ORGANOCHLORINE PESTICIDES AND SEROTONERGIC DEVELOPMENT

本项目的总体目的是确定母亲接触有机氯农药（如狄氏剂）对后代血清素（5-羟色胺）神经元及其受体发育的可能后果。研究狄氏剂对血清素能系统发育影响的基本原理是先前的证据表明狄氏剂在体外强烈抑制胚胎5-羟色胺神经元的生长和存活。狄氏剂似乎通过作为一种有效的GABAA受体拮抗剂来阻断GABA的营养作用而引起这些影响。本实验旨在确定母体在胚胎期E12-E17给予不同剂量的狄氏剂或经典GABAA受体拮抗剂双管碱是否会改变大鼠产前和产后大脑中5-HT神经元及其受体的发育。在Specific Aim 1中，将在胚胎期E18和出生后0、10、30和60天确定处理对后代5-HT神经元发育的影响。标准（ABC过氧化物酶）免疫细胞化学与特异性5-羟色胺抗体将被用来评估对5-羟色胺免疫反应细胞体的数量和它们轴突投射的一般分布的影响。半定量放射标记免疫细胞化学（[125I]蛋白A免疫结合）将用于测量解剖匹配的脑切片中5-羟色胺免疫反应性的相对变化。在Specific Aim 2中，治疗对多种5-HT受体亚型表达的影响将通过测定E18和PND 0、10、30和60处受体mRNA和蛋白的变化来确定。利用5-HT1A、5-HT2A、5-HT2B、5-HT2C和5-HT3受体序列特异性引物和内部标准的定量竞争性RT-PCR，将用于测量解剖脑区域中受体mRNA的变化。[125I]蛋白A与5-HT1A、5-HT2A、5-HT2B、5-HT2C和5-HT3受体特异性抗体的免疫结合将分析解剖匹配的脑切片特定区域受体免疫反应性的相对变化。这些研究将深入了解母体接触有机氯农药对其后代神经发育的潜在风险。这是一个重要的问题，因为在发育障碍和情绪障碍的病因学中怀疑涉及血清素能系统的功能失调个体发生。

The General Aim of this project is to determine possible consequences of maternal exposure to organochlorine pesticides, such as dieldrin, on development of serotonin (5-HT) neurons and their receptors in offspring. The rationale for examining the effects of dieldrin on development of the serotonergic system is previous evidence that dieldrin strongly inhibits the growth and survival of embryonic 5-HT neurons in vitro. Dieldrin appears to cause these effects by acting as as a potent GABAA receptor antagonist to block the trophic effects of GABA. The Specific Aims are designed to determine whether maternal administration of different doses of dieldrin or the classical GABAA receptor antagonist bicuculline from embryonic day E12-E17 alters development of 5-HT neurons and their receptors in prenatal and postnatal rat brain. In Specific Aim 1, effects of treatments on development of 5-HT neurons in offspring will be determined at embryonic day E18 and postnatal days (PND) 0, 10, 30 and 60. Standard (ABC peroxidase) immunocytochemistry with specific 5-HT antibodies will be used to assess effects on numbers of 5-HT immunoreactive cell bodies and the general distribution of their axonal projections. Semi-quantitative radiolabel immunocytochemistry ([125I]protein A immunobinding) will be used to measure relative changes in 5-HT immunoreactivity in anatomically matched brain sections. In Specific Aim 2, effects of treatments on expression of multiple 5-HT receptor subtypes will be determined using assays to measure changes in receptor mRNA and protein at E18 and PND 0, 10, 30 and 60. Quantitative competitive RT-PCR with primers and internal standards specific for 5-HT1A, 5-HT2A, 5-HT2B, 5-HT2C and 5-HT3 receptor sequences will be used to measure changes in receptor mRNA in dissected brain regions. [125I]protein A immunobinding with specific antibodies to 5-HT1A, 5-HT2A, 5-HT2B, 5-HT2C and 5-HT3 receptors will analyze relative changes in receptor immunoreactivity in specific regions of anatomically matched brain sections. These studies will provide insights into the risk potential of maternal exposure to organochlorine pesticides for neural development in their offspring. This is an important question because of the suspected involvement of dysfunctional ontogeny of the serotonergic system in the etiology of developmental disabilities and mood disorders.