家兔下丘脑弓状核区参与从伏核到中脑导水管周围灰质的下行镇痛通路

我们以往的工作表明,中脑导水管周围灰质(PAG)和伏核之间存在着双向性的联系,构成内源性镇痛系统的重要组成部分。本工作探讨下丘脑弓状核区(ARH)和β-内啡肽(β-EP)能纤维在这一神经联系中所起的作用。实验结果表明,在家兔伏核内微量注射吗啡产生的镇痛作用可被PAG内注射β-EP抗血清所对抗,当损毁ARH后上述对抗效应消失。但是,损毁ARH的家兔其基础痛阈仍保持在正常水平,伏核内注射吗啡仍能引起明显的镇痛效应,而且这种效应仍能被PAG内注射纳洛酮所削弱。以上结果提示,(1)从伏核到PAG的下行镇痛通路中有ARH及β-EP能纤维的参与;(2)除β-EP外PAG内尚有其它阿片肽发挥镇痛作用。     

家兔伏核注射吗啡引起的镇痛可被中脑导水管周围灰质注射纳洛酮或甲啡肽抗体所阻断

本实验室以往的资料表明,在家兔中脑导水管周围灰质(PAG)到伏核之间存在一条与镇痛有夫的神经通路,该通路以5-羟色胺(5-HT)和甲啡肽(ME)为其递质。本工作进一步探讨从伏核到PAG的下行镇痛通路。 以辐射热照射家兔嘴侧部皮肤,测量其躲避反应的潜伏期(ERL)作为痛反应阈,简称痛阈。通过预先埋植的慢性套管向伏核内微量注射吗啡,20min后向PAG内双侧注射纳洛酮(NX)或脑啡肽抗血清,观察ERL的变化。(1)伏核内注射吗啡20μg/1μl,引起ERL升高80％以上,作用持续50min以上。(2)PAG内注射NX(每侧0.5、1.0或2.0μg)可不同程度地阻断伏核内注射吗啡的镇痛效应,且呈明显的剂效关系。(3)PAG内注入甲啡肽抗血清(每侧1μl)可部分阻断伏核内注射吗啡的镇痛效应,而注入亮啡肽抗血清或正常兔血清则无效。 实验结果提示,从伏核到PAG存在一条下行镇痛通路,在PAG内可能以ME为其递质。该通路与PAG到伏核的上行镇痛通路构成一个环形的“中脑边缘镇痛回路”,并在针刺镇痛和吗啡镇痛中发挥重要作用。     

下丘脑弓状核区参与由中脑导水管周围灰质到伏核的上行镇痛通路

在家兔中脑导水管周围灰质(PAG)内微量注射吗啡产生的镇痛作用可被伏核内注射β-内啡肽抗体所对抗,在损毁下丘脑弓状核区(ARH)后该对抗效应消失。但损毁ARH并不影响PAG注射吗啡所引起的镇痛作用,也不影响伏核注射纳洛酮对于PAG内注射吗啡引起镇痛的对抗作用,以上结果提示,(1)从PAG到伏核的上行镇痛通路中有ARH及β-内啡肽能纤维参与;(2)除β-内啡肽以外,伏核内尚有其它阿片肽发挥镇痛作用。     

抑制伏核内脑啡肽的降解使电针镇痛和吗啡镇痛得到加强

将“脑啡肽酶”抑制剂 Thiorphan 或氨肽酶抑制剂 Bestatin 经慢性埋植套管注入家兔一侧状核内,观察到明显的镇痛作用,在1—4μg 范围内呈现明确的剂量-效应关系。该作用可为伏核内注射纳洛酮或甲啡肽抗体所完全翻转,亮啡肽抗体则无效。表明伏核内注射 Thior-Phan 或 Bestatn 所产生的镇痛效应主要是通过甲啡肽而完成的。伏核内注射微量 Thiorphan 或 Bestatin 使电针镇痛的后效应明显加强,并能增强吗啡的镇痛作用。表明电针和吗啡的镇痛效果至少有一部分是通过在伏核内释放出脑啡肽(特别是甲啡肽)而实现的。     

家兔杏仁核内的5-羟色胺和内源性吗啡样物质在电针镇痛和吗啡镇痛中起重要作用

本工作的目的是要确定杏仁核内的吗啡样物质(内啡素)和5-羟色胺(5-HT)是否参与电针镇痛和吗啡镇痛。经慢性埋植套管向家兔杏仁核内微量注射阿片受体阻断剂纳洛酮,或5-HT受体阻断剂肉桂硫胺,可使电针的镇痛效果显著减弱,尤以注入中央杏仁核作用最为显著,双侧注射效果大于单侧,注入核外则无效。杏仁核内注入5-HT 的前体5-HTP,或脑啡肽降解酶抑制剂 D-苯丙氨酸可使电针镇痛显著加强。上述措施凡是加强或对抗电针镇痛的,也能加强或对抗吗啡镇痛。以上结果表明,电针刺激或注射吗啡可能在杏仁核内引起5-HT 和内啡素(很可能是脑啡肽)的释放,而发挥镇痛效应。     

家兔中脑导水管周围灰质中甲七肽样免疫活性物质具有镇痛作用并参与电针镇痛

以辐射热照射家兔鼻嘴部皮肤,测定甩头反应潜伏期(ERL)作为痛阈。通过预先埋植的慢性套管向中脑导水管周围灰质(PAG)注射甲七肽降解酶抑制剂 Captopril,观察其镇痛作用以及加强电针镇痛的作用能否被特异的甲七肽抗血清所对抗。(1)单侧 PAG 注射 Captopril240 nmol 的镇痛作用可为同一部位注射甲七肽抗血清(1μl)所翻转,注入甲啡肽抗血清则无效。(2)单侧 PAG 注射 Captopril 60 nmol 有加强电针镇痛的作用。该作用可被1μl 甲七肽抗血清所完全取消,将抗血清量减少到0.1μl 则无效。以上结果说明 PAG 内的甲七肽样免疫活性物质在镇痛和电针镇痛中发挥重要作用。     

家兔隔核中去甲肾上腺素对皮肤与内脏痛阈的影响

本工作以电刺激内脏大神经或耳尖部皮肤测定清醒家兔内脏痛阈或皮肤痛阈,以探讨隔核去甲肾上腺素在内脏镇痛和皮肤镇痛中的作用以及与中脑导水管周围灰质(PAG)中内阿片肽系统的关系。实验观察到,双侧隔核内微量注射α受体激动剂可乐宁(10μg/2μl)或α受体阻断剂酚妥拉明(10μg/2μl)对内脏痛阈无明显影响。注入β受体激动剂异丙肾上腺素(1μg/2μl)使内脏痛阐明显升高;而注入β受体阻断剂心得安(1Cμg/2μl)则内脏痛阈明显降低。隔核内注入酚妥拉明(10μg/2μl)或心得安(10μg/2μl)均可使皮肤痛阈明显提高。提示,隔核内NA通过β受体调制内脏痛;通过α受体和β受体调制皮肤痛。隔核内注入异丙肾上腺素(1μg/2μl)明显地镇内脏痛,此作用可被PAG内注射纳洛酮(1μg/2μl)或注射抗亮啡肽抗血清(1:20,000)所减弱;但可使PAG内亮啡肽样物质释放量增加。这提示,隔核内NA的镇内脏痛作用与PAG的内阿片肽系统有关;其中亮非肽在这一过程中具有重要作用。     

家兔杏仁核内钙、镁离子对电针镇痛和吗啡镇痛的拮抗作用

通过埋植套管向家兔双侧杏仁核内缓慢注射微量氯化钙或氯化镁,对电针镇痛和吗啡镇痛有显著的对抗作用。向杏仁核内注入钙离子螯合剂CDTA或钙通道抑制剂异博定则可显著加强电针镇痛和吗啡镇痛。提示:杏仁核内钙、镁离于浓度是影响吗啡镇痛和电针镇痛的重要因素,同时表明电针和吗啡镇痛的机理可能有相似之处。     

大鼠脊髓蛛网膜下腔注射5-羟色胺和吗啡的镇痛作用

本工作利用大鼠脊髓蛛网膜下腔注射的方法,观察了在脊髓水平5-HT 和吗啡的镇痛作用及其相互关系。结果如下:(1)脊髓蛛网膜下腔注射 200μg 5-HT 或10μg吗啡可产生明显的镇痛作用,并分别被 5-HT受体阻断剂肉桂硫胺和阿片受体阻断剂纳洛酮所对抗。(2)单纯使用肉桂硫胺可产生痛敏。纳洛酮对痛阈无明显影响。(3)5-HT 镇痛作用不能被大剂量纳洛酮(10mg/kg,sc)阻断。吗啡镇痛作用则可被脊髓蛛网膜下腔注射肉桂硫胺所削弱。后一效应可能与肉桂硫胺本身可引起痛敏有关。上述结果表明,在脊髓水平5-HT 和吗啡可通过各自的受体产生镇痛作用,两者间无明显依赖关系。5-HT 可能有紧张性活动,内源性阿片肽似不存在紧张性作用。     

家兔中央灰质内微量注射纳洛酮和p-氯苯丙胺对针刺镇痛的影响

本文用66只家兔进行实验。按 Sawyer 氏图谱,借定向仪将不锈钢套管插入中央灰质(P_(9.5)、L_1、H_(-0.5))和杏仁核(A_2、L_(4.5)、H_(-5))。手术4天以后进行实验。用钾离子透入法测定痛阈。观察结果如下:(1)家兔电针或静脉注射吗啡(5毫克/公斤)20分钟后痛阈明显提高。此时,于中央灰质内注射生理盐水(2微升/3分钟)并不减弱针刺镇痛和吗啡镇痛作用,而注射纳洛酮(2微克/2微升/3分钟)后,则可显著地对抗这两种镇痛作用,这种对抗作用在给药后7分钟最强,与对照组相比,差别有非常显著意义(p<0.01)。然而,等量纳洛酮注射到中脑网状结构或杏仁核却不能对抗针刺镇痛和吗啡镇痛。(2)中央灰质内注射p-氯苯丙胺(PCA,4微克/2微升/3分钟)后24、72、120小时,电针镇痛作用明显减弱,与生理盐水组(2微升/3分钟)相比,差别极为显著(p<0.01)。对 PCA 部分减弱针效的兔子,于中央灰质内注入等量纳洛酮仍可进一步对抗针刺镇痛作用。以上结果提示,家兔中央灰质的内啡肽和5-羟色胺共同参与针刺镇痛。     

Modification of the effects of naloxone in morphine-dependent mice

Mice were rendered dependent on morphine by mixing morphine with their food (2 mg/g) for three days. Increasing doses of naloxone precipitated dose-dependent withdrawal reactions such as weight loss and jumping. These withdrawal reactions were antagonized by morphine pretreatment. Effects of morphine, such as increased locomotor activity, inhibition of intestinal transport, and analgesia were antagonized by naloxone in both non-dependent and dependent subjects. The antagonist actions of naloxone were increased in dependent subjects; lower doses of naloxone were sufficient to antagonize effects of morphine. The present results confirm earlier studies indicating that precipitation of withdrawal can be antagonized by morphine pretreatment suggesting that withdrawal reactions are due to actions of naloxone at the same receptor at which opioid agonists act. The increased antagonist potency of naloxone in dependent subjects extends earlier results obtained with analgesic effects to several other agonist effects of morphine and is consistent with the interpretation that exposure to an opioid agonist induces a change in the conformation of opioid receptors.     

The effect of opiates and opiate antagonists on heat latency response in the parasitic nematode Ascaris suum

The effects of the opiates morphine and morphine-6-glucuronide (M6G), the mu opioid receptor specific antagonist D-Phe-Cys-Tyr-D-Trp-Om-Thr-Pen-Thr-NH(2) (CTOP), and the general opiate antagonist naloxone on the latency of response to thermal stimulation were determined in the parasitic nematode Ascaris suum. Thermal detection and avoidance behaviors of the worms were evaluated with a tail flick analgesia meter using a modification of a technique employed for nociception experiments in rodents. Morphine and M6G were shown to have a dose dependent analgesic effect on A. suum's latency of response to heat with morphine being the most potent. The analgesic effect of morphine was reversed by naloxone but not CTOP. Neither naloxone nor CTOP was able to block the analgesia of M6G. CTOP but not naloxone had significant analgesic effects on its own. These findings are generally consistent with previous results on the effects of opiates and nitric oxide release from A. suum tissue. Apparently these nematodes possess opioid receptors that effect nociception.     

Effects of 5-HT1B receptor ligands microinjected into the accumbal shell or core on the cocaine-induced locomotor hyperactivity in rats.

The present study was designed to examine the effect of 5-HT1B receptor ligands microinjected into the subregions of the nucleus accumbens (the shell and the core) on the locomotor hyperactivity induced by cocaine in rats. Male Wistar rats were implanted bilaterally with cannulae into the accumbens shell or core, and then were locally injected with GR 55562 (an antagonist of 5-HT1B receptors) or CP 93129 (an agonist of 5-HT1B receptors). Given alone to any accumbal subregion, GR 55562 (0.1-10 microg/side) or CP 93129 (0.1-10 microg/side) did not change basal locomotor activity. Systemic cocaine (10 mg/kg) significantly increased the locomotor activity of rats. GR 55562 (0.1-10 microg/side), administered intra-accumbens shell prior to cocaine, dose-dependently attenuated the psychostimulant-induced locomotor hyperactivity. Such attenuation was not found in animals which had been injected with GR 55562 into the accumbens core. When injected into the accumbens shell (but not the core) before cocaine, CP 93129 (0.1-10 microg/side) enhanced the locomotor response to cocaine; the maximum effect being observed after 10 microg/side of the agonist. The later enhancement was attenuated after intra-accumbens shell treatment with GR 55562 (1 microg/side). Our findings indicate that cocaine induced hyperlocomotion is modified by 5-HT1B receptor ligands microinjected into the accumbens shell, but not core, this modification consisting in inhibitory and facilitatory effects of the 5-HT1B receptor antagonist (GR 55562) and agonist (CP 93129), respectively. In other words, the present results suggest that the accumbal shell 5-HT1B receptors play a permissive role in the behavioural response to the psychostimulant.     

Sex differences in morphine-induced analgesia of visceral pain are supraspinally and peripherally mediated

Increasing evidence suggests there is a sex difference in opioid analgesia of pain arising from somatic tissue. However, the existence of a sex difference in visceral pain and opioid analgesia is unclear. This was examined in the colorectal distention (CRD) model of visceral pain in the current study. The visceromotor response (vmr) to noxious CRD was recorded in gonadally intact male and female rats. Subcutaneous injection of morphine dose-dependently decreased the vmr in both groups without affecting colonic compliance. However, morphine was significantly more potent in male rats than females. Because systemic morphine can act at peripheral tissue and in the central nervous system (CNS), the source of the sex difference in morphine analgesia was determined. The peripherally restricted mu-opioid receptor (MOR) antagonist naloxone methiodide dose-dependently attenuated the effects of systemic morphine. Systemic administration of the peripherally restricted MOR agonist loperamide confirmed peripherally mediated morphine analgesia and revealed greater potency in males compared with females. Spinal administration of morphine dose-dependently attenuated the vmr, but there was no sex difference. Intracerebroventricular administration of morphine also dose-dependently attenuated the vmr with significantly greater potency in male rats. The present study documents a sex difference in morphine analgesia of visceral pain that is both peripherally and supraspinally mediated.     

Central integration of cardiovascular and drinking responses elicited by central administration of angiotensin II: divergence of regulation by the ventral tegmental area and nucleus accumbens

Previous studies had implicated the involvement of the ventral tegmental area and its dopamine projections to the nucleus accumbens in goal-directed behavior. This study investigated whether or not the GABAergic inputs to the ventral tegmental area and, in turn, dopaminergic input to the nucleus accumbens from the ventral tegmental area modify drinking and cardiovascular responses elicited by central administration of angiotensin II. Injections of 25 ng of angiotensin II into a lateral cerebral ventricle of the rat elicited water intakes averaging 7-8 mL in 15 min with latencies usually less than 3 min. Pretreatment of the nucleus accumbens with spiperone, a dopamine antagonist, or the ventral tegmental area with gamma-amino butyric acid (GABA) produced dose-dependent reductions in water intake and number of laps taken while increasing the latency to drink. The spiperone injection did not alter the pressor response. On the other hand, the GABA injections attenuated the pressor responses to central angiotensin II administration. These findings suggest that GABA input to the ventral tegmental area modifies both the cardiovascular and drinking responses elicited following central administration of angiotensin II. However, the dopamine projections to the nucleus accumbens appear to be involved only in the drinking responses elicited by central injections of angiotensin II. Divergence for the coordination of the skeletal motor behavioral component and the cardiovascular component elicited by central administration of angiotensin II must occur before the involvement of these dopamine pathways.     

Interactions of galanin and opioids in nociceptive modulation in the arcuate nucleus of hypothalamus in rats

The fact that galanin, beta-endorphin and their receptors are present in the arcuate nucleus of hypothalamus (ARC), coupled with our previous observation that both beta-endorphin and galanin play antinociceptive roles in pain modulation in the ARC, made it of interest to study their interactions. The hindpaw withdrawal latency (HWL) in response to noxious thermal and mechanical stimulation was assessed by the hot-plate test and the Randall Selitto Test. We showed that the antinociceptive effect induced by intra-ARC injection of galanin was dose-dependently attenuated by the following intra-ARC injection of naloxone. Furthermore, intra-ARC administration of the selective mu-opioid receptor antagonist beta-funaltrexamine (beta-FNA) attenuated the increased HWL induced by intra-ARC injection of galanin in a dose-dependent manner, while the delta-opioid receptor antagonist naltrindole or the kappa-opioid receptor antagonist nor-binaltorphimine (nor-BNI) did not. Moreover, intra-ARC injection of a galanin receptor antagonist galantide attenuated intraperitoneal morphine-induced increases in HWLs. These results demonstrate that the antinociceptive effect of galanin was related to the opioid system, especially mu-opioid receptor was involved in, and that systemic morphine induced antinociception involves galanin in the ARC.     

Evidence for a nucleus accumbens CCK2 receptor regulation of rat ventral pallidal GABA levels: a dual probe microdialysis study

We employed dual probe microdialysis in the nucleus accumbens and ipsilateral ventral pallidum of the halothane anaesthetized rat to investigate the effect of intra-accumbens perfusion with the sulphated octapeptide cholecystokinin (CCK-8S, 10-1000 nM, 60 min) alone and in the presence of the selective CCK1 and CCK2 receptor antagonists L-364,718 (10 and 100 nM) and PD134308 (10 nM), tetrodotoxin (TTX, 1000 nM) and the GABA(A) receptor antagonist bicuculline (1000 nM), on dialysate GABA levels in the ventral pallidum. Intra-accumbens perfusion with the 100 and 1000 nM concentration of CCK-8S was associated with a significant decrease (-16+/-3% and -23+/-3% vs basal, respectively) in ventral pallidum GABA levels. The CCK-8S (1000 nM) induced decrease in ventral pallidal dialysate GABA levels was abolished when PD134308, TTX and bicuculline, but not L-364,718, were included into the perfusion medium of the accumbens probe. The data indicate that nucleus accumbens CCK-8S exerts a CCK2 receptor mediated inhibition of ventral pallidal GABA levels. Furthermore, the TTX and bicuculline sensitivity of this effect suggests that this is possibly mediated via CCK2 receptors probably located on local GABA interneurons.     

Blocking TRPV1 in Nucleus Accumbens Inhibits Persistent Morphine Conditioned Place Preference Expression in Rats

The function of TRPV1 (transient receptor potential vanilloid subfamily, member 1) in the central nervous system is gradually elucidated. It has been recently proved to be expressed in nucleus accumbens (NAc), a region playing an essential role in mediating opioid craving and taking behaviors. Based on the general role of TRPV1 antagonist in blocking neural over-excitability by both pre- and post-synaptic mechanisms, TRPV1 antagonist capsazepine (CPZ) was tested for its ability to prohibit persistent opioid craving in rats. In the present study, we assessed the expression of TRPV1 in nucleus accumbens and investigated the effect of CPZ in bilateral nucleus accumbens on persistent morphine conditioned place preference (mCPP) in rats. We also evaluated the side-effect of CPZ on activity by comparing cross-beam times between groups. We found that morphine conditioned place preference increased the TRPV1 expression and CPZ attenuated morphine conditioned place preference in a dose-dependent and target–specific manner after both short- and long-term spontaneous withdrawal, reflected by the reduction of the increased time in morphine-paired side. CPZ (10 nM) could induce prolonged and stable inhibition of morphine conditioned place preference expression. More importantly, CPZ did not cause dysfunction of activity in the subjects tested, which indicates the inhibitory effect was not obtained at the sacrifice of regular movement. Collectively, these results indicated that injection of TRPV1 antagonist in nucleus accumbens is capable of attenuating persistent morphine conditioned place preference without affecting normal activity. Thus, TRPV1 antagonist is one of the promising therapeutic drugs for the treatment of opioid addiction.     

Dynorphin: potent analgesic effect in spinal cord of the rat

Evidence is presented to show a strong and long-lasting analgesic effect after injection of dynorphin into the subarachnoid space of the spinal cord of the rat. Calculating on a molar basis dynorphin was 6-10 times more potent than morphine and 65-100 times more potent than morphiceptin, the specific mu receptor agonist. Dynorphin analgesia was completely reversed by intrathecal injection of anti-dynorphin IgG and partially reversed by naloxone. Acute tolerance to morphine analgesia did not affect the occurrence of dynorphin analgesia. Evidence from different lines of approach suggest that dynorphin may bind with kappa receptors in the spinal cord to exert its analgesic effect.