Yukitoshi Niiyama

Bone cancer increases transient receptor potential vanilloid subfamily 1 expression within distinct subpopulations of dorsal root ganglion neurons.

Bone cancer pain has a strong impact on the quality of life of patients but is difficult to treat. Therefore, the mechanisms of bone cancer pain require elucidation for the purpose of development of new therapeutics. A recent study showed that activation of transient receptor potential vanilloid subfamily 1 (TRPV1) was involved in bone cancer pain. In this study, we re-evaluated the analgesic effects of pharmacological blockade of TRPV1 using the potent TRPV1 antagonist 5-iodoresiniferatoxin (I-RTX) and examined whether bone cancer can change TRPV1 expression and distribution in the primary sensory neurons in a mouse model of bone cancer pain. Implantation of osteosarcoma into the femur induced ongoing and movement-evoked bone cancer-related pain behaviors. These behaviors were significantly reduced by i.p. administration of I-RTX, compared with vehicle. Western blot and reverse transcription-polymerase chain reaction (RT-PCR) analyses revealed that TRPV1 level was significantly increased in dorsal root ganglions (DRGs) ipsilateral to sarcoma implantation. Immunohistochemical analysis showed that implantation of osteosarcoma induced not only an increase in the percentage of TRPV1-positive neurons among DRG neurons (24.3+/-1.3% in sham mice and 31.2+/-1.3% in mice with osteosarcoma implantation, P<0.05) but also an overall shift in the distribution of area of profiles to the right. Colocalization study showed that the percentages of colocalization of TRPV1 with neurofilament 200 kD (NF200) and calcitonin gene-related peptide (CGRP) but not isolectin B4 (IB4) among DRG neurons in mice with osteosarcoma implantation were increased compared with those in sham mice (from 0.8+/-0.1% to 2.1+/-0.3% for TRPV1 and NF200 and from 21.1+/-1.3% to 26.5+/-0.2% for TRPV1 and CGRP). In conclusion, TRPV1 activation plays a critical role in the generation of bone cancer pain, and bone cancer increases TRPV1 expression within distinct subpopulation of DRG neurons. These findings may lead to novel strategies for the treatment of bone cancer pain.

Down-regulation of mu opioid receptor expression within distinct subpopulations of dorsal root ganglion neurons in a murine model of bone cancer pain.

Although micro opioid receptor (MOR) agonists are used for treatment of most types of pain, a recent study has suggested that the sensitivity of bone cancer pain to systemic morphine was lower than that of inflammatory pain. However, the reasons for this have remained unclear. In this study, MOR expression and the analgesic effects of morphine in a bone cancer model were compared with those in an inflammatory pain model. A bone cancer pain model and an inflammatory pain model were made by implantation of sarcoma cells into the intramedullary space of the femur and hind-paw injection of complete Freunds adjuvant (CFA), respectively. In a behavioral study, sarcoma-implanted mice showed flinching behavior of magnitude comparable to that induced by CFA injection. The flinching behavior of sarcoma-implanted mice was less sensitive to intrathecal morphine than that of CFA-injected mice. Western blot analysis showed that MOR expression in the dorsal root ganglion (DRG) ipsilateral to sarcoma implantation was significantly reduced, while that in the DRG ipsilateral to CFA injection was increased. In sarcoma-implanted mice, the percentage of MOR-positive DRG neuronal profiles was lower than that in control mice (30.3% vs. 45.2%). In particular, MOR expression was reduced among calcitonin gene-related peptide- and transient receptor potential vanilloid subfamily 1-positive DRG neuronal profiles, which are considered to be involved in the generation of bone cancer pain (from 61.5% to 41.5% and from 72.1% to 48.4%, respectively). These results suggest that down-regulation of MOR in the distinct populations of DRG neurons contributes to the fact that higher doses of morphine are needed to produce analgesia in bone cancer as compared with those used in non-malignant inflammatory situations.

SB366791, a TRPV1 antagonist, potentiates analgesic effects of systemic morphine in a murine model of bone cancer pain

BACKGROUND Bone cancer pain has a major impact on the quality of life of cancer patients but is difficult to treat. Therefore, development of a novel strategy for bone cancer pain is needed for improvement of the patient quality of life. In this study, we examined the analgesic effects of the combination of a transient receptor potential vanilloid subfamily 1 (TRPV1) antagonist and morphine on pain-related behaviours in a murine model of bone cancer pain. METHODS C3H/HeJ mice underwent injection of osteolytic sarcoma cells into the intramedullary space of the femur. The analgesic effects of intraperitoneal morphine and the analgesic effect of a TRPV1 antagonist, SB366791 [N-(3-methoxyphenyl)-4-chlorocinnamide], on bone cancer pain-related behaviours were examined. The analgesic effects of the combination of SB366791 and morphine on bone cancer pain were also examined. RESULTS Intraperitoneal morphine significantly reduced the number of spontaneous flinches and improved ambulation only at the highest dose of 10 mg kg(-1) whereas weight-bearing was not improved. Intraperitoneal SB366791 at doses of 0.3 and 1.0 mg kg(-1), but not at a dose of 0.1 mg kg(-1), reduced the number of spontaneous flinches, whereas neither weight-bearing nor ambulation was improved. Addition of a sub-analgesic dose of SB366791 (0.1 mg kg(-1)) to morphine significantly reduced the number of flinches and improved weight-bearing compared with the effects of morphine alone. CONCLUSIONS Our findings showed that the combination of morphine and SB366791 has potent analgesic effects on bone cancer pain. The findings of this study may lead to novel strategies for the treatment of bone cancer pain.

Contribution of transient receptor potential vanilloid subfamily 1 to endothelin-1-induced thermal hyperalgesia.

Endothelin-1 (ET-1) plays an important role in peripheral pain processing. However, the mechanisms of the nociceptive action of ET-1 have not been fully elucidated. In this study, we investigated the contribution of transient receptor potential vanilloid subfamily 1 (TRPV1) to ET-1-induced thermal hyperalgesia. Intraplantar ET-1-induced thermal hyperalgesia was examined by assessing the paw withdrawal latency to noxious heat stimuli. In electrophysiological study, whole-cell patch-clamp recordings were performed to investigate the interaction of ET-1 and TRPV1 using human embryonic kidney 293 (HEK293) cells expressing endothelin type A receptor (ET(A)) and TRPV1. Intraplantar ET-1 (3, 10 and 30 pmol) produced thermal hyperalgesia in a dose-dependent manner. Thermal hyperalgesia was attenuated by the inhibition of ET(A) and protein kinase C (PKC) but not that of ET(B). ET-1-induced thermal hyperalgesia was significantly attenuated in TRPV1-deficient mice compared with that in wild-type mice. In voltage-clamp experiments, 10 nM capsaicin evoked small inward currents in HEK293 cells expressing TRPV1 and ET(A). In the presence of ET-1, capsaicin produced much larger current responses (P<0.05). Mutation at PKC-specific TRPV1 phosphorylation sites (S800A/S502A) and PKC inhibitors inhibited the potentiating effect of ET-1. In addition, ET-1 decreased the temperature threshold for TRPV1 activation in a PKC-dependent manner (from 41.0+/-0.4 degrees C to 32.6+/-0.6 degrees C). In addition, Western blot analysis was also performed to confirm ET-1-induced phosphorylation of TRPV1. Incubation of ET-1 and intraplantar ET-1 evoked phosphorylation of TRPV1 in HEK293 cells expressing TRPV1 and ET(A) and the skin, respectively. These results suggest that the sensitization of TRPV1 activity through an ET(A)-PKC pathway contributes to ET-1-induced thermal hyperalgesia.

Suppression of bone marrow–derived microglia in the amygdala improves anxiety-like behavior induced by chronic partial sciatic nerve ligation in mice

Summary Bone marrow–derived microglia aggregated in the central nucleus of the amygdala are involved in the chronic neuropathic pain–induced anxiety. ABSTRACT Chronic neuropathic pain causes abnormal sensitivities such as hyperalgesia and allodynia, and emotional abnormalities such as anxiety and depression. Although spinal cord microglia are involved in abnormal sensitivity to neuropathic pain, no previous studies have examined the mechanism of neuropathic pain–induced anxiety. Here, we examined the involvement of bone marrow (BM)–derived microglia aggregated in the amygdalae of mice with chronic neuropathic pain in the development of anxiety‐like behavior. We prepared partial sciatic nerve ligations (PSNL) in mice that received bone marrow transplantation from green fluorescent protein (GFP)–Tg mice after irradiation with head protection, and examined GFP‐positive microglia in the central nuclei of the amygdalae (CeA). On day 28 after PSNL, BM‐derived microglia aggregated in the CeA concurrent with anxiety‐like behavior. BM‐derived microglia in the CeA highly expressed interleukin (IL)–1&bgr; and C‐C chemokine receptor type 2 (CCR2). In addition, neurons in the CeA highly expressed monocyte chemotactic protein–1 (MCP‐1), a ligand for CCR2, in PSNL‐treated mice compared to sham‐operated mice, suggesting that the MCP‐1/CCR2 axis is involved in the recruitment of BM‐derived microglia. Oral administration of a CCR2 antagonist decreased the number of BM‐derived microglia in the CeA, and successfully reversed the anxiety‐like behavior and hypersensitivity to mechanical stimuli in PSNL‐treated mice. Microinjections of an IL‐1&bgr; receptor antagonist directly into the CeA successfully reversed the anxiety‐like behavior in the PSNL‐treated mice even though the neuropathic pain persisted. These results suggest that the recruitment of BM‐derived microglia to the CeA via the MCP‐1/CCR2 axis and neuron–microglia interactions might be important in the pathogenesis of neuropathic pain–induced anxiety.

Reduction of bone cancer pain by activation of spinal cannabinoid receptor 1 and its expression in the superficial dorsal horn of the spinal cord in a murine model of bone cancer pain.

Background:Bone cancer pain has a strong impact on the quality of life of patients, but it is difficult to treat. Therefore, development of a novel strategy for the treatment of bone cancer pain is needed for improvement of patient quality of life. This study examined whether selective spinal cannabinoid receptor 1 (CB1) activation alleviates bone cancer pain and also examined the spinal expression of CB1. Methods:A bone cancer pain model was made by implantation of sarcoma cells into the intramedullary space of the mouse femur. In behavioral experiments, the authors examined the effects of activation of spinal CB1 and inhibition of metabolism of endocannabinoid on bone cancer-related pain behaviors. Immunohistochemical experiments examined the distribution and localization of CB1 in the superficial dorsal horn of the spinal cord using specific antibodies. Results:Spinal CB1 activation by exogenous administration of a CB1 agonist arachidonyl-2-chloroethylamide reduced bone cancer–related pain behaviors, including behaviors related to spontaneous pain and movement-evoked pain. In immunohistochemical experiments, although &mgr;-opioid receptor 1 expression was reduced in the superficial dorsal horn ipsilateral to the site of implantation of sarcoma cells, CB1 expression was preserved. In addition, CB1 was mainly expressed in the axon terminals, but not in the dendritic process in the superficial dorsal horn. Conclusion:Spinal CB1 activation reduced bone cancer–related pain behavior. Presynaptic inhibition may contribute to the analgesic effects of spinal CB1 activation. These findings may lead to novel strategies for the treatment of bone cancer pain.

The addition of epidural morphine to ropivacaine improves epidural analgesia after lower abdominal surgery

PurposeTo assess the analgesic and side effects of the continuous epidural infusion of 0.2% ropivacaine combined with morphine compared to both drugs alone.MethodsIn this study, both observers and patients were blinded to patient group assignment. Sixty patients scheduled to undergo lower abdominal surgery were enrolled. Patients were randomized to one of three postoperative treatment groups: 1) combination group (a combination of 0.2% ropivacaine and 0.003% morphine); 2) morphine group (0.003% morphine); or 3) ropivacaine group (0.2% ropivacaine). Postoperatively, all solutions were administered epidurally at a rate of 6 mL·hr-1 for 24 hr. Patients were given iv flurbiprofen as a supplemental analgesic on demand.ResultsThe combination group showed lower visual analogue scale scores than those of patients receiving either drug alone, both at rest and on coughing. The combination group showed a slight motor block at two hours after the continuous epidural infusion, while the ropivacaine and morphine groups did not show any motor block. The incidence of itching was significantly increased in the morphine and combination groups, compared to the ropivacaine group. There was no significant difference between the numbers of patients with nausea in the three groups. No hypotension or respiratory complications were observed in the three groups.ConclusionThe combination of epidural 0.2% ropivacaine and 0.003% morphine has more effective analgesic effects than either of the drugs alone for postoperative pain relief after lower abdominal surgery.RésuméObjectifComparer l’effet analgésique et les effets secondaires d’une perfusion péridurale continue de ropivacaïne à 0,2 % combinée à la morphine à ceux des deux médicaments employés seuls.MéthodeDans notre étude, la répartition des sujets s’est faite à l’insu des observateurs et des patients. Nous avons recruté 60 patients devant subir une intervention abdominale basse. Ils ont été répartis en trois groupes: 1) combinaison médicamenteuse (ropivacaïne à 0,2% et morphine à 0,003 %); 2) morphine (morphine à 0,003 %); 3) ropivacaïne (ropivacaïne à 0,2 %). Après l’opération, toutes les solutions ont été administrées par voie péridurale au débit de 6 mL·h-1 pendant 24 h. Les patients ont reçu du flurbiprofène iv comme analgésique complémentaire sur demande.RésultatsAvec la combinaison de médicaments, comparée à chacun employé seul, on a noté des scores plus bas à l’échelle visuelle analogique et ce, au repos et pendant la toux. La combinaison de médicaments a produit un léger blocage moteur deux heures après la perfusion péridurale continue, mais la ropivacaïne et la morphine n’en ont pas provoqué. L’incidence de prurit a été significativement accrue avec la morphine et la combinaison médicamenteuse, comparées à la ropivacaïne. Il n’y a pas eu de différence intergroupe significative pour l’incidence des nausées. Dans les trois groupes, aucun cas d’hypotension ou de complications respiratoires n’a été observé.ConclusionLa combinaison de ropivacaïne à 0,2 % et de morphine à 0,003 % a des effets analgésiques plus efficaces que l’un ou l’autre médicament seul pour le soulagement des douleurs après une opération abdominale basse.

Landiolol, esmolol and propranolol protect from ischemia/reperfusion injury in isolated guinea pig hearts

PurposeBeta blockers are thought to exert beneficial effects on the ischémic heart. The authors examined the effects of landiolol (ONO 1101), a highly selective βl antagonist, propranolol, a nonspecific β blocker, and esmolol, a selective β l antagonist, on postischemic contractile recovery. Drugs were given prophylactically.MethodsIschemia-reperfusion in isolated guinea pig hearts was induced by stopping the perfusion for 45 min and reperfusing for 60 min. Hearts (n = 7 in each group) were treated with or without propranolol (1 or 10μM), esmolol (5 or 50μM), or landiolol (20, 100 or 500 μM) ten minutes before inducing ischemia.ResultsAt the end of reperfusion, left ventricular pressure (LVP) recovered to 64 ± 3% of the baseline value in the control group. With 1 and 10 μM propranolol, LVP recovered to 90 ± 5% and 100 ± 6% of the baseline value at 60 min after reperfusion, respectively. FiftyμM but not 5μM of esmolol resulted in restoration of LVP to 97 ± 17% of the pre-ischemic value at 60 min after reperfusion. In hearts pretreated with 100 and 500 μM landiolol, LVP was restored to 109 ± 5% and 104 ± 596 of the baseline value, respectively. Landiolol 100μM did not depress LVP in the pre-ischemic period.ConclusionsThe present study shows that landiolol, an ultrashort-acting cardioselective β l blocker, has cardioprotective effects on ischemia-reperfusion injury in isolated guinea pig hearts. All three β blockers were equally protective but the intermediate dosage of landiolol preserved LVP during the pre-ischemic period.RésuméObjectifOn croit que les bêtabloquants ont des effets bénéfiques sur le cœur ischémique. Les auteurs ont vérifié les effets du iandioioi (ONO 1101), un antagoniste βl hautement sélectif, du propranolol, un β bloquant non spécifique, et de l’esmolol, un antagoniste β l sélectif, sur la récupération contractile postischémique. Les médicaments ont été administrés préventivement.MéthodeLischémie-reperfusion a été induite dans des cœurs de cobaye isolés en stoppant la perfusion pendant 45 min et en reperfusant pendant 60 min. Les cœurs (n = 7 dans chaque groupe) ont été traités avec ou sans propranolol (1 ou 10 μM), esmolol (5 ou 50 μM) ou landiolol (20, 100 ou 500 μM) dix minutes avant l’induction de l’ischémie.Résultats:ÀLa fin de la reperfusion, la pression ventriculaire gauche (PVG) avait retrouvé 64 ± 3 % de sa valeur de base dans le groupe témoin. Avec I et 10 μM de propranolol, la PVG a été récupérée respectivement à 90 ± 5 % et 100 ± 6 %, 60 min après la reperfusion. La dose de 50 μM, non celle de 5 μM, d’esmolol a été suivie de la restauration de la PVG à 91 ± 17% de la valeur préischémique, 60 min après la reperfusion. Dans les cœurs prétraités avec 100 et 500 μM de landiolol, la PVG a été restaurée à 109 ± 5 % et à 104 ± 5 % de sa valeur de base, respectivement. La dose de 100 μM de landiolol n’a pas causé de baisse de la PVG en période préischémique.ConclusionLa présente étude montre que le landiolol, un β l bloquant cardiosélectif à action très brève, a des effets cardioprotecteurs sur les lésions résultant de l’ischémie-reperfusion dans des cœurs de cobaye isolés. Les trois β bloquants ont été également protecteurs, mais la dose intermédiaire de landiolol a diminué la PVG pendant la période préischémique.

The utility of three-dimensional computed tomography in unanticipated difficult endotracheal intubation.

IMPLICATIONS We experienced a case of unanticipated difficult intubation with direct laryngoscopy because of narrowing of the retropharyngeal air space and laryngeal vestibulum. It is suggested that three-dimensional computed tomography is useful for evaluating both the abnormality of an airway and its relationship to surrounding tissue.

Immunohistochemical analysis of acid-sensing ion channel 2 expression in rat dorsal root ganglion and effects of axotomy.

Several studies have suggested that acid-sensing ion channel 2 (ASIC2) plays a role in mechanoperception and acid sensing in the peripheral nervous system. We examined the expression and distribution of ASIC2 in the rat dorsal root ganglion, the co-localization of ASIC2 with tropomyosin-related kinase (trk) receptors, and the effects of axotomy on ASIC2 expression. ASIC2 immunoreactivity was observed in both neurons and satellite cells. ASIC2-positive neurons accounted for 16.5 +/- 2.4% of the total neurons in normal dorsal root ganglion. Most ASIC2-positive neurons were medium-to-large neurons and were labeled with neurofilament 200 kD (NF200). Within these neurons, ASIC2 was not evenly distributed throughout the cytoplasm, but rather was accumulated prominently in the cytoplasm adjacent to the axon hillock and axonal process. We next examined the co-localization of ASIC2 with trk receptors. trkA was expressed in few ASIC2-positive neurons, and trkB and trkC were observed in 85.2% and 53.4% of ASIC2-positive neurons, respectively, while only 6.9% of ASIC2-positive neurons were co-localized with trkC alone. Peripheral axotomy markedly reduced ASIC2 expression in the axotomized dorsal root ganglion neurons. On the other hand, intense ASIC2 staining was observed in satellite cells. These results show that ASIC2 is expressed in the distinct neurochemical population of sensory neurons as well as satellite cells, and that peripheral axotomy induced marked reductions in ASIC2 in neurons.