SCI. Adult mice were subjected to laminectomy, contusion injuries of different severities, or complete transections to test the hypothesis that the patterns of sensory pathology depend on the extent of tissue damage at the injury site. In the hind paws, hyper-responsiveness to a heat stimulus developed independent of injury severity, while mechanical sensitivity decreased, except after the most severe contusion injuries sparing less than 2% of the white matter at the injury site, when enhanced sensitivity was observed. On the trunk, mechanical and pin prick testing revealed diminished sensitivity at and below the injury level, while responses above the level of the injury were unchanged. The contrast in injury severity threshold for thermal and mechanical hypersensitivity in the hind paws suggests that these responses have different underlying mechanisms. These results establish essential baseline information for murine studies of pain and changes in sensation after SCI. [Copyright &y& Elsevier]

Abstract: The CatWalk gait analysis system has recently been suggested as a rapid and objective alternative method over the von Frey test to assess mechanical allodynia in chronic neuropathic pain models. Our results demonstrate that no correlation exists between the development of mechanical allodynia and changes in CatWalk-gait parameters in a chronic inflammatory pain model. Hence, the use of the CatWalk in assessment of experimental chronic pain is discussed. [Copyright &y& Elsevier]

Abstract: We describe a device which allows the mechanical sensitivity of trigeminovascular sensory neurons to be monitored over extended time periods. The device can be used to stimulate either the skin or dura mater and consists of a solenoid-driven plunger to which are fixed interchangeable von Frey hairs. The solenoid can be attached to a stereotaxic carrier and mounted on a stereotaxic frame to allow precise positioning over the receptive field. The device is driven from the synchronization signals of a standard stimulator via TTL circuitry and a relay driver, to allow stimulation by a single or multiple stimuli. The advantages of the device over manual stimulation include the reproducibility of the site of stimulation; the ability to apply a known force for a known time; the ability to measure response latencies to millisecond precision and to compare them to latencies to other stimuli and; easy interface with computer-control. We discuss some of the drawbacks of the von Frey system as usually used and illustrate the use of the new device with results from experiments on peripherally induced sensitization. [Copyright &y& Elsevier]

Abstract: Tricyclic antidepressants (TCAs) are an important analgesic treatment for neuropathic pain, though the neural substrates mediating these effects are poorly understood. We have used an integrative approach combining behavioural pharmacology with functional magnetic resonance imaging (fMRI) to investigate the effects of chronic treatment with the TCA desipramine, on touch-evoked pain (mechanical allodynia) and brain regional activity in the selective spinal nerve ligation (SNL) model of neuropathic pain. SNL and sham-operated rats received once daily i.p. administration of 10 mg/kg DMI, or saline, for 14 days. Withdrawal responses to the application of a normally non-noxious (10 g) stimulus were recorded in SNL and sham-operated rats over this period. On the final day of the study, SNL and sham-operated rats received a final challenge dose of DMI (10 mg/kg i.p.) during fMRI scanning. Chronic administration of desipramine (DMI) significantly attenuated mechancial allodynia in SNL rats. DMI challenge in chronic DMI-treated neuropathic rats produced significantly greater activation of the deep mesencephalic nucleus, primary somatosensory cortex, insular cortex, medial globus pallidus, inferior colliculus, perirhinal cortex and cerebellum compared to sham-operated rats and saline controls. By contrast, the spatial pattern of brain regional activation by chronic DMI treatment in sham controls encompassed a number of other areas including those associated with learning and memory processes. These novel findings identify key brain regions implicated in the analgesic and mood altering effects associated with chronic treatment with DMI. [Copyright &y& Elsevier]

Abstract: Spinal cord stimulation (SCS) is an established treatment for chronic neuropathic pain. However, in recent studies conflicting results regarding the effect of SCS were noted in a selected group of patients suffering from complex regional pain syndrome and mechanical allodynia. In the present study we investigated the pain relieving effect of SCS in a rat experimental model of neuropathic pain as related to the severity of mechanical allodynia. Adult male rats (n=45) were submitted to a unilateral sciatic nerve ligation. The level of allodynia was tested using the withdrawal response to tactile stimuli with the von Frey test. A portion of these rats developed marked tactile hypersensitivity in the nerve-lesioned paw (von Frey test), similar to “tactile allodynia” observed after nerve injury in humans. Prior to SCS treatment the rats were subdivided into three groups based on the level of allodynia: mild, moderate and severe. All allodynic rats were treated with SCS (n=27) for 30 min (f=50 Hz; pulse width 0.2 ms and stimulation at 2/3 of motor threshold) at 16 days post-injury. Our data demonstrate a differential effect of SCS related to the severity of the mechanical allodynia. SCS leads to a faster and better pain relief in mildly allodynic rats as compared with the more severely allodynic rats. Thus, we suggest that the selection and subdivision of patient groups similar to those defined in our experimental setting (mild, moderate and severe allodynic) may provide better pre-treatment prediction of possible therapeutic benefits of SCS. [Copyright &y& Elsevier]

Abstract: Secondary hyperalgesia in the monoarthritic rat is accompanied by a decrease in nociceptive activation of spinal neurons expressing GABAB receptors and by the opposite effect in the cells expressing neurokinin 1 (NK1)-receptors. In order to ascertain the relative role of each receptor, the effects of intrathecal administration of SP-saporin (SP-SAP), baclofen or both were evaluated, using a model of secondary hyperalgesia that consists of mechanical stimulation of the hindlimb skin close to an inflamed joint. Four days after the induction of monoarthritis by intraarticular injection of Complete Freund’s Adjuvant (CFA), a cannula was implanted at T13–L1 level and 10 μl of saline or SP-SAP (10−6 M) were intrathecally (i.t.) injected. Fourteen days after CFA-injection, half of the animals from each group received i.t. injections of 10 μl saline and the remainder were injected with the same volume of baclofen (1 μg). Ten minutes later, the animals were behaviorally evaluated by the von Frey test or submitted to noxious mechanical stimulation to analyze c-fos expression. The von Frey thresholds increased after the treatments, but more pronouncedly after baclofen or SP-SAP plus baclofen. In segments L2–L3, the spinal area that receives input from the stimulated skin close to the inflamed joint, the numbers of Fos-immunoreactive neurons were reduced after the three treatments both in the superficial and deep dorsal horn. In segments T13–L1, the numbers of Fos-immunoreactive neurons were significantly reduced after treatment with SP-SAP plus baclofen in both dorsal horn regions, and in the deep dorsal horn after baclofen treatment. We conclude that both GABAB and NK1 receptors of spinal dorsal horn neurons participate in secondary hyperalgesia in the monoarthritic rat, although the decrease in GABA inhibition appears to play a more important role than the increase in SP-mediated effects. [Copyright &y& Elsevier]

This study examined the time course of mechanical and cold allodynia in rat hindpaw after spinal cord contusion. Hindpaw withdrawal threshold to graded von Frey hair stimulation and withdrawal frequency to acetone application were measured in rats subjected to contusions of varying severity, produced by a MASCIS impactor device with a 10 g weight dropped from 6.25, 12.5, or 25 mm. Mechanical and cold allodynia developed following the injury, and differences in the incidence of allodynia and in withdrawal threshold were significant among these groups. The least severe injury (6.25 mm) most consistently caused a decreased hindpaw threshold to mechanical stimulation and an increased withdrawal frequency to cold. [ABSTRACT FROM AUTHOR]

Locomotor and/or sensory behaviour is commonly evaluated in laboratory rats in the field of neuroscience. Many strains of rats, however, have been propagated through intensive breeding programs. With any breeding program, traits are selected purposefully or inadvertently. We set out to investigate whether differences in morphology, sensory or motor behaviours exist using five age-matched strains of laboratory rats. Personal observations of morphological differences between different strains of rats led us to hypothesize that Fischer rats were dissimilar to the other strains in each of the parameters investigated. Evaluation of morphology involved measuring long-bone lengths and body weights of each strain. Motor skills were evaluated by measuring paw preferences while rearing, abduction of the distal portion of hindlimbs during locomotion, footfalls through a horizontal ladder during locomotion, and ground reaction forces generated during trotting. Sensory ability was assessed by von Frey testing. Fischer rats had shorter long-bone lengths, weighed less, and had significantly abducted distal portion of their hindlimbs during locomotion compared to the other strains. Lewis and Sprague–Dawley rats were less sensitive to mechanical pedal stimulation compared to Fischer rats. While rearing, all strains of rats tended to use individual forelimbs 25% of the time for each right and left limbs, and both forelimbs together 50% of the time. There were no significant differences in the number of footfalls during the ladder task. Ground reaction force determination revealed that Fischer and Sprague–Dawley rats bore more weight on their hindlimbs compared to forelimbs during locomotion, Long–Evans and Lewis rats bore more weight on their forelimbs compared to their hindlimbs, while Wistar rats distributed weight evenly between forelimbs and hindlimbs during trotting. We conclude that morphologic, sensory and motor differences exist between the five strains of laboratory rats examined and several of these differences are most pronounced in the Fischer strain. [Copyright &y& Elsevier]

A characteristic symptom of neuropathic pain is mechanical allodynia. In animal models of neuropathic pain, mechanical allodynia is often assessed using von Frey filaments. Although the forces applied with these filaments are highly reproducible, there are various disadvantages of using this method. Testing paradigms and definitions of withdrawal threshold are not standardised. Moreover, measurements may be influenced by various conditions, such as ambient temperature, humidity, weight bearing of the limb and stress. We have therefore investigated another technique to assess mechanical allodynia, the ‘CatWalk’ automated quantitative gait analysis. With this computer-assisted method of locomotor analysis, it is possible to objectively and rapidly quantify several gait parameters, including duration of different phases of the step cycle and pressure applied during locomotion. We tested rats with a chronic constriction injury of the sciatic nerve, a model of neuropathic pain, both with von Frey filaments and the CatWalk method. We demonstrate that these rats minimise contact with the affected paw during locomotion, as demonstrated by a reduction in stance phase and pressure applied during stance. Moreover, these parameters show a high degree of correlation with mechanical withdrawal thresholds as determined by von Frey filaments. We therefore suggest that the CatWalk method might serve as an additional tool in the investigation of mechanical allodynia. [Copyright &y& Elsevier]

Chronic pain is a persistent and debilitating health problem. Although the use of analgesics such as opioids is useful in mitigating pain, their prolonged use is associated with unwanted effects including abuse liability. This study assesses the antinociceptive effect of combining subtherapeutic doses of two opioids (morphine or tramadol) with the synthetic cannabinoid CP55940 (2-[(1R,2R,5R)-5-hydroxy-2-(3-hydroxypropyl)cyclohexyl]-5-(2-methyloctan -2-yl)phenol). It also evaluates the associated adverse effects of these drugs and combinations. Adult male rats were injected with intraplantar complete Freund's adjuvant (CFA) to produce mechanical allodyia. Antinociceptive effect of morphine, tramadol, the synthetic cannabinoid CP55940, or their combinations was evaluated three to nine days post-CFA injections. Intracranial self-stimulation (ICSS) was utilized to evaluate the abuse liability of these drugs or their combinations. All drugs alone produced a dose-dependent antinociceptive effect. Morphine produced minimal effect on ICSS, but both tramadol and CP55940 produced dose-dependent depression of ICSS. Morphine at a dose of 0.32 mg/kg enhanced the antinociceptive effects of CP55940, in that, CP55940 produced antinociception at a lower dose (0.1 mg/kg) when compared to the vehicle. The aforementioned combinations did not change CP55940-induced depression of ICSS. On the other hand, tramadol failed to enhance the antinociceptive effect of CP55940. Our data suggest that combining CP55940 with morphine, but not tramadol, shows a better antinociceptive profile with no additional risk of abuse liability, which represents a potential pain management approach. [ABSTRACT FROM AUTHOR]

Capparis erythrocarpos is a medicinal plant used widely in many parts of Africa for the management of pain and inflammatory conditions such as rheumatoid arthritis. Its wide range of use, popularity and high value, make C. erythrocarpos containing products a target for economically driven adulteration. This is made worse, by the use of roots, which are unsustainable plant parts. In addition, the mechanism of anti-inflammation is not clearly understood. Therefore, this study comparatively evaluated the anti-arthritic and analgesic effects of the leaves, stems and roots of C. erythrocarpos, while elucidating the mechanism of anti-inflammation. Using the complete Freund's adjuvant arthritis model, the antiathritic effects were evaluated. The analgesic effects were determined by measuring responses to Von Frey filament number 9. Effects of C. erythrocarpos extracts on the levels of Interleukin-6 (IL-6) and Tumor Necrosis Factor-alpha (TNF-α) were determined using ELISA. Haematological and serum biochemical assays were also carried out. The leaf, stem and root extracts significantly reduced paw volumes with ED 50 values (mg/kg) of 182.5, 181.5 and 36.4 respectively. The leaf extract at a dose of 100 mg/kg showed substantial analgesic activity with a decrease in the percentage response to Von Frey filament 9. However, there was no significant difference in activities of the leaf, stem and root extracts. Results from ELISA assays show that lower doses of the stem extracts reduce levels of IL-6. Lower doses of all extracts also reduce TNF-α levels. Haematological analysis showed extracts reversed elevated WBC and platelet levels. Toxicity evaluation with kidney and liver function tests indicated no significant differences between the treatment and control groups. The leaves and stems, just as the roots, have antiarthritic and analgesic effects and can be used as more sustainable alternatives to the roots. This will support the continuous growth of the industry that has developed around C. erythrocarpos. Image 1 [ABSTRACT FROM AUTHOR]