Role of Nicotine in Regulation of naChRs in Drosophila

描述（由申请人提供）： nAChRs性质的改变与兴奋性突触传递 神经元中的神经元被认为在尼古丁的过程中起着重要作用。 人类的成瘾然而，行为背后的分子机制 对尼古丁的反应仍然知之甚少。在这一补助金中，我们建议 在一个基因可及的动物模型中解决这些问题。我们有 最近描述了介导快速兴奋性的nAChR的性质， 在培养的果蝇神经元中的传递。这些文化提供了一种独特的 有机会探索重要的细胞和分子机制， 调节nAChR和中枢突触的胆碱能传递。我们 第一个目标集中在表征生物物理，药理学， nAChRs介导尼古丁诱发电流的分子特性/钙 果蝇神经元中的瞬变现象。此外，我们还将确定 不同的nAChR基因，通过检查敲除或减少 培养神经元中特异性亚基的表达。这将是 通过检查neurona来完成!对尼古丁的反应 从发现nAChR基因的缺陷或暴露于dsRNA的培养物中 靶向特定的nAChR亚基。 第二个目的是研究尼古丁在调节nAChR中的作用， 胆碱能传递对尼古丁fACh，1125-aBTX的神经元反应 结合，和胆碱能传输的属性将被监测， 神经元长期暴露于尼古丁。我们还将研究 短时间重复尼古丁治疗，模拟暴露 在吸烟过程中发生的。cAMP信号级联在以下方面发挥作用： 调节果蝇和哺乳动物对乙醇和可卡因的反应。在我们 第三个目的，我们将研究nAChRs和胆碱能突触的特性， 电流cAMP信号突变体，以确定是否尼古丁的影响， 也由CAMP调节。最后，我们将使用差异显示和cDNA 微阵列来识别尼古丁差异调节的基因。 我们的研究结果将提供重要的见解莫!分子的 尼古丁诱导的nAChRs调节的潜在机制。这些数据将 也有助于对分子机制的一般理解 在调节兴奋性突触的传递中很重要， 在脊椎动物和无脊椎动物之间高度保守。最后，这些知识 可能有助于设计药物和治疗， 或通过调节nAChR和突触蛋白来预防人类尼古丁成瘾。 功能

DESCRIPTION (provided by applicant): Modifications in the properties of nAChRs and excitatory synaptic transmission in neurons are thought to play an important role in the process of nicotine addiction in humans. However, the molecular mechanisms underlying behavioral responses to nicotine remain poorly understood. In this grant we propose to address these questions in a genetically accessible animal model. We have recently described the properties of nAChRs that mediate fast excitatory transmission in cultured Drosophila neurons. These cultures provide a unique opportunity to explore the cellular and molecular mechanisms important in regulation of nAChRs and cholinergic transmission at central synapses. Our first aim focuses on characterizing the biophysical, pharmacological, and molecular properties of nAChRs mediating nicotine-evoked currents/calcium transients in Drosophila neurons. In addition we will determine the function of different nAChR genes by examining the consequences of knocking out or reducing the expression of specific subunit/s in the cultured neurons. This will be accomplished by examining neurona! responses to nicotine in cultures prepared from deficiencies uncovering nAChR genes or in cultures exposed to dsRNA targeted to particular nAChR subunit/s. The second aim examines the role of nicotine in regulation of nAChRs and cholinergic transmission. Neuronal responses to nicotinefACh, 1125-aBTX binding, and the properties of cholinergic transmission will be monitored in neurons chronically exposed to nicotine. We will also examine the effects of short duration, repetitive treatments with nicotine, mimicking the exposure that occurs during smoking. The cAMP signaling cascade plays a role in regulating responses to ethanol and cocaine in both flies and mammals. In our third aim we will examine the properties of nAChRs and cholinergic synaptic currents in cAMP signaling mutants to determine if the effects of nicotine are also modulated by CAMP. Finally, we will use differential display and cDNA microarrays to identify genes that are differentially regulated by nicotine. The results of our studies will provide important insights into the mo!ecular mechanisms underlying nicotine-induced regulation of nAChRs. These data will also contribute to the general understanding of the molecular mechanisms important in regulating transmission at excitatory synapses, likely to be highly conserved between vertebrates and invertebrates. Finally, this knowledge may be useful in the design of drugs ancVor therapies targeted at ameliorating or preventing nicotine addiction in humans by regulating nAChRs and synaptic function.