NPY AND ATP COTRANMISSION IN CHROMAFFIN CELLS

嗜铬细胞中的 NPY 和 ATP 协同传输

• 批准号: 6530742
• 负责人: TERRY D HEXUM
• 金额: $ 21.9万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-07 至 2004-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6530742
• 关键词: adenosine triphosphate; animal tissue; calcium flux; calcium indicator; catecholamines; chromaffin cells; high performance liquid chromatography; immunoprecipitation; inositol phosphates; neural transmission; neuropeptide Y; neurotransmitter biosynthesis; northern blottings; tissue /cell culture; tyrosine 3 monooxygenase; western blottings

Neuropeptide Y (NPY) exerts a variety of effects including increased blood pressure and food intake and decreased anxiety. The increase in blood pressure occurs in conjunction with co- existing transmitters such as ATP and norepinephrine. The long- term objective of research in this laboratory is to reveal the mechanism(s) by which neuropeptides produce physiological changes. Bovine chromaffin cells provide a useful model with which to study neuropeptide action since these cells synthesize and secrete NPY in response to stimulation with nicotine. The goal of the current proposal is to gain insight into the physiological role of NPY as a co-transmitter with ATP. NPY binds to chromaffin cell receptors of the Y1 subtype and inhibits forskolin stimulated adenylate cyclase activity. Our preliminary data show that it does not alter chromaffin cell intracellular Ca2+ concentration ([Ca2+]i) or inositol phosphate formation unless ATP is added to culture media along with NPY. NPY can significantly enhance both the ATP stimulated increase in inositol phosphate formation and [Ca2+]i. The chromaffin cell response to this action is unknown but may include the regulation of neurotransmitter synthesis since increased inositol phosphate formation and [Ca2+]i could have been shown to result in increased tyrosine hydroxylase synthesis. In addition, an increase in [Ca2+]i could stimulate tyrosine hydroxylase activity acutely due to increased enzyme phosphorylation mediated by CaM kinase II. Our hypothesis is that NPY and ATP are co- transmitters in chormaffin cells that serve to increase [Ca2+]i by increasing inositol phosphate formation with the resultant effect of increased catecholamine biosynthesis. The specific aims are to characterize the enhancing effect of NPY on ATP- stimulated inositol phosphate formation, identify the ATP receptor subtype mediating the stimulation of InsP formation and increased [Ca2+]i, determine the specificity of the ATP potentiating effect of NPY and discover the response of the chromaffin cell to increased intracellular Ca2+. These studies will further our understanding of the physiological role of neuropeptides and their involvement in co-transmission. Information will be provided which may lead to improved therapy of diseases particularly involving aberrations in transmission by catecholamines e.g., epilepsy, anorexia, alcoholism, and hypertension.

神经肽Y （NPY）具有多种作用，包括增加血压和食物摄入量以及减少焦虑。血压升高与共存的递质如ATP和去甲肾上腺素一起发生。本实验室的长期研究目标是揭示神经肽产生生理变化的机制。牛嗜铬细胞在尼古丁刺激下合成和分泌NPY，为研究神经肽的作用提供了一个有用的模型。本研究的目的是深入了解NPY作为ATP共递质的生理作用。NPY与Y1亚型的染色质细胞受体结合，抑制forskolin刺激的腺苷酸环化酶活性。我们的初步数据表明，除非将ATP与NPY一起添加到培养基中，否则它不会改变染色质细胞内Ca2+浓度（[Ca2+]i）或肌醇磷酸的形成。NPY可以显著增强ATP刺激的肌醇磷酸形成和[Ca2+]i的增加。染色质细胞对这一作用的反应尚不清楚，但可能包括神经递质合成的调节，因为肌醇磷酸形成和[Ca2+]i的增加可能导致酪氨酸羟化酶合成的增加。此外，由于CaM激酶II介导的酶磷酸化增加，[Ca2+]i的增加可以急剧刺激酪氨酸羟化酶的活性。我们的假设是，NPY和ATP是染色质细胞中的共递质，通过增加磷酸肌醇形成和增加儿茶酚胺生物合成的结果效应来增加[Ca2+]i。具体目的是表征NPY对ATP刺激的肌醇磷酸形成的增强作用，鉴定介导刺激InsP形成和增加[Ca2+]i的ATP受体亚型，确定NPY ATP增强作用的特异性，并发现染色质细胞对细胞内Ca2+增加的反应。这些研究将进一步加深我们对神经肽的生理作用及其参与共传播的认识。将提供可能导致改进疾病治疗方法的信息，特别是涉及儿茶酚胺传播异常的疾病，例如癫痫、厌食症、酗酒和高血压。