Nicotine Regulation of Developing Brain Catecholamines

尼古丁对发育中的大脑儿茶酚胺的调节

• 批准号: 6497797
• 负责人: FRANCES M. LESLIE
• 金额: $ 37.24万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-05-01 至 2006-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6497797
• 关键词: age difference; behavior test; conditioning; developmental neurobiology; dopamine; embryo /fetus toxicology; genetically modified animals; immunocytochemistry; in situ hybridization; laboratory mouse; laboratory rat; learning; locus coeruleus; mecamylamine; neurochemistry; neurotransmitter transport; nicotine; nicotinic receptors; norepinephrine; radiotracer; receptor expression

In the previous funding period, we have demonstrated that nicotine interacts directly with nicotinic receptors (nAChRs) on catecholamine neurons to stimulate norepinephrine (NE) and dopamine (DA) release in developing rat brain. We now propose a series of studies to examine the physiological and clinical relevance of these observations. We will study the species specificity of nAChR-mediated catecholamine release in C57BL/6J mice, and will use null mutant mouse strains to examine the subunit composition of nAChRs on developing NE and DA terminals. Two types of approach will be used to examine whether nAChRs on developing noradrenergic locus coeruleus (LC) terminals may be activated by endogenous ligand. We will use immunohistochemistry to establish the developmental appearance of cholinergic cells and fibers in relationship to LC and its forebrain and hindbrain terminal fields. Antibodies specific for the vesicular acetylcholine transporter (VAChT) and for dopamine beta hydroxylase (DBH) will be used to visualize developing cholinergic and noradrenergic elements, respectively. We will also determine whether nAChR activation or blockade can modify early learning in paradigms that have been shown to require activation of the LC, including early olfactory learning and somatosensory associative conditioning. Learning in each task has been shown to necessitate LC activation by tactile stimulation and electrical shock, respectively. We will test the hypothesis that peripheral administration of nicotine induces sufficient LC stimulation to facilitate learning in the absence of sensory stimuli. We will also use the nAChR antagonist, mecamylamine, to determine whether endogenous acetylcholine (ACh) influences sensory stimulus-induced LC activation on these tasks. Having established the functional roles of nAChRs in acute regulation of catecholamine release, we will examine the effects of both tonic and phasic modes of chronic prenatal nicotine exposure on the development of central catecholamine systems. Osmotic minipumps will be used for constant infusion and self- administration for pulsatile delivery of nicotine to pregnant rats. Offspring of varying ages will be used for functional and neuroanatomical studies to determine whether prenatal nicotine exposure after the overall excitability, or nicotine-sensitivity, of NE and DA terminal fields. Behavioral tests that involve both central NE (early olfactory learning, associative somatosensory conditioning) and DA systems (locomotion, conditioned place preference, self-administration) will be used to determine whether chronic nicotine-induced changes in catecholamine system development are associated with long-term alterations in behavioral response. These combined in vitro and in vivo approaches are designed to elucidate mechanisms underlying adaptive responses of central catecholamine neurons to chronic prenatal nicotine exposure.

在之前的资助期内，我们已经证明尼古丁直接与儿茶酚胺神经元上的尼古丁受体（nachr）相互作用，以刺激发育中的大鼠大脑中去甲肾上腺素（NE）和多巴胺（DA）的释放。我们现在提出了一系列的研究来检验这些观察的生理和临床相关性。我们将在C57BL/6J小鼠中研究nachr介导的儿茶酚胺释放的物种特异性，并将使用零突变小鼠菌株检测nachr在发育中的NE和DA末端的亚基组成。我们将采用两种方法来检测发育中的去肾上腺素能蓝斑（LC）末端的nachr是否会被内源性配体激活。我们将使用免疫组织化学来建立与LC及其前脑和后脑终端场相关的胆碱能细胞和纤维的发育外观。针对囊泡乙酰胆碱转运蛋白（VAChT）和多巴胺-羟化酶（DBH）的特异性抗体将分别用于观察胆碱能和去甲肾上腺素能元素的形成。我们还将确定nAChR激活或阻断是否可以改变需要LC激活的早期学习模式，包括早期嗅觉学习和体感联想条件反射。在每个任务中学习都需要分别通过触觉刺激和电击激活LC。我们将测试尼古丁外周管理诱导足够的LC刺激以促进在没有感觉刺激的情况下学习的假设。我们还将使用nAChR拮抗剂甲胺来确定内源性乙酰胆碱（ACh）是否会影响感觉刺激诱导的LC激活。在确定了nachr在儿茶酚胺释放的急性调节中的功能作用后，我们将研究慢性产前尼古丁暴露对中枢儿茶酚胺系统发育的强直和相位模式的影响。微型渗透泵将用于持续输注和自我给药，以脉动方式向怀孕大鼠输送尼古丁。不同年龄的后代将被用于功能和神经解剖学研究，以确定产前尼古丁暴露后NE和DA终端场的总体兴奋性或尼古丁敏感性。行为测试包括中央NE（早期嗅觉学习、联想体感条件反射）和DA系统（运动、条件性位置偏好、自我给药），将用于确定慢性尼古丁诱导的儿茶酚胺系统发育的变化是否与行为反应的长期改变有关。这些体外和体内结合的方法旨在阐明中枢儿茶酚胺神经元对慢性产前尼古丁暴露的适应性反应机制。