海洛因对大鼠中脑腹侧被盖区中Nurr1蛋白表达的影响

目的:以Sprague Dawley(SD)大鼠为实验对象,明确海洛因急慢性给药对大鼠的中脑腹侧被盖区(ventral tegmental area,VTA)中孤儿核受体Nurr1表达水平的影响。方法:SD大鼠海洛因(3 mg·kg-1,s.c.)急性给药后15min、12 h、24 h,慢性给药(3 mk·kg-1/day,s.c.×10 d)后24 h断头取脑,蛋白质免疫印记分析(Western-blot)检测药物依赖关键脑区VTA中的Nurr1蛋白表达的变化。结果:海洛因急性给药15 min后,VTA脑区中的Nurr1表达较对照组明显增加(P<0.05);而海洛因慢性给药后,VTA脑区中Nurr1表达水平与对照组相比没有显著性差异。结论:海洛因急性给药可激活大鼠VTA中Nurr1的表达,且Nurr1在VTA中呈现快速表达的特点。     

四氢原小檗碱对吗啡成瘾大鼠脑中腹侧被盖区神经胶质纤维酸性蛋白和酪氨酸羟化酶含量的影响

目的　研究吗啡依赖对大鼠中脑腹侧被盖区(VTA)神经元功能变化以及四氢原小檗碱 (THPB)的干预作用。方法　大鼠ip递增剂量的吗啡 10d建立吗啡依赖模型 ,d 11起分别ipTHPB 30mg·kg- 1或等体积生理盐水 (NS) ,每天 2次 ,治疗 12d或 30d。取脑 ,冰冻切片 ,用ABC法测VTA等脑区神经胶质纤维酸性蛋白 (GFAP)和酪氨酸羟化酶 (TH)的免疫组织化学反应 ,测定其阳性神经元的平均吸光度值表示其含量。结果　吗啡成瘾大鼠VTA中GFAP和TH免疫阳性神经元含量持续高于正常对照组 (P <0 .0 5 ) ;THPB治疗 12d或 30d显著降低GFAP和TH免疫阳性神经元含量的增高 ,与正常对照组比无显著性差异。结论　吗啡成瘾造成大鼠脑VTA“特异性损伤” ,THPB可逆转此损伤 ,对吗啡成瘾大鼠脑有保护作用 ,提示其有可能用于阿片类成瘾的防治。     

中脑腹侧被盖区多巴胺能神经元短暂激活未能诱发小鼠自身给糖行为的重建

目的 研究脑内奖赏系统中脑腹侧被盖区（VTA）多巴胺（DA）能神经元对灯光线索诱发小鼠自身给糖行为重建的调控作用。方法 以转基因DAT-Cre小鼠为研究对象,通过病毒转染的方式,将光敏感通道蛋白特异性表达在小鼠VTA的DA能神经元上。采用10%蔗糖水训练注射过携带光敏感通道蛋白重组腺相关病毒的小鼠建立自身给糖行为,然后进行首次消退训练,达到首次消退期稳定阶段（连续3 d小鼠有效鼻触次数<10%形成期稳定操作值）后,进行线索点燃测试。随后,小鼠进行第2次消退,达到第2次消退期稳定阶段,采用频率20 Hz、脉冲时长15 ms的激光连续100次集中刺激VTA的DA能神经元。最后进行第3次消退,达到第3次消退期稳定阶段,采用频率80 Hz、脉冲时长15 ms的激光连续100次集中刺激VTA的DA能神经元。结果 在相关灯光线索刺激下,小鼠出现蔗糖行为重建,表现为有效鼻触次数较首次消退期稳定阶段有效鼻触次数（首次消退期最后3 d的有效鼻触次数平均值）显著升高（P<0.01）,提示蔗糖相关灯光线索可显著诱发小鼠觅糖行为重建,实验模型建立成功。20 Hz 15 ms或80 Hz 15 ms...     

四氢原小檗碱同类物与多巴胺D2受体作用的定量构效关系

四氢原小檗碱同类物是一类新型的脑内多巴胺受体阻滞剂。本文用Ｆｒｅｅ－Ｗｉｌｓｏｎ法研究了一组四氢原小檗碱同类物与多巴胺Ｄ２受体作用的定量构效关系。结果表明：Ｃ－２位上的ＯＨ和Ｃ－１２位上的Ｃｌ使四氢原小檗碱同类物与多巴胺Ｄ２受本的亲和力增强,Ｃ－１１位上的立体位阻可能使亲和力降低,Ｃ－１０位上的取代基可能对亲和力影响不大。     

神经肽urocortin Ⅱ对吗啡成瘾大鼠中脑腹侧被盖区神经元自发放电的影响

<正>吗啡等阿片类药物反复使用可造成耐受、依赖和成瘾,中断给药出现戒断综合征、强迫觅药行为及戒断后焦虑等精神神经系统症状,对患者的身心有严重影响。目前已知,中脑边缘-多巴胺系统与药物成瘾和强化作用形成密切相关,此系统起自中脑腹侧被盖区(ventral tegmental area,VTA),投射到杏仁中央核(nucleus centralis amygdalae,NAC)、内侧前额叶皮层(mPFC)等前脑区域,并涉及多种神经递质的投射。其中VTA是中脑的重要神经核团,在此系统中发挥非常关键的作用,因此受到人们的关注。神经肽urocortin(UCN)是由40个氨基酸组成的高活性神经肽,属促肾上腺皮质激素释放因子(CRH)家族,与CRF具有相似的生物学     

THPBs同类物对多巴胺D1、D2、D3受体的作用特性

目的：左旋千金藤啶碱（1-SPD）是四氢原小檗碱同类物（THPBs）的先导化合物，本课题组已证明其具有D1受体激动和D2受体阻滞的双重作用，并积极倡导D1激动-D2阻滞双重作用与精神分裂症病因相匹配治疗的设想。新近又发现SPD对D3受体有较强的亲和力，因此本研究SPD的更理想的结构关系。国际上新发现D3受体与精神分裂症、帕金森病和药物成瘾的发病过程均有密切关系。方法：利用体外基因表达技术，构建了表达D1受体、D2受体、D3受体的Sf9细胞和稳定表达D3受体的NG108细胞。     

四氢原小檗碱类与多巴胺受体相互作用的分子模拟

建立多巴胺受体三维结构模型，结合受体模型从分子水平上研究四氢原小檗碱类对多巴胺受体的作用机制。以细菌视紫红质的三维结构为模板，在计算机上建立多巴胺Ｄ１和Ｄ２受体的三维结构模型，选择左旋千金藤立定作为ＴＨＰＢ的先导化合物，将它分别对接到Ｄ１和Ｄ２受体的结合位点。     

Nicotinic and dopamine D2 receptors mediate nicotine-induced changes in ventral tegmental area neurotensin system

Neuropeptides have been implicated in the psychopathology of stimulants of abuse. Neurotensin is a neuropeptide associated with the regulation of the nigrostriatal and mesolimbic dopamine pathways. In addition, the ventral tegmental area, a midbrain region implicated in the rewarding effects of most, if not all, addictive drugs, appears to be a particularly critical target for nicotine action. Because neurotensin has been linked with both mesolimbic and mesocortical dopamine function, we examined the impact of nicotine treatment on central nervous neurotensin systems by measuring changes in neurotensin tissue content because it has been shown that such changes reflect alterations in release and activity of this peptide system. Male Sprague-Dawley rats received multiple administrations of (+/-) nicotine 4.0 mg/kg/day (0.8 mg/kg, i.p.; 5 x 2-h intervals) in the presence or absence of selective dopamine receptor antagonists (dopamine D(1); SCH 23390 or dopamine D(2); eticlopride) or two doses of the non-selective nicotinic acetylcholine receptor antagonist (mecamylamine; 3.0 and 6.0 mg/kg, s.c.). The nicotine treatment significantly decreased neurotensin-like immunoreactivity content in the ventral tegmental area, as well as related regions such as prefrontal cortex, substantia nigra, and anterior striatal region 12-18 h after drug treatment, but not the nucleus accumbens. The nicotine-mediated decrease in the neurotensin-like immunoreactivity of the ventral tegmental area was selectively blocked by a specific dopamine D(2), but not a dopamine D(1), receptor antagonist, while mecamylamine attenuated at the low (3.0 mg/kg) and completely blocked at high (6.0 mg/kg) dose this nicotine effect. These findings with previous studies, suggest that nicotine-mediated dopamine release activates D(2) receptors which in turn increases neurotensin release, turnover and acutely reduces tissue levels in the ventral tegmental area and other limbic and basal ganglia structures.     

Characteristics of tetrahydroprotoberberines on dopamine D1 and D2 receptors in calf striatum.

AIM: To study the characteristics of tetrahydroprotoberberines (THPB) on dopamine D1 and D2 receptors and elucidate their structure-activity relationship. METHODS: Radioligand assay in vitro with a two-site model program analysis. RESULTS: Four THPB with two hydorxyl groups on C2 and C9 or C2 and C10 exhibited RH and RL two binding sites to D1 receptors and guanosine triphosphate regulated the RH binding site of SPD and THPB-132A in competition assay, while eleven THPB including nonhydroxy-THPB, monohydroxy-THPB, and THPB with two hydroxyl groups attaching to C3 and C10 showed one binding site to D1 receptors under the same conditions. However, the tested eleven THPB all manifested one binding site to D2 receptors in competition assay, and the 2-hydroxy-THPB had the most potent affinity for D2 receptors. CONCLUSION: Dihydroxy-THPB with two hydroxyl groups attaching to C2 and C9 or C2 and C10 possess the intrinsic activity of agonist to D1 receptors, while the other THPB do not. The tested eleven THPB all are the antagonists of D2 receptors.     

Clonidine modulates dopamine cell firing in rat ventral tegmental area

The effect of clonidine (5-20 micrograms/kg i.v.) on the activity of single, identified dopamine neurons in the ventral tegmental area of the mesencephalon was studied in chloral hydrate-anesthetized male rats. Clonidine regularized cell firing without affecting the firing rate of the neurons. This effect was blocked by idazoxan (0.5 mg/kg i.v.) or yohimbine (1.0 mg/kg i.v.), but not by phentolamine (1.0 mg/kg i.v.), indicating that clonidine acts at central alpha 2-adrenoceptors. Idazoxan or yohimbine alone produced deregularization and excitation of cell firing. Pretreatment with reserpine (5 mg/kg s.c.) 4 h before the experiment abolished the neuromodulatory effect of clonidine. Thus, the regularization of ventral tegmental area dopamine cell firing by clonidine is indirect and dependent on endogenous monoamines in brain, and, in principle, a tonic adrenergic control of DA cell firing pattern is indicated. The regularization of DA cell activity produced by clonidine may underlie certain therapeutic neuropsychiatric actions of the drug.     

Cholecystokinin octapeptide potentiates the inhibitory response mediated by D2 dopamine receptors in slices of the ventral tegmental area of the brain in the rat

The ability of cholecystokinin octapeptide (CCK8) to modulate dopamine (DA)-induced inhibition of the firing of neurons in the ventral tegmental area of the rat was examined. Extracellular recordings were obtained from putative DA-containing neurons, identified on the basis of their electrophysiological characteristics and response to DA, in an in vitro slice preparation from the ventral tegmental area of the brain. Application of DA produced a concentration-dependent reduction in firing rate. This DA-induced inhibition was mimicked by the D2 selective agonist, LY 171555 (trans-(-)-4,4a,5,6,7,8,8a,9-octahydro-5-propyl-2H- pyrazolo[3,4-g]quinoline), but not by the D1 selective agonist, SKF 38393 (R-(+)-2,3,4,5-tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine). The DA-induced inhibition was antagonized selectively by the D2 antagonist, l-sulpiride, but not by the D1 antagonist, SCH 23390 (R-(+)-8-chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1H-3-benzazepine-7- ol). However, CCK8 elicited a transient increase in firing rate in some neurons and, in addition, potentiated the inhibitory response evoked by DA or LY 171555. Again SKF 38393 was without effect following the administration of CCK8. Taken together, these results suggest that DA-induced inhibition of DA-containing neurons in the ventral tegmental area of the brain of the rat is mediated by activation of D2-receptors and that CCK8 potentiates this D2-mediated inhibition.     

Neuronal excitation in the ventral tegmental area modulates the micturition reflex mediated via the dopamine D1 and D2 receptors in rats

Involvement of the dopamine D(1) and D(2) receptors in the ventral tegmental area (VTA) in the micturition reflex was investigated using female Sprague Dawley rats under urethane anesthesia. Cystometrograms during continuous infusion of warmed saline into the bladder were recorded. When intervals of bladder contraction became constant, the excitatory amino acid DL-homocysteic acid (DLH) was microinjected into the VTA and changes in the cystometric parameters were observed. The selective D(1) antagonist SCH23390 (0.3 mg/kg) or D(2) antagonist eticlopride (0.1 mg/kg) was subcutaneously injected (s.c.) 15 min prior to the DLH treatment. A low dose of DLH (3 microg) facilitated the micturition reflex, whereas a high dose of DLH (30 microg) inhibited the micturition reflex. However, a middle dose of DLH (10 microg) did not show any effect. The facilitated micturition reflex induced by a low dose of DLH was inhibited by the selective dopamine D(2) antagonist eticlopride, but unaltered by SCH23390, a selective dopamine D(1) antagonist. In contrast, the inhibitory effect induced by a high dose of DLH on the micturition reflex was suppressed by SCH23390 but not eticlopride. These results suggested that the facilitated micturition reflex might be mediated via the dopamine D(2) receptors, while the inhibitory effects on the micturition reflex was mediated via the dopamine D(1) receptors.     

Effects of phencyclidine on ventral tegmental A10 dopamine neurons in the rat

Extracellular single unit recordings were used to determine the effects of systemically and iontophoretically applied phencyclidine on electrophysiologically-identified A10 dopamine neurons within the ventral tegmental area of the rat. Intravenous injections of phencyclidine inhibited, as well as excited A10 cells. Approximately 42% of the inhibitions induced by phencyclidine were completely reversed by an injection of haloperidol. The excitatory effects of phencyclidine most often resulted in a state of depolarized inactivation and were not sensitive to haloperidol. The alteration of the activity of A10 cells by systemically-applied phencyclidine was effectively prevented in all animals pretreated with a combination of reserpine and alpha-methyl-p-tyrosine, which depleted stores of central catecholamines by over 90%. In contrast to the data obtained with intravenous injections, iontophoretic applications of phencyclidine produced only inhibition of the activity of the A10 cells and, in a few of these cases the decreased firing rate was accompanied by an increase in the amplitude of the action potential. A comparison of the response patterns of dopaminergic neurons to systemically- and iontophoretically-applied phencyclidine would suggest that excitations induced by phencyclidine are not mediated at the level of the A10 cell bodies but through a site outside the ventral tegmental area. The results of this study also indicate that some of the effects of phencyclidine on the activity of A10 neurons are clearly dependent upon an interaction with dopamine and thus, would support the hypothesis that phencyclidine can act as an indirect dopamine agonist. However, other effects of phencyclidine which are not apparently linked to dopamine may represent the response of a pharmacologically-distinct subpopulation of A10 neurons to phencyclidine.     

Clozapine blocks D-amphetamine-induced excitation of dopamine neurons in the ventral tegmental area.

Current antipsychotic drugs are thought to inhibit central dopamine (DA) transmission by blocking DA receptors. Here, we provide evidence that the atypical antipsychotic drug clozapine may produce part of its effect by inhibiting a subset of excitatory inputs to DA neurons. Thus, in chloral hydrate-anesthetized rats, systemic administration of D-amphetamine produced two opposing effects on DA neurons in the ventral tegmental area. Under control conditions, D-amphetamine inhibited the firing of the cell through D2-like receptors. When D2-like receptors were blocked by raclopride, D-amphetamine excited DA neurons, instead of producing no effect. The excitation, expressed as an increase in firing rate and a slow oscillation in firing pattern, was suppressed by the adrenergic alpha1 receptor antagonist prazosin, suggesting an involvement of alpha1 receptors. In rats pretreated with the typical antipsychotic drug haloperidol, D-amphetamine also excited DA neurons. However, when given after clozapine, D-amphetamine produced no significant effects. The failure of D-amphetamine to produce an excitation is not due to an incomplete blockade of D2-like receptors by clozapine because co-treatment with clozapine and raclopride also failed to enable the excitatory effect of D-amphetamine. The suggestion that clozapine inhibits the excitatory effect of D-amphetamine is further supported by the finding that clozapine, given after D-amphetamine, reliably reversed D-amphetamine-induced excitation in raclopride-treated rats. Thus, different from raclopride and haloperidol, clozapine may inhibit DA transmission through two additive mechanisms: blockade of DA receptors and inhibition of an amphetamine-sensitive, excitatory pathway that innervates DA neurons.     

Inhibitory action of clozapine on rat ventral tegmental area dopamine neurons following increased levels of endogenous kynurenic acid.

The mode of action by which the atypical antipsychotic drug clozapine exerts its superior efficacy to ameliorate both positive and negative symptoms is still unknown. In the present in vivo electrophysiological study, we investigate the effects of haloperidol (a typical antipsychotic drug) and clozapine on ventral tegmental area (VTA) dopamine (DA) neurons in a situation of hyperdopaminergic activity in order to mimic tentatively a condition similar to that seen in schizophrenia. Increased DA transmission was induced by elevating endogenous levels of the N-methyl-D-aspartate receptor and alpha7(*) nicotinic receptor antagonist kynurenic acid (KYNA; by means of PNU 156561A, 40 mg /kg, i.v.). In control rats, i.v. administered haloperidol (0.05-0.8 mg/kg) or clozapine (1.25-10 mg/kg) was associated with increased firing rate and burst firing activity of VTA DA neurons. However, in rats displaying hyperdopaminergia (induced by elevated levels of KYNA), the effects of clozapine on VTA DA neurons were converted into pure inhibitory responses, including decrease in burst firing activity. In contrast, haloperidol still produced an excitatory action on VTA DA neurons in rats with elevated levels of endogenous brain KYNA. The results of the present study suggest that clozapine facilitates or inhibits VTA DA neurotransmission, depending on brain concentration of KYNA. Such an effect of clozapine may be related to its unique effect in also ameliorating negative symptoms of schizophrenia.     

Effects of methamphetamine on dopamine cells in the substantia nigra pars compacta and the ventral tegmental area

The ability of methamphetamine to inhibit the firing rate of dopamine cells in the substantia nigra pars compacta (SNC) and the ventral tegmental area (VTA) was studied. Methamphetamine reduced the firing rates of the dopamine cells in a dose-dependent manner in the SNC and the VTA. The doses of methamphetamine required to produce a 50% inhibition of firing rate in the SNC and the VTA were 0.37 mg/kg and 0.28 mg/kg, respectively.