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: 6693744
• 负责人: FRANCES M. LESLIE
• 金额: $ 37.6万
• 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/6693744
• 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.

在之前的资助期间，我们已经证明尼古丁与儿茶酚胺神经元上的尼古丁受体(NAChRs)直接相互作用，刺激发育中的大鼠脑内去甲肾上腺素(NE)和多巴胺(DA)的释放。我们现在建议进行一系列研究，以检验这些观察结果的生理和临床相关性。我们将研究nAChR介导的儿茶酚胺在C57BL/6J小鼠体内释放的物种特异性，并将使用零突变小鼠品系来检测发育中的NE和DA末梢上nAChRs的亚单位组成。将使用两种类型的方法来研究内源性配体是否可能激活发育中的去甲肾上腺素能蓝斑(LC)终末的nAChRs。我们将用免疫组织化学的方法来确定胆碱能细胞和纤维的发育形态与LC及其前脑和后脑终末区的关系。针对囊泡乙酰胆碱转运蛋白(Vacht)和多巴胺β羟基酶(DBH)的抗体将分别用于显示发育中的胆碱能和去甲肾上腺素能成分。我们还将确定激活或阻断nAChR是否可以改变需要激活LC的范例中的早期学习，包括早期嗅觉学习和体感联合条件反射。在每项任务中的学习都被证明需要分别通过触觉刺激和电击激活LC。我们将测试这样一种假设，即外周注射尼古丁可以诱导足够的LC刺激，从而在没有感觉刺激的情况下促进学习。我们还将使用nAChR拮抗剂mecamylamine来确定内源性乙酰胆碱(ACh)是否影响感觉刺激诱导的LC在这些任务中的激活。在确定了nAChRs在儿茶酚胺释放的急性调节中的功能后，我们将研究产前长期尼古丁暴露的张力性和相性模式对中枢儿茶酚胺系统发育的影响。渗透性微泵将用于持续输注和自我给药，以脉冲式给药给怀孕大鼠尼古丁。不同年龄的后代将被用于功能和神经解剖学研究，以确定在出生前暴露于尼古丁后，NE和DA终末区域的整体兴奋性或尼古丁敏感性。同时涉及中枢NE(早期嗅觉学习、联合体感条件反射)和DA系统(运动、条件性位置偏爱、自我管理)的行为测试将被用来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变化有关。这些体外和体内相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的潜在机制。