Blocking mu opioid receptors in the spinal cord prevents the analgesic action by subsequent systemic opioids

Systemically administered mu opioids may produce analgesia through inhibition of the ascending nociceptive transmission and activation of descending pathways. However, the relative importance of the spinal and supraspinal sites in the analgesic action of systemic opioids remains uncertain. It has been shown that systemic morphine can inhibit dorsal horn neurons independent of the descending system. In this study, we determined the extent to which spinal mu opioid receptors mediate the analgesic effect of systemic mu opioids. Rats were instrumented with an intrathecal catheter with the tip placed in the lumbar spinal cord. Nociception was measured by testing the paw withdrawal threshold in response to a noxious radiant heat or pressure stimulus. Surprisingly, intrathecal pretreatment with naloxone or H-D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP, a specific mu opioid receptor antagonist) completely blocked the inhibitory effect of intravenous morphine on mechanical nociception. Intrathecal naloxone or CTAP also abolished the effect of intravenous morphine on the withdrawal latency of the hindpaw, but not the forepaw, measured with a radiant heat stimulus. Furthermore, the inhibitory effect of subcutaneous fentanyl on mechanical nociception was eliminated by CTAP injected intrathecally. Intrathecal CTAP similarly abolished the effect of subcutaneous fentanyl on thermal nociception of the hindpaw but not the forepaw. Therefore, this study provides new information that when spinal mu opioid receptors are blocked, subsequent systemic administration of mu opioids fails to produce an analgesic effect. This finding highlights the important role of mu opioid receptors in the spinal cord in the antinociceptive action of opioids.     

The effects of aging on mu and delta opioid receptors in the spinal cord of Fischer-344 rats

Previous research has demonstrated that the antinociceptive efficacy of opioids decreases with advancing age. This study utilized radioligand binding techniques to determine if this decline is due to a change in the receptor density (B_max) and/or affinity (measured as K_d) of the mu (μ) and/or delta (δ) opioid receptors in the spinal cord with advancing age. Saturation binding analysis with [³H][ -Ala²,N-methyl-Phe⁴,Gly⁵-ol]enkephalin (DAMGO: a μ-opioid selective agonist) and [³H]naltrindole (a δ-opioid selective antagonist) revealed no age-related changes in B_max for either the μ or δ-opioid receptors. The K_d value for naltrindole was likewise unaffected by age. The K_d value for DAMGO however, was significantly higher in the aged group as compared with the young and mature groups, indicating a decreased affinity of spinal μ-opioid receptors for DAMGO.     

Novel function of bovine milk-derived lactoferrin on antinociception mediated by mu-opioid receptor in the rat spinal cord

Lactoferrin (LF) is a multifunctional protein that is found in milk, neutrophils, and other biological fluids. Although LF and the LF receptor have been identified in the central nervous system (CNS), the physiological role of LF remains unknown. We found that bovine milk-derived LF (BLF) reduces nociception in various pain models, as shown by the formalin test, hot plate test, and acetic acid writhing test in rats. Intraperitoneal (i.p.) administration of BLF significantly inhibited nociception in these pain models. These antinociceptive effects were also confirmed in BLF-fed rats. The antinociceptive effects of BLF were blocked by naloxone treatment, even though prostaglandin E(2) (PGE(2)) production in the ascites fluid that accumulated during the writhing test was not affected by BLF. Intrathecal (i.t.) application of BLF caused marked antinociceptive effects that were reversed by co-administration of a specific mu-opioid receptor antagonist, D-Phe-Cys-Tyr-D-Trp-Orn-Thr-NH(2) (CTOP), or by naloxone during the formalin test. We conclude that LF possesses mu-opioid receptor-mediated antinociceptive activity in the spinal cord.     

Extracellular cholecystokinin levels in the rat spinal cord following chronic morphine exposure: an in vivo microdialysis study

Conflicting results concerning the issue of whether or not chronic morphine exposure induces an increase in CCK biosynthesis have been found in many CNS sites, including the spinal cord, where CCK activity may contribute to the facilitation of the development of opiate tolerance. The present study was undertaken in order to monitor the extracellular level of CCK under spontaneous and stimulus-evoked release in the spinal cord dorsal horn of drug naive and morphine tolerant rats. Tolerance was induced by implantation of two morphine pellets (2x75 mg) which induced a stable morphine plasma concentration after 48 h post-implantation. The tail-flick test and naloxone precipitated withdrawal were used as indexes of tolerance and dependence to morphine. The effect of morphine-pellet implantation on basal and K+-induced release of CCK-like immunoreactivity (CCK-LI) in the rat dorsal horn were monitored with in vivo microdialysis 96 h after implantation of morphine or placebo pellets, when rats showed tolerance and dependence. Basal CCK levels were below the detection limit of the assay (0.6 pM) in both tolerant and normal animals. K+ (100 mM) in the perfusion medium induced a more than 3-fold increase of the extracellular level of CCK-LI in control animals, and a more than 4-fold increase on CCK-LI in morphine-pellet implanted animals. However, this difference was not significant. In addition, naloxone (2 mg/kg; i.v.), did not induce any change in the extracellular level of CCK in either group. The present study suggests that the modulatory interaction between CCK and opioids in the development of tolerance in the spinal cord may occur without necessarily increasing the extracellular level of CCK. Another possible explanation of the finding is that the microdialysis technique is not sensitive enough to detect differences in unstimulated CCK levels in normal and tolerant animals.     

Long-term effects of spinal cord stimulation in chronic pain syndromes

Summary A total of 50 patients with chronic pain syndromes were selected for treatment with spinal cord stimulation. Correct positioning of electrodes was obtained in 44 patients, leading to an initial alleviation of pain in 25 patients. In 6 patients, electrodes (though still effective in 4) had to be removed because of surgical complications within the first 5 months of use. Only 8 patients had at least some beneficial effect lasting for more than 3 years. The long-term results in patients with more severe psychological disturbances were no worse than those of the other patients.     

Relieving effects of electroacupuncture on mechanical allodynia in neuropathic pain model of inferior caudal trunk injury in rat: mediation by spinal opioid receptors

The relieving effects of electroacupuncture (EA) on mechanical allodynia and its mechanism related to the spinal opioid system were investigated in a rat model of neuropathic pain. To produce neuropathic pain in the tail, the right superior caudal trunk was resected between the S1 and S2 spinal nerves. Two weeks after the surgery, EA stimulation (2 or 100 Hz, 0.3 ms, 0.2-0.3 mA) was delivered to Zusanli (ST36) for 30 min. The degree of mechanical allodynia was evaluated quantitatively by touching the tail with von Frey hair (2.0 g) at 10 min intervals. These rats were then subjected to an i.t. injection with one of the three specific opioid agonists in successive ways: the mu agonist (DAMGO 25, 50 and 100 pmol), the delta agonist (DADELT II 0.5, 1 and 2 nmol), and the kappa agonist (U50488H 5, 10 and 20 nmol) separated by 10 min in cumulative doses. During 30 min of EA stimulation, specific opioid antagonists were subjected to i.t. injection: the mu antagonist (beta-FNA 5, 10 and 20 nmol), the delta antagonist (naltrindole 5, 10 and 20 nmol), and the kappa antagonist (nor-BNI 3, 6 and 12 nmol) separated by 10 min in cumulative doses. As a result, EA reduced the behavioral signs of mechanical allodynia. Two Hz EA induced a robust and longer lasting effect than 100 Hz. All three opioid agonists also showed relieving effects on mechanical allodynia. However, nor-BNI could not block the EA effects on mechanical allodynia, whereas beta-FNA or naltrindole significantly blocked EA effects. These results suggest that the mu and delta, but not kappa, opioid receptors in the spinal cord of the rat, play important roles in mediating relieving effects on mechanical allodynia induced by 2 Hz EA.     

Kappa opioid receptor density is consistent along the rostrocaudal axis of the female rat spinal cord

Kappa opioid receptors (KORs) were immunocytochemically localized at four different levels of the spinal cord of normally-cycling female rats in estrus or diestrus. KOR labeling was primarily observed in fine processes and a few neuronal cell bodies in the superficial dorsal horn and the dorsolateral funiculus. Quantitative light microscopic densitometry of the superficial dorsal horn revealed that there were no significant differences in KOR densities among spinal segments C1--C2, T2, T13--L1, and L6--S1 in either the estrus or diestrus phases. These results suggest that the potential for KOR-mediated antinociceptive responses is consistent along the rostrocaudal axis of the female rat spinal cord.     

Effects of opioid receptor antagonists on the effects of i.v. morphine on carrageenin evoked c-Fos expression in the superficial dorsal horn of the rat spinal cord

This study performed in freely moving rats evaluated the ability of specific opioid receptor antagonists to reverse the inhibitory effects of morphine on carrageenin-induced c-Fos expression in the spinal cord. Our study focused on the superficial dorsal horn (laminae I-II), which is the main termination site of nociceptive primary afferent fibers and is rich in opioid receptors. In order to replicate clinical routes of administration, all agents were administered intravenously (i.v.). As previously demonstrated, pre-administered i.v. morphine (3 mg/kg) produced a marked decrease (58+/-5%) in the number of Fos-LI neurones measured at 2 h after intraplantar (i.pl.) carrageenin (6 mg/150 microl) and yet was without influence on peripheral oedema. This decrease in c-Fos expression was completely blocked by combined administration of morphine with the mu-opioid receptor antagonist, [D-Phe-Cys-Tyr-D-Orn-Thr-Pen-Thr-NH2] (CTOP-1+1 mg/kg). Naltrindole (NTI-1+1 mg/kg), a delta-opioid receptor antagonist partially blocked the effects of systemic morphine, so that the inhibitory effects of morphine after NTI injection are now 40+/-4%. However, this effect of NTI was weak since the depressive effects of morphine were still highly significant (p<0.001). In contrast, nor-binaltorphimine (nor-BNI-1+1 mg/kg), a kappa-opioid receptor antagonist, had no significant effect on the effects of morphine. These results indicate the major contribution of mu-opioid receptors to the antinociceptive effects of systemic morphine at the level of the superficial dorsal horn. The observed effect of NTI is not necessarily related to a direct action of morphine on delta-opioid receptors and some possible actions of this antagonist are discussed.     

Opioid use and spinal cord stimulation therapy: The long game

Spinal cord stimulation (SCS) has been considered as an alternative therapy to reduce opioid requirements in certain chronic pain disorders. However, information on long-term opioid consumption patterns and their impact on SCS device explantation is lacking. We conducted a retrospective study of 45 patients to characterize long-term patterns of opioid usage after SCS implantation. Daily morphine equivalent dosage (MED) increased, decreased, and remained the same in 40%, 40%, and 20% of patients at 1-year follow-up, respectively. Twelve (27%) underwent explantation due to treatment failure at a median of 18 months after implantation. Pre-operative opioid status (naïve vs. active use) was not associated with explantation (18% vs. 29%, p = 0.699) and neither was the daily MED change status (i.e. increased, decreased, unchanged) at 1-year (p = 0.499, 1.000, 0.735, respectively). Following explantation, reduction in the daily MED was seen in 92% of patients with dosages falling below pre-operative baseline in nine. Among the opioid naïve patients, 55% were on opioids at last follow-up (average 32.4 ± 14.6 months). Our results indicate that daily opioid consumption does not decrease in most patients 1-year after SCS implantation. Furthermore, post-operative evaluation beyond 1-year is necessary to assess the efficacy and durability of SCS therapy as well as its impact on opioid requirement. Lastly, rigorous patient selection and pre-operative risk assessment for misuse and dependence are paramount to improving outcome after SCS implantation.     

Spinal cord mediation of the opioid analgesia of pregnancy

It has been demonstrated that during pregnancy and labor in rats and humans there is an opioid-mediated elevation in the threshold for responsiveness to aversive stimuli which reaches a maximum at term. Acute administration of the opiate antagonist, naltrexone, into the lumbar intrathecal (i.t.) space of pregnant rats (day 20 of gestation) significantly reduces the threshold for reflexive jumping in response to electric footshock. The i.t. administration of the inactive stereoisomer of a closely related narcotic antagonist, (+)-naloxone, is devoid of any effect on pain threshold. No effect on the pain threshold is observed following intrathecal saline administration to pregnant rats, i.t. naltrexone administration to non-pregnant rats or following systemic administration of an intrathecally effective dose of naltrexone to pregnant rats. These data indicate that the analgesia observed during gestation is mediated, at least in part, via a spinal opioid pathway which is activated by some aspect of the pregnant condition.     

促甲状腺释放激素类似物YM-14673对大鼠脊髓损伤后水肿的影响

目的研究促甲状腺释放激素（ＴＲＨ）类似物,ＹＭ－１４６７３大鼠脊髓损伤后水肿的影响。方法用改良Ａｌｌｅｎ氏法建立大鼠脊髓损伤模型,分设正常组、对照组和治疗组,治疗组在损伤后１５分钟注射ＹＭ－１４６７３,用称重法测量脊髓的水含量,公式：（湿重－干重）÷湿重×１００％。结果对照组示伤后２４小时脊髓水肿,治疗组显示在２４小时脊髓水肿减轻。结论早期应用ＴＲＨ类似物,ＹＭ-１４６７３可减轻脊髓损伤后的脊随水肿。     

Adenosine exerts multiple effects in dorsal horn neurones of the adult rat spinal cord

In the present study, we have examined the effects of adenosine and its analogues on the electrophysiological properties of dorsal horn neurones in the rat adult spinal cord. Adenosine and the A₁ receptor agonist R-phenylisopropyl adenosine (RPIA) reversibly hyperpolarised these neurones via the generation of an outward current at −60 mV that was inhibited by pre-application of barium or Rp-adenosine 3′, 5′-cyclic monophosphothioate triethylamine. In contrast, the A_2a receptor agonist 2-[p-(2-carboxyethyl)phenylethylamino]-5′-N-ethylcarboxamidoadenosine (CGS21680) had no effect on the resting membrane properties of these neurones. Stimulation of the dorsal root evoked non-NMDA receptor-mediated excitatory postsynaptic currents (EPSCs) at −60 mV that were completely abolished by 2,3-dihydroxy-6-nitro-7-sulophamoyl-benzo(F)quinoxalone (NBQX). Bath application of adenosine or RPIA reversibly inhibited these EPSCs in a concentration-dependent manner via a presynaptic action. In contrast, CGS21680 increased the amplitude of the EPSC in 20% of neurones tested and decreased the EPSC amplitude in 30% of neurones tested. It is concluded that adenosine exerts multiple effects upon the electrophysiological properties of dorsal horn neurones in the adult spinal cord via interaction with multiple receptors. These findings have important implications in the understanding of adenosine action in preclinical models of pain.     

A model of chronic pain in the rat: high-resolution neuroanatomical approach identifies alterations in multiple opioid systems in the periaqueductal grey

Inoculation of the tail base of rats with Mycobacterium butyricum led to an arthritic swelling and inflammation of the limbs which displayed a hyperalgesia to noxious pressure: these effects peaked at 3 weeks postinoculation. In vitro autoradiography of coronal sections of rat brain was used for a parallel determination of binding to mu-, delta- and kappa-opioid binding sites. In only two regions, the dorsomedial and dorsolateral parts of the periaqueductal grey (PAG), was a significant change seen: this comprised an increase in binding to kappa-sites, whereas mu- and delta-sites therein were unaffected. This region was analysed for opioid peptides derived from each of the three opioid peptide families known. While no change was seen in levels of immunoreactive (ir)-dynorphin1-17 A (DYN) and ir-Met-enkephalin, a decrease was detected in those of ir-beta-endorphin (beta-EP): this change was restricted to the PAG. These data demonstrate a highly localized and selective influence of chronic arthritic pain upon multiple opioid systems in the PAG of the rat, a structure playing a key role in the control of pain and in the expression of the antinociceptive actions of opioids. The data suggest a possible significance of PAG pools of beta-EP and kappa-receptors in the response to and modulation of chronic pain.     

Intrathecal dynorphin A1–13 and dynorphin A3–13 reduce rat spinal cord blood flow by non-opioid mechanisms

Radiolabeled microspheres were used to examine the effects of paralytic intrathecal doses of dynorphin A (Dyn A_1–13) and Dyn A_3–13 on rat brain and spinal cord blood flows and cardiac output. Dyn A_1–13 produced significant dose-related reductions in blood flow to lumbosacral and thoracic spinal cord without altering cardiac output and blood flow to brain and cervical spinal cord. Naloxone failed to block these effects. Dyn A_3–13, which lacks opioid activity, also significantly reduced blood flow in lumbosacral spinal cord. Thus, the paralytic effects of Dyn A in the rat may involve reductions in spinal cord resulting from non-opioid actions of Dyn A.     

构建脊髓慢性压迫损伤模型大鼠巢蛋白的表达规律*

背景：既往应用的脊髓损伤动物模型难以达到一种慢性渐进性的压迫效果,与人体慢性脊髓压迫损伤机制有很大的不同。 目的：构建一种新的脊髓慢性压迫性损伤模型大鼠,探究慢性压迫损伤后脊髓损伤区域巢蛋白的表达规律及其意义。 方法：Wistar大鼠40只随机分为实验组30只和对照组10只。实验组大鼠取下胸7、8椎板,植入压迫材料,形成慢性压迫脊髓损伤模型。植入后第1,3,7,14,28天,取压迫处脊髓组织,行病理学检查及巢蛋白免疫组织化学染色,半定量反转录PCR反应测定巢蛋白mRNA的表达,同时测量压迫段椎管直径及缓膨胀材料侵占厚度。 结果与结论：随压迫时间的延长,实验组大鼠椎管侵占率逐渐增加,脊髓组织出现坏死等情况,大鼠BBB评分降低,压迫处脊髓组织中Nestin mRNA及蛋白表达至伤后7 d时达到高峰,而后表达逐渐下降,说明实验成功建立慢性脊髓压迫损伤动物模型,且慢性脊髓压迫损伤大鼠脊髓组织Nestin mRNA及蛋白呈动态变化。     

Glucose utilization and reflex activity of the transected rat spinal cord

Spinal cord glucose utilization (SCGU) of gray and white matter was studied with the quantitative autoradiography [¹⁴C]2-deoxyglucose methodology, below and above a complete low thoracic transection. One day after transection, a generalized decrease in SCGU was observed in gray matter, particularly marked in the dorsal horn of the lumbar cord. A progressive increase in SCGU was observed thereafter. Values reached levels greater than those of non-transected controls by 2 and 4 weeks after the intervention in ventral horn of the lumbar cord, and ventral and dorsal horn of the cervical cord. A similar behavior of SCGU was observed in white matter of transected animals. The development of 17 lumbar reflex modalities was quantified between days 1 and 28 after spinal transection. The delay in emergence of these reflexes was related to their complexity. Correlation of reflex scores with SCGU was significant for all lumbar cord regions but linearity of this relationship was only observed in white matter. These results uncover a close relationship between SCGU and reflex activity of the spinal cord below a complete transection, particularly striking in white matter and suggests a role of the fasciculi proprii of the spinal cord in this phenomenon.     

Polyarthritis-associated changes in the opioid control of spinal CGRP release in the rat

As a model of chronic inflammatory pain, Freund's adjuvant-induced polyarthritis has been shown to be associated with marked alterations in the activity of opioid- and calcitonin gene-related peptide (CGRP)-containing neurons in the dorsal horn of the spinal cord in rats. Possible changes in the interactions between these two peptidergic systems in chronic inflammatory pain were investigated by comparing the effects of various opioid receptor ligands on the spinal outflow of CGRP-like material (CGRPLM) in polyarthritic and age-paired control rats. Intrathecal perfusion of an artificial cerebrospinal fluid in halothane-anaesthetized animals allowed the collection of CGRPLM released from the spinal cord and the application of opioid receptor ligands. The blockade of κ-opioid receptors similarly increased CGRPLM release in both groups of rats as expected of a κ-mediated tonic inhibitory control of CGRP-containing fibres in control, as well as in polyarthritic rats. In contrast, the higher increase in CGRPLM outflow due to the preferential blockade of μ opioid receptors by naloxone in polyarthritic rats as compared to non-suffering animals supports the idea of a reinforced μ opioid receptor-mediated tonic inhibitory control of CGRP-containing fibres in rats suffering from chronic pain. Even more strikingly, the differences observed in the effects of ∂-opioid receptor ligands on CGRPLM outflow suggest that ∂ receptors are functionally shifted from a participation in a phasic excitatory control in non-suffering rats to a tonic inhibitory control in polyarthritic rats. These data indicate that agonists acting at the three types of opioid receptors all exert a tonic inhibitory influence on CGRP-containing nociceptive primary afferent fibres within the spinal cord of polyarthritic rats. Such a convergence probably explains why morphine and other opioids are especially potent to reduce pain in subjects suffering from chronic inflammatory diseases.     

胚胎脊髓移植在恢复损伤脊髓传导功能中的作用

目的：探讨胚胎脊髓移植在恢复损伤脊髓传导功能中的作用。方法：将Ｅ１４胚胎脊髓植入成鼠损伤脊髓后３０、４５、６０天时，用单位放电记录技术观察了正常脊髓神经元和移植物神经元的自发放电活动，及其对刺激坐骨神经、红核和同时刺激的反应。结果：正常脊髓神经元的自发单位放电多是一个低频的单发脉冲活动。无论选择那种刺激方式，都可见兴奋、抑制和无反应三种反应。术后３０天时，胚胎神经元的自发单位放电以高频电脉冲活动为主，簇状放电所占比例较大，对刺激有反应的放电单位数也较少；随着动物存活时间的延长，这些单位放电的情况逐渐向着低频、单脉冲以及高反应率的方向发展。至术后６０天时，胚胎神经元单位放电的频率、形式以及对刺激的反应情况都变得和正常神经元的相似。结论：胚胎神经元移植后经历了一个逐渐发育分化过程，在这个过程中它们有可能逐渐和宿主神经元形成了功能性突触连接。     

脊髓型多发性硬化的MRI表现

目的总结分析脊髓型多发性硬化的MRI表现。方法搜集经临床证实的脊髓型多发性硬化11例,均行MRI检查,对其临床及MRI资料进行回顾性分析。结果脊髓型多发性硬化的特征性MRI表现为,11例患者的病灶以颈髓多见,病变脊髓在T1WI像为低或等信号,T2WI像为高信号,病灶位于脊髓两侧和后部,病灶活动期呈斑片状或边缘强化,应用糖皮质激素试验性治疗对脊髓出现的可疑脱髓鞘病灶者有一定的帮助。结论脊髓型多发性硬化有其特征性MRI表现,MRI有助于脊髓型多发性硬化的诊断,是目前诊断脊髓型多发性硬化最敏感的影像学方法 。     

Inhibition of spinal nociceptive responses after intramuscular injection of capsaicin involves activation of noradrenergic and opioid systems

Extracellular recordings of wide dynamic range neurones in the dorsal horn driven by electrical stimulation of the sciatic nerve were performed in intact urethane-anaesthetized Sprague–Dawley rats. The electrically evoked neuronal responses were defined as A- and C-fibres responses according to latencies, and the effect of a deep nociceptive conditioning stimulus induced by 200 μg capsaicin (8-methyl-N-vanillyl-6-noneamide) injected into the contralateral gastrocnemius–soleus muscle was studied for at least 30 min. Independent of the size and location of the receptive field of the neurone under study, a clear inhibition of the neuronal responses was observed. The electrically evoked C-fibre responses were inhibited to 53% of baseline 15–30 min after injection of capsaicin. This inhibition was only slightly attenuated by 125 nmol of the α-adrenoceptor antagonist phentolamine or 250 nmol of the opioid receptor antagonist naloxone applied directly onto the spinal cord when the two compounds were administered separately 5 min before capsaicin. In contrast, when a mixture of the two compounds was given 5 min before capsaicin, the effect of capsaicin was completely abolished. These results indicate that activation of the capsaicin-sensitive afferents in the gastrocnemius–soleus muscle inhibits the electrically evoked C-fibre responses in the dorsal horn by activating noradrenergic and opioidergic inhibitory systems. Moreover, our data indicate that the activation of these two systems following injection of capsaicin has a sub-additive inhibitory effect on the wide dynamic range neurones in the spinal cord. We conclude that only one of these systems is sufficient for the inhibition to occur.