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（由适用提供）：抗精神病药是D2多巴胺受体竞争性拮抗剂。长期以来，人们一直猜测，这些药物是越过血脑屏障的弱胺，通过在包括突触蔬菜在内的酸性细胞器中酸性诱捕来积累。以前解决该假设的实验取决于不是精神病药物和不概括D2接收器依赖性神经传递的细胞培养物的嗜酸染料。因此，我们确定了一种临床使用的抗精神病药，可以通过两光子显微镜直接在脑切片中成像。我们的实验表明，这种抗精神病药受到酸性诱捕，包括在中脑多巴胺神经元突触蔬菜中，其中包含天然发射器。此外，我们表明抗精神病药可以通过作用电位从蔬菜中释放出来，该动作电位需要Ca2+或需要多巴胺转运蛋白和囊泡单胺转运蛋白的苯丙胺。在体外的治疗浓度和动物全身给药时，可以看到囊泡的积累和释放。这是第一次证明抗精神病药在突触时用多巴胺释放的抗精神病药。这一发现很吸引人，因为它暗示局部抗精神病药浓度将在多巴胺突触中精确地缩放多巴胺。鉴于现已建立抗精神病药的合成囊泡释放，该提案决定了囊泡抗精神病药释放对由D2受体介导的多巴胺能传播的功能性影响，由质体Nigra和纹状体中的D2受体介导。我们的工作假设是，囊泡药物的有效性将随着重复的解雇而增加，这与行为有关。由于酸性捕获是被动的，并且在不抑制多巴胺的囊泡储存的情况下无法残疾，因此实验利用了囊泡诱捕动力学的显着差异（即，在数小时内发生，然后发生持续时间），并急性地应用了药物vi（即几分钟之内），这是由两片镜像揭示的。纹状体实验集中在电化学测得的多巴胺溢出上，这受D2自感受器的频率抑制作用。黑质NIGRA实验利用多巴胺神经元的斑块夹具主张赞助商并诱发由体育体D2受体诱导的抑制剂电流。总之，这些完整的方法将确定抗精神病药对突触D2受体功能的活性依赖性囊泡释放的影响。