KAPPA OPIOID RECEPTORS AND TURNOVER IN THE BRAIN

这项提议的总体目标是研究kappa阿片类药物 受体刺激的磷脂酰肌醇(PI)在体内几个区域的周转 老鼠的大脑。这项建议源于最近在这方面的发现 Kappa(但既不是Mu也不是Delta)阿片受体的实验室 刺激海马片中PI的周转。 这一建议旨在扩大kappa受体介导的PI 对大脑其他区域的周转反应。拟议的工作将 阿片受体介导的猪的首次表征 并可能提供第一个生化证据，证明PI的周转 是与kappa受体相关的初级第二信使系统 刺激大脑，也许也刺激其他组织。 这种反应的药理作用将在不同的大脑中进行比较 测量kappa选择性激动剂和拮抗剂的能力 激活或抑制反应的激活。Mu和Delta- 选择性激动剂和拮抗剂也将进行测试，以确定 选择性。Kappa受体介导的PI周转水平 反应将与kappa受体在脑内的分布进行比较。 大脑。建议使用最近可用的选择性kappa配体 用于放射自显影和最先进的图像分析系统 确定kappa受体的分布，这一直是个问题。 Kappa阿片受体介导的心肌细胞PI周转的测定 海马区切片需要对分析方法进行广泛的优化。至 进一步提高了分析的灵敏度，分离和 高效液相色谱法和放射化学法测定肌醇磷酸盐的含量 提出了一种流量检测方法。自动取样器和放化学流 要实现这一目标，就需要探测器。 最后，作为衡量G蛋白与kappa受体偶联的指标，它是 建议在不同的疾病中检测kappa受体刺激的低Km GTP酶 脑区，比较GTP酶的反应与PI周转率和 受体分布。拟议工作的总体意义是 它将在kappa受体和PI周转之间建立联系， 它以前没有与任何阿片受体亚型相关。 这项工作可能为生物化学反应提供证据。 中枢神经系统kappa受体激活的行为效应。

The overall objective of this proposal is to investigate kappa opioid receptor-stimulated Phosphoinositide (PI) turnover in several areas in the rat brain. The proposal derives from the recent discovery in this laboratory that kappa (but neither mu nor delta) opioid receptors stimulate PI turnover in hippocampal slices. This proposal is aimed at extending the kappa receptor-mediated PI turnover response to other areas of the brain. The proposed work will constitute the first characterization of opioid receptor-mediated PI turnover and may provide the first biochemical evidence that PI turnover is the primary second messenger system associated with kappa receptor stimulation in the brain and perhaps in other tissues as well. The pharmacology of the response will be compared in different brain areas, measuring the ability of kappa-selective agonists and antagonists to activate or inhibit the activation of the response. Mu and delta- selective agonists and antagonists will be tested as well to establish selectivity. The level of the kappa receptor-mediated PI turnover response will be compared with the distribution of kappa receptor in the brain. It is proposed to use recently available selective kappa ligands for autoradiography and a state-of-the-art image analysis system to determine kappa receptor distribution, which has been problematic. The measurement of kappa opioid receptor-mediated PI turnover in hippocampal slices required extensive optimization of the assay. To increase further the sensitivity of the assay, the separation and quantitation of individual inositol phosphates by HPLC and radiochemical flow detection is proposed. An autosampler and radiochemical flow detector are requested to accomplish this aim. Finally, as a measure of G-protein coupling to kappa receptors, it is proposed to measure kappa receptor-stimulated, low Km GTPase in different areas of the brain, comparing the GTPase response with PI turnover and receptor distribution. The overall significance of the proposed work is that it will establish a link between kappa receptors and PI turnover, which previously has not been associated with any opioid receptor subtype. The work may provide evidence for a biochemical response underlying the behavioral effects of kappa receptor activation in the CNS.