MDA7: a novel selective agonist for CB2 receptors that prevents allodynia in rat neuropathic pain models

Background and purpose:

There is growing interest in using cannabinoid type 2 (CB₂) receptor agonists for the treatment of neuropathic pain. In this report, we describe the pharmacological characteristics of MDA7 (1-[(3-benzyl-3-methyl-2,3-dihydro-1-benzofuran-6-yl)carbonyl]piperidine), a novel CB₂ receptor agonist.

Experimental approach:

We characterized the pharmacological profile of MDA7 by using radioligand-binding assays and in vitro functional assays at human cannabinoid type 1 (CB₁) and CB₂ receptors. In vitro functional assays were performed at rat CB₁ and CB₂ receptors. The effects of MDA7 in reversing neuropathic pain were assessed in spinal nerve ligation and paclitaxel-induced neuropathy models in rats.

Key results:

MDA7 exhibited selectivity and agonist affinity at human and rat CB₂ receptors. MDA7 treatment attenuated tactile allodynia produced by spinal nerve ligation or by paclitaxel in a dose-related manner. These effects were selectively antagonized by a CB₂ receptor antagonist but not by CB₁ or opioid receptor antagonists. MDA7 did not affect rat locomotor activity.

Conclusion and implications:

MDA7, a novel selective CB₂ agonist, was effective in suppressing neuropathic nociception in two rat models without affecting locomotor behaviour. These results confirm the potential for CB₂ agonists in the treatment of neuropathic pain.     

Central and peripheral sites of action for CB₂ receptor mediated analgesic activity in chronic inflammatory and neuropathic pain models in rats

BACKGROUND AND PURPOSE

Cannabinoid CB₂ receptor activation by selective agonists has been shown to produce analgesic effects in preclinical models of inflammatory and neuropathic pain. However, mechanisms underlying CB₂-mediated analgesic effects remain largely unknown. The present study was conducted to elucidate the CB₂ receptor expression in ‘pain relevant’ tissues and the potential sites of action of CB₂ agonism in rats.

EXPERIMENTAL APPROACH

Expression of cannabinoid receptor mRNA was evaluated by quantitative RT-PCR in dorsal root ganglia (DRGs), spinal cords, paws and several brain regions of sham, chronic inflammatory pain (CFA) and neuropathic pain (spinal nerve ligation, SNL) rats. The sites of CB₂ mediated antinociception were evaluated in vivo following intra-DRG, intrathecal (i.t.) or intraplantar (i.paw) administration of potent CB₂-selective agonists A-836339 and AM1241.

KEY RESULTS

CB₂ receptor gene expression was significantly up-regulated in DRGs (SNL and CFA), spinal cords (SNL) or paws (CFA) ipsilateral to injury under inflammatory and neuropathic pain conditions. Systemic A-836339 and AM1241 produced dose-dependent efficacy in both inflammatory and neuropathic pain models. Local administration of CB₂ agonists also produced significant analgesic effects in SNL (intra-DRG and i.t.) and CFA (intra-DRG) pain models. In contrast to A-836339, i.paw administration of AM-1241 dose-relatedly reversed the CFA-induced thermal hyperalgesia, suggesting that different mechanisms may be contributing to its in vivo properties.

CONCLUSIONS AND IMPLICATIONS

These results demonstrate that both DRG and spinal cord are important sites contributing to CB₂ receptor-mediated analgesia and that the changes in CB₂ receptor expression play a crucial role for the sites of action in regulating pain perception.     

D-Amino acid oxidase-mediated increase in spinal hydrogen peroxide is mainly responsible for formalin-induced tonic pain

BACKGROUND AND PURPOSE

Spinal reactive oxygen species (ROS) are critically involved in chronic pain. d-Amino acid oxidase (DAAO) oxidizes d-amino acids such as d-serine to form the byproduct hydrogen peroxide without producing other ROS. DAAO inhibitors are specifically analgesic in tonic pain, neuropathic pain and cancer pain. This study examined the role of spinal hydrogen peroxide in pain and the mechanism of the analgesic effects of DAAO inhibitors.

EXPERIMENTAL APPROACH

Formalin-induced pain behaviours and spinal hydrogen peroxide levels were measured in rodents.

KEY RESULTS

Formalin injected into the paw increased spinal hydrogen peroxide synchronously with enhanced tonic pain; both were effectively prevented by i.t. fluorocitrate, a selective astrocyte metabolic inhibitor. Given systemically, the potent DAAO inhibitor CBIO (5-chloro-benzo[d]isoxazol-3-ol) blocked spinal DAAO enzymatic activity and specifically prevented formalin-induced tonic pain in a dose-dependent manner. Although CBIO maximally inhibited tonic pain by 62%, it completely prevented the increase in spinal hydrogen peroxide. I.t. catalase, an enzyme specific for decomposition of hydrogen peroxide, completely depleted spinal hydrogen peroxide and prevented formalin-induced tonic pain by 65%. Given systemically, the ROS scavenger PBN (phenyl-N-tert-butylnitrone) also inhibited formalin-induced tonic pain and increase in spinal hydrogen peroxide. Formalin-induced tonic pain was potentiated by i.t. exogenous hydrogen peroxide. CBIO did not increase spinal d-serine level, and i.t. d-serine did not alter either formalin-induced tonic pain or CBIO''s analgesic effect.

CONCLUSIONS AND IMPLICATIONS

Spinal hydrogen peroxide is specifically and largely responsible for formalin-induced pain, and DAAO inhibitors produce analgesia by blocking spinal hydrogen peroxide production rather than interacting with spinal d-serine.     

Evidence for a differential opioidergic involvement in the analgesic effect of antidepressants: prediction for efficacy in animal models of neuropathic pain?

BACKGROUND AND PURPOSE

Antidepressants are one of the recommended treatments for neuropathic pain. However, their analgesic action remains unpredictable, and there are no selection criteria for clinical use. Better knowledge of their mechanism of action could help highlight differences underlying their unequal efficacy.

EXPERIMENTAL APPROACH

We compared the activity of a tricyclic antidepressant (clomipramine) with selective 5-HT and noradrenaline reuptake inhibitors (milnacipran and duloxetine) in streptozocin-induced diabetic and chronic constriction nerve injury-induced neuropathic rats, after repeated injections. We looked for an opioidergic mechanism in their action.

KEY RESULTS

Abolition of mechanical hyperalgesia was observed in mononeuropathic rats after five injections of clomipramine (5 mg·kg⁻¹, s.c.) and milnacipran (10 or 20 mg·kg⁻¹, i.p.) and in diabetic rats after clomipramine. An additional antinociceptive effect was obtained with five injections of duloxetine (3 mg·kg⁻¹, i.p.) in both models and milnacipran (10 mg·kg⁻¹, i.p.) in diabetic rats. These effects were observed with plasma antidepressant concentrations similar to those found in patients treated for neuropathic pain. Naloxone (1 mg·kg⁻¹, i.v.) only suppressed the anti-hyperalgesic effects of clomipramine in both models of pain and of milnacipran in the traumatic model.

CONCLUSIONS AND IMPLICATIONS

The opioid system appears to be involved in the mechanism of action of antidepressants that only have an anti-hyperalgesic effect but not in those that have a stronger (i.e. antinociceptive) effect. These differences between the antidepressants occurred whatever the aetiology of the neuropathy and, if confirmed in clinical trials, could be used to decide which antidepressant is administered to a patient with neuropathic pain.     

Spinal actions of ω-conotoxins,CVID, MVIIA and related peptides in a rat neuropathic pain model

BACKGROUND AND PURPOSE

Antagonists of the N-type voltage gated calcium channel (VGCC), Ca_v2.2, have a potentially important role in the treatment of chronic neuropathic pain. ω-conotoxins, such MVIIA and CVID are effective in neuropathic pain models. CVID is reported to have a greater therapeutic index than MVIIA in neuropathic pain models, and it has been suggested that this is due to faster reversibility of binding, but it is not known whether this can be improved further.

EXPERIMENTAL APPROACH

We examined the potency of CVID, MVIIA and two intermediate hybrids ([K10R]CVID and [R10K]MVIIA) to reverse signs of neuropathic pain in a rat nerve ligation model in parallel with production of side effects. We also examined the potency and reversibility to inhibit primary afferent synaptic neurotransmission in rat spinal cord slices.

KEY RESULTS

All ω-conotoxins produced dose-dependent reduction in mechanical allodynia. They also produced side effects on the rotarod test and in a visual side-effect score. CVID displayed a marginally better therapeutic index than MVIIA. The hybrids had a lesser effect in the rotarod test than either of their parent peptides. Finally, the conotoxins all presynaptically inhibited excitatory synaptic neurotransmission into the dorsal horn and displayed recovery that was largely dependent upon the magnitude of inhibition and not the conotoxin type.

CONCLUSIONS AND IMPLICATIONS

These findings indicate that CVID provides only a marginal improvement over MVIIA in a preclinical model of neuropathic pain, which appears to be unrelated to reversibility from binding. Hybrids of these conotoxins might provide viable alternative treatments.     

Activation of NK₁ receptors in the locus coeruleus induces analgesia through noradrenergic-mediated descending inhibition in a rat model of neuropathic pain

BACKGROUND AND PURPOSE

The locus coeruleus (LC) is a major source of noradrenergic projections to the dorsal spinal cord, and thereby plays an important role in the modulation of nociceptive information. The LC receives inputs from substance P (SP)-containing fibres from other regions, and expresses the NK₁ tachykinin receptor, a functional receptor for SP. In the present study, we investigated the roles of SP in the LC in neuropathic pain.

EXPERIMENTAL APPROACH

Chronic constriction injury (CCI) of the left sciatic nerve was performed in rats to induce neuropathic pain. After development of neuropathic pain, SP was injected into the LC and the nocifensive behaviours were assessed. The involvement of noradrenergic descending inhibition in SP-induced analgesia was examined by i.t. administration of yohimbine, an α₂-adrenoceptor antagonist. NK₁ receptor expression in the LC was examined by immunohistochemistry.

KEY RESULTS

In CCI rats, mechanical allodynia was alleviated by SP injection into the LC. These effects were abolished by prior injection of WIN 51708, an NK₁ receptor antagonist, into the LC or i.t. treatment with yohimbine. NK₁ receptor-like immunoreactivity was observed in noradrenergic neurons throughout the LC in intact rats, and remained unchanged after CCI.

CONCLUSION AND IMPLICATIONS

SP in the LC exerted analgesic effects on neuropathic pain through NK₁ receptor activation and resulted in facilitation of spinal noradrenergic transmission. Accordingly, manipulation of the SP/NK₁ receptor signalling pathway in the LC may be a promising strategy for effective treatment of neuropathic pain.     

Topical application of disodium isostearyl 2-O-L-ascorbyl phosphate, an amphiphilic ascorbic acid derivative, reduces neuropathic hyperalgesia in rats

BACKGROUND AND PURPOSE

Ca_v3.2 T-type calcium channels, targeted by H₂S, are involved in neuropathic hyperalgesia in rats and ascorbic acid inhibits Ca_v3.2 channels. Therefore, we evaluated the effects of intraplantar (i.pl.) administration of ascorbic acid or topical application of disodium isostearyl 2-O-L-ascorbyl phosphate (DI-VCP), a skin-permeable ascorbate derivative on hyperalgesia induced by NaHS, an H₂S donor, and on neuropathic hyperalgesia.

EXPERIMENTAL APPROACH

In rats mechanical hyperalgesia was evoked by i.pl. NaHS, and neuropathic hyperalgesia was induced by L5 spinal nerve cutting (L5SNC) or by repeated administration of paclitaxel, an anti-cancer drug. Dermal ascorbic acid levels were determined colorimetrically.

KEY RESULTS

The NaHS-evoked Ca_v3.2 channel-dependent hyperalgesia was inhibited by co-administered ascorbic acid. Topical application of DI-VCP, but not ascorbic acid, prevented the NaHS-evoked hyperalgesia, and also increased dermal ascorbic acid levels. Neuropathic hyperalgesia induced by L5SNC or paclitaxel was reversed by i.pl. NNC 55–0396, a selective T-type calcium channel blocker, ascorbic acid or DI-VCP, and by topical DI-VCP, but not by topical ascorbic acid. The effects of i.pl. ascorbic acid and topical DI-VCP in the paclitaxel-treated rats were characterized by the faster onset and greater magnitude, compared with their effects in the L5SNC rats. Dermal ascorbic acid levels in the hindpaw significantly decreased after paclitaxel treatment, but not L5SNC, which was reversed by topical DI-VCP.

CONCLUSIONS AND IMPLICATIONS

Ascorbic acid, known to inhibit Ca_v3.2 channels, suppressed neuropathic hyperalgesia. DI-VCP ointment for topical application may be of benefit in the treatment of neuropathic pain.     

Gelsemine alleviates both neuropathic pain and sleep disturbance in partial sciatic nerve ligation mice

Aim:

Gelsemine, an alkaloid from the Chinese herb Gelsemium elegans (Gardn & Champ) Benth., is effective in mitigating chronic pain in rats. In the present study we investigated whether the alkaloid improved sleep disturbance, the most common comorbid symptoms of chronic pain, in a mouse model of neuropathic pain.

Methods:

Mice were subjected to partial sciatic nerve ligation (PSNL). After the mice were injected with gelsemine or pregabalin (the positive control) intraperitoneally, mechanical allodynia and thermal hyperalgesia were assessed, and electroencephalogram (EEG)/electromyogram (EMG) recording was performed. Motor performance of the mice was assessed using rota-rod test. c-Fos expression in the brain was analyzed with immunohistochemical staining.

Results:

In PSNL mice, gelsemine (2 and 4 mg/kg) increased the mechanical threshold for 4 h and prolonged the thermal latencies for 3 h. Furthermore, gelsemine (4 mg/kg, administered at 6:30 AM) increased non-rapid eye movement (non-REM, NREM) sleep, decreased wakefulness, but did not affect REM sleep during the first 3 h in PSNL mice. Sleep architecture analysis showed that gelsemine decreased the mean duration of wakefulness and increased the total number of episodes of NREM sleep during the first 3 h after the dosing. Gelsemine (4 mg/kg) did not impair motor coordination in PSNL mice. Immunohistochemical study showed that PSNL increased c-Fos expression in the neurons of the anterior cingulate cortex, and gelsemine (4 mg/kg) decreased c-Fos expression by 58%. Gelsemine (4 mg/kg, administered at either 6:30 AM or 8:30 PM) did not produce hypnotic effect in normal mice. Pregabalin produced similar antinociceptive and hypnotic effects, but impaired motor coordination in PSNL mice.

Conclusion:

Gelsemine is an effective agent for treatment of both neuropathic pain and sleep disturbance in PSNL mice; anterior cingulate cortex might play a role in the hypnotic effects of gelsemine.     

Increase in hemokinin-1 mRNA in the spinal cord during the early phase of a neuropathic pain state

Background and purpose:

Substance P (SP), a representative member of the tachykinin family, is involved in nociception under physiological and pathological conditions. Recently, hemokinin-1 (HK-1) was identified as a new member of this family. Although HK-1 acts on NK₁ tachykinin receptors that are thought to be innate for SP, the roles of HK-1 in neuropathic pain are still unknown.

Experimental approach:

Using rats that had been subjected to chronic constrictive injury (CCI) of the sciatic nerve as a neuropathic pain model, we examined the changes in expression of SP- and HK-1-encoding genes (TAC1 and TAC4, respectively) in the L4/L5 spinal cord and L4/L5 dorsal root ganglia (DRGs) in association with changes in pain-related behaviours in this neuropathic pain state.

Key results:

The TAC4 mRNA level was increased on the ipsilateral side of the dorsal spinal cord, but not in DRGs, at day 3 after CCI. In contrast, the TAC1 mRNA level was significantly increased in the DRGs at day 3 after CCI without any changes in the dorsal spinal cord. Analysis of a cultured microglial cell line revealed the presence of TAC4 mRNA in microglial cells. Minocycline, an inhibitor of microglial activation, blocked the increased expression of TAC4 mRNA after CCI and inhibited the associated pain-related behaviours and microglial activation in the spinal cord.

Conclusions and implications:

The present results suggest that HK-1 expression is increased at least partly in activated microglial cells after nerve injury and is clearly involved in the early phase of neuropathic pain.     

HDAC inhibitors restore C-fibre sensitivity in experimental neuropathic pain model

Background and Purpose

Hypoesthesia is a clinical feature of neuropathic pain. The feature is partly explained by the evidence of epigenetic repression of Na_v1.8 sodium channel in the dorsal root ganglion (DRG).

Experimental Approach

We investigated the possibility of trichostatin A (TSA), valproic acid (VPA) and suberoylanilide hydroxamic acid (SAHA) to reverse the unique C-fibre sensitivity observed following partial ligation of sciatic nerve in mice.

Key Results

Nerve injury-induced down-regulation of DRG Na_v1.8 sodium channel and C-fibre-related hypoesthesia were reversed by TSA, VPA and SAHA treatments, which inhibit histone deacetylase (HDAC), and increase histone acetylation at the regulatory sequence of Na_v1.8.

Conclusions and Implications

Taken together, these studies provide the evidence that hypoesthesia and underlying down-regulation of Na_v1.8, negative symptoms observed in nerve injury-induced neuropathic pain models are regulated by an epigenetic chromatin remodelling through HDAC-related machineries.     

A spinal mechanism of action for duloxetine in a rat model of painful diabetic neuropathy

BACKGROUND AND PURPOSE

This study was designed to clarify mechanisms responsible for the anti-allodynic effects of duloxetine in diabetes.

EXPERIMENTAL APPROACH

The streptozotocin-induced diabetic rat model was used to compare the efficacy of duloxetine, 5-HT, the 5-HT_2A receptor agonist [1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI)] and two antagonists (ketanserin and pruvanserin) on tactile allodynia.

KEY RESULTS

Systemic or intrathecal injection of duloxetine alleviated tactile allodynia in diabetic rats. The effect of systemic duloxetine was reduced by intrathecal administration of ketanserin or pruvanserin, indicating participation of spinal 5-HT_2A receptors in the mechanism of action of duloxetine. In contrast to spinal delivery, systemic and local peripheral injections of ketanserin or pruvanserin alleviated tactile allodynia in diabetic rats. This effect was reversed immediately after systemic or local DOI injection.

CONCLUSIONS AND IMPLICATIONS

These results support the involvement of spinal 5-HT_2A receptors in the ability of duloxetine to ameliorate painful diabetic neuropathy. Our data also suggest that the role of 5-HT_2A receptors depends on the level of the neuraxis at which activation takes place, with peripheral activation contributing to tactile allodynia in diabetic rats, whereas spinal activation of this receptor alleviates tactile allodynia. The development of selective peripheral 5-HT_2A receptor antagonists may offer a novel approach for the treatment of diabetic neuropathic pain.     

Inhibitors of catechol-O-methyltransferase sensitize mice to pain

BACKGROUND AND PURPOSE

Catechol-O-methyltransferase (COMT) inhibitors are used in Parkinson''s disease in which pain is an important symptom. COMT polymorphisms modulate pain and opioid analgesia in humans. In rats, COMT inhibitors have been shown to be pro-nociceptive in acute pain models, but also to attenuate allodynia and hyperalgesia in a model of diabetic neuropathy. Here, we have assessed the effects of acute and repeated administrations of COMT inhibitors on mechanical, thermal and carrageenan-induced nociception in male mice.

EXPERIMENTAL APPROACH

We used single and repeated administration of a peripherally restricted, short-acting (nitecapone) and also a centrally acting (3,5-dinitrocatechol, OR-486) COMT inhibitor. We also tested CGP 28014, an indirect inhibitor of COMT enzyme. Effects of OR-486 on thermal nociception were also studied in COMT deficient mice. Effects on spinal pathways were assessed in rats given intrathecal nitecapone.

KEY RESULTS

After single administration, both nitecapone and OR-486 reduced mechanical nociceptive thresholds and thermal nociceptive latencies (hot plate test) at 2 and 3 h, regardless of their brain penetration. These effects were still present after chronic treatment with COMT inhibitors for 5 days. Intraplantar injection of carrageenan reduced nociceptive latencies and both COMT inhibitors potentiated this reduction without modifying inflammation. CGP 28014 shortened paw flick latencies. OR-486 did not modify hot plate times in Comt gene deficient mice. Intrathecal nitecapone modified neither thermal nor mechanical nociception.

CONCLUSIONS AND IMPLICATIONS

Pro-nociceptive effects of COMT inhibitors were confirmed. The pro-nociceptive effects were primarily mediated via mechanisms acting outside the brain and spinal cord. COMT protein was required for these actions.     

Inhibition of nociceptive responses after systemic administration of amidated kyotorphin

BACKGROUND AND PURPOSE

Kyotorphin (KTP; L-Tyr-L-Arg), an endogenous neuropeptide, is potently analgesic when delivered directly to the central nervous system. Its weak analgesic effects after systemic administration have been explained by inability to cross the blood–brain barrier (BBB) and detract from the possible clinical use of KTP as an analgesic. In this study, we aimed to increase the lipophilicity of KTP by amidation and to evaluate the analgesic efficacy of a new KTP derivative (KTP-amide – KTP-NH₂).

EXPERIMENTAL APPROACH

We synthesized KTP-NH₂. This peptide was given systemically to assess its ability to cross the BBB. A wide range of pain models, including acute, sustained and chronic inflammatory and neuropathic pain, were used to characterize analgesic efficacies of KTP-NH₂. Binding to opioid receptors and toxicity were also measured.

KEY RESULTS

KTP-NH₂, unlike its precursor KTP, was lipophilic and highly analgesic following systemic administration in several acute and chronic pain models, without inducing toxic effects or affecting motor responses and blood pressure. Binding to opioid receptors was minimal. KTP-NH₂ inhibited nociceptive responses of spinal neurons. Its analgesic effects were prevented by intrathecal or i.p. administration of naloxone.

CONCLUSIONS AND IMPLICATIONS

Amidation allowed KTP to show good analgesic ability after systemic delivery in acute and chronic pain models. The indirect opioid-mediated actions of KTP-NH₂ may explain why this compound retained its analgesic effects although the usual side effects of opioids were absent, which is a desired feature in next-generation pain medications.     

Glial cell line-derived neurotrophic factormediated enhancement of noradrenergic descending inhibition in the locus coeruleus exerts prolonged analgesia in neuropathic pain

Background and Purpose

The locus coeruleus (LC) is the principal nucleus containing the noradrenergic neurons and is a major endogenous source of pain modulation in the brain. Glial cell line-derived neurotrophic factor (GDNF), a well-established neurotrophic factor for noradrenergic neurons, is a major pain modulator in the spinal cord and primary sensory neurons. However, it is unknown whether GDNF is involved in pain modulation in the LC.

Experimental Approach

Rats with chronic constriction injury (CCI) of the left sciatic nerve were used as a model of neuropathic pain. GDNF was injected into the left LC of rats with CCI for 3 consecutive days and changes in mechanical allodynia and thermal hyperalgesia were assessed. The α₂-adrenoceptor antagonist yohimbine was injected intrathecally to assess the involvement of descending inhibition in GDNF-mediated analgesia. The MEK inhibitor U0126 was used to investigate whether the ERK signalling pathway plays a role in the analgesic effects of GDNF.

Key Results

Both mechanical allodynia and thermal hyperalgesia were attenuated 24 h after the first GDNF injection. GDNF increased the noradrenaline content in the dorsal spinal cord. The analgesic effects continued for at least 3 days after the last injection. Yohimbine abolished these effects of GDNF. The analgesic effects of GDNF were partly, but significantly, inhibited by prior injection of U0126 into the LC.

Conclusions and Implications

GDNF injection into the LC exerts prolonged analgesic effects on neuropathic pain in rats by enhancing descending noradrenergic inhibition.     

LASSBio-881: an N-acylhydrazone transient receptor potential vanilloid subfamily type 1 antagonist orally effective against the hypernociception induced by capsaicin or partial sciatic ligation

Background and purpose:

Compound LASSBio-881 is an orally effective antinociceptive that binds to cannabinoid receptors and is active mainly on the neurogenic component of pain models. We investigated whether transient receptor potential vanilloid subfamily type 1 (TRPV1) channels are involved in the effects of LASSBio-881.

Experimental approach:

Modulation of capsaicin (CAP)- and low pH-induced currents was evaluated in TRPV1-expressing Xenopus oocytes. In vivo effects were evaluated in CAP-induced acute and inflammatory changes in nociception, as well as in partial sciatic ligation-induced thermal hypernociception.

Key results:

LASSBio-881 inhibited TRPV1 currents elicited by CAP with an IC₅₀ of 14 µM, and inhibited proton-gated currents by 70% at 20 µM. Functional interaction with CAP was surmountable. Locally applied LASSBio-881 decreased time spent in CAP-elicited nocifensive behaviour by 30%, and given orally it reduced measures of CAP- or carrageenan-evoked thermal hypernociception by 60 and 40% respectively. In addition, LASSBio-881 decreased the paw withdrawal responses to thermal stimuli of animals with sciatic neuropathy 7–11 days after nerve ligation, at a dose of 300 µmol·kg⁻¹·day⁻¹ p.o. At this dose, hyperthermia was not observed within 4 h following oral administration.

Conclusions and implications:

LASSBio-881 is a TRPV1 antagonist that apparently competes with CAP. Accordingly, LASSBio-881 inhibited nociception in models of acute, inflammatory and neuropathic pain presumed to involve TRPV1 signalling. These in vivo actions were not hindered by hyperthermia, a common side effect of other TRPV1 antagonists. We propose that the antinociceptive properties of LASSBio-881 are due to TRPV1 antagonism, although other molecular interactions may contribute to the effects of this multi-target drug candidate.     

Differential inhibitory effects of drugs acting at the noradrenaline and 5-hydroxytryptamine transporters in rat and human neocortical synaptosomes

Background and purpose:

Although the amino acid sequences of rat and human 5-hydroxytryptamine (5-HT) and noradrenaline (NA) transporters (i.e. SERT and NET) are highly homologous, species differences exist in the inhibitory effects of drugs acting at these transporters. Therefore, comparison of the potencies of drugs acting at SERT and NET in native human and rat neocortex may serve to more accurately predict their clinical profile.

Experimental approach:

Synaptosomes prepared from fresh human and rat neocortical tissues were used for [³H]-5-HT and [³H]-NA saturation and competition uptake experiments. The drugs tested included NA reuptake inhibitors (desipramine, atomoxetine and (S,S)-reboxetine), 5-HT reuptake blockers (citalopram, fluoxetine and fluvoxamine) and dual 5-HT/NA reuptake inhibitors (duloxetine and milnacipran).

Key results:

In saturation experiments on synaptosomal [³H]-5-HT and [³H]-NA uptake, the dissociation constants did not indicate species differences although a smaller density of both SERT and NET was observed in human tissues. In competition experiments with the various drugs, marked species differences in their potencies were observed, especially at SERT. The rank order of selectivity ratios (SERT/NET) in human neocortex was as follows: citalopram ≥ duloxetine = fluvoxamine ≥ fluoxetine > milnacipran > desipramine = atomoxetine > (S,S)-reboxetine. Significant species differences in these ratios were observed for duloxetine, atomoxetine and desipramine.

Conclusions and implications:

This study provides the first compilation of drug potency at native human neocortical SERT and NET. The significant species differences (viz., human vs. rat) in drug potency suggest that the general use of rodent data should be limited to predict clinical efficacy or profile.     

Antihyperalgesic effects of imidazoline I2 receptor ligands in rat models of inflammatory and neuropathic pain

Background and Purpose

A new imidazoline I₂ receptor ligand, CR4056, is effective for chronic inflammatory pain and diabetic neuropathy. However, it is unclear whether other I₂ receptor ligands have similar effects and whether antinociceptive tolerance develops with repeated treatment.

Experimental Approach

The Von Frey filament test was used to measure mechanical hyperalgesia and the plantar test to measure thermal hyperalgesia in rats injected with complete Freund''s adjuvant (CFA) treatment or had undergone surgery to induce chronic constriction injury (CCI), models of inflammatory pain and peripheral neuropathic pain respectively. The effects of morphine and I₂ receptor ligands, 2-BFI, BU224, tracizoline and CR4056, 3.2–32 mg·kg−1, i.p., on hyperalgesia or affective pain (as measured by a place escape/avoidance paradigm) were studied in separate experiments.

Key Results

Morphine and the I₂ receptor ligands (2-BFI, BU224 and tracizoline) all dose-dependently attenuated mechanical and thermal hyperalgesia in CFA-treated rats. The anti-hyperalgesic effects of 2-BFI in CFA-treated and CCI rats were attenuated by the I₂ receptor antagonist idazoxan. The combination of 2-BFI and morphine produced additive effects against mechanical hyperalgesia in CFA-treated rats. Repeated treatment (daily for 7–9 days) with 2-BFI or CR4056 did not produce antinociceptive tolerance in CFA-treated or CCI rats. Morphine and the I₂ receptor ligands (2-BFI, BU224 and CR4056) were all effective at attenuating place escape/avoidance behaviour in CFA-treated rats.

Conclusions and Implications

Imidazoline I₂ receptor ligands have antihyperalgesic effects in rat models of inflammatory and neuropathic pain and may represent a new class of pharmacotherapeutics for the management of chronic pain.     

Combined inhibition of monoacylglycerol lipase and cyclooxygenases synergistically reduces neuropathic pain in mice

Background and Purpose

Neuropathic pain is commonly treated with GABA analogues, steroids or non-steroidal anti-inflammatory drugs (NSAIDs). NSAIDs inhibit one or more COX isozymes but chronic COX inhibition paradoxically increases gastrointestinal inflammation and risk of unwanted cardiovascular events. The cannabinoids also have analgesic and anti-inflammatory properties and reduce neuropathic pain in animal models. The present study investigated the analgesic effects of inhibiting both monoacylglycerol lipase (MAGL) and COX enzymes, using low doses of both inhibitors.

Experimental Approach

Mice subjected to chronic constriction injury (CCI) were tested for mechanical and cold allodynia after administration of the MAGL inhibitor, JZL184, or the non-selective COX inhibitor diclofenac. Then, both drugs were co-administered at fixed dose proportions of 1:3, 1:1 and 3:1, based on their ED₅₀ values. PGs, endocannabinoids and related lipids were quantified in lumbar spinal cord.

Key Results

Combining low doses of JZL184 and diclofenac synergistically attenuated mechanical allodynia and additively reduced cold allodynia. The cannabinoid CB₁ receptor antagonist, rimonabant, but not the CB₂ receptor antagonist, SR144528, blocked the analgesic effects of the JZL184 and diclofenac combination on mechanical allodynia, implying that CB₁ receptors were primarily responsible for the anti-allodynia. Diclofenac alone and with JZL184 significantly reduced PGE₂ and PGF_2α in lumbar spinal cord tissue, whereas JZL184 alone caused significant increases in the endocannabinoid metabolite, N-arachidonoyl glycine.

Conclusions and Implications

Combining COX and MAGL inhibition is a promising therapeutic approach for reducing neuropathic pain with minimal side effects.     

Milnacipran Remediates Impulsive Deficits in Rats with Lesions of the Ventromedial Prefrontal Cortex

Background:

Deficits in impulse control are often observed in psychiatric disorders in which abnormalities of the prefrontal cortex are observed, including attention-deficit/hyperactivity disorder and bipolar disorder. We recently found that milnacipran, a serotonin/noradrenaline reuptake inhibitor, could suppress impulsive action in normal rats. However, whether milnacipran could suppress elevated impulsive action in rats with lesions of the ventromedial prefrontal cortex, which is functionally comparable with the human prefrontal cortex, remains unknown.

Methods:

Selective lesions of the ventromedial prefrontal cortex were made using quinolinic acid in rats previously trained on a 3-choice serial reaction time task. Sham rats received phosphate buffered saline. Following a period of recovery, milnacipran (0 or 10mg/kg/d × 14 days) was orally administered 60 minutes prior to testing on the 3-choice task. After 7 days of drug cessation, Western blotting, immunohistochemistry, electrophysiological analysis, and morphological analysis were conducted.

Results:

Lesions of the ventromedial prefrontal cortex induced impulsive deficits, and repeated milnacipran ameliorated the impulsive deficit both during the dosing period and after the cessation of the drug. Repeated milnacipran remediated the protein levels of mature brain-derived neurotrophic factor and postsynaptic density-95, dendritic spine density, and excitatory currents in the few surviving neurons in the ventromedial prefrontal cortex of ventromedial prefrontal cortex-lesioned rats.

Conclusions:

The findings of this study suggest that milnacipran treatment could be a novel strategy for the treatment of psychiatric disorders that are associated with a lack of impulse control.     

Enriched enrollment: definition and effects of enrichment and dose in trials of pregabalin and gabapentin in neuropathic pain. A systematic review

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• Enriched enrolment (the exclusion of non-responders or specific inclusion of responders) is believed to add both to trial sensitivity and to the measured effect of an intervention.
• Enriched enrolment lacks specific definition, and the extent of any differences between results with non-enriched recruitment and enriched enrolment is not known.
• Enriched enrolment is thought to have influenced neuropathic pain trials.

WHAT THIS STUDY ADDS

• The paper suggests definitions for complete and partial enriched enrolment, and applies those definitions to trials of pregabalin and gabapentin in neuropathic pain.
• The effect of enrichment was small, and especially in pregabalin trials with the best data, no difference was found between partial enrichment and no enrichment.
• The effects of complete enrichment are unknown.

AIMS

Enriched enrolment study designs have been suggested to be useful for proof of concept when only a proportion of the diseased population responds to a treatment intervention. We aim to investigate whether this really is the case in trials of pregabalin and gabapentin in neuropathic pain.

METHODS

We defined ‘complete’, ‘partial’ and ‘non-enriched’ enrolment, and examined pregabalin and gabapentin trials for the extent of enrichment and for effects of enrichment on efficacy and adverse event outcomes.

RESULTS

There were no studies using complete enriched enrolment; seven trials used partial enriched enrolment and 14 non-enriched enrolment. In pregabalin trials the maximum extent of enrichment was estimated at about 12%. Partial enriched enrolment did not change estimates of efficacy or harm. Over 150–600 mg maximum daily dose there was strong dose dependence for pregabalin.

CONCLUSIONS

A benefit of partial over non-enriched enrolment could not be demonstrated because the degree of enrichment was rather small, and possibly because enrichment produced little enhancement of treatment effect. Whether a greater degree of enrichment would result in important differences is unknown. Researchers reporting clinical trials with any enrichment must describe both process and extent of enrichment. As things stand, the effects of enriched enrolment remain unknown for neuropathic pain trials.