Morphine tolerance in live neurons

活神经元的吗啡耐受性

• 批准号: 8249261
• 负责人: Erica Sawyer Levitt
• 金额: $ 4.92万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8249261
• 关键词: ARRB2; Agonist; Animals; Brain; Cells; Chronic; Clinic; Coupled; Development; Dose; Effectiveness; Electrophysiology (science); Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Excision; G protein-coupled inwardly-rectifying potassium channel; GTP-Binding Proteins; Goals; Hour; In Vitro; Injection of therapeutic agent; Knockout Mice; Learning; Life; Ligands; Long-Term Effects; Measures; Mediating; Morphine; Mus; Neurons; Okadaic Acid; Opioid; Pain; Patients; Pharmaceutical Preparations; Phosphoric Monoester Hydrolases; Phosphorylation; Potassium; Property; Protein Dephosphorylation; Rattus; Recovery; Regimen; Risk; Role; Signal Transduction; Slice; Testing; Therapeutic; Time; Tissues; Work; addiction; arrestin 2; chronic pain; clinically relevant; desensitization; locus ceruleus structure; mu opioid receptors; novel therapeutics; phosphatase inhibitor; prevent; receptor; research study; response; voltage clamp

Morphine is one of the most commonly prescribed opioids to treat pain. However, prolonged use often leads to the development of tolerance characterized by loss of effectiveness and dose escalation. Morphine tolerance is in part due to a loss of signaling from mu opioid receptors (MOR) independent of a change in receptor levels. Despite years of study, the mechanisms causing this loss of signaling remain unsolved. Traditionally, tolerance is measured in cells or tissue removed from a chronically treated animal and washed free of morphine for several hours. Preliminary experiments have discovered that if brain slices from a morphine-treated rat are instead maintained in morphine continuously, the morphine response is significantly smaller compared to slices in which morphine was washed out, indicating that a portion of tolerance can recover within a relatively short wash. This rapid recovery following morphine removal is newly identified and may have appreciable importance to the persistence of tolerance. The goal of this proposal is to better understand morphine tolerance by carefully examining the often overlooked desensitization that is maintained by the continuous presence of morphine. The overall hypothesis is that two types of tolerance (long-lasting tolerance and chronic desensitization) develop during long-term morphine treatment distinguished by the time-course of recovery, which have different mechanisms and depend on treatment paradigm. This hypothesis will be tested using locus coeruleus neurons contained in brain slices from naove or morphine-treated animals. Morphine effects will be measured using whole-cell voltage-clamp electrophysiology to detect MOR-mediated activation of G protein-coupled inwardly rectifying potassium (GIRK) channels. The specific aims of this proposal are as follows: (1) to identify the long-lasting tolerance and chronic desensitization that results from long-term continuous morphine treatment of rats and mice, (2) to compare this to tolerance and desensitization following long-term intermittent morphine treatment and (3) to determine the role of 2-arrestin 2 in the development of long-lasting tolerance (using 2-arrestin 2 knockout mice) and the role of ser/thr phosphatases in development of and recovery from chronic desensitization (using okadaic acid). Together, this proposal will provide new information about chronic desensitization that develops after long-term morphine treatment. Focusing on morphine specifically will help identify mechanisms involved in tolerance to this most commonly used opioid agonist.

吗啡是治疗疼痛最常用的阿片类药物之一。然而，长期使用通常会导致耐受性的产生，其特征是有效性丧失和剂量增加。吗啡耐受的部分原因是 mu 阿片受体 (MOR) 信号丢失，与受体水平的变化无关。尽管经过多年的研究，导致这种信号丢失的机制仍未解决。传统上，耐受性是通过从长期治疗的动物身上取出的细胞或组织来测量的，并清洗数小时以去除吗啡。初步实验发现，如果将接受吗啡治疗的大鼠脑切片持续置于吗啡中，与洗掉吗啡的切片相比，吗啡反应明显较小，这表明部分耐受性可以在相对较短的清洗时间内恢复。吗啡去除后的这种快速恢复是新发现的，可能对耐受性的持久性具有相当重要的意义。该提案的目标是通过仔细检查吗啡持续存在所维持的经常被忽视的脱敏作用，更好地了解吗啡耐受性。总体假设是，在长期吗啡治疗过程中会出现两种类型的耐受性（长期耐受性和慢性脱敏），其区别在于恢复的时间过程，这两种耐受性具有不同的机制并取决于治疗模式。这一假设将使用幼稚动物或吗啡治疗动物脑切片中包含的蓝斑神经元进行测试。将使用全细胞电压钳电生理学来测量吗啡效应，以检测 MOR 介导的 G 蛋白偶联内向整流钾 (GIRK) 通道的激活。该提案的具体目的如下：（1）确定大鼠和小鼠长期连续吗啡治疗导致的持久耐受和慢性脱敏，（2）将其与长期间歇吗啡治疗后的耐受和脱敏进行比较，以及（3）确定2-arrestin 2在长期耐受发展中的作用（使用2-arrestin 2敲除小鼠）和 丝氨酸/苏氨酸磷酸酶在慢性脱敏（使用冈田酸）的发展和恢复中的作用。总之，该提案将提供有关长期吗啡治疗后出现的慢性脱敏的新信息。特别关注吗啡将有助于确定对这种最常用的阿片类激动剂的耐受性机制。