THERAPEUTIC PROFILES OF OPIOID DELTA RECEPTORS--NEW MODALITIES

这个项目的目标是基于三个假设。 假设1阿片受体可能以 子类型。在NG 108-15中将采用反义策略 在原代培养的DRG和抗伤害性模型中的细胞 在活体内。反义寡核苷酸(ODN)“敲除” 将使用ODN研究NG108-15细胞中的Delta受体 针对DOR-1克隆和 阿片受体(保守的阿片受体； COR)。细胞摄取和ODN转位，时间进程， β受体的浓度依赖性和抑制作用 表达将用荧光显微镜和 放射性配基结合；将重点介绍控制ODN序列。 在DRG培养中，将使用DOR或COR反义ODN来评估 ODN摄取与形态分布及DOR的验证 ODN对Delta Mu和Kappa受体的选择性。在老鼠身上， DOR/COR ODN将用于阐明脊髓的分布 ODN与阿片受体的“击倒”及其可能 假定的三角洲亚型选择性的差异敏感性 DOR-1反义ODN激动剂(但不是Mu或Kappa激动剂) 的影响以及时间进程和恢复 抗伤害作用。假设2是激活 脊髓上阿片受体将产生抗伤害作用 不一定依赖于所描述的下行路径 用于阿片类u受体激动剂。局部微量注射 阿片亚型选择性激动剂和拮抗剂入选 将利用脊髓上的基因座来建立选择性受体 抗伤害性作用。下行通路的参与 在确认的棘上三角区激活后 抗伤害性作用的研究将通过检查抑制 脊髓伤害性行为和c-fos的表达 对照大鼠的脊髓，或有脊髓损伤的动物的脊髓 背外侧索(DLF)。假设3是激活 将导致脊髓上和脊髓阿片三角洲受体的 具有协同的抗伤害作用。分析 脊柱上/脊柱联合给药引起的抗伤害效应 亚型选择性激动剂在固定比例的范例中 将使用等同相学方法。这些实验应该 提供了关于三角洲作用的重要新信息 伤害性感受中的受体及其亚型的易感性 体外和体内受体。

The aims of this project are based on three hypothesis. Hypothesis 1 is that the opioid delta receptor may exist as subtypes. An antisense strategy will be employed in NG 108-15 cells, in primary DRG cultures and in an antinociceptive model in vivo. Antisense oligodeoxynucleotide (ODN) "knock-down" of delta receptors in NG108-15 cells will be studied using ODNs directed at the DOR-1 clone and at a conserved sequence in the opioid mu, delta and kappa receptors (conserved opioid receptor; COR). Cellular uptake and ODN translocation, time-course, concentration-dependence and inhibition of the delta receptor expression will be determined with fluorescence microscopy and radioligand binding; control ODN sequences will be emphasized. In DRG cultures DOR or COR antisense ODN will be used to evaluate ODN uptake and morphological distribution and to validate the DOR ODN selectivity against delta mu and kappa receptors. In rats, DOR/COR ODNs will be used to elucidate the distribution of spinal ODNs and "knock-down" of opioid receptors and possible differential sensitivity of putative delta subtype selective agonists (but not mu or kappa agonists), to DOR-1 antisense ODN effects as well as time-course and recovery of the antinociceptive effect. Hypothesis 2 is that activation of supraspinal opioid delta receptors will produce antinociception not necessarily dependent on the descending pathway characterized for opioid mu receptor agonists. Localized microinjection of opioid subtype selective agonists and antagonists into selected supraspinal loci will be employed to establish receptor selective antinociceptive activity. The involvement of descending pathways following activation of confirmed supraspinal delta antinociception will be studied by examining suppression of nociceptive induced behavior and c-FOS expression in the spinal cord in control rats, or in animals with lesions of the dorsolateral funiculus (DLF). Hypothesis 3 is that activation of both supraspinal and spinal opioid delta receptors will result in a synergistic antinociceptive effect. Analysis of antinociception elicited by supraspinal/spinal co-administration of subtype-selective agonists in a fixed-ratio paradigm with isobolographic methods will be used. These experiments should yield significant new information about the role of the delta receptor in nociception and the bossibility of subtypes of this receptor in vitro and in vivo.