Dong K. Ahn

Intratrigeminal ganglionic injection of LPA causes neuropathic pain-like behavior and demyelination in rats

ABSTRACT We have previously reported a novel method for producing chronic nociceptive behavior in rats following compression of the trigeminal ganglion. In the present study, we have further studied the role of demyelination in the development of prolonged nociceptive behavior in the trigeminal territory. For this purpose, lysophosphatidic acid (LPA) was injected into the trigeminal ganglia of male Sprague–Dawley rats weighing between 250 and 260 g. Under pentobarbital sodium anesthesia, the rats were mounted onto a stereotaxic frame and 3 μL of LPA (1 nmol) solution was injected into the trigeminal ganglion to produce demyelination. This treatment decreased the air‐puff thresholds both ipsilateral and contralateral to the injection site, which persisted until postoperative day 100 and returned to the preoperative levels 130 days after the LPA injection. The LPA injection also produced a significant ipsilateral hyper‐responsiveness to pin‐prick stimulation. The effects of DGPP, an LPA1/3 receptor antagonist, and Y‐27632, a Rho kinase inhibitor, upon LPA‐induced mechanical allodynia and hyperalgesia were also investigated. Pretreatment with DGPP blocked both mechanical allodynia and ipsilateral hyperalgesia. However, pretreatment with Y‐27632 blocked only ipsilateral and contralateral mechanical allodynia. These results thus indicate that a targeted blockade of LPA receptor and Rho kinase pathways are potentially important new treatments for demyelination‐induced trigeminal neuralgia‐like nociception.

Intracisternal administration of mitogen-activated protein kinase inhibitors reduced mechanical allodynia following chronic constriction injury of infraorbital nerve in rats.

The present study investigated the role of mitogen-activated protein kinase (MAPK) in orofacial neuropathic pain following chronic constriction injury of the infraorbital nerve (ION-CCI). Experiments were carried out on male Sprague-Dawley rats weighing between 200 and 230 g. The ION was separated from adhering tissue, and two ligatures (5-0 chromic gut) were tied loosely around it. We examined the air-puff thresholds (mechanical allodynia), scores of pinprick (mechanical hyperalgesia), and face grooming frequency for acetone application (hypersensitivity for cold stimulation) - 3, 3, 6, 9, 12, 15, 20, 25, 30, and 40 days after surgery. ION-CCI produced mechanical allodynia, hyperalgesia, and cold hypersensitivity. We investigated whether administration of MAPKs inhibitors blocks ION-CCI-induced mechanical allodynia. Intracisternal administration with PD98059 or SB203580, a MEK inhibitor or a p38 MAPK inhibitor, respectively, significantly inhibited ION-CCI-induced mechanical allodynia in the orofacial area. These results indicate that the ION-CCI produced behavioral alterations in the orofacial area and those central MAPKs pathways contribute to orofacial neuropathic pain. Our findings suggest that MAPKs inhibitors have a potential role in treatment for orofacial neuropathic pain.

Blockade of central cyclooxygenase (COX) pathways enhances the cannabinoid-induced antinociceptive effects on inflammatory temporomandibular joint (TMJ) nociception

Abstract The present study is the first to investigate the participation of central cyclooxygenase (COX) pathways in modulating the antinociceptive effects of intracisternally administered cannabinoid on nociception induced by inflammation of the temporomandibular joint (TMJ) in freely moving rats. Following intra‐articular injection of 5% formalin in the TMJ, nociceptive scratching behavior was recorded for nine successive 5‐min intervals in Sprague–Dawley rats. Intracisternal injection of 30 μg of WIN 55,212‐2, a synthetic non‐subtype‐selective CB1/2 agonist, administered 20 min prior to formalin injection significantly reduced the number of scratches and duration of scratching induced by formalin compared with the vehicle‐treated group. Antinociceptive effect of WIN 55,212‐2 was blocked by intracisternal injection of 10 μg of AM251, a CB1 receptor‐selective antagonist, but not by AM630, a CB2 receptor‐selective antagonist. A 10 μg dose of WIN 55,212‐2 that was ineffective in producing antinociception became effective following intracisternal administration of NS‐398, a selective COX‐2 inhibitor; indomethacin, a non‐selective COX 1/2 inhibitor; acetaminophen, a putative COX‐3 inhibitor, but not following pretreatment with the selective COX‐1 inhibitor, SC‐560. The ED50 value of WIN 55,212‐2 in the NS‐398‐treated group was significantly lower than that in the vehicle‐treated group. Importantly, administration of low doses of COX inhibitors alone did not attenuate nociception. These results indicate that inhibition of central COX pathways, presumably via COX‐2 inhibition, reduces inflammatory pain by enhancing the cannabinoid‐induced antinociceptive effect. Based on our observations, combined administration of cannabinoids with COX inhibitors may hold a therapeutic promise in the treatment of inflammatory TMJ pain.

Role of peripheral group I and II metabotropic glutamate receptors in IL-1β-induced mechanical allodynia in the orofacial area of conscious rats

&NA; The present study investigated the role of peripheral group I and II metabotropic glutamate receptors (mGluRs) in interleukin‐1β (IL‐1β)‐induced mechanical allodynia in the orofacial area. Experiments were carried out on Sprague–Dawley rats weighing between 230 and 280 g. After subcutaneous administration of 0.01, 0.1, 1, or 10 pg of IL‐1β, we examined withdrawal behavioral responses produced by 10 successive trials of a ramp of air‐puffs pressure applied ipsilaterally or contralaterally to the IL‐1β injection site. The thresholds of air puffs were measured 10, 30, 60, 120, or 180 min after 25 μl of IL‐1β was administered through an implanted tube. Subcutaneous injection of IL‐1β produced bilateral mechanical allodynia. While the IL‐1β‐induced mechanical allodynia was blocked by pretreatment with an IL‐1 receptor antagonist, the IL‐1β‐induced mirror‐image mechanical allodynia was not blocked by an IL‐1 receptor antagonist injected into the contralateral side. Subcutaneous administration of CPCCOEt or LY367385, an mGluR1 antagonist, or MPEP or SIB1893, an mGluR5 antagonist, 10 min prior to injection of IL‐1β abolished IL‐1β‐induced mechanical allodynia. Pretreatment with APDC or DCG4, a group II mGluR agonist, blocked the IL‐1β‐induced mechanical allodynia. The anti‐allodynic effect induced by APDC was inhibited by pretreatment with LY341495, a group II mGluR antagonist. These results suggest that peripheral group I and II mGluRs participate in IL‐1β‐induced mechanical allodynia in the orofacial area. Peripheral group I mGluR antagonists blocked the IL‐1β‐induced mechanical allodynia, while peripheral group II mGluR agonists produced anti‐allodynic effects on IL‐1β‐induced mechanical allodynia in the orofacial area of rats.

Behavioral evidence for the differential regulation of p-p38 MAPK and p-NF-κB in rats with trigeminal neuropathic pain

BackgroundWe investigated the differential regulation of p-p38 MAPK or p-NF-κB in male Sprague-Dawley rats with inferior alveolar nerve injury resulting from mal-positioned dental implants. For this purpose, we characterized the temporal expression of p-p38 MAPK or p-NF-κB in the medullary dorsal horn and examined changes in nociceptive behavior after a blockade of p-p38 MAPK or p-NF-κB pathways in rats with trigeminal neuropathic pain.ResultsUnder anesthesia, the left lower second molar was extracted and replaced with a mini dental implant to intentionally injure the inferior alveolar nerve. Western and immunofluorescence analysis revealed that p-p38 MAPK is upregulated in microglia following nerve injury and that this expression peaked on postoperative day (POD) 3 through 7. However, the activation of p-NF-κB in astrocyte peaked on POD 7 through 21. The intracisternal administration of SB203580 (1 or 10 μg), a p38 MAPK inhibitor, on POD 3 but not on POD 21 markedly inhibits mechanical allodynia and the p-p38 MAPK expression. However, the intracisternal administration of SN50 (0.2 or 2 ng), an NF-κB inhibitor, on POD 21 but not on POD 3 attenuates mechanical allodynia and p-NF-κB expression. Dexamethasone (25 mg/kg) decreases not only the activation of p38 MAPK but also that of NF-κB on POD 7.ConclusionsThese results suggest that early expression of p-p38 MAPK in the microglia and late induction of p-NF-κB in astrocyte play an important role in trigeminal neuropathic pain and that a blockade of p-p38 MAPK at an early stage and p-NF-κB at a late stage might be a potential therapeutic strategy for treatment of trigeminal neuropathic pain.

Compression of the trigeminal ganglion produces prolonged nociceptive behavior in rats

The present study is the first demonstration of prolonged nociceptive behavior in the trigeminal region following compression of the trigeminal ganglion in rats. Experiments were carried out on male Sprague–Dawley rats mounted onto a stereotaxic frame under pentobarbital sodium anesthesia. For compression of the trigeminal ganglion, a 4% agar solution (8μl) was injected into the trigeminal ganglion through a stainless steel injector (24 gauge), which extended 2mm beyond the end of a guide cannula (21 gauge). Following agar injection, the injector and guide cannula were removed. In the control group, rats were sham operated without agar injection. Air‐puff thresholds (mechanical allodynia), pin prick responses (mechanical hyperalgesia), and spontaneous scratching behavior were examined 3 days before surgery and at 3, 7, 10, 14, 17, 21, 24, 30, and 40 days after surgery. Data were analyzed using a repeated measures ANOVA followed by multiple group comparisons using the LSD post‐hoc test. Air‐puff thresholds significantly decreased after compression of the trigeminal ganglion. Mechanical allodynia was established within 3 days and lasted beyond postoperative day 24. Mechanical hyperalgesia was also evident 3 days after compression and persisted until the 40th postoperative day. Although mechanical allodynia and hyperalgesia appeared bilaterally, the ipsilateral side was significantly more sensitive. Intraperitoneal treatment with carbamazepine significantly blocked mechanical allodynia produced by compression of the trigeminal ganglion. These findings suggest that prolonged nociceptive behavior following compression of the trigeminal ganglion may mimic trigeminal neuralgia in this animal model.

Central cyclooxygenase inhibitors reduced IL-1β-induced hyperalgesia in temporomandibular joint of freely moving rats

&NA; Microinjection of formalin (5%, 50 μl) into a temporomandibular joint (TMJ) causes noxious behavioral responses in freely moving rats. In the present study, we investigated the role of central cyclooxygenase (COX) pathways in IL‐1β‐induced hyperalgesia with formalin‐induced TMJ pain model. Intra‐articular injection of 100 pg or 1 ng of IL‐1β significantly facilitated formalin‐induced behavior by 130 or 174% in the number of scratches. Intracisternal administration of 100 pg or 1 ng of IL‐1β also significantly increased formalin‐induced behavior by 166 or 82% in the number of scratches. IL‐1β‐induced hyperalgesia was blocked by pretreatment with IL‐1 receptor antagonist. Intracisternal pretreatment with SC‐560, a selective COX‐1 inhibitor, or NS‐398, a selective COX‐2 inhibitor, abolished intra‐articular administration of IL‐1β‐induced hyperalgesic response. Intracisternal pretreatment with NS‐398, a selective COX‐2 inhibitor, abolished the intracisternal administration of IL‐1β‐induced hyperalgesic response, while pretreatment with SC‐560, a selective COX‐1 inhibitor, did not change IL‐1β‐induced hyperalgesic responses. On the other hand, pretreatment with acetaminophen, a tentative COX‐3 inhibitor, also abolished both intra‐articular and intracisternal administration of IL‐1β‐induced hyperalgesic responses. These results indicate that central COX‐2 plays important role in the central administration of IL‐1β‐induced hyperalgesia and that central COX‐1/2 pathways mediate peripheral administration of IL‐1β‐induced hyperalgesia in the TMJ. Central COX‐3 inhibitor seems to play an important role in the nociceptive process associated with both peripheral and central administration of IL‐1β‐induced hyperalgesia in TMJ. It is concluded that central acting of COX‐3 inhibitors may be of therapeutic value in the treatment of inflammatory pain in TMJ.

Low doses of cannabinoids enhance the antinociceptive effects of intracisternally administered mGluRs groups II and III agonists in formalin-induced TMJ nociception in rats

Abstract This study provides the first demonstration that central cannabinoids modulate the antinociceptive actions of metabotropic glutamate receptors (mGluRs) on formalin‐induced temporomandibular joint (TMJ) nociception. Noxious scratching behavior induced by formalin injection in the TMJ was used as a model of pain. Intracisternal injection of 30 μg of WIN 55,212‐2, a non‐subtype selective cannabinoid receptor agonist, attenuated the number of scratches by 75% as compared with the vehicle‐treated group, whereas vehicle alone or 3 or 10 μg of WIN 55,212‐2 had no effect. To explore the postulated interaction between central cannabinoid receptors and mGluRs, effects of combined administration of sub‐analgesic doses of WIN 55,212‐2 and group II or III mGluR agonists were tested. Group II or III mGluRs agonists were administered intracisternally 10 min after intracisternal administration of WIN 55,212‐2. Neither 100 nmol APDC, a group II mGluRs agonist, nor L‐AP4, a group III mGluR agonist, altered nociceptive behavior when given alone but significantly inhibited the formalin‐induced nociceptive behavior in the presence of a sub‐threshold dose (3 μg) of WIN 55,212‐2. The ED50 value of APDC or L‐AP4 was significantly reduced upon co‐treatment with WIN 55,212‐2 than in the vehicle‐treated group, highlighting the important therapeutic potential of the combined administration of group II or III mGluR agonists with cannabinoids to effectively treat inflammatory pain associated with the TMJ. Potentiating effects of group II or III mGluRs agonists will likely permit the administration of cannabinoids at doses that do not achieve significant accumulation to produce undesirable motor dysfunction.

Participation of peripheral group I and II metabotropic glutamate receptors in the development or maintenance of IL-1β-induced mechanical allodynia in the orofacial area of conscious rats

The present study investigated the role of peripheral groups I and II metabotropic glutamate receptors (mGluRs) in interleukin (IL)-1beta-induced mechanical allodynia in the orofacial area of rats. Subcutaneous injection of 10 pg of IL-1beta decreased air-puff thresholds ipsilateral or contralateral to the injection site. The decrease in air-puff thresholds appeared 10 min after the injection of IL-1beta and IL-1beta-induced mechanical allodynia persisted for over 3 h. Pre-treatment with 7-(hydroxyimino) cyclopropa[b] chromen-1a-carboxylate ethyl ester (CPCCOEt) or 2-methyl-6-(phenylethynyl)-pyridine hydrochloride (MPEP), a mGluR1 or mGluR5 antagonist, blocked IL-1beta-induced mechanical allodynia and mirror-image mechanical allodynia produced by a subcutaneous injection of 10 pg of IL-1beta. However, post-treatment with CPCCOEt or MPEP did not affect changes in behavioral responses, which were produced by the IL-1beta injection. Pre-treatment, as well as post-treatment with (2R,4R)-4-aminopyrrolidine-2,4-dicarboxylate (APDC), a group II mGluR agonist, blocked either IL-1beta-induced mechanical allodynia or mirror-image mechanical allodynia. The anti-allodynic effects of APDC were abolished by pre-treatment with (2S)-2-amino-2[(1S,2S)-2-carboxycycloprop-1-yl]-3-(xanth-9-yl) propanoic acid (LY341495), a group II mGluR antagonist. These results indicate that peripheral group II mGluRs are involved in the development and maintenance of IL-1beta-induced mechanical allodynia, while peripheral group I mGluRs are involved in the development of IL-1beta-induced mechanical allodynia. Based on our observations, the peripheral application of group II mGluR agonists may be of therapeutic value in treating inflammatory pain.

Intramuscular administration of morphine reduces mustard oil‐induced craniofacial muscle pain behavior in lightly anesthetized rats

The present study investigated the role of peripheral opioid receptors in mustard oil‐induced nociceptive behavior and inflammation in the masseter muscles of lightly anesthetized rats. Experiments were carried out on male Sprague–Dawley rats weighing between 300 and 400g. After initial anesthesia with sodium pentobarbital (40mg/kg, i.p.), one femoral vein was cannulated and connected to an infusion pump for the intravenous infusion of sodium pentobarbital. The rate of infusion was adjusted to provide a constant level of anesthesia. Mustard oil (MO, 30μl) was injected into the mid‐region of the left masseter muscle via a 30‐gauge needle. Intramuscularly‐administered morphine significantly reduced shaking behavior but not MO‐induced inflammation. Intramuscular pretreatment with naloxone, an opioid receptor antagonist, reversed antinociception produced by intramuscularly‐administered morphine, while intracisternal administration of naloxone did not affect the antinociception of peripheral morphine. Pretreatment with d‐Pen‐Cys‐Tyr‐d‐Trp‐Orn‐Thr‐Pen‐Thr‐NH2 (CTOP), a μ opioid receptor antagonist, but not naltrindole, a δ opioid receptor antagonist, nor norbinaltorphimine (nor‐BNI), a κ opioid receptor antagonist, reversed intramuscularly‐administered morphine‐induced antinociception. These results indicate that intramuscularly‐administered morphine produces antinociception in craniofacial muscle nociception and that this intramuscularly‐administered morphine‐induced antinociception is mediated by a peripheral μ opioid receptor. Our observations further support the clinical approach of administering opioids in the periphery for the treatment of craniofacial muscle nociception.