Yoshi Tsukiyama

Sleep disturbances in a neuropathic pain-like condition in the mouse are associated with altered GABAergic transmission in the cingulate cortex

&NA; Insomnia is a common problem for people with chronic pain. Cortical GABAergic neurons are part of the neurobiological substrate that underlies homeostatic sleep regulation. In the present study, we confirmed that sciatic nerve ligation caused thermal hyperalgesia and tactile allodynia in mice. In this experimental model for neuropathic pain, we found an increase in wakefulness and a decrease in non‐rapid eye movement sleep under a neuropathic pain‐like state. Under these conditions, membrane‐bound GABA (γ‐aminobutyric acid) transporters (GATs) on activated glial fibrillary acidic protein‐positive astrocytes were significantly increased in the cingulate cortex, and extracellular GABA levels in this area after depolarization were rapidly decreased by nerve injury. Furthermore, sleep disturbance induced by sciatic nerve ligation was improved by the intracingulate cortex injection of a GAT‐3 inhibitor. These findings provide novel evidence that sciatic nerve ligation decreases extracellular‐released GABA in the cingulate cortex of mice. These phenomena may, at least in part, explain the insomnia in patients with neuropathic pain. Neuropathic pain‐like stimuli suppress the GABAergic transmission with increased GABA (γ‐aminobutyric acid) transporters located on activated astrocytes in the cingulate cortex related to sleep disturbance.

Effects of gabapentin on brain hyperactivity related to pain and sleep disturbance under a neuropathic pain-like state using fMRI and brain wave analysis.

Neuropathic pain is the most difficult pain to manage in the pain clinic, and sleep problems are common among patients with chronic pain including neuropathic pain. In the present study, we tried to visualize the intensity of pain by assessing neuronal activity and investigated sleep disturbance under a neuropathic pain‐like state in mice using functional magnetic resonance imaging (fMRI) and electroencephalogram (EEG)/electromyogram (EMG), respectively. Furthermore, we investigated the effect of gabapentin (GBP) on these phenomena. In a model of neuropathic pain, sciatic nerve ligation caused a marked decrease in the latency of paw withdrawal in response to a thermal stimulus only on the ipsilateral side. Under this condition, fMRI showed that sciatic nerve ligation produced a significant increase in the blood oxygenation level‐dependent (BOLD) signal intensity in the pain matrix, which was significantly decreased 2 h after the i.p. injection of GBP. Based on the results of an EEG/EMG analysis, sciatic nerve‐ligated animals showed a statistically significant increase in wakefulness and a decrease in nonrapid eye movement (NREM) sleep during the light phase, and the sleep disturbance was almost completely alleviated by a higher dose of GBP in nerve‐ligated mice. These findings suggest that neuropathic pain associated with sleep disturbance can be objectively assessed by fMRI and EEG/EMG analysis in animal models. Furthermore, GBP may improve the quality of sleep as well as control pain in patients with neuropathic pain. Synapse 2011.

A beta-adrenoceptor agonist evokes a nitric oxide-cGMP relaxation mechanism modulated by adenylyl cyclase in rat aorta : Halothane does not inhibit this mechanism

Background The objective of this study was to characterize the effects of halothane on the agonist-induced nitric oxide-cyclic GMP (NO-cGMP) mechanisms by comparing the intracellular signal transduction mediating isoproterenol- and acetylcholine-induced nitric oxide formation. Methods Isoproterenol-induced relaxations of rat aortic rings with and without endothelia were examined in the absence and presence of halothane. Studies were also done in the presence of inhibitors of nitric oxide-synthase, adenylyl cyclase, calmodulin, protein kinase A, and intracellular Calcium2+ release mechanism. The relaxations under some of these conditions were compared with those induced by acetylcholine. Cyclic nucleotide contents of the rings were also measured. Results Isoproterenol relaxed aortic rings via the endothelium-dependent nitric oxide-cyclic GMP mechanism. Inhibition of adenylyl cyclase or of protein kinase A attenuated the isoproterenol-induced relaxations significantly but did not affect those induced by acetylcholine. Inhibition of intracellular Calcium2+ release abolished the acetylcholine-induced relaxations but did not affect those induced by isoproterenol. Calmodulin inhibition attenuated both agonist-induced relaxations significantly. Unlike acetylcholine-induced relaxation, that induced by isoproterenol was not affected by halothane. Isoproterenol increased both the cyclic adenosine monophosphate and cGMP contents of rings significantly when endothelia were intact. Inhibition of nitric oxide synthase attenuated the isoproterenol-induced cGMP content increases significantly but did not affect the cyclic adenosine monophosphate content increases. Halothane (2%) did not affect isoproterenol-induced increases in nucleotide content. Conclusions Isoproterenol-induced nitric oxide formation requires the activation of constitutive nitric oxide synthase, but the Calcium sup 2+ release mechanism is not involved in activating this enzyme. Halothane can inhibit the nitric oxide-cyclic GMP mechanism only when Calcium2+ release is greatly involved in the activation of constitutive nitric oxide synthase.

Halothane inhibition of acetylcholine-induced relaxation in rat mesenteric artery and aorta.

PurposeT1he effect of halothane was compared on acetylcholine (ACh)-induced relaxation of the mesentenc artery and the aorta in ratsMethodsThe responses of isolated rat aortic and mesentenc artenal nng segments precontracted with phenylephnne to ACh (10−8– 10−5M), in the presence of halothane 0–3%. were compared using isometric force tension recordings. Effects of NG -nitro-l-arginine (L-NOARG, 3 × 10−5), methylene blue (MB, 5 × 10−6 M), oxyhaemoglobin (OxyHB, 10−7 M), and vanous potassium channel inhibitors; tetraethylammonium (TEA, 10−5 M. 10−3 M), apamin (AP, 10−7 M), charybdotoxin (ChTx, 10−7M) and glibenclamide (GC, 10−5M) on ACh-induced relaxation in mesentenc artery were tested. Using radioimmunoassay, ACh (10−6M)-induced guanosine 3′:5′-cyclic monophosphate (cGMP) accumulation of mesentenc arterial rings pretreated with L-NAORG were also measured.ResultsL-NOARG partially inhibited ACh-induced relaxation in mesentenc arterial rings (P < 0.05. maximum relaxation reduced by approximately 50%), whereas it abolished them in aortic rings. The remaining relaxation resistant to L-NOARG in mesentenc artenal rings was insensitive to additional MB or OxyHB, and was not accompanied by increases in cGMP contents of rings. Halothane inhibited endothelium-dependent relaxation in aorta and mesentenc artenal rings. This inhibitory effect was larger in aorta. Halothane also inhibited NO independent EDHF-dependent relaxation in the mesentenc arterial rings.ConclusionDespite a similar inhibitory effect on the EDHF relaxing pathway, halothane has a larger effect on endothelium-dependent relaxation in the aorta (NO dependent mainly) than in the mesenteric rings (NO and EDHF dependent).RésuméObjectifComparer sur l’artère mésenténque et sur l’aorte du rat l’influence de l’halothane sur la vasodilatation induite par l’acétylcholine (ACh).MéthodesOn a comparé les réactions d’anneaux arténels isolés de segments d’aorte et d’artère mésentérique préalablement contractés avec de la phényléphnne à l’ACh (10−8–10−5M), en présence d’halothane à 0 à 3% sur des enregistrements de la force de la tension isométrique. On a vérifié les effets de la NG-nitro-l-arginine (L-NOARG, 3 × 10−5 M), du bleu de méthylène (MB. 5 × 10−6 M), de l’oxyhémoglobine (OxyHB, 10−7 M) et de plusieurs inhibiteurs des canaux potassiques: le tétraéthylammonium (TEA, 10−5M, 10−3 M), l’apamine (AP, 10−7M), la charybdotoxine (ChTx, 10−7 M) et le glibenclamide (GC, 10−5M) sur la vasodilatation de l’artère mésentérique induite par l’ACh. Le radioxmmunodosage a servi en outre à mesurer l’accumulation de guanosine 3′:′5 — monophosphatase cyclique (cGMP) dans les anneaux arténels mésentériques prétraités à la L-NOARG.RésultatsLa L-NOARG n’abolissart que partiellement la vasodilatation des anneaux arténels mésentériques induite par l’ACh (P < 0,05. réduction de la dilatation maximale d’environ 50%) alors qu’elle abolissait celle des anneaux aortiques. La vasodilatation résiduelle L-NOARG-résistante des anneaux artériels mésentériques ne réagissait pas à l’ajout de MB ou d’OxyHB et ne s’accompagnait pas d’une augmentation du contenu en cGMP des anneaux. L’inhibition par l’halothane de la vasodilatation endothélium-dépendante des anneaux aortiques et artériels mésentériques était plus importante dans l’aorte. L’halothane inhibait aussi la vasodilatation NO-indépendante EDHP-dépendante des anneaux mésentériques.ConclusionMalgré des effets inhibiteurs similaires sur les voies de la vasodilation EDHF, l’halothane a un effet plus prononcé sur la vasodilatation endothélium-dépendante au niveau de l’aorte (principalement NO-dépendante) qu’au niveau des anneaux mésentériques (NO-et EDHF-dépendante).

Sigma‐1 receptor function is critical for both the discriminative stimulus and aversive effects of the kappa‐opioid receptor agonist U‐50488H

The present study was undertaken to identify possible similarities between the effects of kappa‐opioid receptor agonist, N‐methyl‐D‐aspartate‐receptor antagonist, and sigma receptor agonist on the discriminative stimulus effects of U‐50488H, and the possible involvement of sigma receptors in the discriminative stimulus and aversive effects of U‐50488H. The kappa‐opioid receptor agonist U‐50488H produced significant place aversion as measured by the conditioned place preference procedure, and this effect was completely abolished by treatment with the putative sigma‐1 receptor antagonist NE‐100. In addition, phencyclidine (+)‐SKF‐10047 and (+)‐pentazocine, which are sigma receptor agonists, generalized to the discriminative stimulus effects of U‐50488H in rats that had been trained to discriminate between U‐50488H (3.0 mg/kg) and saline. Furthermore, NE‐100 significantly attenuated the discriminative stimulus effects of U‐50488H and the U‐50488H‐like discriminative stimulus effects of phencyclidine. These results suggest that the sigma‐1 receptor is responsible for both the discriminative stimulus effects and aversive effects of U‐50488H.

Role of dopamine D2 and D3 receptors in mediating the U-50,488H discriminative cue: comparison with methamphetamine and cocaine

Substitutions of the dopamine D2 or D3 receptor agonists for the discriminative stimulus effect induced by U‐50,488H, methamphetamine (METH) and cocaine in rats were examined. The D2 receptor agonist R‐propylnorapomorphine [(−)‐NPA] failed to substitute for U‐50,488H cue, while the D3 receptor‐preferred agonist (+/–)‐7‐hydroxy‐dipropylaminotetralin hydrobromide (7‐OH‐DPAT) produced dose‐related increases in drug‐appropriate responding up to 0.03 mg/kg, which fully substituted. At doses greater than 0.03 mg/kg of 7‐OH‐DPAT, there was a dose‐dependent decrease in the percentage of responses on the U‐50,488H‐appropriate lever. Furthermore (−)‐NPA and 7‐OH‐DPAT at high doses substituted for the discriminative stimulus effect induced by both METH and cocaine, indicating that 7‐OH‐DPAT at high doses may interact with D2 receptors. These results suggest that the stimulation of D2 receptor may be critical for the production of the discriminative stimulus effect induced by METH and cocaine, whereas the stimulation of D3 receptor may contribute to the production of the U‐50,488H cue.

Upregulation of bradykinin receptors is implicated in the pain associated with caerulein-induced acute pancreatitis.

Although the way for pain management associated with acute pancreatitis has been searched for, there are not enough medications available for it. The aim of the present study was to investigate the role of bradykinin (BK) in pain related to acute pancreatitis. After repeated injections of caerulein (50 μg/kg and 6 times), mice showed edema in the pancreas, and blood concentrations of pancreatic enzymes (amylase and lipase) were clearly elevated. A histopathological study demonstrated that caerulein caused tissue damage characterized by edema, acinar cell necrosis, interstitial hemorrhage, and inflammatory cell infiltrates. Furthermore, the mRNA levels of interleukin‐1β and monocyte chemotactic protein (MCP)‐1 were significantly increased in the pancreas of caerulein‐treated mice. The sensitivity of abdominal organs as measured by abdominal balloon distension was enhanced in caerulein‐injected mice, suggesting that caerulein caused pancreatic hyperalgesia. Moreover, repeated treatment with caerulein resulted in cutaneous tactile allodynia of the upper abdominal region as demonstrated by the use of von Frey filaments, indicating that caerulein‐treated mice exhibited referred pain. Under this condition, the mRNA levels of bradykinin B1 receptor (BKB1R) and bradykinin B2 receptor (BKB2R) were significantly increased in the dorsal root ganglion (DRG). Finally, we found that des‐Arg9‐(Leu8)‐bradykinin (BKB1R antagonist) and HOE‐140 (BKB2R antagonist) attenuated the acute pancreatitis pain‐like state in caerulein‐treated mice. These findings suggest that the upregulation of BK receptors in the DRG may, at least in part, contribute to the development of the acute pancreatitis pain‐like state in mice. Synapse, 2011.

Effects of halothane and isoflurane on acetylcholine-induced, endothelium-dependent vasodilation in perfused rat mesenteric arterial beds

AbstractPurpose. The present study was designed to examine the effects of halothane and isoflurane on acetylcholine-induced, endothelium-dependent vasodilation in rat mesenteric arterial beds perfused at a constant flow both in vitro and in situ. Methods. In the in-vitro preparation, the mesenteric artery was cannulated and perfused (5 ml·min−1). The perfusion pressure was continuously monitored. Under active tone induced by methoxamine, the effects of halothane and isoflurane on the vasodilator response to acethylcholine in either the presence or absence of NG-nitro-L-arginine (L-NA), tetraethylammonium (TEA), or KCl (30 mM)-depolarization were examined. All experiments in these preparations were performed in the presence of indomethacin (10 mM). In the in-situ experimental model, rats were anesthetized with pentobarbital and the lungs were mechanically ventilated via a tracheostomy with a ventilator. The superior mesenteric artery was cannulated and used for the monitoring of the perfusion pressure. Blood shunting with constant flow (2 ml·min−1) from the carotid artery to the superior mesenteric artery was introduced with clamping at the immediately distal portion of the mesenteric artery branching. Following 20-min ventilation with halothane or isoflurane at 1 minimum alveolar concentration (MAC) in oxygen, acetylcholine was given from the mesenteric artery, under active tone induced by norepinephrine (100 mg·kg−1·hr−1). Results. In the in-vitro preparation, the nitric oxide synthase inhibitor, L-NA (100 μM) did not affect vasodilations to acetylcholine (1, 10 nM), while the K+ channel inhibitor TEA (10 mM), as well as KCl (30 mM), significantly reduced these vasodilations. However, only in the presence of L-NA, TEA and KCl completely abolished the vasodilations produced by acetylcholine. The higher concentrations of halothane (2.0%, 3.0%), but neither isoflurane (3.0%) nor the lower concentration of halothane (1.0%), significantly impaired vasodilator responses to acetylcholine in the presence of L-NA, whereas the volatile anesthetics did not affect these vasodilations in the absence of L-NA. Halothane (2.0%) did not alter the vasodilation produced by acetylcholine in the presence of TEA or KCl. In the in-vivo preparation, the vasodilator effects of acetylcholine (1 and 10 nmol) were not affected by the inhalation of halothane (1.0%) or isoflurane (1.3%). Conclusion. These results suggest that, in resistance arteries in conditions of constant flow, halothane and isoflurane do not affect vasodilations in response to an endothelium-dependent agonist. However, in these preparations, once the enzymatic activity of nitric oxide synthase is inhibited, higher concentrations of halothane, but neither isoflurane nor the lower concentration of halothane, appear to impair endothelium-dependent relaxations, probably mediated by TEA-sensitive K+ channels.