CORE 3: RAT ANTIPSYCH. DRUG TESTING CORE

核心 3：老鼠反心理。

• 批准号: 7451388
• 负责人: Bryan L. Roth
• 金额: --
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-11-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7451388
• 关键词: Acclimatization; Acoustics; Adverse effects; Agonist; Amphetamines; Animal Model; Animals; Antipsychotic Agents; Apomorphine; Attenuated; Auditory; Behavior; Behavioral; Behavioral Assay; Behavioral Model; Biological Assay; Caliber; Catalepsy; Complement; Computer software; Computers; Condition; Conditioned Stimulus; Count; Data; Development; Dextroamphetamine; Dopamine; Dopamine Agonists; Dopamine D2 Receptor; Dose; Drug Kinetics; Drug effect disorder; Ensure; Exhibits; Failure; Figs - dietary; Floor; Functional disorder; Glutamates; Goals; HTR2A gene; Haloperidol; Hour; Hyperactive behavior; Individual; Injection of therapeutic agent; Light; MDL 100907; Maintenance; Measurement; Measures; Modeling; Monitor; Motor; Motor Activity; Movement; Mus; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; NMDA receptor antagonist; Neurons; Noise; Numbers; Pathway interactions; Patients; Performance; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacology; Phencyclidine; Physiologic pulse; Plexiglas; Plexiglass; Positioning Attribute; Posture; Pre-Clinical Model; Pulse taking; Range; Rattus; Reading; Records; Reporting; Rodent; Rodent Model; Role; Schedule; Schizophrenia; Score; Sedation procedure; Serotonin Receptor 5-HT2A; Shock; Side; Source; Stainless Steel; Stimulus; Symptoms; Syndrome; System; Testing; Time; Training; Transgenic Organisms; Week; atypical antipsychotic; avoidance behavior; behavior measurement; day; detector; drug discovery; drug efficacy; drug testing; experience; foot; in vitro Assay; in vivo; instrument; member; mouse model; neurotransmission; postsynaptic; pre-clinical; predictive modeling; prepulse inhibition; receptor; receptor internalization; response; retinal rods; stereotypy

The NCDDG proposes to ascertain the extent to which compounds that differentially display functional selectivity in a range of in vitro assays can be differentiated in a battery of in vivo tests that predict antipsychotic drug-like efficacy. The central role of dopamine in the pathophysiology of schizophrenia and in the modes of action of clinically efficacious anti-psychotics has led to the development and common use of rodent behavioral models that are highly predictive of anti-psychotic drug efficacy primarily as a result of blockade of dopamine D2 receptors. Several behavioral animal models that are predictive of antipsychotic drug efficacy will be employed to profile the functionally selective compounds identified by the NCDDG, including conditioned avoidance responding, antagonism of apomorphine-induced climbing and stereotypy (AICS), blockade of PCP/amphetamine induced locomotor activity, prepulse inhibition (PPI) of the acoustic startle response. Measurements of drug-induced catalepsy will be performed to assess the potential for extrapyramidal motor side-effects liability. Where appropriate, routine pharmacokinetic measurements will be employed in order to ensure that the compound exposure is sufficient. Selective 5-HT2A antagonists such as M100907 have shown activity in some, but not all, of these models. While M100907 is reported to reduce amphetamine and PCPstimulated locomotor activity (Sorensen et al., 1993; Maurel-Remy et al., 1995; Gleason and Shannon, 1997) and attenuate MK-801 induced prepulse inhibition deficits (Varty et al., 1999), the compound does not block apomorphine-induced climbing (Sorensen et al., 1993) and there are no reports of efficacy in the conditioned avoidance response assay, despite reports of M100907 enhancing the efficacy of typical antipsychotics like haloperidol (Wadenberg et al., 2001). These data underscore the potential for the selected behavioral assays to further extend and contrast the antipsychotic profiles of compounds varying in effects on 5-HT2A receptor internalization identified as part of Project 3. In addition, as many dopamine D2 partial agonists show activity in the preclinical models, these assays will be used to profile compounds with D2 functional selectivity profiles identified in Project 1. These studies are proposed to complement the mechanistic studies proposed in Project 2 that will employ transgenic mouse models of NMDA receptor hypofunction and dopamine hyperactivity. Conditioned avoidance responding. The conditioned avoidance responding model has been shown to be highly predictive of anti-psychotic drug efficacy (Wadenburg and Hicks, 1999). The model is performed using Shuttlebox test chambers (MED Associates, St Albans, VT) divided into two compartments by an archway. Each chamber floor half is composed of thirteen 3/16" diameter stainless steel grid rods placed on 1/2" centers wired for the presentation of an electric foot shock (0.5 mA). In addition, each side of the chamber is equipped with a stimulus light, tone generator and two infrared beam source/detectors used to locate the rat within the chamber. Rats trained to avoid the foot shock are placed in the chambers for a 4-minute habituation period followed by 50 trials presented on a 15-second variable interval schedule (range = 7.5 - 22.5 seconds). Each trial consists of a 10-second warning tone and stimulus light (conditioned stimulus) followed by a 10- second shock (unconditioned stimulus), presented through the grid floor on the side where the rat is located, in the presence of the tone and Conditioned avoidance responding. The conditioned avoidance responding model has been shown to be highly predictive of anti-psychotic drug efficacy (Wadenburg and Hicks, 1999). The model is performed using Shuttlebox test chambers (MED Associates, St Albans, VT) divided into two compartments by an archway. Each chamber floor half is composed of thirteen 3/16" diameter stainless steel grid rods placed on 1/2" centers wired for the presentation of an electric foot shock (0.5 mA). In addition, each side of the chamber is equipped with a stimulus light, tone generator and two infrared beam source/detectors used to locate the rat within the chamber. Rats trained to avoid the foot shock are placed in the chambers for a 4-minute habituation period followed by 50 trials presented on a 15-second variable interval schedule (range = 7.5 - 22.5 seconds). Each trial consists of a 10-second warning tone and stimulus light (conditioned stimulus) followed by a 10- second shock (unconditioned stimulus), presented through the grid floor on the side where the rat is located, in the presence of the tone and light. If an animal crosses hrough the archway during the nitial 10 seconds of the trial, the one and light are terminated and he response is considered an jvoidance response. If an animal Bosses through the archway after a foot shock is initiated, the one, light and shock are erminated and the response is considered an escape response, f a response is made during an nter-trial interval (ITI), the animal s punished with a 0.5-second shock (0.5 mA). A MED \ssociates computer with dEDSTATE NOTATION¿ software controls the test session and counts the number of trials in which the animal avoids shock, escapes Only animals displaying stable performance (approximately 90% avoidance responding on the training session prior to test day) are considered "trained" and included on test day. Training is maintained by at least one nondrug test session each week. On test days, drugs are typically administered 30 minutes prior to testing. Clinically effective antipsychotics reduce avoidance behavior and increase escape behavior without increasing failures to respond that are indicative of sedation (Fig. 1). Antagonism of apomorphine-induced climbing and stereotypy (AICS). Stimulation of dopaminergic neurotransmission by administration of the dopamine D1/D2 agonist, apomorphine, produces sets of stereotypic behaviors indicative of activation of distinct dopaminergic neuronal pathways. Stimulation by apomorphine of mesolimbic DA receptors induces climbing behavior while activation of nigrostriatal DA receptors produces a stereotypic syndrome of sniffing-licking-gnawing behaviors termed stereotypy (Costall, et al. 1975 and 1980). Testing of anti-psychotic In order to measure these behaviors, drugs are administered thirty minutes prior to challenge with apomorphine (1 mg/kg sc). Five minutes after the apomorphine injection, the climbing behavior exhibited on a wire cage and stereotypy are scored and recorded for each animal. Readings are repeated every 5 minutes during a 30-minute test session. Scores for each animal are totaled for each set of behaviors and expressed as a percent of control values observed in vehicle-treated mice. In this model, typical anti-psychotics block both climbing and stereotypy at similar doses while atypical antipsychotics produce a block of climbing behavior at doses that do not inhibit stereotypy. As a result, this commonly used preclinical screen is useful for predicting efficacy in treating the positive symptoms of schizophrenia and provides an indication of the potential for motoric side-effects liability (Fig. 2). PCP/Amphetamine induced locomotor activity. Amphetamine induced locomotor activity represents a model for hyperdopaminergia and has been commonly used to predict anti-psychotic efficacy (Arnt et al. 1997). In this model the amphetamine administration induces dopamine release resulting in increased locomotor activity. Antipsychotics exert their effects by competing for postsynaptic dopamine D2 receptors in the mesolimbic pathway reducing the induced locomotor response. In similar fashion, NMDA antagonists, e.g. phencyclidine (PCP), which stimulate locomotor activity, have also been employed to assess the impact of glutamatergic hypofunction in schizophrenia (Coyle et al, 2003; Lewis et at 2003). Atypical antipsychotics have been shown to block hyperactivity induced with NMDA antagonists in rodents (Gleason and Shannon, 1997). In this model the capacity of anti-psychotic drugs to block spontaneous, amphetamine - induced and PGP - induced locomotor activity are compared. Drugs are administered to the mice concurrently with d-amphetamine (3 mg/kg, sc), phencyclidine (3 mg/kg, sc) or vehicle. The animals are then immediately placed in locomotor activity monitoring chambers for 70-minute tests during which time locomotor activity data is recorded under room light and white noise using Accuscan infrared beam activity monitors with enclosed Plexiglas chambers (8 in. x 8 in.). Accuscan Versamax and Versadat software (Columbus, OH) are used to convert the infrared beam breaks into horizontal activity counts in 5-minute bins (Fig. 3). Prepulse inhibition (PPI) of the acoustic startle response. PPI is a measure of sensorimotor gating that is commonly used to characterize antipsychotic compounds. PPI is reduced in schizophrenia patients and in rodents treated with dopamine agonists or NMDA receptor antagonists (Geyer et al. 2001). The PPI response in normal individuals is displayed when subjects are exposed to a startling auditory pulse a diminished startle response to a loud noise if it is preceded by a non-startling auditory tone (prepulse). Schizophrenic patients show a response whether stimulus is prepulse. Schizophrenic patients show a response whether stimulus is prepulse. Animals are subjected paradigm similar startle regardless of the startling preceded by a Animals are to the PPI using testing chambers (SR-LAB system, San Diego Instruments, San Diego, CA) that consist of a Plexiglas cylinder fitted with a piezoelectric accelerometer that detects movement of animal within the cylinder. A loudspeaker mounted above the cylinder provides background white noise, acoustic noise bursts and acoustic prepulses. Presentation of acoustic pulse and prepulse stimuli are controlled by the SR-LAB software and interface system, which also digitizes, rectifies and records the responses from the accelerometer. Mean startle amplitude is determined by averaging 100 1-ms readings taken from the beginning of the pulse stimulus onset. Test sessions consist of 60 total trials with a 15 sec inter-trial interval. Following a 5 min acclimation to a 64 dB background noise, four trial types (120 dB pulse, or a 69, 74, or 79 dB prepulse paired with a 120 dB pulse) are presented in a pseudorandom order. Drug is typically administered 30 min prior to test with disrupting agents, d-amphetamine (4 mg/kg, sc) and MK-801 (0.15 mg/kg, ip), administered afterwards. Prepulse inhibition is defined as 100-[(startle amplitude on prepulse trials/startle amplitude on pulse alone trials) x 100] (Fig. 4). This model measures the ability of anti-psychotic compounds to reverse deficits in auditory sensorimotor gating in rats induced by the indirect DA agonist amphetamine and the NMDA antagonist MK-801. Catalepsy. Assessment of drug-induced catalepsy is widely used as a preclinical screen for potential extrapyramidal motor side effect liability of anti-psychotic drug candidates (Hoffman and Donovan, 1995). High levels of blockade of dopamine D2 receptor, typically exceeding 80 %, results in catalepsy, which is defined as a lack of movement and maintenance of awkward posture. While typical anti-psychotic compounds produce catalepsy at doses comparable to their efficacious dose, atypical antipsychotic drugs exhibit a distinct separation between cataleptogenic and efficacious doses in models predictive of antipsychotic activity. In order to measure the cataleptogenic potential of anti-psychotic compounds, animals are monitored for cataleptic behavior at 30-minute intervals for 2 hours post dosing with drug. The cataleptic behavior measurement consists of draping the forelegs of the treated animals a thin horizontal rod 1 3/4" high. The amount of time (in seconds) for which the animal maintains this awkward position is recorded (60 second maximum). Maintenance of this position is considered catalepsy. Mean seconds spent in the catalepsy position for each dose at each time point is calculated. (Fig. 5).

NCDDG 提议确定化合物差异的程度 在一系列体外测定中显示的功能选择性可以在一系列体内测试中进行区分 预测抗精神病药物的类似疗效。 多巴胺在精神分裂症的病理生理学及其作用方式中的核心作用 临床有效的抗精神病药物促进了啮齿动物行为模型的开发和普遍使用 高度预测抗精神病药物疗效主要是由于多巴胺 D2 的阻断 将采用几种预测抗精神病药物疗效的行为动物模型。 分析 NCDDG 鉴定的功能选择性化合物，包括条件性回避 反应，拮抗阿朴吗啡引起的攀爬和刻板印象（AICS），封锁 PCP/苯丙胺诱导运动活动、声惊吓反应的前脉冲抑制 (PPI)。 将进行药物引起的僵直测量，以评估锥体外系运动的潜力 在适当的情况下，将采用常规药代动力学测量，以便 确保化合物暴露充足，如 M100907 等选择性 5-HT2A 拮抗剂。 据报道，M100907 可以减少安非他明和 PCP 刺激，但在某些模型（但不是全部）中具有活性。 运动活动（Sorensen 等人，1993；Maurel-Remy 等人，1995；Gleason 和 Shannon，1997） 并减弱 MK-801 诱导的前脉冲抑制缺陷（Varty 等，1999），该化合物不会阻断 阿朴吗啡引起的攀爬（Sorensen 等人，1993），并且没有关于在条件反射中的有效性的报告 尽管有报道称 M100907 可以增强典型抗精神病药物（如 氟哌啶醇（Wadenberg 等，2001）。这些数据强调了所选行为测定的潜力。 进一步扩展和对比对 5-HT2A 受体作用不同的化合物的抗精神病特性 内化被确定为项目 3 的一部分。此外，许多多巴胺 D2 部分激动剂显示出活性 在临床前模型中，这些测定将用于分析具有 D2 功能选择性特征的化合物 项目 1 中确定。这些研究旨在补充项目 1 中提出的机制研究 2 将采用 NMDA 受体功能低下和多巴胺过度活跃的转基因小鼠模型。 条件性回避反应模型已被证明可以 可以高度预测抗精神病药物的疗效（Wadenburg 和 Hicks，1999）。 使用 Shuttlebox 测试室（MED Associates，St Albans，VT），通过 每个房间的一半地板由十三根直径为 3/16 英寸的不锈钢网格杆组成。 1/2" 中心接线，用于提供足部电击 (0.5 mA)。此外，腔室的每一侧 配备刺激光、音频发生器和两个红外光束源/探测器，用于定位大鼠 将经过避免足部电击训练的大鼠放入室内进行 4 分钟的适应。 随后按 15 秒可变间隔计划（范围 = 7.5 - 22.5 秒）进行 50 次试验。 每次试验均包含 10 秒警告音和刺激光（条件刺激），然后是 10 秒警告音和刺激光（条件刺激）。 第二次电击（无条件刺激），通过大鼠所在一侧的网格地板呈现， 语气和条件性回避反应的存在已被证明是条件性回避反应模型。 可以高度预测抗精神病药物的疗效（Wadenburg 和 Hicks，1999）。 使用 Shuttlebox 测试室（MED Associates，圣奥尔本斯，佛蒙特州）将其分为两个隔间 每个房间的一半地板由十三根直径为 3/16 英寸的不锈钢网格杆组成。 1/2" 中心接线，用于提供足部电击 (0.5 mA)。此外，腔室的每一侧 配备刺激光、音频发生器和两个红外光束源/探测器，用于定位大鼠 将经过避免足部电击训练的大鼠放入室内进行 4 分钟的适应。 随后按 15 秒可变间隔计划（范围 = 7.5 - 22.5 秒）进行 50 次试验。 每次试验均包含 10 秒警告音和刺激光（条件刺激），然后是 10 秒警告音和刺激光（条件刺激）。 第二次电击（无条件刺激），通过大鼠所在一侧的网格地板呈现， 如果动物在试验的最初 10 秒内穿过拱门，则声音和灯光的存在。 如果动物头目通过了，则一和光被终止，并且他的反应被认为是回避反应。 足部冲击启动后，消除光和冲击并考虑响应 逃跑反应，如果在试验间隔（ITI）期间做出反应，动物将受到 0.5 秒的惩罚 电击（0.5 mA）。MED 将计算机与 dEDSTATE NOTATION 关联起来。软件控制测试会话并计算 动物避免电击、逃脱的试验次数 仅表现出稳定表现的动物 （在测试日之前的培训课程中，大约 90% 的回避反应被考虑） “接受过培训”并在测试日进行，培训由至少一名非药物人员维持。 每周进行一次测试，在测试日，通常在测试前 30 分钟给药。 临床上有效的抗精神病药物可减少回避行为并增加逃避行为而不增加 没有反应表明镇静（图 1）。 拮抗阿扑吗啡引起的攀爬和刻板印象（AICS）的多巴胺能刺激。 通过施用多巴胺 D1/D2 激动剂阿扑吗啡来进行神经传递，产生组 刻板行为表明不同的多巴胺能神经通路的刺激被激活。 中脑边缘 DA 受体的阿朴吗啡诱导攀爬行为，同时激活黑质纹状体 DA 受体产生嗅-舔-啃行为的刻板综合症，称为刻板印象（Costall 等，2016）。 al. 1975 和 1980) 抗精神病药物测试 为了测量这些行为，需要服用药物。 在用阿扑吗啡（1 mg/kg sc）攻击前三十分钟，在注射阿扑吗啡后五分钟， 攀爬行为显示在铁丝笼上，并对每只动物的刻板印象进行评分和记录。 在 30 分钟的测试过程中，每 5 分钟重复一次读数，对每只动物的分数进行总计。 对于每组行为，并表示为在媒介物处理的小鼠中观察到的对照值的百分比。 在这个模型中，典型的抗精神病药物在相似的剂量下可以阻止攀爬和刻板印象，而非典型的药物则可以阻止攀爬和刻板印象。 抗精神病药物在不抑制刻板印象的剂量下会产生攀爬行为障碍。 常用的临床前筛查可用于预测治疗阳性症状的疗效 精神分裂症并提供了潜在的运动副作用倾向的指示（图 2）。 PCP/安非他明诱导的运动活动代表了苯丙胺诱导的运动活动模型。 用于治疗多巴胺能亢进，并常用于预测抗精神病药的疗效（Arnt 等人，1997 年）。 在该模型中，安非他明给药诱导多巴胺释放，导致多巴胺增加 运动活动。 抗精神病药通过竞争中脑边缘的突触后多巴胺 D2 受体发挥作用 以类似的方式，NMDA 拮抗剂，例如苯环己哌啶，可以减少诱导的运动反应。 （PCP），刺激运动活动，也被用来评估谷氨酸能的影响 已证明非典型抗精神病药会导致精神分裂症的功能减退（Coyle 等人，2003 年；Lewis 等人，2003 年）。 阻断啮齿动物中 NMDA 拮抗剂诱导的过度活跃（Gleason 和 Shannon，1997）。 抗精神病药物阻断自发性、安非他明诱导和 PGP 诱导的能力 比较小鼠的运动活性和 d-苯丙胺 (3)。 mg/kg，皮下注射)、苯环己哌啶(3mg/kg，皮下注射)或媒介物然后立即将动物置于运动状态。 活动室监测 70 分钟测试，在此期间运动活动数据记录在 使用带有封闭有机玻璃室的 Accuscan 红外光束活动监视器测量室内光线和白噪声 （8 英寸 x 8 英寸） Accuscan Versamax 和 Versadat 软件（俄亥俄州哥伦布市）用于转换红外线。 光束分为 5 分钟内的水平活动计数（图 3）。 声惊吓反应的前脉冲抑制 (PPI) PPI 是感觉运动门控的一种测量方法。 通常用于表征抗精神病药物的 PPI 在精神分裂症患者和患者中降低。 用多巴胺激动剂或 NMDA 受体拮抗剂治疗的啮齿动物（Geyer 等，2001）。 在正常个体中，当受试者暴露于令人震惊的听觉脉冲时，会表现出惊吓减弱 精神分裂症患者对巨大噪音的反应，如果其之前是非令人惊异的听觉音调（前脉冲）。 显示刺激是否为前脉冲的反应 精神分裂症患者显示 a 无论刺激是否是前脉冲，动物都会受到类似的惊吓反应，无论刺激如何。 令人震惊的是，动物是使用测试室的 PPI（SR-LAB 系统、圣地亚哥仪器、 加利福尼亚州圣地亚哥），由一个装有压电加速度计的有机玻璃圆柱体组成，该压电加速度计可检测 安装在圆柱体上方的扬声器提供了动物在圆柱体内的移动背景。 白噪声、声噪声突发和声前脉冲 声脉冲和前脉冲的呈现。 刺激由 SR-LAB 软件和接口系统控制，该系统还可以数字化、校正和 记录来自加速度计的响应，通过平均 100 个 1 毫秒来确定。 从脉冲刺激开始时获取的读数总共包括 60 个。 试验间间隔为 15 秒，适应 64 dB 背景噪声 5 分钟后， 四种试验类型（120 dB 脉冲，或 69、74 或 79 dB 预脉冲与 120 dB 配对 脉冲）以伪随机顺序呈现，药物通常在测试前 30 分钟给药。 随后给予干扰剂 d-苯丙胺（4 mg/kg，皮下注射）和 MK-801（0.15 mg/kg，腹膜内注射）。 前脉冲抑制定义为 100-[（前脉冲试验的惊跳幅度/单独脉冲的惊跳幅度） 试验）x 100]（图 4）。 间接 DA 激动剂安非他明和 NMDA 诱导大鼠的听觉感觉运动门控 拮抗剂MK-801。 僵直症的评估被广泛用作潜在的临床前筛查。 抗精神病药物候选者的锥体外系运动副作用责任（Hoffman 和 Donovan，1995）。 多巴胺 D2 受体阻断水平通常超过 80%，会导致僵住症，其定义为 缺乏运动和维持尴尬的姿势，而典型的抗精神病药物会产生化合物。 在与有效剂量相当的剂量下，非典型抗精神病药物表现出明显的强直性昏厥症状。 在预测抗精神病活性的模型中区分致昏迷剂量和有效剂量。 为了测量抗精神病化合物的致昏迷症潜力，对动物进行监测 服药后 2 小时内每隔 30 分钟出现强直行为。 测量包括在接受治疗的动物的前腿上覆盖一根 1 3/4 英寸高的细水平杆。 记录动物保持这种尴尬姿势的时间（以秒为单位）（60 秒） 保持该姿势被认为是僵住所花费的平均秒数。 计算每个时间点每个剂量的位置（图5）。