Neonatal Anesthetic Neurotoxicity

描述（由申请人提供）：全身麻醉的前提是麻醉剂产生无毒和可逆的无意识状态。最近的数据表明，新生动物暴露于麻醉剂会引发广泛的神经变性，导致成年期持续记忆和学习异常。麻醉神经毒性引起了人们对全身麻醉对人类胎儿、新生儿和婴儿潜在不良影响的关注。虽然确切的机制尚不清楚，但毒性发生在突触形成过程中，本质上是细胞凋亡[1,2]。在发育中的大脑中，与适当靶标的突触连接对于神经元的存活至关重要，因为神经元依赖于其靶标的营养支持[13-15]。突触连接缺失导致细胞凋亡。神经营养因子BDNF有助于神经元存活和突触发生，以及突触[16]的巩固和成熟。然而，BDNF也可以导致神经元凋亡[17,18]。BDNF作为前分子（proBDNF）从突触囊泡分泌，并在突触间隙中被纤溶蛋白裂解生成成熟的BDNF (mBDNF)[19]。纤溶酶原，纤溶酶的前体，被tPA（一种从突触前囊泡释放的蛋白酶）水解裂解。mBDNF通过TrkB受体发出信号，促进神经元存活和突触发生。在缺乏tPA的情况下，proBDNF不断裂，并优先通过p75NTR受体发出信号，导致突触发生减少，树突棘退缩和神经元凋亡[20]。我们实验室的初步数据表明，挥发性麻醉剂异氟醚可以减少tPA的释放；这导致proBDNF通过p75NTR优先信号传导，导致JNK激活，神经元凋亡和树突棘减少。重要的是，异氟醚诱导的神经元死亡可以通过外源性重组tPA减轻；tPA通过TrkB受体恢复信号，导致Akt激活，增加树突棘形成和神经元存活。基于这些数据，我们提出了一种假设，即麻醉神经毒性是通过p75NTR降低神经元活性、减少突触tPA释放、增强proBDNF信号、减少树突棘形成和神经元凋亡的功能。为了验证这一假设，我们提出将在三个具体目标下进行的研究。首先，麻醉药的毒性将在体内和体外通过评估麻醉诱导的细胞凋亡，抑制突触发生和减少树突棘来表征。还将评估突触和脊柱的恢复程度。此后，BDNF-TrkB信号和tpa -纤溶酶系统在上述毒性中的作用将被确定。最后，将评估新生儿期麻醉暴露对成年期认知功能的影响。总的来说，拟议研究的积极结果将为麻醉剂损伤发育中的大脑的机制以及减轻这种毒性的具体机制提供新的见解。重要的是，我们已经开发了一种新的治疗方法来预防异氟烷神经毒性。因此，所提出的工作具有明确的翻译应用。

DESCRIPTION (provided by applicant): A premise of general anesthesia is that anesthetics produce a non-toxic and reversible state of unconsciousness. Recent data indicate that exposure of neonatal animals to anesthetics triggers widespread neurodegeneration leading to persistent memory and learning abnormalities during adulthood [1]. Anesthetic neurotoxicity has raised concerns about the potential adverse impact of general anesthesia in the human fetus, neonate and infant. Although the precise mechanism is not clear, the toxicity occurs during synaptogenesis and is apoptotic in nature [1,2]. In the developing brain, synaptic connections with appropriate targets are essential for neuronal survival as neurons are dependent upon trophic support from their targets [13-15]. Loss of synaptic connection leads to apoptosis. The neurotrophin BDNF contributes to neuronal survival and synaptogenesis, and to the consolidation and maturation of synapses [16]. BDNF can, however, also result in neuronal apoptosis [17,18]. BDNF is secreted from synaptic vesicles as a pro molecule (proBDNF) and undergoes proteolytic cleavage in the synaptic cleft by plasmin to generate mature BDNF (mBDNF) [19]. Plasminogen, the precursor to plasmin, is proteolytically cleaved by tPA, a protease released from pre-synaptic vesicles. The mBDNF signals through TrkB receptors to promote neuronal survival and synaptogenesis. In the absence of tPA, proBDNF is uncleaved and preferentially signals through p75NTR receptors, resulting in reduced synaptogenesis, withdrawal of dendritic spines and neuronal apoptosis [20]. Preliminary data from our laboratory indicate that the volatile anesthetic, isoflurane, reduces tPA release; this results in the preferential signaling of proBDNF through p75NTR, leading to JNK activation, neuronal apoptosis and reduction in dendritic spines. Importantly, isoflurane-induced neuronal death can be mitigated by administration of exogenous recombinant tPA; tPA restores signaling through TrkB receptors, leading to the activation of Akt, increased dendritic spine formation and neuronal survival. Based on these data, we advance the hypothesis that anesthetic neurotoxicity is a function of reduced neuronal activity, decreased synaptic tPA release, enhanced proBDNF signaling, reduced dendritic spine formation and neuronal apoptosis via p75NTR. To test this hypothesis, we propose studies that will be conducted within three specific aims. First, the toxicity of anesthetics will be characterized in vivo and in vitro with the evaluation of anesthetic induced apoptosis, suppression of synaptogenesis and reduction in dendritic spines. The extent of recovery of synapses and spines will also be evaluated. Thereafter, the role of BDNF-TrkB signaling and of the tPA-plasmin system on the aforementioned mentioned toxicity will be determined. Finally, the effects of anesthetic exposure during the neonatal period on cognitive function during adulthood will be evaluated. Collectively, the positive outcome of the proposed studies will provide novel insights into the mechanisms by which anesthetic agents injure the developing brain and into specific mechanisms by which this toxicity can be mitigated. Importantly, we have developed a novel therapeutic approach to the prevention of isoflurane neurotoxicity. As such, the proposed work has clear translational application. PUBLIC HEALTH RELEVANCE: Recent data have indicated that anesthetics can produce widespread neurodegeneration in the developing brain and this leads to cognitive dysfunction during adulthood. This has provoked concern about the possibility that anesthesia in neonates might lead to brain injury. The proposed research will attempt to characterize the mechanisms by which anesthetics injure the developing brain and the means by which this toxicity can be prevented or treated.