PICK1 knockout mice: role for D-serine in neonatal forebrains

描述(申请人提供)：与C-Kinase相互作用的蛋白(PICK1)是一种多功能支架蛋白，与神经元和神经胶质细胞中的许多蛋白相互作用。我们最近报道了PICK1直接与D-丝氨酸合成酶，丝氨酸消旋酶(SR)结合。PICK1基因敲除表达降低了细胞培养中SR的D-丝氨酸水平。我们发表了在PICK1基因敲除(PICK1 KO)中观察到新生儿前脑中D-丝氨酸水平降低的结果，这与细胞生物学的观察结果一致。最近的几项研究表明，D-丝氨酸在与大脑皮层回路相关的精神障碍中发挥了作用，包括精神分裂症。然而，据我们所知，PICK1的作用在很大程度上是在突触可塑性的背景下进行的，主要是通过使用小脑和海马区的切片来研究的。因此，在这项拟议的研究中，我们将从行为分析和电生理方法方面研究PICK1基因敲除小鼠，重点关注该蛋白对D-丝氨酸处置和脑功能的影响，特别是与皮质电路相关的功能。在我们的初步研究中，我们获得了(1)新生期间前脑中D-丝氨酸水平的下降，但在成年期间不会；(2)成年期的几种行为缺陷，包括空间工作记忆和前脉冲抑制；以及(3)成年期PICK1 KO小鼠皮质锥体神经元对NMDA的反应降低。根据初步数据，我们的总体假设是，新生儿D-丝氨酸缺乏，可能与谷氨酸功能异常有关，影响前额叶皮质回路的成熟，并导致成年后锥体神经元和中间神经元突触和NMDA依赖性反应的缺陷，而此时D-丝氨酸水平正常。在目标1中，我们将检测发育过程中前脑中D-丝氨酸的水平，特别是在PICK1 KO小鼠的新生阶段。在目标2中，我们将通过一组行为分析，特别是那些用于测量皮质功能的行为分析，以及电生理方法来描述成年PICK1 KO小鼠。在目标3中，我们将使PICK1 KO小鼠的D-丝氨酸水平正常化，如有必要，在新生期给予D-丝氨酸和DAAO抑制剂。然后，我们将检查新生儿期的D-丝氨酸治疗是否影响成年PICK1 KO小鼠可能的异常行为和生理表型。通过这些实验，我们将研究PICK1与D-丝氨酸在新生儿前脑中的作用，希望从这项研究中获得的信息将成为理解精神障碍和脑发育的重要基础。 公共卫生相关性：PICK1基因敲除小鼠最近的证据表明，PICK1可能通过调节新生儿前脑中的D-丝氨酸代谢来调节谷氨酸的神经传递。新生阶段神经元回路的形成对大脑的正常发育和成人大脑的功能至关重要。我们将通过对PICK1基因敲除小鼠的研究，研究PICK1与D-丝氨酸在新生儿前脑中的作用，希望从这项研究中获得的信息将为理解精神障碍和脑发育提供重要基础。

DESCRIPTION (provided by applicant): Protein Interacting with C-Kinase (PICK1) is a multifunctional scaffold protein that interacts with many proteins in neurons and glial cells. We recently reported that PICK1 binds directly to the D-serine synthesizing enzyme, serine racemase (SR). Knockdown expression of PICK1 decreased levels of D-serine from SR in cell cultures. We published that decreased levels of D-serine were observed in neonatal forebrains in PICK1 knockout (PICK1 KO), consistent with the observation from cell biology. Several recent studies have suggested roles for D-serine in mental disturbances associated with cortical circuitry, including schizophrenia. To our knowledge, however, roles for PICK1 have largely been studied in the context of synaptic plasticity, mainly by using slices from the cerebellum and hippocampus. Therefore, in this proposed study, we will examine PICK1 knockout mice in behavioral assays and electrophysiological approaches and focus on the impact of the protein on D- serine disposition and brain functions, especially those associated with cortical circuitry. In our preliminary studies, we obtained (1) decreased levels of D-serine in the forebrain during the neonatal period, but not in the adulthood; (2) several behavioral deficits in adulthood, which includes those in spatial working memory and prepulse inhibition; and (3) decreased response of the cortical pyramidal neurons to NMDA in adulthood, in PICK1 KO mice. Based on preliminary data, our overall hypothesis is that neonatal deficits in D-serine, which is likely to associate with abnormal glutamate function, affect the maturation of prefrontal cortical circuits and result in deficits in synaptic and NMDA-dependent responses in pyramidal neurons and interneurons in adulthood, whereas at this time the levels of D-serine are normal. In Aim 1, we will examine levels of D-serine in the forebrain during development, especially in neonatal stages in PICK1 KO mice. In Aim 2, we will characterize adult PICK1 KO mice by a set of behavioral assays, in particular those for measuring cortical functions, and also by electrophysiological approaches. In Aim 3, we will normalize the levels of D-serine in PICK1 KO mice by administering D-serine, if necessary, combined with a DAAO inhibitor during the neonatal stage. We will then examine whether the D-serine treatment during neonatal periods influences possible abnormal behaviors and physiological phenotypes in adult PICK1 KO mice. Through these experiments, we will study a role for PICK1 in the neonatal forebrain in conjunction with D-serine, hoping that the information obtained from this study will be an important basis in understanding mental disorders and brain development. PUBLIC HEALTH RELEVANCE: PICK1 knockout mice Recent evidence has suggested that PICK1 may regulate glutamate neurotransmission via regulating D-serine metabolism in the neonatal forebrain. Formation of neuronal circuitry during neonatal stages is crucial for proper brain development and functions of adult brain. We will study role for PICK1 in the neonatal forebrain in conjunction with D- serine by characterizing PICK1 knockout mice, hoping that the information obtained from this study will be an important basis in understanding mental disorders and brain development.