Wu Ning, Su Rui-Bin, Xu Bo, Lu Xin-Qiang, Liu Yin, Zheng Jian-Quan, Piletz John E, Li Jin, Qin Bo-Yi
Beijing Institute of Pharmacology and Toxicology, Beijing 100850, China.
Biochem Pharmacol. 2005 Oct 1;70(7):1079-87. doi: 10.1016/j.bcp.2005.07.006.
Agmatine, an endogenous ligand for the I1-imidazoline receptor, has previously been shown to prevent morphine dependence in rats and mice. To investigate the role of imidazoline receptor antisera-selected protein (IRAS), a strong candidate for I1R, in morphine dependence, two CHO cell lines were created, in which mu opioid receptor (MOR) was stably expressed alone (CHO-mu) or MOR and IRAS were stably co-expressed (CHO-mu/IRAS). After 48 h administration of morphine (10 microM), naloxone induced a cAMP overshoot in both cell lines, suggesting cellular morphine dependence had been produced. Agmatine (0.1-2.5 microM) concentration-dependently inhibited the naloxone-precipitated cAMP overshoot when co-pretreated with morphine in CHO-mu/IRAS, but not in CHO-mu. Agmatine at 5-100 microM also inhibited the cAMP overshoot in CHO/mu and CHO-mu/IRAS. Efaroxan, an I1R-preferential antagonist, completely blocked the effect of agmatine on the cAMP overshoot at 0.1-2.5 microM in CHO-mu/IRAS, while partially reversing the effects of agmatine at 5-100 microM. L-type calcium channel blocker nifedipine entirely mimicked the effects of agmatine at high concentrations on forskolin-stimulated cAMP formation in CHO-mu and naloxone-precipitated cAMP overshoot in morphine-pretreated CHO-mu. Therefore, IRAS, in the co-transfected CHO-mu/IRAS cell line, appears necessary for low concentrations of agmatine to cause attenuation of cellular morphine dependence. An additional effect of agmatine at higher concentrations seems to relate to both transfected IRAS and some naive elements in CHO cells, and L-type voltage-gated calcium channels are not ruled out. This study suggests that IRAS mediates agmatine's high affinity effects on cellular morphine dependence and may play a role in opioid dependence.
胍丁胺是一种咪唑啉I1受体的内源性配体，此前已被证明可预防大鼠和小鼠的吗啡依赖。为了研究咪唑啉受体抗血清选择蛋白（IRAS）（一种I1受体的有力候选蛋白）在吗啡依赖中的作用，构建了两种CHO细胞系，其中μ阿片受体（MOR）单独稳定表达（CHO-μ）或MOR和IRAS稳定共表达（CHO-μ/IRAS）。给予吗啡（10μM）48小时后，纳洛酮在两种细胞系中均诱导了cAMP超调，表明已产生细胞吗啡依赖。在CHO-μ/IRAS中，与吗啡共同预处理时，胍丁胺（0.1 - 2.5μM）浓度依赖性地抑制了纳洛酮诱发的cAMP超调，但在CHO-μ中则无此作用。5 - 100μM的胍丁胺也抑制了CHO/μ和CHO-μ/IRAS中的cAMP超调。I1受体选择性拮抗剂依酚氯铵在0.1 - 2.5μM时完全阻断了胍丁胺对CHO-μ/IRAS中cAMP超调的作用，而在5 - 100μM时部分逆转了胍丁胺的作用。L型钙通道阻滞剂硝苯地平在高浓度时完全模拟了胍丁胺对CHO-μ中福司柯林刺激的cAMP形成以及对吗啡预处理的CHO-μ中纳洛酮诱发的cAMP超调的作用。因此，在共转染的CHO-μ/IRAS细胞系中，IRAS似乎是低浓度胍丁胺减轻细胞吗啡依赖所必需的。胍丁胺在较高浓度时的额外作用似乎与转染的IRAS和CHO细胞中的一些天然成分有关，并且不排除L型电压门控钙通道的作用。本研究表明，IRAS介导了胍丁胺对细胞吗啡依赖的高亲和力作用，可能在阿片类药物依赖中发挥作用。
