Targeting glutamate carboxypeptidase in perinatal brain injury

项目摘要/摘要 尽管在新生儿护理和实施治疗性低温方面取得了重大进展，但足月和 早产儿脑病幸存者患认知和学习障碍的风险很高 即使在没有功能性运动缺陷的情况下。最近的研究表明，小脑功能障碍与 这些儿童中出现的长期学习障碍。浦肯野细胞发育受损与 小脑联想学习障碍。小胶质细胞已被证明在 小脑皮质初级输出神经元浦肯野细胞的发育。宫内炎症 导致小胶质细胞激活，导致浦肯野细胞发育不良和小脑学习 赤字。小脑中激活的小胶质细胞过表达谷氨酸羧肽酶II(GCPII)， 它能水解丰富的神经肽N-乙酰天冬氨酸(NAAG)，NAAG是一种 代谢性谷氨酸受体，mGluR3)为N-乙酰天冬氨酸和谷氨酸。NAAG水平下降 与认知功能受损有关。这项研究建议特异性靶向小胶质细胞GCPII酶 树枝状大分子偶联到纳摩尔有效但脑穿透能力差的GCPII抑制剂2PMPA(2- (磷酸亚甲基)戊烷-1，5-二酸)(D-2PMPA)。中心假设是正常的小胶质细胞 功能和动力学在小脑发育中起关键作用，抑制小脑GCPII表达上调 D-2PMPA激活的小胶质细胞将导致小脑NAAG的增加，使正常的浦肯野细胞成为可能 小剂量内毒素对幼兔小脑学习记忆能力的影响 在子宫里。这将通过以下方式进行测试：(1)评估低剂量宫内内毒素暴露对 浦肯野细胞发育、小脑小胶质反应和小脑学习(用经典眨眼测试 条件反射)在母体炎症诱导脑损伤的兔模型中。(2)评价D-半乳糖的疗效。 2PMPA介导的小胶质细胞GCPII抑制6~8周浦肯野细胞发育和小脑学习 宫内炎症对兔年龄的影响及(3)小脑机制的研究 NAAG介导的D-2PMPA对mGluR3的神经保护作用这项研究将提供一个 更好地了解围产期轻度母婴炎症是如何导致 长期的小脑学习障碍。这项拟议的工作具有创新性，因为它使用了一种新颖而有力的D- 2PMPA偶联靶向小胶质细胞GCPII，用于解决认知和学习障碍并使正常 围产期脑损伤兔模型的小脑发育。这一多PI提案将带来 在围产期/新生儿脑损伤、神经药理学/GCP2和纳米医学方面的协同专业知识，以及 临床翻译来实现这些目标。这项工作可以导致开发新的治疗方法 解决新生儿脑损伤常见的学习障碍问题。

Project summary/abstract Despite significant advances in neonatal care and implementation of therapeutic hypothermia, term and preterm survivors of neonatal encephalopathy are at high risk for developing cognitive and learning deficits even in the absence of functional motor deficits. Recent studies have implicated cerebellar dysfunction with the long-term learning disorders seen in these children. Impaired Purkinje cell development has been linked to impairment in cerebellar associative learning. Microglia have been shown to play a major role in the development of Purkinje cells, the primary output neurons of the cerebellar cortex. Intrauterine inflammation leads to microglial activation that results in maldevelopment of the Purkinje cells and cerebellar learning deficits. Activated microglia in the cerebellum overexpress the enzyme glutamate carboxypeptidase II (GCPII), which hydrolyzes the abundant neuropeptide N-acetylaspartylglutamate (NAAG, a specific agonist of the metabotropic glutamate receptor, mGluR3) to N-acetyl-aspartate and glutamate. Reduced NAAG levels have been linked to impaired cognition. This study proposes to specifically target microglial GCPII enzyme using dendrimer conjugated to the nanomolar potent but poorly brain penetrating GCPII inhibitor 2PMPA (2- (phosphonomethyl) pentane-1,5-dioic acid) (D-2PMPA). The central hypothesis is that normal microglial function and dynamics play a critical role in cerebellar development, and inhibiting upregulated GCPII in activated microglia with D-2PMPA will lead to increased NAAG in the cerebellum enabling normal Purkinje cell development and improving cerebellar learning and memory in juvenile rabbits exposed to low dose endotoxin in utero. This will be tested by (1) Evaluating the effects of low dose intrauterine endotoxin exposure on Purkinje cell development, cerebellar microglial response and cerebellar learning (tested by classical eyeblink conditioning reflex) in a rabbit model of maternal inflammation induced brain injury. (2) Evaluate efficacy of D- 2PMPA mediated microglial GCPII inhibition on Purkinje cell development and cerebellar learning at 6-8 weeks of age in rabbits exposed to intrauterine inflammation and (3) Determine mechanisms of cerebellar neuroprotection by D-2PMPA mediated by NAAG induced activation of mGluR3. This study will provide a better understanding of how low-grade maternal-fetal inflammation in the perinatal period can lead to cerebellar learning deficits long term. The proposed work is innovative because it uses a novel and potent D- 2PMPA conjugate to target microglial GCPII for addressing cognitive and learning deficits and enable normal development of the cerebellum in a rabbit model of perinatal brain injury. This Multi-PI proposal will bring synergistic expertise in perinatal/neonatal brain injury, neuropharmacology/GCP2 and nanomedicine, and clinical translation to accomplish these aims. This work can lead to the development of novel therapies to address learning deficits commonly seen with neonatal brain injury.