Role of GABA Interneurons in the Rapid Antidepressant Actions of NMDA Receptor Blockade

严重抑郁障碍(MDD)是世界范围内最流行和最令人衰弱的疾病之一，影响 约占人口的17%，造成巨大的个人和经济负担。MDD的影响是 强调了现有药物的局限性，包括低应答率、治疗 耐药患者，以及时间滞后(几周至几个月)。这些数据突显了对更有效的药物的主要需求尚未得到满足 以及见效更快的抗抑郁剂。最近的研究表明，一次小剂量的氯胺酮， 谷氨酸-NMDA受体拮抗剂，产生快速的抗抑郁作用(2小时)，在 抵抗治疗的患者。这种快速的作用是通过一种与典型单胺完全不同的机制 重摄取抑制剂，是过去6年来抑郁症领域最显著的进展之一。 几十年。我们已经报道，氯胺酮引起内侧突触连接的快速增加。 前额叶皮质(MPFC)，靶向并纠正慢性应激和 抑郁症。尽管取得了这些进展，但氯胺酮突触作用的细胞机制，以及 谷氨酸转移率的增加还没有确定。我们假设该事件的初始触发因素 氯胺酮的快速作用是阻断刺激GABA中间神经元紧张性放电的NMDA受体， 从而解除对谷氨酸传递的抑制，增加mPFC中突触的形成。或者， 氯胺酮可直接作用于锥体神经元。此应用程序描述了一种集成的多学科 方法，包括分子、生化、电生理、形态和行为研究 这种去抑制假说。AIM 1将使用细胞类型特异性的NMDA受体敲除GABA MPFC内可见中间神经元亚型和锥体神经元。根据药理学证据， GluN2B亚单位将通过FLOXED-GluN2B shRNA和细胞特异性CRE的病毒表达而成为靶点 重组酶转基因株系。初步结果表明，氯胺酮的快速行为作用是 被GluN2B基因敲除的GABA中间神经元阻断，与去抑制假说一致。目标2将 通过表征NMDA/GluN2B受体对生长抑素(SST)和GluN2B受体的调节来扩展这些研究 通过斑片记录小鼠的小白蛋白(PV)中间神经元亚型以确定细胞基础 这些中间神经元对NMDA和氯胺酮的敏感性存在差异。氯胺酮对抑制作用的影响 可塑性也将被确定，GABA中间神经元亚型在氯胺酮作用中的作用将是 使用细胞特有的光遗传方法进行测试。据报道，压力和抑郁会改变GABA 神经传递对女性的影响比男性更大。AIM 3将使用报告老鼠来确定角色 GABA中间神经元亚型在慢性应激和卵巢类固醇激素影响中的作用，包括分子， 细胞反应和转录反应。对细胞机制的表征，这些细胞机制构成了 氯胺酮、慢性应激和卵巢类固醇将为更安全、快速起效的抗抑郁药物提供新的靶点。

Major depressive disorder (MDD) is one of the most prevalent and debilitating illnesses world wide, affecting ~17 percent of the population and causing enormous personal and economic burden. The impact of MDD is underscored by the limitations of currently available medications, including low response rates, treatment resistant patients, and time-lag (weeks-months). These data highlight a major unmet need for more efficacious and faster-acting antidepressant agents. Recent studies demonstrate that a single low dose of ketamine, a glutamate-NMDA receptor antagonist, produces rapid antidepressant actions (2 hr) that last for up to 7 days in treatment resistant patients. This rapid action, by a mechanism completely different from typical monoamine reuptake inhibitors, represents one of the most significant advances in the field of depression over the past 6 decades. We have reported that ketamine causes a rapid increase of synaptic connections in the medial prefrontal cortex (mPFC), which targets and corrects the synaptic deficits caused by chronic stress and depression. Despite this progress, the cellular mechanisms underlying the synaptic actions of ketamine, and increased glutamate transmission have not been determined. We hypothesize that the initial trigger for the rapid actions of ketamine is blockade of NMDA receptors that stimulate tonic firing of GABA interneurons, resulting in disinhibition of glutamate transmission and increased synapse formation in mPFC. Alternatively, ketamine could act directly on pyramidal neurons. This application describes an integrated multidisciplinary approach, including molecular, biochemical, electrophysiological, morphological, and behavioral studies to test this disinhibition hypothesis. Aim 1 will use cell type specific knockdown of NMDA receptors on GABA interneuron subtypes as well as pyramidal neurons in the mPFC. Based on pharmacological evidence, the GluN2B subunit will be targeted using viral expression of floxed-GluN2B shRNA and cell specific Cre recombinase transgenic lines. Preliminary results indicate that the rapid behavioral actions of ketamine are blocked by GluN2B knockdown on GABA interneurons, consistent with the disinhibition hypothesis. Aim 2 will extend these studies by characterizing NMDA/GluN2B receptor regulation of somatostatin (SST) and parvalbumin (PV) interneuron subtypes by patch recordings in reporter mice to identify the cellular basis for differences in NMDA and ketamine sensitivity of these interneurons. The influence of ketamine on inhibitory plasticity will also be determined, and the role of GABA interneuron subtypes in the actions of ketamine will be tested using cell specific optogenetic approaches. Stress and depression are reported to alter GABA neurotransmission with greater effects in women than men. Aim 3 will use reporter mice to determine the role of GABA interneuron subtypes in the effects of chronic stress and ovarian steroids, including molecular, cellular, and transcriptional responses. Characterization of the cellular mechanisms that underlie the actions of ketamine, chronic stress, and ovarian steroids will provide novel targets for safer, rapid-acting antidepressants.