Yoburn B C, Gomes B A, Rajashekara V, Patel C, Patel M
Department of Pharmaceutical Sciences, College of Pharmacy and Allied Health Professions, St. John's University, Queens, New York 11439, USA.
Synapse. 2003 Feb;47(2):109-16. doi: 10.1002/syn.10149.
Although opioid receptors are G-protein coupled, the role that specific G-protein subunits play in the development of opioid tolerance and the regulation of opioid receptor number is not well understood. In the present study, we used a G((i)alpha2) antisense oligodeoxynucleotide (ODN) to examine the contribution of G((i)alpha2) proteins to mu-opioid tolerance and receptor downregulation in the mouse. Mice were injected intracerebroventricularly (ICV) and into the spinal intrathecal space (IT) for 4-5 consecutive days (30 microg/site/day), with an antisense ODN or a mismatch ODN directed at mRNA for the G((i)alpha2) subunit of G-proteins. Controls were treated with dH(2)O. On the second day of ODN treatment continuous subcutaneous (SC) infusion of etorphine (200 microg/kg/day) or morphine (40 mg/kg/day + 25 mg pellet) was begun. Control mice were implanted with inert placebo pellets. Three days later, pumps and pellets were removed and mice were tested for morphine analgesia or mu-opioid receptor density was determined in whole brain. Etorphine produced significant tolerance (ED(50) shift = approximately 11-fold) and downregulation of mu-opioid receptors (approximately 25%). Morphine treatment produced significant tolerance (ED(50) shift approximately 9-fold), but no mu-opioid receptor downregulation. Antisense treatment reduced G((i)alpha2) protein levels in striatum and spinal cord by approximately 25%. G((i)alpha2) antisense reduced the acute potency of morphine. G((i)alpha2) antisense blocked the development of tolerance to morphine treatment and reduced the development of tolerance to etorphine treatment. Antisense did not have any effect on etorphine-induced mu-opioid receptor downregulation. In another experiment, 7-day treatment with morphine or etorphine similarly increased G((i)alpha2) mRNA and protein abundance in spinal cord. Overall, these results support an important role for G((i)alpha2)-protein in the acute effects of opioids and opioid tolerance. However, G((i)alpha2) is not required for agonist-induced mu-opioid receptor density regulation in vivo.
尽管阿片受体是G蛋白偶联受体，但特定G蛋白亚基在阿片耐受形成及阿片受体数量调节中所起的作用尚未完全明确。在本研究中，我们使用G（i）α2反义寡脱氧核苷酸（ODN）来研究G（i）α2蛋白在小鼠μ-阿片耐受及受体下调中的作用。小鼠连续4 - 5天经脑室（ICV）和脊髓鞘内（IT）注射（30μg/部位/天）针对G蛋白G（i）α2亚基mRNA的反义ODN或错配ODN。对照组用蒸馏水处理。在ODN处理的第二天开始持续皮下（SC）输注埃托啡（200μg/kg/天）或吗啡（40mg/kg/天 + 25mg药粒）。对照小鼠植入惰性安慰剂药粒。三天后，取出泵和药粒，检测小鼠的吗啡镇痛效果或测定全脑μ-阿片受体密度。埃托啡产生显著耐受（半数有效剂量（ED50）变化约11倍）和μ-阿片受体下调（约25%）。吗啡处理产生显著耐受（ED50变化约9倍），但无μ-阿片受体下调。反义处理使纹状体和脊髓中的G（i）α2蛋白水平降低约25%。G（i）α2反义降低了吗啡的急性效力。G（i）α2反义阻断了对吗啡处理的耐受形成，并降低了对埃托啡处理的耐受形成。反义对埃托啡诱导的μ-阿片受体下调无任何影响。在另一实验中，吗啡或埃托啡7天处理同样增加了脊髓中G（i）α2 mRNA和蛋白丰度。总体而言，这些结果支持G（i）α2蛋白在阿片类药物的急性效应及阿片耐受中起重要作用。然而，体内激动剂诱导的μ-阿片受体密度调节并不需要G（i）α2。
