NEURODEVELOPMENTAL STUDIES OF LEAD POISONING

在三年的时间里，我们将研究体外细胞培养， 系统的神经毒性和神经发育的影响， 无机和有机铅（Pb）化合物通常存在于 环境保护 我们打算研究这种重金属对体外细胞的影响， 准确反映体内CNS的实验系统 功能与发展。 这种细胞培养系统提供了 将神经元和神经胶质细胞暴露于已知的 不同发育阶段的潜在毒素浓度 阶段，并允许直接评估神经损伤， 恢复以及探索潜在的分子 机制和可能的干预方法。 将来自16天龄胚胎小鼠脑的细胞解离 转化成单细胞悬浮液并置于旋转培养中， 它们通过重新聚集来恢复它们的生长和分化， 发展轴突网络，髓鞘形成， 神经递质的合成、储存、释放和摄取。 在四 体外发育的阶段（相当于prenate， 新生儿、青少年和年轻成人CNS）， 无机和有机铅化合物对后续 发育事件和神经元和神经胶质细胞的组织 细胞应通过将重新聚集体暴露于四种 不同浓度的铅化合物24小时。 在 收获时，应取再集料的等分试样， 确定这些先导化合物对1） 通过以下方法测定的再聚集体的细胞组织学 常规和电子显微镜，2）蛋白质和 磷蛋白组成3）髓鞘膜合成，和4） 第二信使信号转导机制 系统. 对照细胞应从处理过的培养瓶中采集 相同地，但其中没有添加铅化合物。 特定CNS解剖学、生理学和 铅在不同发育阶段引起的生化变化 使用这种体外实验系统的阶段将提供 用于预防和可能治疗的信息 干预导致铅中毒的分子过程 神经毒性作用。

Over a three year period we will examine an in vitro cell culture system for the neurotoxic and neurodevelopmental effects caused by inorganic and organic lead (Pb) compounds commonly found in the environment. We intend to study the influence of this heavy metal on an in vitro experimental system that accurately reflects in vivo CNS function and development. This cell culture system offers the advantage of exposing neurons and glial cells to known concentrations of potential toxins at diffferent developmental stages and permits direct assessment of neural damage and recovery as well as exploration of the underlying molecular mechanisms and possible intervention methods. Cells from 16 day old embryonic mouse brains shall be dissociated into single cell suspensions and placed into rotary culture where they resume their growth and differentiation by reaggregating, developing axonal networks, myelinating, and exhibiting neurotransmitter synthesis, storage, release and uptake. At four stages of in vitro development (equivalent to the prenate, neonate, adolescent and young adult CNS), the effects of inorganic and organic lead compounds on subsequent developmental events and the organization of neuronal and glial cells shall be evaluated by exposing the reaggregates to four different concentrations of lead compounds for 24 hours. At the time of harvesting, aliquots of reaggregates shall be taken to determine the influence of these lead compounds on 1) the cellular organizaiton of the reaggregates as determined by conventional and electron microscopy, 2) protein and phosphoprotein composition 3) myelin membrane synthesis, and 4) signal tranduction mechanisms involving second messenger systems. Control cells shall be taken from flasks treated identically, but in which no lead compound has been added. Definition of the specific CNS anatomical, physiological and biochemical changes caused by lead at various developmental stages using this in vitro experimental system will provide information for preventing and possibly therapeutically intervening in the molecular processes responsible for lead's neurotoxic effects.