损毁弓状核区对大鼠脑内β-内啡肽、5-羟色胺、去甲肾上腺素含量及其对针刺镇痛的影响

下丘脑弓状核区是脑内合成β-内啡肽等物质的主要部位,并与脑内中缝背核、蓝斑等结构有密切的交互纤维支配。本实验用新生期大鼠注射谷氨酸—钠(MSG)损毁弓状核区的方法,观察对脑内β-内啡肽、5-羟色胺、去甲肾上腺素含量及针刺镇痛的影响。MSG 处理组大鼠下丘脑弓状核神经元减少72%左右,脑β-内啡肽含量降低67%,针刺镇痛效应明显下降,电针后脑去甲肾上腺素含量明显高于电针对照组;将 MSG 处理大鼠的垂体摘除后,针刺镇痛效应几乎消失,同时电针后脑去甲肾上腺素含量则显著高于单纯 MSG处理组。本文对可能的机理进行了讨论。     

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。     

针刺镇痛过程中中缝大核内5-羟色胺和去甲肾上腺素荧光强度的变化

本实验用荧光组织化学和显微荧光定量法测定了针刺镇痛过程中大自鼠中缝大核内5-羟色胺(5-HT)胞体和去甲肾上腺素(NA)末梢荧光强度的变化。电针组在电针后,痛阈均有不同程度的提高。33只接受电针刺激的动物中,有22只为强针效,6只弱针效,7只无针效。实验表明,针效强的动物,中缝大核内5-HT胞体荧光增强,NA末梢荧光强度减弱;针效弱的动物,5-HT胞体的荧光强度也有所增强,NA末梢的荧光强度略有减弱,但无统计意义。无针效动物,两者均无明显变化。本结果表明,针刺镇痛过程中,中缝大核内5-HT和NA含量都发生变化,其变化程度与针刺镇痛效应的强弱密切相关。本文对中缝大核5-HT和NA在针刺镇痛过程中的相互关系进行了讨论。     

注射6-羟多巴胺于中缝背核后对针刺镇痛和单胺能神经元荧光组织化学反应的影响

新近的研究资料表明,在针刺镇痛过程中,中缝背核的5-羟色胺(5-HT)荧光增强,蓝斑核的去甲肾上腺素(NA)荧光减弱;分别损毁中缝背核和蓝斑核或从蓝斑核发出的上行背束时,前者减弱针刺镇痛效应,后者加强这个效应;若刺激中缝背核和蓝斑核,它们对针刺镇痛效应分别表现为易化和压抑的作用。这些结果从正反两方面肯定了中缝背核和蓝斑核在针刺镇痛中的作用,并揭示它们的作用是拮抗性的。但迄今为止,没有资料证明这两个核团及其所含的5-HT 和 NA 在针刺镇痛中的相互关系。因     

视前区儿茶酚胺神经末梢损毁对针刺镇痛的影响

本实验在家兔双侧视前区内注入6-羟基多巴胺(6-OH-DA,4μg/每侧),选择性损毁该区儿茶酚胺(CA)神经末梢后,观察对针刺镇痛的影响。实验结果表明,注药后第二、四天,针刺镇痛效应较针前或对照组均有明显提高。注药后第十天,针刺镇痛效应基本恢复到对照组水平。用甲醛诱发荧光组织化学法显示视前区 CA 末梢,可见6-羟基多巴胺组的 CA 末梢明显减少。结果提示,视前区 CA 含量减少有利于针刺镇痛作用。     

家兔若干与下行抑制有关的中枢部位中去甲肾上腺素、5-羟色胺和脑啡肽在针刺镇痛中的变化

应用推挽灌流技术、去甲肾上腺素(NA)放射酶学法和亮-脑啡肽放射免疫法观察不同脑区 NA 和脊髓背角亮-脑啡肽的释放。应用分子筛柱层析分离家兔不同脑区的5-羟色胺(5-HT)和5-羟吲哚乙酸(5-HIAA),并对它们进行荧光微量测定。以此来阐明针刺镇痛时 NA、5-HT 和亮-脑啡肽在下行抑制中的作用。1.家兔电针20 min,痛阈显著提高,此时中脑导水管周围灰质(PAG)和中缝大核(NRM)的 NA 释放显著减少,而 Al 核团和脊髓背角的 NA释放显著增加。2.电针镇痛时,PAG、延脑中缝核区和脊髓的5-HT 和5-HIAA 含量均有显著增加,除 PAG 外,这种增加的出现较 NA 为晚。提示可能在针刺镇痛的下行抑制中,NA 的参予较5-HT 为早。3.针刺镇痛时脊髓背角亮-脑啡肽的释放也明显增加。     

5-羟色胺对电针抑制青霉素痫样放电的作用

本工作研究电针抑制青霉素所致大脑皮层痫样放电中5-羟色胺的作用。大鼠腹腔注射对氯苯丙氨酸,注射后第4天麻醉开颅,皮层施加青霉素引起痫样放电,并记录逆向刺激锥体央所引起的皮层回返抑制电位(SN)。在注药后动物,电针制痫作用明显减弱,痫波幅度及频率减小缓慢,SN 的恢复也较对照缓慢。电解损毁中缝背核后电针制痫作用亦减弱,痫波发作时程显著延长。电刺激中缝背核可促进痫波振幅及频率的衰减,发作时程缩短。电刺激中脑导水中管周围 灰质有遏制痫波的作用,但在注射对氯苯丙氨酸后此作用亦趋减弱。以上结果表明,在电针制痫中5-HT及5-HT能神经元集中的中缝背核起重要作用。     

用6-羟基多巴胺损毁去甲肾上腺素能下行通路对针刺镇痛的影响和脊髓荧光组织化学的变化

本文用行为和单胺荧光组织化学相结合的方法研究下行去甲肾上腺素(NA)神经通路在针刺镇痛中的作用。用6-羟基多巴胺(6-OH-DA)选择性地损毁大白鼠 NA 下行纤维7天后,脊髓背、腹角中 NA 末梢荧光大部分消退,针刺镇痛效应亦明显下降。而不注药的对照组动物,脊髓灰质内 NA 本梢荧光不受影响,针效无明显变化。实验结果奉明:下行到脊髓的 NA 纤维在针刺镇痛过程中起重要作用。     

脑内5-羟色胺减少对致痫大鼠癫痫发作及学习记忆的影响

目的:于中脑正中中缝核局部微量注射5,7-二羟色胺(5,7-DHT),探讨5-羟色胺(5-HT)与癫痫的关系及匹罗卡品(PILO)致痫大鼠学习记忆改变的可能机制。方法:成年SD大鼠随机分为PILO组、PILO+5,7-DHT组、空白对照组三组,然后根据是否出现癫痫持续状态(SE)再将PILO组分成:PILO+SE组和PILO-SE组两亚组;利用视频脑电图观察大鼠癫痫发作及皮层脑电变化;运用Morris水迷宫测评大鼠空间学习记忆水平;最后运用免疫组化法观察大鼠中缝核5-HT能神经元。结果:大鼠予以5,7-DHT(PILO+5,7-DHT组)处理后造模成功率、死亡率及慢性期自发性发作频率均增高;与空白组比较PILO+SE组中缝核5-HT能神经元数目有所下降(P<0.05),而PILO+5,7-DHT组下降更明显(P<0.01);与空白组比较PILO+SE组平均逃避潜伏期延长、穿越平台次数减少、原平台象限停留时间缩短(P<0.05),而与PILO+SE组比较PILO+5,7-DHT组变化不明显。结论:脑内5-HT水平的降低容易诱发癫痫发作,尚不能认为癫痫大鼠合并认知功能障碍与脑内5-HT水平下降有关。     

大鼠下丘脑弓状核抑制蓝斑神经元放电的机制及其在电针效应中的作用

本工作在清醒箭毒化大鼠人工呼吸条件下进行。实验显示下丘脑弓状核(AR)内微量注射谷氨酸钠(Glut)所兴奋的 AR 神经元可以显著抑制蓝斑(LC)神经元放电。这一效应可被 LC内注射纳洛酮(Nx)或抗β-内啡肽(β-End)血清,以及切断支配 LC 的β-End 能束取消。这些结果提示它主要是通过兴奋β-End 能神经元,使其在 LC 内的轴突末梢释放β-End 而实现的。由于 LC 内单纯注射 Nx 时 LC 神经元放电率无可见变化,似可认为 AR 对 LC 无明显紧张性抑制作用。实验还显示 LC 内注抗β-End 血清后,电针兴奋 LC 的后期效应增加;如在电针的同时向 AR 内注 Glut 则电针兴奋 LC 的后期效应被取消;表明电针过程中有弓状核β-End 能神经元对 LC 的抑制作用参与。但此抑制作用似弱于电针对 LC 的兴奋效应,故只能取消(较弱的)电针的后期效应。不能取消(较强的)电针的早期效应。     

Electrophysiological study of 5,6-dihydroxytryptamine effect on defensive reflex conditioning in the snail

The role of serotonin in expression of membrane properties of identified neurons was studied during defensive reflex conditioning using the neurotoxic analogue of serotonin 5,6-dihydroxytryptamine (5,6-DHT). The defensive reflex conditioning in snails was destroyed on the second day after second injection of 5,6-DHT. Through the 1st weeks after second injection of 5,6-DHT the snails were learned but worse than snails after injection of saline solution. This result shows the recovery of snail's learning ability within 2 weeks after the second injection of 5,6-DHT. It was found that injection of 5,6-DHT prevented the decrease of membrane and threshold potentials of command neurons during defensive reflex conditioning as compared with the snails injected with 5,6-DHT without learning.     

The action of the neurotoxins 5,6-dihydroxytryptamine and p-chlorophenylalanine on the electrical activity parameters of the command neurons during long-term sensitization and learning in the snail]

The influence of 5,6-dihydroxytryptamine (5,6-DHT), which selectively destroyed serotonin terminals, and p-chlorphenylalanine, which inhibited serotonin synthesis, was studied on the long-term sensitization (LTS) in a snail. The membrane mechanisms were analyzed by measuring electrical characteristics of command neurons of defensive behavior LPa3, RPa3, LPa2, and RPa2. Snails injected with saline served as an active control. It was shown that the injected drugs inhibited the LTS in certain concentrations. A significant increase was observed in the membrane potential and the threshold of the action potential generation in the command neurons after 5,6-DHT injection in the doses of 20 and 30 mg/kg (in comparison with the active control). Sensitization of snails injected with saline solution led to the LTS and decrease in the membrane and threshold potentials of the command neurons. After the LTS, changes in membrane and threshold potentials in snails injected with 5,6-DHT were negligible in comparison with those injected with 5,6-DHT but without the LTS. Neither the LTS nor subsequent learning resulted in a further decrease in membrane and threshold potentials. Thus, the neurotoxin injection led to an increase in excitability of command neurons and their depolarization, and the LDS did not elicit further excitability increase. Since the shifts of the threshold and membrane potentials were the same, it was concluded that the increase in membrane excitability was induced by the depolarizing shift of the membrane potential.     

Comparative investigation of influence of injections of chlorpromazine and 5,6-dihydroxytryptamine on locomotion, defensive reactions of snall helix lucorum and excitability of command neurons in long-term sensitization

Comparative analysis of the action of chlorpromazine (CPZ) and neurotoxin 5,6-dihydroxytryptamine (5,6-DHT) on defensive reactions and locomotion of grape snail and elaboration of long-term sensitization (LTS), was carried out. Long-term (chronic) injection of chlorpromazine led to significant increasing of a pneumostome closing time and to changing of motor behaviour towards decrease of the velocity of the locomotion. Daily injections of 5,6-DHT in small doses within a week were accompanied by the gradual decrease of the velocity of snails locomotion, which was kept for a week. Similar effect was observed in injection of neurotoxin (30 mgs/kg). Injections of CPZ prevents elaboration of LTS, as well as injections of 5,6-DHT. After the action of CPZ, LTS, LTS followed by CPZ, and also during elaboration of LTS after injection of CPZ, the velocity of locomotion directly depended on the length of leg. During elaboration of LTS after injection of 5,6-DHT, such dependency is not retained. Electrophysiological study revealed that chronic injections of CPZ led to depolarizing shift of membrane potential and decrease of the threshold of action potential generation in command neurons as after injection of neurotoxin 5,6-DHT. Therefore, the action of neuroleptic drug CPZ on the defensive behaviour, locomotion of grape snail and electrical characteristics of identifying neurons is comparable with the action of toxic analogue of serotonin.     

Functional recuperation of the serotoninergic innervation in the rat locus coeruleus

1. 5,6-dihydroxytryptamine (5,6-DHT) or a lesion of the raphe centralis superior (RCS) cause significant decreases in the serotonin (5-HT) content and significant increases in the tyrosine hydroxylase activity in the locus coeruleus (LC) of the rat. This suggests that noradrenaline (NA) synthesis is controlled by serotonin-containing neurons in the raphe system via their terminals in the LC. 2. Radioautography after intraventricular infusion of tritiated serotonin (3H-5-HT) and biochemical determinations of endogenous 5-HT content showed an almost complete disappearance of serotoninergic axonal varicosities and content in the LC region 10-15 days after intraventricular administration of 75 micrograms of 5,6-DHT. Two to 4 months after neurotoxin administration, 5-HT fibers had regrown in the LC but, contrary to the normal innervation pattern, the majority of them invaded the medial most portion of the nucleus and the adjacent subependymal region. The LC region regained almost all of its endogenous 5-HT content in the same time period. 3. Functional recuperation of these 5-HT fibers was demonstrated by the fact that the RCS had, after regeneration, the same functional control on NA synthesis as in the normal animal.     

False transmitter, 5,6-dihydroxytryptamine as a tool in selecting serotonergic Helix neurons for studies with isolated,dialysed cells

1. Dialysed serotonergic neurons were identified, isolated from the ganglia of 5,6-dihydroxytryptamine (5,6-DHT) treated snail, Helix pomatia L. Twenty-four to 40 days after injection of 5,6-DHT into the animal, serotonergic neurons show a specific brown pigmentation, which stays there for several weeks. After protease digestion (0.5–1.0 mg/ml for 10–12 min) the labelled neurons can be easily separated. This method ensures the reliable identification of serotonergic neurons for intracellular dialysis.2. We showed that isolated serotonergic neurons maintain their membrane characteristics, and ion-currents can be registered under voltage clamp, just as from neurons of untreated animals. The threshold concentration of serotonin (10 ⁻⁷ M) and the survival time of pigment labelled dissociated cells were the same as for the control cells.3. Following 5-HT application, the voltage activated Ca-currents were either increased or decreased, depending on the neuron used.4. The different responses are probably caused by different receptors on the cell membrane or by the presence of different types of Ca-channels.5. The deactivation time constant of the Ca-current, calculated from the tail current, was also altered in the pigment labelled neuron following serotonin treatment.     

The effect of 5,6-dihydroxytryptamine on post-decapitation convulsions

Previous data (1) have shown that L-DOPA increases the duration of the clonic phase of post-decapitation convulsions (PDC) in mice. It was suggested that this effect is produced by depleting 5-hydroxytryptamine (5-HT) in the inhibitory bulbospinal pathways and thus enhancing reflex activity in the spinal cord. If this were true then L-DOPA administration should not influence clonic PDC in animals whose 5-HT pathways were destroyed. We therefore tested the effects of L-DOPA on mice 3 weeks after pretreatment with the 5-HT neurotoxin, 5,6-dihydroxytryptamine (5, 6-DHT) (50 μg/kg, intracerebroventricularly). All mice were given the peripheral decarboxylase inhibitor, Ro 4-4602. 5,6-DHT halved the brain 5-HT levels and significantly increased the duration of clonic PDC. The administration of L-DOPA (320 mg/kg i.p.) to 5,6 DHT treated mice did not produce any further significant increases in duration. The administration of 5-hydroxytryptophan (5-HTP) (100 mg/kg, i.v.) to 5,6-DHT treated mice, however, increased 5-HT to above control levels and reduced convulsions to control levels. Administration of both 5-HTP and L-DOPA to 5,6-DHT treated mice resulted in 5-HT levels and convulsion times which were also not significantly different from the controls. These data give additional indication that intact 5-HT nerve terminals are necessary for L-DOPA to prolong the duration of clonic PDC.     

Polarographic Measurements of Spontaneous and Mitochondria-Promoted Oxidation of 5,6-and 5,7-Dihydroxytryptamine

Abstract: Spontaneous oxygen consumption by 5,6- and 5,7-DHT (dihydroxytryptamine), related indoleethylamines, and 6-hydroxydopamine and oxygen consumption by these compounds in the presence of rat liver mitochondria were measured by the polarographic oxygen electrode technique. 5,6- and 5,7-DHT react with oxygen at very different rates (2.7 nmol O₂/min and 33.4 nmol O₂/min, respectively) when incubated in buffer, pH 7.2, at a concentration of 1 mm and with different kínetic characteristics. While the oxidation of 5,7-DHT obeys a reaction of second-order type, the oxidation of 5,6-DHT is more complex and characterized by autocatalytic promotion. Coloured quinoidal oxidation products appeared during the degradation of both indoleamines. Glutathione, ascorbate, dithiothreitol, cysteine, albumin, and superoxide dismutase partially prevented 5,6- and 5,7-DHT from oxidative destruction. Catalase saved oxygen only in the case of 5,6-DHT by recycling of O₂ released from near-stoichiometrically formed H₂O₂ during oxidation of 5,6-DHT: 5,7-DHT did not generate H₂O₂ in measurable amounts. Oxygen consumption rates of 5,6- and 5,7-DHT were enhanced after addition of rat liver mitochondria to the incubation medium; this resulted in an accelerated formation of quinoidal products. This stimulatory effect on the oxidation rates of both 5,6- and 5,7-DHT was blocked by cyanide, but not rotenone, and was abolished by boiling of the mitochondria fraction. The observed increase in oxygen consumption in the presence of mitochondria was found not to be influenced by monoamine oxidase-dependent deamination of 5,6- and 5,7-DHT. It is postulated that 5,6- and 5,7-DHT are capable of participating in the electron transfer of the mitochondrial respiration chain beyond complex III. Results obtained in determinations of ADP:0 ratios in respiratory control experiments exclude a possible interference of 5,6-DHT, 5,7-DHT, and 6-OH-DA with phosphorylating sites. During the activated state of respiration, no signs of electron transfer inhibition by 5,6- and 5,7-DHT were detectable. A comparison and evaluation of the autoxidation rates of various hydroxylated indoleethylamines, of their affinity to the 5-HT transport sites, and their neurotoxic potency in vivo reveals that interaction of these compounds with oxygen at restricted reaction velocity is a prerequisite for efficient toxicity in monoaminergic neurons following active accumulation in these neurons via the high-affinity uptake systems.     

Central oxytocin enhances antinociception in the rat

The study aimed to investigate the effect of oxytocin on antinociception in the rat. The pain threshold was elevated by oxytocin following intraventricular (icv) or intrathecal injection (ith), and reduced by anti-oxytocin serum (icv or ith). But the pain threshold was not altered by intravenous injection (iv) of oxytocin or anti-oxytocin serum. Pain stimulation induced oxytocin concentration decrease in the hypothalamic supraoptic nucleus, and increase in the locus coeruleus, raphe magnus nucleus, caudate nucleus and spinal cord, but no change in the hypothalamic paraventricular nucleus and plasma. The results indicated that central, not peripheral oxytocin could enhance antinociception.     

Single cell shape and population densities of indoleamine-accumulating and displaced bipolar cells in Reeves' turtle retina

Two types of bipolar cell in the Geoclemys reevesii retina were studied quantitatively by means of specific cell labelling with an indoleamine derivative (5,6-dihydroxytryptamine, 5,6-DHT), a nucleic acid stain (4,6-diamidino-2-phenylindole, DAPI) and Lucifer yellow CH. Indoleamine-accumulating (IA) bipolar cells were selectively labelled with 5,6-DHT applied intraocularly. After the cells accumulated 5,6-DHT, the indoleamine fluorescence was photoconverted to diaminobenzidine products to allow observation of morphological details. Close examination of many cells (cell number; n = 120) showed that the IA bipolar cells consist of a single morphological type whose axon collaterals ramify sublaminae 1, 4 and 5 respectively. This terminal branching pattern corresponds to cells that hyperpolarize when their receptive field centres are illuminated (Weiler 1981). The density of IA bipolar cells was highest in the visual streak (4130 cells mm-2) and lowest at the peripheral margin (1970 cells mm-2). By applying a small amount of DAPI to the eye, nuclei located in the most proximal row of the outer nuclear layer were labelled selectively. By using selective intracellular dye injection into DAPI-labelled cells under fluorescence microscope (Tauchi & Masland 1984, 1985), these cells were found to have Landolt's clubs and single descending axons. Dye injections into more than fifty DAPI-labelled somata showed that they belonged exclusively to displaced bipolar cells. These comprised at least two subtypes that differ in the ramification pattern of their axon terminals within the inner plexiform layer: one was monostratified, whereas the other was bistratified. The displaced bipolar cell density was as high as 9400 cells mm-2 in the central retina, falling to 2000 cells mm-2 in the superior margin. In vitro Lucifer labelling revealed that the overall bipolar cell density in the central retina was as high as 39,300 cells mm-2. Both the conventionally located and displaced bipolar cells were included in this population. About 11% of the total bipolar cell population consisted of IA bipolar cells. Assuming that one half of the conventionally located bipolar cells are the centre-hyperpolarizing type, IA bipolar cells represent approximately 28% of the total. As displaced bipolar cells represent almost one quarter of the total bipolar population, the dislocation of their somata stands out morphologically, inviting investigation of possible functional correlates.     

Neonatal 6-hydroxydopamine prevents adaptation to chemical disruption of the pituitary-adrenal system in the rat

Three days after the subcutaneous implant of a dexamethasone pellet, which depletes both corticosterone and ACTH, normal rats showed impaired acquisition of a two-way avoidance task. Rats who had received systemic 6-hydroxydopamine at birth to lesion the forebrain noradrenergic terminals from the locus coeruleus did not show this impairment. After a single injection of metyrapone, which inhibits corticosterone synthesis and increases ACTH release, both intact and norepinephrine (NE)-depleted rats showed impaired avoidance acquisition. After the seventh injection, however, acquisition in normal rats was no longer impaired by the drug while the NE-depleted rats were still deficient. These results indicate that the simple combination of forebrain NE loss with reduced corticosterone levels does not necessarily retard avoidance acquisition. Rather, they suggest that the NE efferents from the locus coeruleus are essential for the brain's adaptation to at least some behavioral consequences of changes in the circulating level of ACTH.