Presynaptic Mechanisms of Lead Neurotoxicity

描述（由申请人提供）：生命早期暴露于铅（Pb2+）的严重神经系统发育风险是众所周知的。现在越来越清楚的是，低浓度的Pb2+会对儿童产生严重的有害影响，这就更加需要了解Pb2+对大脑作用的性质和程度。我们的实验室最近发现，在体外暴露于极低水平的Pb2+会对培养的海马神经元突触前递质释放产生长期损伤，并且这些作用类似于在缺乏营养因子脑源性神经营养因子（BDNF）的动物中观察到的情况。我们建议在发育期间暴露于低水平Pb2+的大鼠海马切片中进行研究，以利用1)最先进的双光子成像方法来评估完整突触中突触前Ca2+内流和囊泡性递质释放的长期影响，以及2)CA1锥体神经元的全细胞膜片钳记录来表征低水平Pb2+暴露对突触后n -甲基- d -天冬氨酸受体（NMDAR）门控电流的长期影响。我们的工作假设是，早期Pb2+暴露会导致NMDAR功能受损，从而导致BDNF合成和释放减少，进而导致对正常认知功能至关重要的突触前传递受损。已知增加BDNF水平和释放的一种操作是丰富的环境。为了验证我们的假设并确定保护大脑免受Pb2+发育损伤的潜在方法，我们提出：1)表征低[Pb2+]暴露对BDNF基因表达、启动子甲基化和TrkB受体激活的影响；2)测试富集环境、外源性脑室内BDNF输注或TrkB受体拮抗剂7,8-二羟黄酮预防Pb2+诱导的NMDAR功能和递质释放的长期损伤的能力。我们提出的研究计划解决了早期发育暴露于低水平Pb2+是否比以前认为的对脑功能有长期有害影响的关键问题。这些研究将为Pb2+对认知和记忆储存至关重要的突触前和突触后机制的影响提供新的信息。此外，我们将研究提高BDNF释放和TrKB受体激活的新型治疗方法，以防止Pb2+暴露的长期有害影响。

DESCRIPTION (provided by applicant): The severe nervous system developmental risks of early life exposure to lead (Pb2+) are well known. It is now becoming increasingly clear that much lower concentrations of Pb2+ can produce significant detrimental effects in children, heightening the need to understand the properties and extent of Pb2+ actions on the brain. Our laboratory has recently discovered that in vitro exposure to very low levels of Pb2+ produces long-term impairments in presynaptic transmitter release in cultured hippocampal neurons, and that these actions mimic those observed in animals lacking the trophic factor brain-derived neurotrophic factor (BDNF). We propose studies in hippocampal slices from rats exposed to low levels of Pb2+ during development to utilize 1) state-of- the-art two-photon imaging methods to assess long-term effects on presynaptic Ca2+ influx and vesicular transmitter release in intact synapses, and 2) whole-cell patch-clamp recording from CA1 pyramidal neurons to characterize the long-term effects of low level Pb2+ exposure on postsynaptic N-methyl-D-aspartate receptor (NMDAR)-gated currents. Our working hypothesis is that early Pb2+ exposure produces impairments in NMDAR function that leads to reduced BDNF synthesis and release and subsequent impairments of presynaptic transmission that are critical to normal cognitive function. One manipulation known to increase BDNF levels and release is an enriched environment. To test our hypothesis and identify potential methods of protecting the brain from developmental damage from Pb2+, we propose to 1) characterize the effects of low [Pb2+] exposure on BDNF gene expression, promoter methylation and TrkB receptor activation, and 2) test the ability of an enriched environment, exogenous intraventricular BDNF infusion or administration of the TrkB agonist 7,8-dihydroxyflavone to prevent Pb2+-induced long-term damage to NMDAR function and transmitter release. The research program we propose addresses the critical question of whether early developmental exposure to low-levels of Pb2+ than previously thought have long-term detrimental effects on brain function. These studies will provide novel information about Pb2+ effects on both presynaptic and postsynaptic mechanisms critical to cognition and memory storage. Further, we will examine novel therapeutic manipulations that elevate BDNF release and TrKB receptor activation in order to protect against the long-term detrimental effects of Pb2+ exposure.