Epitranscriptomic mechanisms of the antidepressant response to ketamine in human neurons

项目摘要 尽管急性增强单胺功能，长期服用单胺抗抑郁药， 是临床反应所必需的，这表明增强的单胺能信号传导下游的适应 是其治疗功效的核心。一种这样的下游适应是大脑的表观遗传上调- 神经源性营养因子（BDNF）是神经营养过程中的关键蛋白。这是符合神经营养 抑郁症的假说，假定神经元生长不足导致MDD的发病机制， 抑郁症患者死后大脑中海马体积的减少证明了这一点， 神经营养的恢复对抗抑郁药的功效至关重要。除了需要长期治疗外， 传统的抗抑郁药物在很大一部分抑郁症患者中也是无效的， 产生各种各样的不良副作用。另一方面，氯胺酮，一种N-甲基-D-天冬氨酸受体 （NMDAR）拮抗剂和麻醉剂，其单剂量已被证明在不到20分钟内缓解症状。 一个小时和至少一个星期的病人谁没有回应至少两个典型的抗抑郁药。 与传统的抗抑郁药不同，氯胺酮似乎能迅速促进BDNF介导的神经营养， 但优化这种麻醉剂的抗抑郁药的应用和设计新的药物，其形象需要 进一步阐明其靶点及其治疗作用的机制。我们假设 而不是间接地改变神经营养基因，就像慢性施用典型的 作为抗抑郁药，氯胺酮绕过了这一漫长的转录调控过程， 快速上调神经营养蛋白翻译的RNA修饰。应力引起的变化 在MDD中观察到最丰富的RNA修饰N6，2'-O-二甲基腺苷（m6A）水平 患者和m6A脱甲基酶、脂肪量和肥胖相关蛋白的单核苷酸多态性 (FTO)与MDD风险增加有关。此外，抑制糖原合成酶激酶3（GSK-1）， 3）氯胺酮通过NMDAR拮抗作用增加FTO浓度， 通过基质金属蛋白酶9（MMP-9）的去甲基化将促凋亡proBDNF转化为BDNF， 9）。总之，这些发现提出了一个合理的epitranscriptomic机制的快速神经营养 以及克他命的抗抑郁作用因为m6A甲基化在物种之间是不同的， 相关基因的位点，如MMP-9，在小鼠和人之间不同，我们将使用人诱导的多能 干细胞（hiPSC）衍生的皮质多巴胺能神经元，以描述氯胺酮对 epitranscriptome，m6A机制，以及神经元的结构和功能。成功完成这些 目的是阐明氯胺酮快速和强大的抗抑郁作用的机制， 从而能够鉴定和优化新型抗抑郁药。

PROJECT SUMMARY Despite acutely enhancing monoamine function, chronic administration of monoamine antidepressants is required for clinical response, suggesting that adaptations downstream of enhanced monoaminergic signaling are central to their therapeutic efficacy. One such downstream adaption is the epigenetic upregulation of brain- derived neurotrophic factor (BDNF), a key protein in the neurotrophic process. This is in line with the neurotrophic hypothesis of depression, which posits that insufficient neuronal growth contributes to the pathogenesis of MDD, as evidenced by reduced hippocampal volume in the postmortem brains of depressed patients, and thus the restoration of neurotrophy is central to the efficacy of antidepressants. In addition to requiring chronic treatment, traditional antidepressants drugs are also ineffective in a significant proportion of depressed patients and produce a wide variety of undesirable side effects. On the other hand, ketamine, a N-methyl-D-aspartate receptor (NMDAR) antagonist and anesthetic, a single dosage of which has been shown to relieve symptoms in less than an hour and for at least one week in patients who failed to response to at least two typical antidepressants. Unlike traditional antidepressants, ketamine appears to promote BDNF-mediated neurotrophy rapidly and directly but optimizing this anesthetic for antidepressant application and designing novel drugs its image requires further elucidation of its targets and the mechanisms underlying its therapeutic effects. We hypothesize that rather than rather than indirectly altering neurotrophic genes, as with chronic administration of typical antidepressants, ketamine bypasses this prolonged process of transcriptional regulation and instead induces RNA modifications that rapidly upregulates the translation of neurotrophic proteins. stress-induced changes in levels of N6,2’-O-dimethyladenosine (m6A), the most abundant RNA modification, have been observed in MDD patients and single-nucleotide polymorphisms in the m6A demethylase, fat mass and obesity associated protein (FTO), are associated with increased risk of MDD. Furthermore, inhibition of glycogen synthase kinase 3 (GSK- 3), which ketamine does through NMDAR antagonism, increases FTO concentrations which promotes conversion of the pro-apoptotic proBDNF to BDNF through demethylation of matrix metalloprotease 9 (MMP- 9). Taken together, these findings present a plausible epitranscriptomic mechanism for the rapid neurotrophic and antidepressant effects of ketamine. Because m6A methylation varies between species, and the methylation sites of relevant genes, such as MMP-9, differ between mice and humans, we will use human induced pluripotent stem cell (hiPSC)-derived cortical glutamatergic neurons to delineate the effects of ketamine on the epitranscriptome, m6A machinery, as well as neuronal structure and function. The successful completion of these aims will elucidate the mechanisms underlying the rapid and robust antidepressant effects of ketamine thus enabling the identification and optimization of novel antidepressants.