Yong-Xiang Wang

Spinal transient receptor potential ankyrin 1 channel contributes to central pain hypersensitivity in various pathophysiological conditions in the rat.

&NA; The transient receptor potential ankyrin 1 (TRPA1) ion channel is expressed on nociceptive primary afferent neurons. On the proximal nerve ending within the spinal dorsal horn, TRPA1 regulates transmission to spinal interneurons, and thereby pain hypersensitivity. Here we assessed whether the contribution of the spinal TRPA1 channel to pain hypersensitivity varies with the experimental pain model, properties of test stimulation or the behavioral pain response. The antihypersensitivity effect of intrathecally (i.t.) administered Chembridge‐5861528 (CHEM; a selective TRPA1 channel antagonist; 5–10 μg) was determined in various experimental models of pain hypersensitivity in the rat. In spinal nerve ligation and rapid eye movement (REM) sleep deprivation models, i.t. CHEM attenuated mechanical hypersensitivity. Capsaicin‐induced secondary (central) but not primary (peripheral) mechanical hypersensitivity was also reduced by i.t. administration of CHEM or A‐967079, another TRPA1 channel antagonist. Formalin‐induced secondary mechanical hypersensitivity, but not spontaneous pain, was suppressed by i.t. CHEM. Moreover, mechanical hypersensitivity induced by cholekystokinin in the rostroventromedial medulla was attenuated by i.t. pretreatment with CHEM. Independent of the model, the antihypersensitivity effect induced by i.t. CHEM was predominant on responses evoked by low‐intensity stimuli (⩽6 g). CHEM (10 μg i.t.) failed to attenuate pain behavior in healthy controls or mechanical hypersensitivities induced by i.t. administrations of a GABAA receptor antagonist, or NMDA or 5‐HT3 receptor agonists. Conversely, i.t. administration of a TRPA1 channel agonist, cinnamon aldehyde, induced mechanical hypersensitivity. The results indicate that the spinal TRPA1 channel exerts an important role in secondary (central) pain hypersensitivity to low‐intensity mechanical stimulation in various pain hypersensitivity conditions. The spinal TRPA1 channel provides a promising target for the selective attenuation of a central mechanism contributing to pathophysiological pain.

Gelsemine, a principal alkaloid from Gelsemium sempervirens Ait., exhibits potent and specific antinociception in chronic pain by acting at spinal α3 glycine receptors

TOC summary Gelsemine produces potent and specific antinociception in chronic pain states by activating spinal &agr;3 glycine receptors without inducing tolerance. Abstract The present study examined the antinociceptive effects of gelsemine, the principal alkaloid in Gelsemium sempervirens Ait. A single intrathecal injection of gelsemine produced potent and specific antinociception in formalin‐induced tonic pain, bone cancer‐induced mechanical allodynia, and spinal nerve ligation‐induced painful neuropathy. The antinociception was dose‐dependent, with maximal inhibition of 50% to 60% and ED50 values of 0.5 to 0.6 &mgr;g. Multiple daily intrathecal injections of gelsemine for 7 days induced no tolerance to antinociception in the rat model of bone cancer pain. Spinal gelsemine was not effective in altering contralateral paw withdrawal thresholds, and had only a slight inhibitory effect on formalin‐induced acute nociception. The specific antinociception of gelsemine in chronic pain was blocked dose‐dependently by the glycine receptor (GlyR) antagonist strychnine with an apparent ID50 value of 3.8 &mgr;g. Gelsemine concentration‐dependently displaced H3‐strychnine binding to the membrane fraction of rat spinal cord homogenates, with a 100% displacement and a Ki of 21.9 &mgr;M. Gene ablation of the GlyR &agr;3 subunit (&agr;3 GlyR) but not &agr;1 GlyR, by a 7‐day intrathecal injection of small interfering RNA (siRNA) targeting &agr;3 GlyR or &agr;1 GlyR, nearly completely prevented gelsemine‐induced antinociception in neuropathic pain. Our results demonstrate that gelsemine produces potent and specific antinociception in chronic pain states without induction of apparent tolerance. The results also suggest that gelsemine produces antinociception by activation of spinal &agr;3 glycine receptors, and support the notion that spinal &agr;3 glycine receptors are a potential therapeutic target molecule for the management of chronic pain.

A Series of d-Amino Acid Oxidase Inhibitors Specifically Prevents and Reverses Formalin-Induced Tonic Pain in Rats

We have found that mutation of d-amino acid oxidase (DAO) diminished formalin-induced tonic pain. The present research further studied the analgesic effects of a series of DAO inhibitors in this model. 5-Chlorobenzo[d]isoxazol-3-ol (CBIO), 4H-thieno[3,2-b]pyrrole-5-carboxylic acid (compound 8), 5-methylpyrazole-3-carboxylic acid (AS057278), sodium benzoate, and 4-nitro-3-pyrazole carboxylic acid (NPCA) inhibited rat spinal cord-derived DAO activity in a concentration-dependent manner, with maximal inhibition of 100% and potency rank of CBIO > compound 8 > AS057278 > sodium benzoate > NPCA. In rats, intrathecal injections of CBIO, compound 8, AS057278, and sodium benzoate but not NPCA specifically prevented formalin-induced tonic pain but not acute nociception, with the same potency order as in the DAO activity assay. The highly potent analgesia of DAO inhibitors was evidenced by CBIO, which prevented 50% pain at 0.06 μg, approximately 5-fold the potency of morphine. CBIO given after formalin challenge also reversed the established pain state to the same degree as prevention. The antihyperalgesic potencies of these DAO inhibitors were highly correlated to their inhibitions of spinal DAO activity. Maximum inhibition of pain by these compounds was approximately 60%, comparable with that of the N-methyl-d-aspartic acid receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801), suggesting that a larger portion of formalin-induced tonic pain is “DAO-sensitive,” whereas the remaining 40% of tonic pain and acute nociception is “DAO-insensitive.” These findings, combined with our previous DAO gene mutation and induction results, indicate spinal DAO mediates both induction and maintenance of formalin-induced tonic pain and further validate spinal DAO as a novel and efficacious target molecule for the treatment of chronic pain.

Site‐specific PEGylation of exenatide analogues markedly improved their glucoregulatory activity

Exenatide is a 39‐amino‐acid peptide widely used to manage type 2 diabetes mellitus. However, it has a short plasma half‐life and requires a twice daily injection regime. To overcome these drawbacks we used maleimide‐polyethylene glycol to induce site‐specific PEGylation.

Pain-related behavior following REM sleep deprivation in the rat : Influence of peripheral nerve injury, spinal glutamatergic receptors and nitric oxide

We assessed whether pain-related behavior in neuropathic or control rats is changed following rapid eye movement sleep deprivation (REMSD). Furthermore, we determined the contribution of spinal glutamatergic receptors and nitric oxide to sensitivity changes following REMSD versus peripheral nerve injury. Pain behavior was assessed in Sprague-Dawley (SD) and Hannover-Wistar (HW) rats with a spinal nerve ligation or a sham operation. Nerve ligation produced mechanical hypersensitivity of the injured dermatome in all animals. Baseline sensitivity to mechanical stimulation was higher in the HW than the SD group, independent of nerve injury. In both strains, mechanical sensitivity of neuropathic and sham-operated animals was increased following 48 h of REMSD. Heat sensitivity of an uninjured dermatome was not different among experimental conditions. Reversal of mechanical hypersensitivity was attempted in HW rats by spinal administration of an antagonist of the metabotropic glutamate receptor 5 (mGluR(5)) or the NMDA receptor and a nitric oxide synthase (NOS) inhibitor. Mechanical hypersensitivity induced by REMSD in unoperated rats was attenuated by all three drugs, while in neuropathic animals the mechanical anti-hypersensitive effect was most pronounced with the antagonist of the mGluR(5) or a NOS inhibitor. The results indicate that the strain of the animals markedly influences baseline withdrawal threshold to mechanical stimulation. Mechanical hypersensitivity following REMSD, however, is similarly increased in HW and SD strains, and the REMSD-associated increase in mechanical sensitivity is independent of nerve injury. Furthermore, mechanical hypersensitivities following REMSD and peripheral nerve injury share common spinal mechanisms involving, at least, the mGluR(5) and nitric oxide.

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.

EGT1442, a potent and selective SGLT2 inhibitor, attenuates blood glucose and HbA1c levels in db/db mice and prolongs the survival of stroke-prone rats

Sodium glucose co-transporter 2 (SGLT2) is a renal type III integral membrane protein that co-transports sodium and glucose from filtrate to epithelium in the proximal tubule. Human subjects with homozygous or compound heterozygous mutations in SLC5A2 exhibit glucosuria without hypoglycemia or other obvious morbidity, suggesting that blockade of SGLT2 has the potential to promote normalization of blood glucose without hypoglycemia in the setting of type 2 diabetes. This report presents the in vitro and in vivo pharmacological activities of EGT1442, a recently discovered SGLT2 inhibitor in the C-aryl glucoside class. The inhibitory effects of EGT1442 for human SGLT1 and SGLT2 were evaluated in an AMG uptake assay and the in vivo efficacy of treatment with EGT1442 was investigated in rats and dogs after a single dose and in db/db mice after chronic administration. The effect of EGT1442 on median survival of SHRSP rats was also evaluated. The IC(50) values for EGT1442 against human SGLT1 and SGLT2 are 5.6μM and 2nM, respectively. In normal rats and dogs a saturable urinary glucose excretion was produced with an ED(50) of 0.38 and 0.09mg/kg, respectively. Following chronic administration to db/db mice, EGT1442 dose-dependently reduced HbA(1c) and blood glucose concentration without affecting body mass or insulin level. Additionally, EGT1442 significantly prolonged the median survival of SHRSP rats. EGT1442 showed favorable properties both in vitro and in vivo and could be beneficial to the management of type 2 diabetic patients.

Role of spinal 5-HT receptors in cutaneous hypersensitivity induced by REM sleep deprivation.

Previous studies indicate that rapid eye movement (REM) sleep deprivation facilitates pain sensitivity. Since serotoninergic raphe neurons are involved both in regulation of sleep and descending pain modulation, we studied whether spinal 5-HT receptors have a role in sleep deprivation-induced facilitation of pain-related behavior. REM sleep deprivation of 48h was induced by the flower pot method in the rat. The pain modulatory influence of various serotoninergic compounds administered intrathecally was assessed by determining limb withdrawal response to monofilaments. REM sleep deprivation produced a marked hypersensitivity. Sleep deprivation-induced hypersensitivity and normal sensitivity in controls were reduced both by a 5-HT(1A) receptor antagonist (WAY-100635) and a 5-HT(2C) receptor antagonist (RS-102221). An antagonist of the 5-HT(3) receptor (LY-278584) failed to modulate hypersensitivity in sleep-deprived or control animals. Paradoxically, sensitivity in sleep-deprived and control animals was reduced not only by a 5-HT(1A) receptor antagonist but also by a 5-HT(1A) receptor agonist (8-OHDPAT). The results indicate that serotoninergic receptors in the spinal cord have a complex role in the control of sleep-deprivation induced cutaneous hypersensitivity as well as baseline sensitivity in control conditions. While endogenous serotonin acting on 5-HT(1A) and 5-HT(2C) receptors may facilitate mechanical sensitivity in animals with a sleep deprivation-induced hypersensitivity as well as in controls, increased activation of spinal 5-HT(1A) receptors by an exogenous agonist leads to suppression of mechanical sensitivity in both conditions. Spinal 5-HT(3) receptors do not contribute to cutaneous hypersensitivity induced by sleep deprivation.

Lamiophlomis rotata, an Orally Available Tibetan Herbal Painkiller, Specifically Reduces Pain Hypersensitivity States through the Activation of Spinal Glucagon-like Peptide-1 Receptors

Background:Lamiophlomis rotata is an orally available Tibetan herb prescribed for the management of pain, with shanzhiside methylester (SM) and 8-O-acetyl-SM as quality control ingredients. This study aimed to evaluate the antinociceptive properties of L. rotata, determine whether SM and 8-O-acetyl-SM are principle effective ingredients, and explore whether L. rotata produces antinociception through activation of spinal glucagon-like peptide-1 receptors (GLP-1Rs). Methods:Formalin test, neuropathic pain, and bone cancer pain models were used, and the animal sample size was 5 to 6 in each group. Hydrogen peroxide–induced oxidative damage was also assayed. Results:The L. rotata aqueous extract blocked formalin-induced tonic hyperalgesia and peripheral nerve injury– and bone cancer–induced mechanical allodynia by 50 to 80%, with half-effective doses of 130 to 250 mg/kg, close to the human dosage. The herb was not effective in alleviating acute nociceptive pain. A 7-day gavage with L. rotata aqueous extract did not lead to antiallodynic tolerance. Total iridoid glycosides, rather than total flavonoids, were identified by the activity-tracking method as effective ingredients for antihyperalgesia, whereas both SM and 8-O-acetyl-SM were principal components. Further demonstrations using the GLP-1R antagonist and gene silencer against GLP-1R at both the spinal and the cellular levels indicated that L. rotata inhibited pain hyperactivity by activation of spinal GLP-1Rs, and SM and 8-O-acetyl-SM appeared to be orthosteric, reversible, and fully intrinsic agonists of both rat and human GLP-1Rs. Conclusions:Results support the notion that the activation of spinal GLP-1Rs leads to specific antinociception in pain hypersensitivity and further suggest that GLP-1R is a human-validated target molecule for the treatment of chronic pain.

The non-peptide GLP-1 receptor agonist WB4-24 blocks inflammatory nociception by stimulating β-endorphin release from spinal microglia

Two peptide agonists of the glucagon‐like peptide‐1 (GLP‐1) receptor, exenatide and GLP‐1 itself, exert anti‐hypersensitive effects in neuropathic, cancer and diabetic pain. In this study, we have assessed the anti‐allodynic and anti‐hyperalgesic effects of the non‐peptide agonist WB4‐24 in inflammatory nociception and the possible involvement of microglial β‐endorphin and pro‐inflammatory cytokines.