Ross R A, Brockie H C, Stevenson L A, Murphy V L, Templeton F, Makriyannis A, Pertwee R G
Department of Biomedical Sciences, Institute of Medical Sciences, University of Aberdeen, Scotland.
Br J Pharmacol. 1999 Feb;126(3):665-72. doi: 10.1038/sj.bjp.0702351.
We have tested our prediction that AM630 is a CB2 cannabinoid receptor ligand and also investigated whether L759633 and L759656, are CB2 receptor agonists. Binding assays with membranes from CHO cells stably transfected with human CB1 or CB2 receptors using [3H]-CP55940, confirmed the CB2-selectivity of L759633 and L759656 (CB2/CB1 affinity ratios = 163 and 414 respectively) and showed AM630 to have a Ki at CB2 receptors of 31.2 nM and a CB2/CB1 affinity ratio of 165. In CB2-transfected cells, L759633 and L759656 were potent inhibitors of forskolin-stimulated cyclic AMP production, with EC50 values of 8.1 and 3.1 nM respectively and CB1/CB2 EC50 ratios of > 1000 and > 3000 respectively. AM630 inhibited [35S]-GTPgammaS binding to CB2 receptor membranes (EC50 = 76.6 nM), enhanced forskolin-stimulated cyclic AMP production in CB2-transfected cells (5.2 fold by 1 microM), and antagonized the inhibition of forskolin-stimulated cyclic AMP production in this cell line induced by CP55940. In CB1-transfected cells, forskolin-stimulated cyclic AMP production was significantly inhibited by AM630 (22.6% at 1 microM and 45.9% at 10 microM) and by L759633 at 10 microM (48%) but not 1 microM. L759656 (10 microM) was not inhibitory. AM630 also produced a slight decrease in the mean inhibitory effect of CP55940 on cyclic AMP production which was not statistically significant. We conclude that AM630 is a CB2-selective ligand that behaves as an inverse agonist at CB2 receptors and as a weak partial agonist at CB1 receptors. L759633 and L759656 are both potent CB2-selective agonists.
我们验证了AM630是一种CB2大麻素受体配体的预测，并研究了L759633和L759656是否为CB2受体激动剂。使用[3H]-CP55940对稳定转染人CB1或CB2受体的CHO细胞膜进行结合试验，证实了L759633和L759656对CB2的选择性（CB2/CB1亲和力比值分别为163和414），并表明AM630对CB2受体的Ki为31.2 nM，CB2/CB1亲和力比值为165。在转染CB2的细胞中，L759633和L759656是福司可林刺激的环磷酸腺苷（cAMP）产生的强效抑制剂，EC50值分别为8.1和3.1 nM，CB1/CB2的EC50比值分别>1000和>3000。AM630抑制[35S]-GTPγS与CB2受体膜的结合（EC50 = 76.6 nM），增强转染CB2的细胞中福司可林刺激的cAMP产生（1 μM时增加5.2倍），并拮抗CP55940对该细胞系中福司可林刺激的cAMP产生的抑制作用。在转染CB1的细胞中，福司可林刺激的cAMP产生受到AM630（1 μM时为22.6%，10 μM时为45.9%）和10 μM的L759633（48%）的显著抑制，但1 μM时不受抑制。L759656（10 μM）无抑制作用。AM630还使CP55940对cAMP产生的平均抑制作用略有降低，但无统计学意义。我们得出结论，AM630是一种CB2选择性配体，在CB2受体上表现为反向激动剂，在CB1受体上表现为弱部分激动剂。L759633和L759656均为强效CB2选择性激动剂。
