The role of neurotrophins in anesthesia-induced developmental neuroapoptosis

描述(由申请人提供)：这是FIRCA奖的研究支持申请，将主要在塞尔维亚和黑山，在分子神经生物学实验室，贝尔格莱德大学神经生物学系与Vesna Jevtovic-Todorovic，弗吉尼亚大学麻醉学系，作为美国国立卫生研究院/美国国立卫生研究院/美国国立卫生研究院家长补助金#R01 HD 044517的延伸。早产儿、新生儿和幼儿经常暴露在全身麻醉中。然而，最近的一些发现表明，麻醉药物通过短暂地抑制发育中的哺乳动物大脑中的神经元活动，干扰正常的大脑发育(例如突触发生)，从而促进未成熟神经元的凋亡细胞死亡。麻醉诱导幼年大鼠神经细胞的凋亡性变性，不仅表现为发育中的大脑中广泛存在的严重的神经细胞凋亡性变性(如神经细胞凋亡)，而且还表现为在生命后期检测到的显著且持续的学习/记忆缺陷。神经营养因子是一个生长因子家族，支持神经元的存活、分化和多种形式的突触可塑性，因此在哺乳动物大脑的突触发生中起着重要的作用。因此，神经元活动的广泛抑制可能会削弱由神经营养因子调节的促进生存的信号。为了探讨神经营养素介导的麻醉诱导发育性神经细胞凋亡的潜在相关性，我们对新生大鼠(在其突触发生的高峰期-7日龄)进行了一项初步研究，这些大鼠吸入临床相关浓度的异氟醚(空气中1.5%-VOL%)，持续2、4或6小时。我们发现，在两个最脆弱的大脑区域-大脑皮层和丘脑-异氟烷调节神经营养素调节的细胞凋亡级联中的关键步骤。也就是说，大脑皮质和丘脑中激活的Akt蛋白水平显著降低，Akt是阻止细胞凋亡级联反应的关键成分。然而，脑源性神经营养因子(BDNF)的蛋白水平在皮质中受到不同程度的调节；它们显著增加，其中在丘脑中BDNF水平迅速而显著下降。此外，虽然皮质中激活的神经酰胺上调，但丘脑中神经酰胺的激活没有变化：这些发现表明，异氟醚不仅激活Akt调节的凋亡通路，而且在不同的脑区以不同的方式激活，很可能是通过激活Trk依赖和Trk非依赖的、p75NTR依赖的神经营养生存通路。在本次拨款申请中，申请者计划通过对神经营养素调节的麻醉诱导神经细胞凋亡的机制(S)进行详细研究，来扩展父拨款中提出的麻醉诱导发育凋亡机制的研究。通过扩大我们对麻醉诱导的细胞凋亡级联过程中所有关键步骤的理解，我们希望提高我们预防全身麻醉药对发育中的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.