VGLUT2 Transmission in Prefrontal Cortex and Working Memory

描述（由申请人提供）：精神分裂症是一种发育障碍，假设包括谷氨酸能功能改变、n -甲基- d -天冬氨酸受体（NMDAR）信号异常和内侧前额叶皮层（mPFC）兴奋/抑制（E/I）平衡改变。改善精神分裂症治疗和预防策略的一个关键障碍是缺乏对突触前谷氨酸释放到gaba能中间神经元的作用的理解，通过内在皮质水泡谷氨酸转运蛋白-2 （VGLUT2）突触调节E/I平衡，以及这如何影响含有PV的中间神经元的成熟。我们的目标是通过发展和提供vglut2介导的谷氨酸释放在pv抑制系统功能障碍中的作用和影响的理解，改善治疗和预防精神分裂症的策略。我们的中心假设是VGLUT2向mPFC的新皮质PV+抑制性中间神经元提供谷氨酸，这对兴奋/抑制（E/I）平衡的成熟和工作记忆很重要。我们的目标是：(1)在体内和体外建立功能性VGLUT2突触靶向mPFC的PV+神经元间系统，(2)证明VGLUT2突触的缺失减少了E-I突触的谷氨酸释放并抑制了抑制性传递，以及(3)证明条件VGLUT2敲除（KO）小鼠表现出工作记忆缺陷。我们的预期结果将是：(1)了解皮质内生性VGLUT2突触与E/I平衡相关，因为它们靶向复发性抑制性反馈神经元，并表现出谷氨酸的活性依赖性释放；(2)条件性VGLUT2 KO小鼠中皮质内生性VGLUT2表达的缺失导致PV+中间神经元中GAD67的减少，并抑制抑制性突触向锥体神经元的传递。(3)确认vglut2编码的传导在皮质边缘回路中对工作记忆至关重要。这些结果对治疗和预防精神分裂症策略的影响，我们的结果将提供重要的一步，以治疗性地了解与精神分裂症相关的认知障碍，并了解vglut2介导的释放在pv -神经元间系统成熟和E/I平衡中的作用和影响。目的1将验证皮层内生性VGLUT2突触与E/I平衡相关的假设，因为a)它们靶向PV+ gaba能中间神经元，b)它们表现出谷氨酸的活性依赖性释放，以及c)它们的缺失抑制了mPFC中的抑制性突触传递。我们将利用GAD67gfp+转基因小鼠在PV+中间神经元中选择性表达EGFP进行免疫荧光和电生理研究。目的2将测试发育过程中VGLUT2编码的皮质边缘回路兴奋传递对成人工作记忆至关重要的假设，通过在t迷宫中使用连续延迟交替任务评估工作记忆功能，在发育早期VGLUT2失活的两种小鼠和青春期小鼠中。

DESCRIPTION (provided by applicant): Schizophrenia is a developmental disorder that is hypothesized to include alterations in glutamatergic function, aberrant N-methyl-D-aspartic acid receptor (NMDAR) signaling and altered excitatory/inhibitory (E/I) balance in the medial prefrontal cortex (mPFC). A critical barrier to progress in improving strategies for the treatment and prevention of schizophrenia is a lack of understanding of the role of presynaptic glutamate release onto GABAergic interneurons by intrinsic cortical vesicular glutamate transporter-2 (VGLUT2) synapses in regulating E/I balance and how this may impact the maturation of PV- containing interneurons. Our goal is to improve strategies for the treatment and prevention of schizophrenia by developing and providing an understanding of the role and influence of VGLUT2-mediated glutamate release on the dysfunction of the PV-inhibitory system. Our central hypothesis is that VGLUT2 provides glutamate to neocortical PV+ inhibitory interneurons in the mPFC that is important for the maturation of excitatory/inhibitory (E/I) balance and to working memory. Our objectives are to (1) establish that functional VGLUT2 synapses target the PV+ interneuronal system in mPFC in vivo and in vitro, (2) demonstrate that the loss of VGLUT2 synapses decreases glutamate release at E-I synapses and suppresses inhibitory transmission, and (3) show that conditional VGLUT2 knockout (KO) mice display deficits in working memory. Our expected outcomes will be (1) an understanding that intrinsic cortical VGLUT2 synapses are relevant to E/I balance because they target recurrent inhibitory feedback neurons and exhibit activity-dependent release of glutamate, (2) the knowledge that loss of intrinsic cortical VGLUT2 expression in conditional VGLUT2 KO mice leads to a reduction in GAD67 in PV+ interneurons and suppresses inhibitory synaptic transmission onto pyramidal neurons, and (3) to affirm that VGLUT2-encoded transmission in corticolimbic circuits is critical to working memory. The impact of these results on strategies for treating and preventing schizophrenia that our outcomes will provide is an important step towards therapeutic insight into the cognitive impairment associated with schizophrenia and an understanding of the role and influence of VGLUT2-mediated release on the maturation of the PV-interneuronal system and E/I balance. Aim 1 will test the hypothesis that intrinsic cortical VGLUT2 synapses are relevant to E/I balance because a) they target PV+ GABAergic interneurons, b) they exhibit activity-dependent release of glutamate, and c) their loss suppresses inhibitory synaptic transmission in the mPFC. We will utilize GAD67gfp+ transgenic mice that selectively express EGFP in PV+ interneurons for immunofluorescence and electrophysiologic studies. Aim 2 will test the hypothesis that VGLUT2-encoded excitatory transmission in corticolimbic circuits during development is critical to working memory in adults, by assessing working memory function using the continuous delayed alternation task in a T-maze in two lines of mice where VGLUT2 inactivation occurs early in development and in adolescent mice.