Neal Charles R, Owens Constance E, Taylor Larry P, Hoversten Mary T, Akil Huda, Watson Stanley J
Mental Health Research Institute, University of Michigan Medical Center, 205 Zina Pitcher Place, Ann Arbor, MI 48109-0720, USA.
J Chem Neuroanat. 2003 Jul;25(4):233-47. doi: 10.1016/s0891-0618(03)00032-2.
Utilizing agonist-stimulated GTPgammaS autoradiography, we analyzed the ability of preproorphanin FQ (ppOFQ) peptides to stimulate [35S]-GTPgammaS binding in adult rat brain. Orphanin FQ (OFQ) stimulated [35S]-GTPgammaS binding in a pattern similar to that described for [125I]-OFQ at the endogenous opioid receptor-like (ORL1) receptor. The ppOFQ peptides nocistatin and orphanin FQ2 (OFQ II(1-17)) had no effect, suggesting that they do not mediate their reported analgesic effects via a G(i/o)-coupled receptor (i.e. opioid or ORL1). Unlike OFQ II(1-17), high concentrations of its C-terminal extension, OFQ II(1-28), stimulated [35S]-GTPgammaS binding in a mu (mu) opioid receptor-like distribution and the effect was blocked by naloxone. To explore these observations, we evaluated the receptor binding profile of OFQ II(1-28) at the cloned ORL1 and mu opioid receptors. OFQ II(1-28) had no specific binding at either ORL1 or mu opioid receptors at concentrations up to 50 microM. This lack of affinity was not consistent with a mu-mediated effect, as suggested by preliminary observation using functional autoradiography in rat brain sections. Although behavioral studies suggest that OFQ II(1-28) possesses analgesic activity, this effect does not appear to be mediated via direct binding at the mu opioid receptor. Taken together, these findings support the view that (1) OFQ is the only ppOFQ peptide that binds to and activates the ORL1 receptor and (2) OFQ II(1-28) does not bind or stimulate [35S]-GTPgammaS binding in cells expressing the mu opioid receptor.
利用激动剂刺激的GTPγS放射自显影技术，我们分析了前阿片样肽FQ（ppOFQ）肽刺激成年大鼠脑中[35S]-GTPγS结合的能力。孤啡肽FQ（OFQ）刺激[35S]-GTPγS结合的模式与内源性阿片受体样（ORL1）受体上[125I]-OFQ所描述的模式相似。ppOFQ肽nocistatin和孤啡肽FQ2（OFQ II(1-17)）没有作用，这表明它们并非通过G(i/o)偶联受体（即阿片受体或ORL1）介导其已报道的镇痛作用。与OFQ II(1-17)不同，其C末端延伸肽高浓度的OFQ II(1-28)以μ（mu）阿片受体样分布刺激[35S]-GTPγS结合，且该效应被纳洛酮阻断。为探究这些观察结果，我们评估了OFQ II(1-28)在克隆的ORL1和μ阿片受体上的受体结合情况。OFQ II(1-28)在浓度高达50μM时，在ORL1或μ阿片受体上均无特异性结合。这种缺乏亲和力与μ介导的效应不一致，大鼠脑切片中功能放射自显影的初步观察结果表明了这一点。尽管行为学研究表明OFQ II(1-28)具有镇痛活性，但这种效应似乎并非通过直接结合μ阿片受体介导。综上所述，这些发现支持以下观点：（1）OFQ是唯一能结合并激活ORL1受体的ppOFQ肽；（2）OFQ II(1-28)在表达μ阿片受体的细胞中不结合或刺激[35S]-GTPγS结合。
