Antipsychotic Drug Vesicular Release at Dopamine Synapses

多巴胺突触的抗精神病药物囊泡释放

基本信息

批准号：
9273637
负责人：
EDWIN S LEVITAN
金额：
$ 19.38万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-05-15 至 2018-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9273637
关键词：
Action Potentials Acute Address Affect Agonist Amination Amines Amphetamines Animals Antidepressive Agents Antipsychotic Agents Attenuated Autoreceptors Bathing Behavior Biological Assay Blood - brain barrier anatomy Brain Cell Culture Techniques Charge Clinical Corpus striatum structure Dopamine Dopamine D2 Receptor Dopamine Receptor Dorsal Dose Drug Design Drug effect disorder Dyes Electrodes Employee Strikes Exocytosis Frequencies Hippocampus (Brain)Hour Image In Vitro Kinetics Lead LysoTracker Measures Mediating Membrane Mental Depression Microscopy Midbrain structure Mus Neurons Organelles Pattern Periodicity Pharmaceutical Preparations Pharmacology Physiological Predisposition Proteins Protocols documentation Quinpirole Rattus Slice Substantia nigra structure Synapses Synaptic Cleft Synaptic Vesicles Testing Therapeutic Time Uncertainty Vesicle Wit base carbon fiber design dopamine transporter dopaminergic neuron experimental study extracellular in vivo monoamine nanomolar neurotransmission novel therapeutics patch clamp postsynaptic protonation public health relevance receptor function response transmission process two-photon uptake vesicular monoamine transporter vesicular release

项目摘要

DESCRIPTION (provided by applicant): Antipsychotic drugs are D2 dopamine receptor competitive antagonists. It has long been speculated that these drugs, which are weak base amines that cross the blood brain barrier, accumulate by acidic trapping in acidic organelles including synaptic vesicles. Previous experiments addressing this hypothesis relied on acidophilic dyes that are not psychiatric drugs and cell cultures that do not recapitulate D2 receptor-dependent neurotransmission. Therefore, we identified a clinically used antipsychotic drug that can be directly imaged in the brain slice by two-photon microscopy. Our experiments established that this antipsychotic is subject to acidic trapping, including in midbrain dopamine neuron synaptic vesicles, which contain the native transmitter. Furthermore, we showed that the antipsychotic drug can be released from vesicles by action potentials, which requires Ca2+, or an amphetamine, which requires the dopamine transporter and the vesicular monoamine transporter. Vesicular accumulation and release were seen at a therapeutic concentration in vitro and with systemic administration in animals. Thus, these results demonstrated for the first time that an antipsychotic drug is subject to vesicular release with dopamine at synapses. This finding is intriguing because it implies that local antipsychotic drug concentration will scale wit dopamine exactly where and when dopamine synapses are active. Given that synaptic vesicular release of an antipsychotic drug has now been established, this proposal determines the functional impact of vesicular antipsychotic drug release on dopaminergic transmission mediated by D2 receptors in the substantia nigra and the striatum. Our working hypothesis is that efficacy of the vesicular drug will increase with repetitive firing, which is relevant for behavior. As acidic trapping is passive and cannot be disrupted without inhibiting vesicular storage of dopamine, experiments make use of the striking difference in kinetics of vesicular trapping (i.e., it occurs over hours and then is long lasting) and acute application of the drug vi the bath (i.e., in minutes) that was revealed by two-photon microscopy. Striatum experiments are focused on electrochemically measured dopamine overflow, which is subject to frequency dependent inhibition by D2 autoreceptors. Substantia nigra experiments utilize patch clamping of dopamine neurons to assay spontaneous miniature and evoked inhibitory currents that are induced by somatodendritic D2 receptors. Together, these complementary approaches will ascertain the impact of activity-dependent vesicular release of antipsychotic drug on synaptic D2 receptor function.

描述（申请人提供）：抗精神病药物是 D2 多巴胺受体竞争性拮抗剂，长期以来人们推测这些药物是穿过血脑屏障的弱碱胺，通过酸性捕获在包括突触小泡在内的酸性细胞器中积累。解决这一假设依赖于非精神药物的嗜酸染料和不重现 D2 受体依赖性神经传递的细胞培养物，因此，我们确定了一种临床使用的药物。可以通过双光子显微镜在脑切片中直接成像的抗精神病药物，我们的实验表明，这种抗精神病药物会受到酸性捕获，包括在含有天然递质的中脑多巴胺神经元突触小泡中。药物可以通过动作电位从囊泡中释放，这需要 Ca2+，或安非他明，这需要多巴胺转运蛋白和囊泡单胺在体外和在动物体内全身给药时观察到囊泡积聚和释放，因此，这些结果首次证明抗精神病药物在突触处会与多巴胺一起发生囊泡释放。意味着局部抗精神病药物浓度将在多巴胺突触活跃的地方和时间与多巴胺成比例。鉴于现已确定抗精神病药物的突触小泡释放，该提议决定了功能。囊泡抗精神病药物释放对黑质和纹状体中 D2 受体介导的多巴胺能传递的影响我们的工作假设是，囊泡药物的功效会随着重复发射而增加，这与行为相关，因为酸性捕获是被动的，不能。被破坏而不抑制多巴胺的囊泡储存，实验利用了囊泡捕获动力学的显着差异（即，它发生在双光子显微镜揭示了纹状体实验的重点是电化学测量的多巴胺溢出，该溢出受到 D2 的频率依赖性抑制。黑质实验利用多巴胺神经元的膜片钳来测定由体树突 D2 受体诱导的自发微型电流和诱发抑制电流。确定抗精神病药物活性依赖性囊泡释放对突触 D2 受体功能的影响。