Project II- Impact of Hypoxia-Ischemia and/or inflammation on Metabolism in Cerebellum

新生儿缺氧缺血(H/I)导致皮质、皮质下和海马区能量迅速衰竭 由于缺乏氧气和葡萄糖输送到大脑。这之后是期间的瞬时正规化 再灌注和随后的继发性能量衰竭(H/I后约2-6小时)，最终导致脑损伤，以及 神经发育不良。H/I前发生的炎症可加重损伤。剧烈的变化 新生儿缺氧缺氧性脑损伤后新陈代谢和长期代谢紊乱 支持对正常发展至关重要的进程。我们将使用一个描述良好的围产期H/I模型 在足月(出生后(PN)第10天的大鼠幼鼠)有和没有先前的炎症。缺氧缺氧性脑损伤对小脑的影响 尚未被仔细研究，尽管快速发展的小脑特别容易 炎症和围产期损伤。我们将结合能量的高场1H-核磁共振和13C-核磁共振波谱 基质辅助激光解吸/电离-质谱法用于代谢和神经递质合成 成像(MALDI-MSI)测定小脑特定层中的代谢物、脂肪和蛋白质。我们的 这项研究将首次确定通过星形胶质细胞和神经元特异性通路进行的新陈代谢变化。 和小脑谷氨酸和GABA的合成。分子和代谢改变及长时间脑缺血 长期的功能结果将决定低温和药物治疗的潜力。我们 假设新生儿缺氧缺血导致小脑能量衰竭，从而导致脑组织功能受损。 脑细胞包括能量代谢和神经递质合成的急性和长期改变。我们 炎症和缺氧/缺氧的综合作用加剧小脑损伤和调节失调的假说 男性和女性大脑的新陈代谢。我们的具体目标是检验这些假设：1.确定H/I的效果 PN10对雌雄幼鼠小脑能量代谢的影响。 2.测定浦肯野的神经递质、代谢物、血脂和蛋白质的变化，分子和 雄性和雌性幼鼠H/I后小脑颗粒细胞层、深核和白质 没有先前的炎症。3.测定亚低温、胆碱和头孢曲松单独应用的保护作用。 而这些化合物与低温相结合，可以对抗神经元和神经胶质代谢的变化以及 浦肯野细胞层、分子细胞层、颗粒细胞层、深核和髓鞘的特异性分子变化。小说， 有关神经元和星形胶质细胞损伤和改变的时间和靶点的临床相关信息 脑内特定代谢途径及代谢物、核苷酸、膜和信号的变化 小脑层和深核团中的脂类和蛋白质将使用1H-1的独特组合获得。 核磁共振和13C-核磁共振结合MALDI-MSI和LC/MS-MS分析H/I后的变化 没有炎症，没有炎症，没有单独的炎症，对神经保护治疗也没有反应。

Neonatal hypoxia-ischemia (H/I) leads to rapid energy failure in cortical, subcortical and hippocampal regions due to lack of oxygen and glucose delivery to brain. This is followed by a transient normalization during reperfusion and a later secondary energy failure (~2-6 hours after H/I) that ultimately leads to brain injury, and poor neurodevelopmental outcome. Inflammation occurring prior to H/I can exacerbate injury. Acute alterations in metabolism and prolonged metabolic dysregulation after neonatal H/I leave the brain vulnerable and unable to support processes essential for normal development. We will use a well-characterized model of perinatal H/I at term (postnatal (PN) day 10 rat pup) with and without prior inflammation. The effects of H/I on cerebellum have not been carefully explored even though the rapidly developing cerebellum is particularly vulnerable to inflammation and perinatal injury. We will combine high field 1H-NMR and 13C-NMR spectroscopy of energy metabolism and neurotransmitter synthesis with matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) determination of metabolites, lipids, proteins in specific layers of the cerebellum. Our studies will be the first determination of alterations in metabolism via astrocyte and neuron specific pathways and synthesis of glutamate and GABA in cerebellum after H/I. Molecular and metabolic alterations and long- term functional outcomes will determine the therapeutic potential of hypothermia and drug therapy. We hypothesize that neonatal H/I leads to energy failure in the cerebellum that contributes to impaired function of brain cells including acute and long-term alterations in energy metabolism and neurotransmitter synthesis. We hypothesize that the combined effect of inflammation and H/I exacerbates cerebellar damage and dysregulated metabolism in male and female brain. Our Specific Aims test these hypotheses: 1. Determine the effect of H/I at PN10 with and without prior inflammation on energy metabolism in cerebellum of male and female rat pups. 2. Determine the alterations in neurotransmitters, metabolites, lipids and proteins in Purkinje, molecular and granule cell layers, deep nuclei and white matter of the cerebellum of male and female pups after H/I with and without prior inflammation. 3. Determine the protective effect of hypothermia, choline and ceftriaxone alone, and these compounds in combination with hypothermia against changes in neuronal and glial metabolism and specific molecular changes in the Purkinje, molecular and granule cell layers, deep nuclei and myelin. Novel, clinically relevant information about the timing and targets of injury and alterations in neuron and astrocyte specific metabolic pathways in brain and alterations of metabolites, nucleotides, membrane and signaling lipids, and proteins in cerebellar layers and deep nuclei will be obtained using the unique combination of 1H- NMR and 13C-NMR in conjunction with MALDI-MSI and LC/MS-MS for profiling the changes after H/I with and without inflammation, inflammation alone and response to neuroprotective therapy.