Evidence that gabapentin reduces neuropathic pain by inhibiting the spinal release of glutamate

Gabapentin is an anticonvulsant that successfully treats many neuropathic pain syndromes, although the mechanism of its antihyperalgesic action remains elusive. This study aims to help delineate gabapentin's antihyperalgesic mechanisms. We assessed the effectiveness of gabapentin at decreasing mechanical and cold hypersensitivity induced in a rat model of neuropathic pain. Thus, we compared the effectiveness of pre‐ or post‐treatment with systemic or intrathecal (i.t.) gabapentin at reducing the development and maintenance of the neuropathic pain symptoms. Gabapentin successfully decreased mechanical and cold hypersensitivity, both as a pretreatment and post‐treatment. Furthermore, both i.t. and systemic administration of gabapentin were effective in reducing the behavioral hypersensitivity; however, the i.t. administration was superior to the systemic. We also examined gabapentin's effects at inhibiting hindpaw formalin‐induced release of excitatory amino acids (EAAs) in the spinal cord dorsal horn (SCDH) both in naïve rats and in rats with neuropathic pain. We present the first evidence that gabapentin reduces the formalin‐induced release of both glutamate and aspartate in SCDH. Furthermore, i.t. gabapentin reduces the enhanced noxious stimulus‐induced spinal release of glutamate seen in neuropathic rats. These data suggest that gabapentin reduces neuropathic pain symptoms by inhibiting the release of glutamate in the SCDH.     

Endogenous methionine enkephalin may play an anticonvulsant role in the seizure-susceptible El mouse

After the intracisternal injection of three protease inhibitors which prevent the degradation of methionine enkephalin (amastatin, Des-Pro²-bradykinin, and phosphoramidon) and a mixture of these protease inhibitors, we investigated the effect on convulsive seizures in the seizure-susceptible El mouse. We also measured the cerebral methionine enkephalin content by high-performance liquid chromatography coupled with radioimmunoassay. Protease inhibitors significantly decreased both the incidence of seizures and the seizure score in El mice in a dose-dependent manner. This anticonvulsant effect was reversed by naloxone (2 mg/kg, sc). The cerebral methionine enkephalin content increased significantly after the administration of protease inhibitors in comparison with saline injection. These findings suggest that it was not protease inhibitors but instead increase of endogenous methionine enkephalin that reduced the incidence of seizures and the seizure score in El mice. Together with our previous data, the present findings support our hypothesis that a deficit in anticonvulsant endogenous methionine enkephalin is involved in the pathogenesis of seizures in the El mouse.     

Measurement of leucine enkephalin in caudate nucleus tissue with fast atom bombardment-collision activated dissociation-linked field scanning mass spectrometry

The endogenous amount of the opioid pentapeptide leucine enkephalin was measured in a canine caudate nucleus tissue extract using mass spectral analytical methods which retain absolute molecular specificity. Fast atom bombardment mass spectrometry generation of the protonated molecular ion of leucine enkephalin followed by collision activated dissociation produced amino acid sequence-determining ions. These amino acid sequence-determining ions were analyzed by a linked field (B/E) scan. One amino acid sequence-determining ion was selected to measure endogenous leucine enkephalin. This novel measurement mode offers optimal molecular specificity for quantification of an endogenous amount (451 pmol g-1 tissue) of leucine enkephalin in a biologic tissue extract of canine caudate nucleus.     

A comparative study of the effect of methionine enkephalin upon the stereospecific binding of an opiate agonist and an antagonist in mice and rats.

The synthetic enkephalins especially methionine enkephalin are more potent in inhibiting the stereospecific binding of ³H-dihydromorphine than that of ³H-naloxone in mouse brain homogenates. Methionine enkephalin is a more potent inhibitor of ³H-dihydromorphine binding in whole mouse brain homogenates than in washed mouse brain membranes. No difference was observed with regard to the inhibitory effect of methionine enkephalin on the binding of ³H-dihydromorphine in whole rat brain homogenates or washed rat brain membranes. The use of different radiolabelled drugs (agonist versus antagonist), different species (mouse versus rat) and/or the variation in the preparation (brain homogenates versus washed membranes) may account for the difference between the IC₅₀ of methionine enkephalin versus ³H-dihydromorphine and versus ³H-naloxone stereospecific binding. The increased inhibitory effect of methionine enkephalin when the supernatant was added to the washed brain membranes supports the hypothesis that methionine enkephalin may be one part of the real endogenous morphine ligand.     

Regulation of hypophysial secretion by endogenous opiates: Humoral endorphin stimulates the release of growth hormone

Humural endorphin, a recently discovered endogenous opioid factor stimulates the release of growth hormone and, to some extent of prolactin, similarly to other endogenous (enkephalin, β-endorphin) and exogenous (morphine) opiates. This stimulatory effect is dose-dependent with peak values at 30 minutes following intraventricular injection to newborn rats. However, in contrast to the other opioid ligands, the effect of humoral endorphin is not blocked in a dose-dependent fashion by naloxone, the potent opiate antagonist. Thus, while moderate doses of naloxone partially inhibit the stimulatory effect, higher doses which completely block morphine, enkephalin and β-endorphin, are ineffective in antagonizing humoral endorphin. This peculiar interaction between naloxone and humoral endorphin resembles the effect of the opiate antagonist on spontaneous release of growth hormone and prolactin, suggesting the involvement of humoral endorphin in the physiological regulation of hypophysial secretion.     

Role of endogenous opioids in soman (pinacolyl methylphosphonofluoridate)-induced antinociception

The effect of soman poisoning on the levels of methionine enkephalin and beta-endorphin in mice and rats were determined. Soman poisoning produced no significant effect on methionine enkephalin levels in the striatum of rats or mice or beta-endorphin levels in the pituitary gland of mice. In rats beta-endorphin levels were significantly reduced 24 hr post soman poisoning, but returned to control levels by 48 hr. In vitro, the hydrolysis of leucine enkephalin by aminopeptidase was virtually complete by 30 min and found to be the major route of degradation. The release of TYR-GLY-GLY in the presence or absence of puromycin (10 microM) was found to be low (less than or equal to 2.0%). A minor effect on TYR release in the presence of GLY-GLY-PHE-MET (50 microM) was insignificant. Preincubation of mouse striatum homogenates with soman (1 or 10 microM) did not inhibit the hydrolysis of leucine enkephalin. These results suggest that the long term antinociception following soman exposure is not due to either altered concentration of endogenous opioid-like substances or inhibition of the enzymes responsible for their degradation.     

生物活性肽——蛋氨酸脑啡肽的免疫

源自肾上腺前脑啡肽原的具有吗啡样生物活性的内源性神经肽蛋氨酸脑啡肽(methionine enkephalin,MENK),由５个氨基酸残基Tyr Gly Gly Phe Met组成,与G蛋白偶联的７次跨膜特异性受体结合后发挥不同的生物学功能。目前,对蛋氨酸脑啡肽的作用及其机制研究已经取得了很大进展。本文就MENK对于免疫系统的调节及其对肿瘤、自身免疫病、艾滋病等免疫系统相关疾病的治疗作用予以综述。     

Dopaminergic Regulation of Enkephalin Release

The effects of dopamine receptor stimulation on enkephalin release were evaluated in vitro and in vivo by measuring the changes in the levels of [Met5]enkephalin (YGGFM) and Tyr-Gly-Gly (YGG), a characteristic extracellular enkephalin metabolite produced under the action of enkephalinase. In rat striatal slices, D1-receptor agonists or antagonists did not modify enkephalin release. By contrast, D2-receptor agonists enhanced the potassium-induced release of YGGFM and YGG without affecting spontaneous release from nondepolarized slices. This response was prevented by the D2-receptor antagonists haloperidol and RIV 2093, the latter compound being more potent, which suggested the involvement of a putative D2-receptor subtype. Acute administration of apomorphine or selective D2-receptor agonists, but not that of a D1-receptor agonist, enhanced the steady-state level of YGG without affecting the YGGFM level in rat striatum. The effect was blocked selectively by D2-receptor antagonists which, administered alone, had no effect. These observations indicate that D2-receptor stimulation in vitro or in vivo facilitates enkephalin release from striatal neurons, but that endogenous dopamine does not exert any tonic influence upon the opioid peptide neuron activity under basal conditions. However, chronic administration of haloperidol resulted in increases in striatal YGGFM and YGG, an effect presumably reflecting a long-term adaptive process.     

抗阿片肽血清对催产素增强电针镇痛的影响

本工作观察了大鼠侧脑室注射抗阿片肽血清对催产素增强电针镇痛作用的影响。脑室注射抗Beta-内啡肽血清(AEPS)使电针镇痛效应明显降低,但在注射催产素前预先注射AEPS对催产素增强电针镇痛作用无明显影响。注射抗强啡肽Al-13血清使电针镇痛明显降低,但却使催产素增强电针镇痛作用明显增强。无论抗甲-脑啡肽血清还是抗亮-脑啡肽血清对电针镇痛都无明显影响,对催产素增强电针镇痛的作用也不产生影响。上述结果说明,催产素增强电针镇痛的作用虽被脑内强啡肽Al-13部分对抗,但不受Beta-内啡肽和脑啡肽的影响,表明催产素增强针刺镇痛的作用不依赖于脑内内源性阿片肽系统。     

Heterogeneity of endogeneous opiates: H-endorphin is not correlated with enkephalin or with beta-endorphin

H-endorphin, an endogenous opiate present in brain, blood and cerebrospinal fluid of rats and humans is distinguishable from both enkephalin and β-endorphin. Its distribution within the rats' brain resembles, but does not overlap that of enkephalin. Its stability and presence in blood is similar to that of β-endorphin; however, its blood level is not altered under extreme conditions (stress, hypophysectomy) which change dramatically the level of β-endorphin. H-endorphin is almost equally potent in three bio-assay systems (guinea pig ileum, mouse vas deferans and rat vas deferens) thus dissociable from both typical μ (morphine-like) and δ (enkephalin-like) ligands. The distinct characteristics of H-endorphin, enkephalin and β-endorphin, differing in their anatomical distribution, physiological response and pharmacological effect, indicate the heterogeneity of the internal opioid system which consists of at least three different mediators.     

Differential Effects of Long-Term Electroconvulsive Shock on Brain Levels of Enkephalin and Humoral-Endorphin

Abstract: Electroconvulsive shock (ECS) administrations repeated for 10 consecutive days cause an elevation in the opioid content of the rat brain. Two different endogenous opioids, enkephalin and humoral-endorphin, undergo independent changes that differ in both their time course and intracerebral localization. These metabolic changes parallel long-term behavioral modifications such as the development and dissipation of tolerance to the analgesic effect of ECS. The activation of two different, independent, endogenous opioid systems by ECS is in agreement with previous behavioral and pharmacological studies.     

贝类等无脊椎动物脑啡肽研究进展

贝类等许多无脊椎动物体内含有脑啡肽等阿片肽系统,并且在其生长、繁殖等生理生化过程中发挥着重要的作用。本文从脑啡肽在贝类等无脊椎动物体内的定位、分离提取、测序鉴定及其相关酶的研究等方面,对贝类等无脊椎动物脑啡肽的研究进展进行了概述。目前来看,脑啡肽物质在无脊椎动物中有关各种功能和机理研究仍然处于初步阶段,对于其深层次的分子作用机理还需要作进一步的研究。     

Analysis of interaction between etoricoxib and tramadol against mechanical hyperalgesia of spinal cord injury in rats

Drug combinations have the potential advantage of greater analgesia over monotherapy. The present study was aimed to assess any possible interaction (additive or potentiation) in the antinociceptive effects of etoricoxib; a novel cyclooxygenase-2 inhibitor, and tramadol; a typical opioid agonist when administered in combination against mechanical hyperalgesia induced by spinal cord injury in rats. The nature of interaction was analyzed using surface of synergistic interaction (SSI) analysis and an isobolographic analysis. Etoricoxib or tramadol when administered alone to rats, exhibited different antihyperalgesic potencies (ED50 etoricoxib: 0.58+/-0.19 mg/kg, po; ED50 tramadol: 9.85+/-0.57 mg/kg, po). However, both the drugs were found to be long acting against this model of hyperalgesia. Further, etoricoxib and tramadol were co-administered in fixed ratios of ED50 fractions. One combination (0.29/4.79 mg/kg, po: etoricoxib/tramadol) exhibited additivity and other three combinations (0.15/2.39, 0.08/1.19, and 0.04/0.59 mg/kg, po: etoricoxib/tramadol) resulted in potentiation when analyzed by SSI. The SSI was calculated from the total antihyperalgesic effect produced by the combination after the subtraction of the antihyperalgesic effect produced by each of the individual drug. In the isobolographic analysis, the experimental ED50 was found to be far below the line of additivity also indicating a significant (P < 0.05) synergistic antihyperalgesic effect when etoricoxib and tramadol was co-administered to rats. The synergistic antihyperalgesic effect of etoricoxib and tramadol combination suggests that these combinations may have clinical utility in mechanical hyperalgesia associated with spinal injury.     

Motilin and a synthetic enkephalin induce colonic motor complexes (CMC) in the conscious dog

In the conscious dog migrating and nonmigrating colonic motor complexes (CMC) were recorded by means of chronically implanted strain gauge force transducers. Intravenous injection of a synthetic enkephalin analogue immediately induced a premature CMC at all three recording sites of the colon. Naloxone inhibited the enkephalin- but not motilin-induced CMC. We therefore exclude that motilin acts by release of endogenous enkephalins. The two peptides stimulate CMC in the canine colon by different pathways. Naloxone alone had no effect on normal colonic motility, suggesting strongly that endogenous enkephalins do not modulate regular CMC in the dog.     

Ethanol-Induced Increase in Endogenous Dopamine Release May Involve Endogenous Opiates

The effect of opiate peptides on basal and potassium-stimulated endogenous dopamine (DA) release from striatal slices was studied in vitro. Dual stimulation of the striatal slices gave a reproducible increase in DA release that was calcium dependent. Addition of the delta-opiate receptor agonists Met5-enkephalin, [D-Ala2,D-Leu5]enkephalin (DADLE), and [D-Ser2]Leu-enkephalin-Thr (DSLET), increased the basal DA release without affecting potassium-stimulated release in a dose-dependent manner. The effect of DADLE was antagonized by the addition of naloxone. In contrast, the mu-opioid receptor agonist [D-Ala2,N-MePhe4,Gly-ol5]enkephalin (DAGO) and the epsilon-opioid agonist beta-endorphin inhibited the stimulated DA release without changing the basal release. The inhibitory effect of DAGO on potassium-stimulated release was antagonized by naloxone. The addition of ethanol (75 mM) to the incubation media produced a delayed increase of both the basal and stimulated DA release. There was no change in stimulated DA release when the change in basal release was subtracted, suggesting that ethanol produced a dose-dependent, selective increase in basal DA release. Naloxone and the selective delta-opiate antagonist ICI 174864 inhibited the ethanol-induced increase in basal DA release. Naloxone and ICI 174864 added alone did not alter either basal or stimulated DA release. We therefore suggest that the ethanol-induced increase in basal DA release is an indirect effect involving an endogenous delta-opiate agonist.     

生物活性肽—蛋氨酸脑啡肽免疫调节作用研究进展

蛋氨酸脑啡肽( methionine-enkephaIin,Met-ENK)是具有内源性吗啡样活性物质,它是由5个氨基酸残基组成的多肽,其第5位氨基酸残基为蛋氨酸.目前,对Met-ENK的生物学功能及其作用机制的研究已取得了显著进展.就Met-ENK对免疫细胞调控、抗肿瘤作用等方面做一综述.     

Central administration of oxytocin reduces hyperalgesia in mice: Implication for cannabinoid and opioid systems

The neuropeptide oxytocin (OXT) contributes to the regulation of diverse cognitive and physiological functions including nociception. Indeed, OXT has been reported to be analgesic when administered directly into the brain, the spinal cord, or systemically. Although many authors have reported the analgesic effects of OXT, its mechanism has not been well elucidated. Recently, it has been also hypothesize that OXT, increasing intracellular concentration of calcium, could regulate the production of mediators, like endocannabinoids (eCB). It has been well documented that eCB are able to suppress pain pathways. The present study investigates the effect of OXT in paw carrageenan-induced pain. Intracerebroventricular (icv) administration of OXT, but neither intraperitoneal nor intraplantar route, induces an antihyperalgesic effect increasing paw withdrawal latency to mechanical or thermal stimuli. Our results clearly demonstrate that 3 and 6 h following carrageenan challenge, central administration of OXT (30 ng/mouse) shows a significant antihyperalgesic activity. Moreover, for the first time, we demonstrate that CB1 receptor plays a key role in the antihyperalgesic effect of OXT. In fact our results show CB1 antagonist, but not the specific CB2 antagonist reduce OXT-induced antihyperalgesic effect. In addition, our data show that central OXT administration is able to reduce carrageenan-induced hyperalgesia but does not modify carrageenan-induced paw edema. Finally, using opioid antagonists we confirm an important role of opioid receptors. In conclusion, our experiments suggest that central administration of OXT reduces hyperalgesia induced by intraplantar injection of carrageenan, and this effect may work via cannabinoid and opioid systems.     

Opioid peptides and primary biliary cirrhosis.

Patients with liver disease have increased plasma concentrations of the endogenous opioid peptides methionine enkephalin and leucine enkephalin. As an initial investigation to determine whether opioid peptides contribute to any of the clinical manifestations of hepatic disease nalmefene, a specific opioid antagonist devoid of agonist activity, was given to 11 patients with cirrhosis. They all experienced a severe opioid withdrawal reaction on starting the drug. In the nine patients with primary biliary cirrhosis pruritus was greatly alleviated, fatigue seemed to improve, and plasma bilirubin concentration, which had been rising, showed a modest fall in all except one patient. These results indicate that blocking opioid receptors has an effect on some of the metabolic abnormalities of liver disease.     

Partial depletion of endogenous zinc level by (D-Pen2, D-Pen5)enkephalin in the rat brain.

The effects of the potent delta opioid agonist (D-Pen2, D-Pen5)enkephalin (DPDPE) were studied on the endogenous levels and regional distribution of Zn2+ in rat central nervous system by means of flame atomic absorption spectrophotometry. The olfactory bulb exhibited the highest Zn2+ level, followed by the frontal and parietal cortices, striatum and hippocampus; the lowest ion levels were found in the medulla and thoracic spinal cord. Intracerebroventricular administration of DPDPE resulted in significant, time- and dose-dependent decreases in endogenous Zn2+ contents in the parietal cortex, hippocampus and striatum. The action of DPDPE was antagonized by a 30 min naloxone pretreatment. Naloxone alone was without effect in eliciting these responses. Thus, delta opioid receptors may regulate or modulate endogenous Zn2+ levels in the rat brain.     

The metabolism of neuropeptides. Both phosphoramidon-sensitive and captopril-sensitive metallopeptidases are present in the electric organ of Torpedo marmorata.

A membrane fraction from the electric organ of Torpedo marmorata hydrolyses the Gly3-Phe4 bond of [D-Ala2, Leu5]enkephalin as well as the Gly-His bond of benzoyl-Gly-His-Leu. The hydrolysis of benzoyl-Gly-His-Leu is completely inhibitable by Captopril (I50 = 19nM), consistent with peptidyl dipeptidase activity, but enkephalin hydrolysis is inhibited to a maximum of only 70%. The residual activity hydrolysing enkephalin is inhibited by phosphoramidon (I50 = 15nM) and therefore resembles endopeptidase-24.11, a mammalian plasma-membrane enzyme implicated in the metabolism of neuropeptides. Both enkephalin-hydrolysing activities in Torpedo electric organ are inhibited by 1,10-phenanthroline, like their mammalian counterparts. The peptidases may function in the hydrolysis of endogenous peptides or in neurotransmitter exocytosis in the electric organ.