Unterwald E M, Anton B, To T, Lam H, Evans C J
Department of Psychiatry, New York University Medical Center, New York, USA.
Neuroscience. 1998 Aug;85(3):897-905. doi: 10.1016/s0306-4522(97)00659-3.
The present study utilized a newly developed quantitative immunohistochemical assay to measure changes in mu opioid receptor abundance following chronic administration of the opioid receptor antagonist naltrexone. These data were compared with those obtained from mu receptor radioligand binding on adjacent tissue sections, in order to determine whether the characteristic antagonist-induced increase in radioligand binding is due to an increase in the total number of mu receptors and/or to an increase in the proportion of receptors that are in an active binding conformation in the absence of a change in the total number of receptors. Adult male Sprague-Dawley rats were administered naltrexone, 7-8 mg/kg per day, or saline continuously for seven days by osmotic minipumps, after which time their brains were processed for immunohistochemistry and receptor autoradiography on adjacent fresh frozen tissue sections. Semiquantitative immunohistochemistry was performed using a radiolabelled secondary antibody for autoradiographic determination and a set of radioactive standards. Results demonstrate an overall concordance between the distribution of mu opioid receptors as measured by the two different methods with a few exceptions. Following naltrexone administration, mu receptor immunoreactivity was significantly higher in the amygdala, thalamus, hippocampus, and interpeduncular nucleus as compared with the saline-treated control animals. [3H]D-Ala2,N-Me-Phe4,Gly-ol5-enkephalin binding to mu opioid receptors was significantly higher in the globus pallidus, amygdala, thalamus, hypothalamus, hippocampus, substantia nigra, ventral tegmental area, central gray, and interpeduncular nucleus of the naltrexone-treated rats. These findings indicate that in some brain regions chronic naltrexone exposure increases the total number of mu opioid receptors, while in other regions there is an increase in the percent of active receptors without an observable change in the total number of receptors. Quantitative receptor immunodetection together with ligand autoradiography provides a new approach for investigating the regulation of mu opioid receptors on tissue sections.
本研究采用一种新开发的定量免疫组织化学检测方法，来测量阿片受体拮抗剂纳曲酮长期给药后μ阿片受体丰度的变化。将这些数据与在相邻组织切片上进行的μ受体放射性配体结合实验所获得的数据进行比较，以确定拮抗剂诱导的放射性配体结合增加这一特征，是由于μ受体总数增加，和/或在受体总数不变的情况下，处于活性结合构象的受体比例增加所致。成年雄性Sprague-Dawley大鼠通过渗透微型泵连续7天给予纳曲酮（每天7 - 8毫克/千克）或生理盐水，之后将它们的大脑用于免疫组织化学和相邻新鲜冷冻组织切片上的受体放射自显影。使用放射性标记的二抗进行半定量免疫组织化学，用于放射自显影测定和一组放射性标准品。结果表明，除少数例外情况外，两种不同方法测量的μ阿片受体分布总体一致。与生理盐水处理的对照动物相比，给予纳曲酮后，杏仁核、丘脑、海马体和脚间核中的μ受体免疫反应性显著更高。在纳曲酮处理的大鼠的苍白球、杏仁核、丘脑、下丘脑、海马体、黑质、腹侧被盖区、中央灰质和脚间核中，[³H]D-Ala²，N-Me-Phe⁴，Gly-ol⁵-脑啡肽与μ阿片受体的结合显著更高。这些发现表明，在某些脑区，长期暴露于纳曲酮会增加μ阿片受体的总数，而在其他区域，活性受体的百分比增加，而受体总数没有明显变化。定量受体免疫检测与配体放射自显影相结合，为研究组织切片上μ阿片受体的调节提供了一种新方法。
