RESEARCH PROJECT: "Mechanisms of B(a)P Induced Neurotoxicity"

拟议研究的长期目标是确定妊娠期B(A)P暴露 C57BL小鼠行为和神经功能正常的干扰素 “脂肪海马区突触的可塑性和行为。需要检验的中心假设是妊娠 在某些环境中，暴露在人类可见水平的B(A)P气溶胶(PM2.5亩)会导致终生 学习和记忆缺陷，这些缺陷至少部分是通过调节 发育中NMDA/AMPA谷氨酸受体亚单位的表达和功能 C57BL小鼠的海马区正在形成兴奋性突触。目标1将决定 B(A)P在孕14、16、18和出生后F1胎脑中的分布 GD14-17吸入剂量分别为0、50、100和200微克/立方米时，海马区的(PND)0 同时测定谷氨酸受体亚单位在子宫内表达的影响 使用体外原代神经元培养进行。Aim#2将测试怀孕期间是否暴露于 吸入相同剂量的B(A)P会导致学习障碍，使用以前被证明依赖于 对海马体功能的影响。目标#3将孕期暴露于苯并(A)P与海马区的测量相关联 长时程增强(LTP)，在PND 60和120对F1代小鼠进行评估。参与其中的 本LTP中的NMDA和/或AMPA受体及其因暴露于B(A)P而发生的变化将通过以下方式进行评估 在对照和B(A)P的LTP分析之前应用NMDA或AMPA选择性受体拮抗剂 暴露F1代小鼠。目标4将测试妊娠期是否暴露于剂量为 导致行为学习和生理缺陷也调节各种NMDA和 AMPA受体亚基，实时分析GD18和PND 0、5、10、20和60上的表达水平 对照和对照的聚合酶链式反应(用于信使核糖核酸的评估)和蛋白质印迹分析(用于亚单位蛋白的评估) B(A)P暴露于F1代C57BL小鼠。这些研究将作为第一步，用 孕期接触B(A)P后谷氨酸受体亚单位表达改变的因果关系 成年动物学习行为的减弱，这将在未来的研究中通过利用小鼠来扩展 基因改变以改变相关AMPA或NMDA受体亚单位的表达，或其 下游效应器分子。这些研究与苯并(A)磷的人类后果直接相关。 我们预计，我们的研究将导致对一种重要动物的严格描述 了解环境毒素所致神经功能障碍的病因和测试的模型 关于有效治疗或其他干预措施的假设。

The long-term objective of the proposed studies is to determine whether gestational B(a)P exposure nterferes with normal behavior and neural function in C57BL mice as demonstrated by significant deficits in "lippocampal synaptic plasticity and behavior. The central hypothesis to be tested is that gestational exposure to B(a)P aerosol (PM2.5mu) at levels seen in humans in certain environments result in lifelong learning and memory deficits, and that these deficits that are mediated, at least in part, through modulation of developmental NMDA/AMPA glutamate receptor subunit expression and function at a time when excitatory synapses are being formed in the hippocampus of C57BL mice. AIM #1 will determine the disposition of B(a)P in F1 fetal brain (in C57BL mice) on gestational day (GD) 14, 16, 18, and postnatal day (PND) 0 in the hippocampus after GD14-17 inhalation exposure at doses of 0, 50, 100 and 200 mu g/m3A simultaneous determination of the effect on glutamate receptor subunit expression in utero will be conducted, using ex vivo primary neuronal cultures. AIM #2 will test whether gestational exposure to the same inhalational doses of B(a)P results in deficits in learning, using behaviors previously shown to depend on hippocampal function. AIM #3 will correlate gestational exposure to B(a)P with measures of hippocampal long-term potentiation (LTP), evaluated at PND 60 and 120 in F1 generation mice. The involvement of NMDA and/or, AMPA receptors in this LTP, and its changes due to B(a)P exposure, will be assessed by application of NMDA or AMPA-selective receptor antagonists prior to LTP analysis of control and B(a)P exposed F1 generation mice. AIM #4 will test whether gestational exposure to the doses of B(a)P which lead to behavioral learning and physiological deficits also modulate the expression of various NMDA and AMPA receptor subnits, profiled for expression levels on GD18 and PND 0, 5, 10, 20, and 60 using real time PCR (for mRNA assessment) and Western blot analyses( for subunit protein assessment) in control and B(a)P exposed F1 generation C57BL mice. These studies will serve as the first step in querying the with the causality of altered glutamate receptor subunit expression subsequent to gestational exposure to B(a)P in attenuation of learning behaviors in adult animals, which will be extended in future studies by exploiting mice genetically altered to modify the expression of relevant AMPA or NMDA receptor subunits, or their downstream effector molecules. These studies are directly relevant to human consequences of B(a)P exposure, and we anticipate that our studies will lead to the rigorous characterization of an important animal model both to understand the etiology of environmental toxin-induced neurological dysfunction and to test hypotheses regarding effective therapeutic or other interventions .