Inhibition of P2X7 receptors by Lu AF27139 diminishes colonic hypersensitivity and CNS prostanoid levels in a rat model of visceral pain.

Visceral pain is a prominent feature of various gastrointestinal diseases. The P2X7 receptor is expressed by multiple cell types including dorsal root ganglion satellite glial cells, macrophages, and spinal microglia, all of which have been implicated in nociceptive sensitization. We have used the selective and CNS penetrant P2X7 receptor antagonist Lu AF27139 to explore this receptor's role in distinct rat models of inflammatory and visceral hypersensitivity. Rats injected with CFA in the hindpaw displayed a marked reduction in hindpaw mechanical threshold, which was dose-dependently reversed by Lu AF27139 (3-30 mg/kg, p.o.). In rats injected with TNBS in the proximal colon, the colorectal distension threshold measured distally was significantly lower than sham treated rats at 7 days post-injection (P < 0.001), indicative of a marked central sensitization. Colonic hypersensitivity was also reversed by Lu AF27139 (10-100 mg/kg) and by the κ-opioid receptor agonist U-50,488H (3 mg/kg, s.c.). Moreover, both Lu AF27139 and U-50,488H prevented a TNBS-induced increase in spinal and brain levels of PGE2 and LTB4, as well as an increase in brain levels of PGF2α and TXB2. Lu AF27139 was well tolerated as revealed by a lack of significant effect on rotarod motor function and coordination at all doses tested up to 300 mg/kg. Thus, P2X7 receptor antagonism is efficacious in a rat model of visceral pain, via a mechanism which potentially involves attenuation of microglial function within spinal and/or supraspinal pain circuits, albeit a peripheral site of action cannot be excluded.

ATP sensitive potassium (KATP) channel inhibition: A promising new drug target for migraine.

BACKGROUND: Recently, the adenosine triphosphate (ATP) sensitive potassium channel opener levcromakalim was shown to induce migraine attacks with a far higher incidence than any previous provoking agent such as calcitonin gene-related peptide. Here, we show efficacy of ATP sensitive potassium channel inhibitors in two validated rodent models of migraine. METHODS: In female spontaneous trigeminal allodynic rats, the sensitivity of the frontal region of the head was tested by an electronic von Frey filament device. In mice, cutaneous hypersensitivity was induced by repeated glyceryl trinitrate or levcromakalim injections over nine days, as measured with von Frey filaments in the hindpaw. Release of calcitonin gene-related peptide from dura mater and trigeminal ganglion was studied ex vivo. RESULTS: The ATP sensitive potassium channel inhibitor glibenclamide attenuated the spontaneous cephalic hypersensitivity in spontaneous trigeminal allodynic rats and glyceryl trinitrate-induced hypersensitivity of the hindpaw in mice. It also inhibited CGRP release from dura mater and the trigeminal ganglion isolated from spontaneous trigeminal allodynic rats. The hypersensitivity was also diminished by the structurally different ATP sensitive potassium channel inhibitor gliquidone. Mice injected with the ATP sensitive potassium channel opener levcromakalim developed a progressive hypersensitivity that was completely blocked by glibenclamide, confirming target engagement. CONCLUSION: The results suggest that ATP sensitive potassium channel inhibitors could be novel and highly effective drugs in the treatment of migraine.

Targeting CGRP via receptor antagonism and antibody neutralisation in two distinct rodent models of migraine-like pain.

INTRODUCTION: Rodent disease models can play an indispensable role in drug development. Confirming that translationally-relevant disease mechanisms are engaged in such models is a crucial facet of this process. Accordingly, we have validated the role of calcitonin gene-related peptide signaling in a mouse model of glyceryl trinitrate-provoked migraine-like pain and a spontaneous rat model of migraine-like pain by assessing their pharmacological responsiveness to the small molecule calcitonin gene-related peptide receptor antagonist olcegepant, and the humanised monoclonal calcitonin gene-related peptide antibody ALD405. METHODS: Cutaneous sensitivity to hind paw, and periorbital mechanical stimulation were used as surrogate markers of activation of relevant pain pathways in each respective model. Separate experiments were performed to identify the time-course of treatment response to olcegepant (1 mg/kg i.p.) and ALD405 (10 mg/kg i.p.). RESULTS: Olcegepant and ALD405 significantly alleviated cutaneous mechanical hypersensitivity in both models compared with corresponding control treatments (saline and IgG control antibody respectively). As expected, the duration of anti-nociceptive action obtained with ALD405 was considerably longer than that associated with olcegepant. Surprisingly, in the spontaneous rat model the onset of action of ALD405 occurred within just 4 hours after administration. DISCUSSION: The current data clearly show that calcitonin gene-related peptide-mediated signaling is critically involved in the manifestation of cutaneous hypersensitivity in distinct rodent models of migraine-like pain and emphasise their translational relevance. Moreover, the unexpected rapidity of onset observed for ALD405 supports i) a probable site of action outside the blood-brain barrier, and ii) a potential clinical utility of specific monoclonal calcitonin gene-related peptide antibodies in the abortive treatment of migraine.

Pharmacokinetic/Pharmacodynamic Relationship of Gabapentin in a CFA-induced Inflammatory Hyperalgesia Rat Model.

PURPOSE: Gabapentin displays non-linear drug disposition, which complicates dosing for optimal therapeutic effect. Thus, the current study was performed to elucidate the pharmacokinetic/pharmacodynamic (PKPD) relationship of gabapentin's effect on mechanical hypersensitivity in a rat model of CFA-induced inflammatory hyperalgesia. METHODS: A semi-mechanistic population-based PKPD model was developed using nonlinear mixed-effects modelling, based on gabapentin plasma and brain extracellular fluid (ECF) time-concentration data and measurements of CFA-evoked mechanical hyperalgesia following administration of a range of gabapentin doses (oral and intravenous). RESULTS: The plasma/brain ECF concentration-time profiles of gabapentin were adequately described with a two-compartment plasma model with saturable intestinal absorption rate (K m = 44.1 mg/kg, V max = 41.9 mg/h∙kg) and dose-dependent oral bioavailability linked to brain ECF concentration through a transit compartment. Brain ECF concentration was directly linked to a sigmoid E max function describing reversal of hyperalgesia (EC 50, plasma = 16.7 µg/mL, EC 50, brain = 3.3 µg/mL). CONCLUSIONS: The proposed semi-mechanistic population-based PKPD model provides further knowledge into the understanding of gabapentin's non-linear pharmacokinetics and the link between plasma/brain disposition and anti-hyperalgesic effects. The model suggests that intestinal absorption is the primary source of non-linearity and that the investigated rat model provides reasonable predictions of clinically effective plasma concentrations for gabapentin.

Finding new analgesics: Computational pharmacology faces drug discovery challenges.

Despite the worldwide prevalence and huge burden of pain, pain is an undertreated phenomenon. Currently used analgesics have several limitations regarding their efficacy and safety. The discovery of analgesics possessing a novel mechanism of action has faced multiple challenges, including a limited understanding of biological processes underpinning pain and analgesia and poor animal-to-human translation. Computational pharmacology is currently employed to face these challenges. In this review, we discuss the theory, methods, and applications of computational pharmacology in pain research. Computational pharmacology encompasses a wide variety of theoretical concepts and practical methodological approaches, with the overall aim of gaining biological insight through data acquisition and analysis. Data are acquired from patients or animal models with pain or analgesic treatment, at different levels of biological organization (molecular, cellular, physiological, and behavioral). Distinct methodological algorithms can then be used to analyze and integrate data. This helps to facilitate the identification of biological molecules and processes associated with pain phenotype, build quantitative models of pain signaling, and extract translatable features between humans and animals. However, computational pharmacology has several limitations, and its predictions can provide false positive and negative findings. Therefore, computational predictions are required to be validated experimentally before drawing solid conclusions. In this review, we discuss several case study examples of combining and integrating computational tools with experimental pain research tools to meet drug discovery challenges.

A novel reuptake inhibitor, IP2015, induces erection by increasing central dopamine and peripheral nitric oxide release.

BACKGROUND AND PURPOSE: An estimated 40% of patients with erectile dysfunction have a poor prognosis for improvement with currently available treatments. The present study investigated whether a newly developed monoamine transport inhibitor, IP2015, improves erectile function. EXPERIMENTAL APPROACH: We investigated the effects of IP2015 on monoamine uptake and binding, erectile function in rats and diabetic mice and the effect on corpus cavernosum contractility. KEY RESULTS: IP2015 inhibited the uptake of 5-HT, noradrenaline and dopamine by human monoamine transporters expressed in cells and in rat brain synaptosomes. Intracavernosal pressure measurement in anaesthetized rats revealed that IP2015 dose-dependently increased the number and the duration of spontaneous erections. Whereas pretreatment with the dopamine D2-like receptor antagonists, clozapine and (-)-sulpiride, or cutting the cavernosal nerve inhibited IP2015-induced erectile responses, the phosphodiesterase type 5 inhibitor sildenafil further enhanced the IP2015-mediated increase in intracavernosal pressure. IP2015 also increased the number of erections in type 2 diabetic db/db mice. Direct intracavernosal injection of IP2015 increased penile pressure, and in corpus cavernosum strips, IP2015 induced concentration-dependent relaxations. These relaxations were enhanced by sildenafil and blunted by endothelial cell removal, a nitric oxide synthase inhibitor, NG-nitro-l-arginine and a D1-like receptor antagonist, SCH23390. Quantitative polymerase chain reaction (qPCR) showed the expression of the dopamine transporter in the rat corpus cavernosum. CONCLUSION AND IMPLICATIONS: Our findings suggest that IP2015 stimulates erectile function by a central mechanism involving dopamine reuptake inhibition and direct NO-mediated relaxation of the erectile tissue. This novel multi-modal mechanism of action could offer a new treatment approach to erectile dysfunction.

"Of mice and men": the relevance of Cometin and Erythropoietin origin for its effects on murine spiral ganglion neuron survival and neurite outgrowth in vitro.

Neurotrophic factors (NTF) play key roles in the survival of neurons, making them promising candidates for therapy of neurodegenerative diseases. In the case of the inner ear, sensorineural hearing loss (SNHL) is characterized over time by a degeneration of the primary auditory neurons, the spiral ganglion neurons (SGN). It is well known that selected NTF can protect SGN from degeneration, which positively influences the outcome of cochlear implants, the treatment of choice for patients with profound to severe SNHL. However, the outcome of studies investigating protective effects of NTF on auditory neurons are in some cases of high variability. We hypothesize that the factor origin may be one aspect that affects the neuroprotective potential. The aim of this study was to investigate the neuroprotective potential of human and mouse Erythropoietin (EPO) and Cometin on rat SGN. SGN were isolated from neonatal rats (P 2-5) and cultured in serum-free medium. EPO and Cometin of mouse and human origin were added in concentrations of 0.1, 1, and 10 ng/mL and 0.1, 1, and 10 µg/mL, respectively. The SGN survival rate and morphology, and the neurite outgrowth were determined and compared to negative (no additives) and positive (brain-derived neurotrophic factor, BDNF) controls. A neuroprotective effect of 10 µg/mL human Cometin comparable to that obtained with BDNF was observed in the SGN-culture. In contrast, mouse Cometin was ineffective. A similar influence of 10 µg/mL human and mouse and 1 µg/mL human Cometin on the length of regenerated neurites compared to BDNF was also detected. No other Cometin-conditions, and none of the EPO-conditions tested had neuroprotective or neuritogenic effects or influenced the neuronal morphology of the SGN. The neuroprotective effect of 10 µg/mL human Cometin on SGN indicates it is a potentially interesting protein for the supportive treatment of inner ear disorders. The finding that mouse Cometin had no effect on the SGN in the parallel-performed experiments underlines the importance of species origin of molecules being screened for therapeutic purpose.

Meteorin Alleviates Paclitaxel-Induced Peripheral Neuropathic Pain in Mice.

Chemotherapy-induced peripheral neuropathy is a challenging condition to treat, and arises due to severe, dose-limiting toxicity of chemotherapeutic drugs such as paclitaxel. This often results in debilitating sensory and motor deficits that are not effectively prevented or alleviated by existing therapeutic interventions. Recent studies have demonstrated the therapeutic effects of Meteorin, a neurotrophic factor, in reversing neuropathic pain in rodent models of peripheral nerve injury induced by physical trauma. Here, we sought to investigate the potential antinociceptive effects of recombinant mouse Meteorin (rmMeteorin) using a paclitaxel-induced peripheral neuropathy model in male and female mice. Paclitaxel treatment (4 × 4 mg/kg, i.p.) induced hind paw mechanical hypersensitivity by day 8 after treatment. Thereafter, in a reversal dosing paradigm, five repeated injections of rmMeteorin (.5 and 1.8 mg/kg s.c. respectively) administered over 9 days produced a significant and long-lasting attenuation of mechanical hypersensitivity in both sexes. Additionally, administration of rmMeteorin ( .5 and 1.8 mg/kg), initiated before and during paclitaxel treatment (prevention dosing paradigm), reduced the establishment of hind paw mechanical hypersensitivity. Repeated systemic administration of rmMeteorin in both dosing paradigms decreased histochemical signs of satellite glial cell reactivity as measured by glutamine synthetase and connexin 43 protein expression in the dorsal root ganglion. Additionally, in the prevention administration paradigm rmMeteorin had a protective effect against paclitaxel-induced loss of intraepidermal nerve fibers. Our findings indicate that rmMeteorin has a robust and sustained antinociceptive effect in the paclitaxel-induced peripheral neuropathy model and the development of recombinant human Meteorin could be a novel and effective therapeutic for chemotherapy-induced peripheral neuropathy treatment. PERSPECTIVE: Chemotherapy neuropathy is a major clinical problem that decreases quality of life for cancer patients and survivors. Our experiments demonstrate that Meteorin treatment alleviates pain-related behaviors, and signs of neurotoxicity in a mouse model of paclitaxel neuropathy.

P2X7 receptor-mediated release of microglial prostanoids and miRNAs correlates with reversal of neuropathic hypersensitivity in rats.

BACKGROUND: P2X7 receptor antagonists have potential for treating various central nervous system (CNS) diseases, including neuropathic pain, although none have been approved for clinical use. Reasons may include insufficient understanding of P2X7 receptor signalling in pain, and the lack of a corresponding preclinical mechanistic biomarker. METHODS: Lu AF27139 is a highly selective and potent small molecule antagonist at rat, mouse and human forms of the P2X7 receptor, with excellent pharmacokinetic and CNS permeability properties. In the current experiments, we probed the utility of previously characterized and novel signalling cascades exposed to Lu AF27139 using cultured microglia combined with release assays. Subsequently, we assessed the biomarker potential of identified candidate molecules in the rat chronic constriction injury (CCI) model of neuropathic pain; study design limitations precluded their assessment in spared nerve injury (SNI) rats. RESULTS: Lu AF27139 blocked several pain-relevant pathways downstream of P2X7 receptors in vitro. At brain and spinal cord receptor occupancy levels capable of functionally blocking P2X7 receptors, it diminished neuropathic hypersensitivity in SNI rats, and less potently in CCI rats. Although tissue levels of numerous molecules previously linked to neuropathic pain and P2X7 receptor function (e.g. IL-6, IL-1ß, cathepsin-S, 2-AG) were unaffected by CCI, Lu AF27139-mediated regulation of spinal PGE2 and miRNA (e.g. rno-miR-93-5p) levels increased by CCI aligned with its ability to diminish neuropathic hypersensitivity. CONCLUSIONS: We have identified a pain-relevant P2X7 receptor-regulated mechanism in neuropathic rats, which could hold promise as a translatable biomarker and by association enhance the clinical progression of P2X7 receptor antagonists in neuropathic pain. SIGNIFICANCE: Sub-optimal translation of preclinical molecules has hindered the clinical development of novel mechanism of action analgesics. We have undertaken a comprehensive in vitro analysis of migroglial signalling mechanisms recruited upon P2X7 receptor activation, a number of which were shown to be modulated by a selective P2X7 receptor antagonist in a well characterized animal model of neuropathic pain. Subject to further confirmation in other neuropathic models, this opens up the possibility to investigate their clinical utility as potential pain biomarkers in patients.

Preclinical target validation for non-addictive therapeutics development for pain.

INTRODUCTION: The Helping to End Addiction Long-termSM Initiative supports a wide range of programs to develop new or improved prevention and opioid addiction treatment strategies. An essential component of this effort is to accelerate development of non-opioid pain therapeutics. In all fields of medicine, therapeutics development is an arduous process and late-stage translational efforts such as clinical trials to validate targets are particularly complex and costly. While there are plentiful novel targets for pain treatment, successful clinical validation is rare. It is therefore crucial to develop processes whereby therapeutic targets can be reasonably 'de-risked' prior to substantial late-stage validation efforts. Such rigorous validation of novel therapeutic targets in the preclinical space will give potential private sector partners the confidence to pursue clinical validation of promising therapeutic concepts and compounds. AREAS COVERED: In 2020, the National Institutes of Health (NIH) held the Target Validation for Non-Addictive Therapeutics Development for Pain workshop to gather insights from key opinion leaders in academia, industry, and venture-financing. EXPERT OPINION: The result was a roadmap for pain target validation focusing on three modalities: 1) human evidence; 2) assay development in vitro; 3) assay development in vivo.

The role of GABA(A) receptor subtypes as analgesic targets.

Recent GABA(A) receptor drug discovery efforts have culminated in the development of transgenic mice and subtype-selective pharmacological tools, enhancing our understanding of the major inhibitory neural system in the central nervous system. Notably, subtype-selective tools have demonstrated in both preclinical studies and, to some extent, in man that it is possible to develop drugs that share the clinical benefits of benzodiazepines (e.g., anxiolysis) while obviating some adverse effects of these clinically important drugs. Here, we highlight chronic pain as another therapeutic area in which subtype-selective GABA(A) receptor drugs might have clinical utility. Specifically, based on research in animal models of inflammatory/neuropathic pain, we suggest that subtype-selective positive modulators of GABA(A) alpha(2/3) receptors might reverse a loss of postsynaptic GABA(A) receptor-mediated inhibitory function in spinal cord, leading to analgesia. However, alteration of presynaptic inhibitory neural transmission in chronic pain suggests that drugs that negatively modulate GABA(A) receptors might also be effective analgesics. For example, the non-selective negative allosteric modulator FG-7142 reverses allodynia in an animal model of neuropathic pain. Importantly, these two mechanisms are not mutually exclusive. Further clinical exploration in pain of available positive and negative subtype-selective modulators that have been administered to humans would considerably aid back translation, allowing for improved therapeutic development.

The influence of rat strain on the development of neuropathic pain and comorbid anxio-depressive behaviour after nerve injury.

Back-translating the clinical manifestations of human disease burden into animal models is increasingly recognized as an important facet of preclinical drug discovery. We hypothesized that inbred rat strains possessing stress hyper-reactive-, depressive- or anxiety-like phenotypes may possess more translational value than common outbred strains for modeling neuropathic pain. Rats (inbred: LEW, WKY, F344/ICO and F344/DU, outbred: Crl:SD) were exposed to Spared Nerve Injury (SNI) and evaluated routinely for 6 months on behaviours related to pain (von Frey stimulation and CatWalk-gait analysis), anxiety (elevated plus maze, EPM) and depression (sucrose preference test, SPT). Markers of stress reactivity together with spinal/brain opioid receptor expression were also measured. All strains variously developed mechanical allodynia after SNI with the exception of stress-hyporesponsive LEW rats, despite all strains displaying similar functional gait-deficits after injury. However, affective changes reflective of anxiety- and depressive-like behaviour were only observed for F344/DU in the EPM, and for Crl:SD in SPT. Although differences in stress reactivity and opioid receptor expression occurred, overall they were relatively unaffected by SNI. Thus, anxio-depressive behaviours did not develop in all strains after nerve injury, and correlated only modestly with degree of pain sensitivity or with genetic predisposition to stress and/or affective disturbances.

Workplace responsibility, stress, alcohol availability and norms as predictors of alcohol consumption-related problems among employed workers.

The objectives of this study were to determine the prevalence of alcohol use and problems among employed individuals in Alberta, Canada (N = 1,890), and to conduct a multivariate examination of predictors of alcohol consumption-related problems. General alcohol problems were identified by 10%, although very few workers described any specific work-related alcohol problems (1%). Structural equation modeling revealed that, as hypothesized, workplace alcohol availability predicted general alcohol problems. Job responsibility and workplace norms also predicted alcohol problems but only for men. Perceived work stress did not predict alcohol problems. Results support the development of interventions that focus on re-shaping alcohol use norms.

Stress sensitivity and cutaneous sensory thresholds before and after neuropathic injury in various inbred and outbred rat strains.

Chronic pain is associated with altered affective state, stress, anxiety and depression. Conversely, stress, anxiety and depression can all modulate pain perception. The relative link between these behavioural constructs in different inbred and outbred rat strains, known to be variously hypo/hyperresponsive to stress has not been determined. Hindpaw sensory thresholds to repeated mechanical (von Frey filament and electronic Randall Selitto) and thermal (Hargreaves, cold plate and hot plate) stimulation were routinely assessed over three weeks in non-injured male rats of the following strains; WKY, LEW, F344, Hsd:SD and Crl:SD. Thereafter, threshold responses to Spared Nerve Injury (SNI) were assessed using von Frey, pin prick and Hargreaves testing in the same strains over a three month period. Finally, anxiolytic efficacy of the benzodiazepine drug diazepam was assessed using the Elevated Plus Maze (EPM), as a surrogate index of functional plasticity of circuits involved in affective processing. Repeated nociceptive testing was associated with distinct strain-dependent changes in sensory thresholds in naïve rats; stress-hyporesponsive LEW rats presented with a mechanical/thermal hyperalgesia phenotype, whereas stress-hyperresponsive WKY rats presented with an unexpected heat/cold hypoalgesia phenotype. After SNI, LEW rats showed minimal signs of neuropathic sensitivity. Diazepam was anxiolytic in all tested strains with the exception of LEW rats reflecting distinct inherent affective processing only in this strain. The contribution of stress reactivity to nociceptive sensory profiles appears to vary in the absence or presence of neuropathic injury. Intriguingly, the functional responsiveness of affective state prior to injury may be a predisposing factor to developing chronic pain.

Antihyperalgesic effects of Meteorin in the rat chronic constriction injury model: a replication study.

Data from preclinical research have been suggested to suffer from a lack of inherent reproducibility across laboratories. The goal of our study was to replicate findings from a previous report that demonstrated positive effects of Meteorin, a novel neurotrophic factor, in a rat model of neuropathic pain induced by chronic constriction injury (CCI). Notably, 5 to 6 intermittent subcutaneous (s.c.) injections of Meteorin had been reported to produce reversal of mechanical allodynia/thermal hyperalgesia after injury, wherein maximum efficacy of Meteorin was reached slowly and outlasted the elimination of the compound from the blood by several weeks. Here, we evaluated the efficacy of Meteorin in reversing hindpaw mechanical hyperalgesia and cold allodynia in male, Sprague-Dawley rats with CCI. Nociceptive behavior was monitored before and after CCI, and after drug treatment until day 42 after injury. Systemic administration of recombinant mouse Meteorin (0.5 and 1.8 mg/kg, s.c.) at days 10, 12, 14, 17, and 19 after CCI produced a prolonged reversal of neuropathic hypersensitivity with efficacy comparable with that obtained with gabapentin (100 mg/kg, orally). Despite some protocol deviations (eg, nociceptive endpoint, animal vendor, testing laboratory, investigator, etc.) being incurred, these did not affect study outcome. By paying careful attention to key facets of study design, using bioactive material, and confirming drug exposure, the current data have replicated the salient findings of the previous study, promoting confidence in further advancement of this novel molecule as a potential therapy for neuropathic pain.

The analgesic efficacy of morphine varies with rat strain and experimental pain model: implications for target validation efforts in pain drug discovery.

BACKGROUND: Translating efficacy of analgesic drugs from animal models to humans remains challenging. Reasons are multifaceted, but lack of sufficiently rigorous preclinical study design criteria and phenotypically relevant models may be partly responsible. To begin to address this fundamental issue, we assessed the analgesic efficacy of morphine in three inbred rat strains (selected based on stress reactivity and affective/pain phenotypes), and outbred Sprague Dawley (SD) rats supplied from two vendors. METHODS: Sensitivity to morphine (0.3-6.0 mg/kg, s.c.) was evaluated in the hot plate test of acute thermal nociception, the Complete Freund's Adjuvant (CFA) model of inflammatory-induced mechanical hyperalgesia, and in a locomotor motility assay in male rats from the following strains; Lewis (LEW), Fischer (F344), Wistar Kyoto (WKY), and SD's from Envigo and Charles River. RESULTS: F344 and SD rats were similarly sensitive to morphine in hot plate and CFA-induced inflammatory hyperalgesia (Minimum Effective Dose (MED) = 3.0 mg/kg). WKY rats developed a less robust mechanical hypersensitivity after CFA injection, and were less sensitive to morphine in both pain tests (MED = 6.0 mg/kg). LEW rats were completely insensitive to morphine in the hot plate test, in contrast to the reversal of CFA-induced hyperalgesia (MED = 3.0 mg/kg). All strains exhibited a dose-dependent reduction in locomotor activity at 3.0-6.0 mg/kg. CONCLUSION: Sensory phenotyping in response to acute thermal and inflammatory-induced pain, and sensitivity to morphine in various inbred and outbred rat strains indicates that different pathophysiological mechanisms are engaged after injury. This could have profound implications for translating preclinical drug discovery efforts into pain patients. SIGNIFICANCE: The choice of rat strain used in preclinical pain research can profoundly affect the outcome of experiments in relation to (a) nociceptive threshold responses, and (b) efficacy to analgesic treatment, in assays of acute and tonic inflammatory nociceptive pain.

The novel compound (+/-)-1-[10-((E)-3-Phenyl-allyl)-3,10-diaza-bicyclo[4.3.1]dec-3-yl]-propan-1-one (NS7051) attenuates nociceptive transmission in animal models of experimental pain; a pharmacological comparison with the combined mu-opioid receptor agonist and monoamine reuptake inhibitor tramadol.

Tramadol is an atypical analgesic with a unique dual mechanism of action. It acts on monoamine transporters to inhibit reuptake of noradrenaline (NA) and serotonin (5-HT), and consequent upon metabolism, displays potent agonist activity at micro-opioid receptors. Here, we present data for the novel compound NS7051, which was shown to have sub-micromolar affinity (Ki=0.034microM) for micro-opioid receptors and inhibited reuptake of 5-HT, NA and DA (IC(50)=4.2, 3.3 and 3.5microM in cortex, hippocampus and striatum respectively). NS7051 (1-30mg/kg, s.c.) produced a dose-dependent naloxone-reversible increase in the hot plate withdrawal latency, and was also analgesic in the tail flick test. In models of persistent and chronic inflammatory nociception, NS7051 reversed flinching behaviours during interphase and second phase of the formalin test (ED(50)=1.7 and 1.8mg/kg, s.c.), and hindpaw weight-bearing deficits induced by complete Freund's adjuvant injection (ED50=1.2mg/kg, s.c.). In the chronic constriction injury model of neuropathic pain, mechanical allodynia and hyperalgesia were both reversed by NS7051 (ED50=6.7 and 4.9mg/kg, s.c.). Tramadol was also active in all pain models although at considerably higher doses (20-160mg/kg, s.c.). No ataxia was observed at antiallodynic doses giving therapeutic indices of 19 and 3 for NS7051 and tramadol. The combined opioid receptor agonism and monoamine reuptake inhibitory properties of NS7051 inferred from behavioural studies appear to contribute to its well tolerated antinociceptive profile in rats. However, unlike tramadol this did not correlate with the ability to increase hippocampal monoamine levels measured by microdialysis in anesthetised rats.

A unique inbred rat strain with sustained cephalic hypersensitivity as a model of chronic migraine-like pain.

Animal models of migraine-like pain enabling ongoing study of behaviour typically involve the systemic administration of chemical vasodilators or dural administration of inflammatory algogens. However, neither method mediates prolonged effects on behavior indicative of enduring pathophysiological changes occurring within dural or trigeminal pain circuits. We generated successive generations of a unique inbred rat strain, spontaneous trigeminal allodynia (STA) rats, previously reported to exhibit an episodic migraine-like behavioural phenotype. We show that both male and female STA rats display robust and sustained reductions in periorbital thresholds to cutaneous mechanical stimulation. Otherwise, the general behavior (e.g. locomotor, grooming) of these rats appeared normal. In female STA rats, the mechanical hypersensitivity was confined to the cephalic region, manifested after puberty through adolescence, and was sustained into adulthood recapitulating the clinical manifestation of migraine. We exploited this hitherto unidentified chronic phenotype to show that the migraine-specific drugs sumatriptan (5-HT1B/1D receptor agonist) and olcegepant (CGRP receptor antagonist) could completely reverse cephalic hypersensitivity using a within subject cross-over paradigm. Our findings indicate that STA rats actually possess a phenotype indicative of migraine chronicity which is exquisitely sensitive to migraine therapeutics. This unique strain could prove to be an invaluable resource in preclinical migraine drug discovery.