The role of neurotrophins in anesthesia-induced developmental neuroapoptosis

描述（由申请人提供）：这是FIRCA奖申请，用于支持主要在塞尔维亚和黑山进行的研究，贝尔格莱德大学神经生物学系分子神经生物学实验室与弗吉尼亚大学麻醉学系Vesna Jevtovic-Todorovic合作，作为NIH/NICHD母基金#R01 HD 044517的延伸。早产儿、新生儿和幼儿经常暴露于全身麻醉。然而，最近的一些研究结果表明，麻醉药物，通过短暂抑制神经元活动在发育中的哺乳动物的大脑，干扰正常的大脑发育（如突触），从而促进未成熟的神经元的凋亡性细胞死亡。麻醉诱导的未成熟大鼠（本提议的感兴趣物种）的凋亡性神经变性不仅表现为发育中大脑中严重的广泛凋亡性神经变性（例如神经凋亡），而且表现为在生命后期检测到的显著和持续的学习/记忆缺陷。神经营养因子是一类支持神经元存活、分化和突触可塑性的生长因子，在哺乳动物脑突触发生中起重要作用。因此，神经元活动的广泛抑制可以损害由神经营养因子调节的促进存活的信号。为了研究神经营养因子介导的麻醉诱导的发育性神经细胞凋亡的潜在相关性，我们对暴露于临床相关浓度（空气中1.5体积%）的吸入性麻醉剂异氟烷的幼鼠（处于其突触发生的高峰期-7天大）进行了为期2、4或6小时的初步研究。我们发现，异氟烷在两个最脆弱的大脑区域-大脑皮层和丘脑中调节神经营养因子调节的凋亡级联反应的关键步骤。也就是说，激活Akt的蛋白水平，在阻断凋亡级联反应的关键组成部分，在皮质和丘脑显着下降。然而，BDNF（脑源性神经营养因子）蛋白水平在皮质中被差异调节;它们显著增加，其中在丘脑中BDNF水平快速且显著降低。此外，虽然皮质中活化的神经酰胺上调，但丘脑中的神经酰胺活化没有变化：这些结果表明，异氟烷不仅激活Akt调节的细胞凋亡途径，而且在不同脑区以不同的方式激活，最有可能是通过激活Trk依赖性和Trk非依赖性、p75 NTR依赖性神经营养存活途径。在本资助申请中，申请人计划通过对神经营养因子调节的麻醉诱导的神经细胞凋亡的机制进行详细研究，扩展母资助中提出的麻醉诱导的发育细胞凋亡机制的研究。通过扩大我们对麻醉诱导的细胞凋亡级联反应中所有关键步骤的理解，我们希望提高我们预防全身麻醉剂对发育中哺乳动物大脑的潜在有害神经毒性作用的机会。

DESCRIPTION (provided by applicant): This is the FIRCA Award application for the support of the research that will be done primarily in Serbia and Montenegro, in Laboratory for Molecular Neurobiology, the Department of Neurobiology at the University of Belgrade, Belgrade in collaboration with Vesna Jevtovic-Todorovic, Department of Anesthesiology, University of Virginia, as an extension of NIH/NICHD parent grant #R01 HD 044517. Premature infants, neonates and very young children are frequently exposed to general anesthesia. However, some recent findings indicate that anesthetic drugs, by transiently suppressing neuronal activity in the developing mammalian brain, disturb normal brain development (e.g. synaptogenesis) thus promoting apoptotic cell death of the immature neurons. Anesthesia-induced apoptotic neurodegeneration in the immature rats, a species of interest for this proposal, was manifested not only as a severe widespread apoptotic neurodegeneration (e.g. neuroapoptosis) in the developing brain but also as a significant and persistent learning/memory deficits detected latter on in life. The neurotrophins, a family of growth factors, support neuronal survival, differentiation and several forms of synaptic plasticity and therefore play an important role in synaptogenesis of the mammalian brain. Consequently, extensive depression of neuronal activity can impair survival-promoting signals that are regulated by neurotrophins. To investigate the potential relevance of neurotrophin-mediated anesthesia-induced developmental neuroapoptosis we conducted a pilot study on infant rats (at the peak of their synaptogenesis-7 days old) that were exposed to an inhalational anesthetic, isoflurane, at clinically relevant concentration (1.5-vol% in air), for a period of 2, 4 or 6 hrs. We found that isoflurane modulates the key steps in neurotrophin-modulated apoptotic cascade in two most vulnerable brain regions - the cerebral cortex and the thalamus. Namely, the protein levels of activated Akt, a key component in blocking apoptotic cascade, were significantly decreased in both the cortex and the thalamus. However, the BDNF (brain derived neurotrophic factor) protein levels were differentially modulated in the cortex; they were significantly increased, wherein in the thalamus there was a rapid and significant decrease in BDNF levels. In addition, while there was an up-regulation of the activated ceramide in the cortex there were no changes in ceramide activation in the thalamus: These findings suggest that isoflurane not only activates Akt-regulated apoptotic pathways, but does so in differential fashion in different brain regions most likely by activating both Trk dependent and Trk independent, p75NTR dependent neurotrophic survival pathways. In this grant application, the applicants plan to extend the studies of the mechanism of anesthesia-induced developmental apoptosis proposed in the parent grant by performing detailed studies of the mechanism(s) of neurotrophin-modulated anesthesia-induced neuroapoptosis. By expanding our understanding of all the crucial steps in the anesthesia-induced apoptotic cascade we hope to improve our chances of preventing potentially detrimental neurotoxic effects of general anesthetics to the developing mammalian brain.