Unmasking a Role for Cortical Dopamine D4 Receptors in Controlling Circuit Dynamics and Behavior

摘要 前额叶皮层（PFC）是皮质-纹状体-丘脑-皮质（CSTC）回路的一部分，是认知/执行功能和社会过程的关键调节器。这一回路和这些行为领域的中断通常与几种神经精神疾病有关。尽管这些损伤具有使人衰弱的性质，但开发有效管理这种无数缺陷的疗法的挑战仍未得到满足。尽管目前治疗神经精神病症的疗法主要靶向多巴胺受体（DAR），特别是D2 R，但它们也与DAR的其他D2亚类（D3 R，D4 R）相互作用。D4 R特别令人感兴趣，因为它在关键的PFC神经元群体中的表达水平高于D2 R。此外，一些临床上更有效的药物也表现出高D4 R亲和力。 然而，先前开发靶向D4 R的临床有效疗法的尝试已被证明是不成功的。有趣的是，这些努力发生在我们对GPCR偏置信号传导概念的最新进展之前，即通过G蛋白或β-抑制蛋白的信号传导，其引起不同的细胞和生理结果。这一点，再加上工程细胞特异性遗传操作的能力和在真实的时间内以及响应药理学或行为操作而询问大脑神经网络功能的能力，都为我们提供了一个令人兴奋的机会来解决这一重要挑战。我们的合作者最近开发了一种纯D4 R选择性部分激动剂（UCSF 924），我们发现它在药理学和遗传动物模型中表现出显着的抗精神病样特性。该提案的目的是提供概念验证，即D4 R的功能选择性靶向对于减轻在神经精神障碍中观察到的认知/执行功能和社交障碍是必要的。这是在以下具体目标中实现的。目的1：系统测定PFC神经元亚型中D4 R介导的信号传导。目的2：阐明选择性D4 R信号在神经网络调控中的作用。在目标1中，我们将利用我们验证的CRISPR/Cas9交叉系统选择性地和系统性地删除PFC锥体神经元或中间神经元中的D4 R或β-arrestin 2，并评估UCSF 924如何调节选择的行为域。这将在目标2中扩展，我们的合作研究者实验室将利用体内电生理学合奏记录来评估UCSF 924如何参与神经网络并调节行为结果。解决这些目标将测试我们的中心假设，选择性靶向D4 R介导的信号在关键PFC神经元具有治疗潜力。我们提出的工作应该提供一个概念验证的发展战略，以纠正行为领域，目前难治性的现有疗法。

ABSTRACT The prefrontal cortex (PFC), part of the cortico-striato-thalamo-cortical (CSTC) circuit, is a critical regulator of cognition/executive function and social processes. Disruptions to this circuit and these behavioral domains are often associated with several neuropsychiatric disorders. Despite the debilitating nature of these impairments, the challenge to develop therapies that effectively manage this myriad of deficits remains unmet. Although current therapeutics to treat neuropsychiatric conditions primarily target dopamine receptors (DARs), specifically the D2R, they also interact with other D2 subclass of DARs (D3R, D4R). The D4R is of particular interest as it is expressed at higher levels than the D2R in key PFC neuronal populations. Additionally, some of the more clinically effective agents also exhibit high D4R affinity. However, previous attempts to develop clinically effective therapies targeting the D4R have proved unsuccessful. Intriguingly, these efforts occurred prior to recent advances in our understanding of the concepts of GPCR biased signaling i.e. signaling through G proteins or β-arrestins, which elicit different cellular and physiological outcomes. This, coupled with the ability to engineer cell specific genetic manipulations and the capacity to interrogate brain neural network function in real time and in response to pharmacological or behavioral manipulations, all provide us with an exciting opportunity to address this important challenge. Our collaborators have recently developed a purely D4R-selective partial agonist (UCSF924), and we have discovered that it exhibits remarkable antipsychotic-like properties in pharmacological and genetic animal models. The objective of this proposal is to provide proof-of-concept that functionally-selective targeting of the D4R is necessary to mitigate both the cognitive/executive function and social impairments observed in neuropsychiatric disorders. This is accomplished in the following Specific Aims. Aim 1: Systematic determination of D4R-mediated signaling in PFC neuronal subtypes. Aim 2: Elucidating the effect of selective D4R signaling in the regulation of neuronal networks. In Aim 1, we will utilize our validated CRISPR/Cas9 intersectional system to selectively and systematically delete D4R or β-arrestin2 in PFC pyramidal neurons or interneurons, and assess how UCSF924 modulates select behavioral domains. This will be expanded in Aim 2, where our co-investigator’s laboratory will utilize in vivo electrophysiological ensemble recordings to assess how UCSF924 engages neural networks and modulates behavioral outcomes. Addressing these aims will test our central hypothesis that selective targeting of D4R-mediated signaling in key PFC neurons has therapeutic potential. Our proposed work should provide a proof-of-concept for the development of a strategy to correct behavioral domains that are currently refractory to existing therapies.