Samways Damien S K, Li Wen-hong, Conway Stuart J, Holmes Andrew B, Bootman Martin D, Henderson Graeme
Department of Pharmacology, University of Bristol, University Walk, Bristol BS8 1TD, UK.
Biochem J. 2003 Nov 1;375(Pt 3):713-20. doi: 10.1042/BJ20030508.
Activation of G(i)/G(o)-coupled opioid receptors increases [Ca2+]i (intracellular free-Ca2+ concentration), but only if there is concomitant G(q)-coupled receptor activation. This G(i)/G(o)-coupled receptor-mediated [Ca2+]i increase does not appear to result from further production of Ins P3 [Ins(1,4,5) P3] in SH-SY5Y cells. In the present study, fast-scanning confocal microscopy revealed that activation of mu-opioid receptors alone by 1 muM DAMGO ([L-Ala, NMe-Phe, Gly-ol]-enkephalin) did not stimulate the Ins P3-dependent elementary Ca2+-signalling events (Ca2+ puffs), whereas DAMGO did evoke Ca2+ puffs when applied during concomitant activation of M3 muscarinic receptors with 1 muM carbachol. We next determined whether mu-opioid receptor activation might increase [Ca2+]i by sensitizing the Ins P3 receptor to Ins P3. DAMGO did not potentiate the amplitude of the [Ca2+]i increase evoked by flash photolysis of the caged Ins P3 receptor agonist, caged 2,3-isopropylidene-Ins P3, whereas the Ins P3 receptor sensitizing agent, thimerosal (10 muM), did potentiate this response. DAMGO also did not prolong the rate of decay of the increase in [Ca2+]i evoked by flash photolysis of caged 2,3-isopropylidene-Ins P3. Furthermore, DAMGO did not increase [Ca2+]i in the presence of the cell-membrane-permeable Ins P3 receptor agonist, Ins P3 hexakis(butyryloxymethyl) ester. Therefore it appears that mu-opioid receptors do not increase [Ca2+]i through either Ins P3 receptor sensitization, enhancing the releasable pool of Ca2+ or inhibition of Ca2+ removal from the cytoplasm.
G(i)/G(o)偶联的阿片受体激活会增加细胞内游离钙离子浓度（[Ca2+]i），但前提是同时存在G(q)偶联受体激活。这种由G(i)/G(o)偶联受体介导的[Ca2+]i增加似乎并非源于SH-SY5Y细胞中肌醇三磷酸（Ins P3 [Ins(1,4,5) P3]）的进一步生成。在本研究中，快速扫描共聚焦显微镜显示，单独用1 μM的DAMGO（[L-丙氨酸，N-甲基苯丙氨酸，甘醇]-脑啡肽）激活μ-阿片受体会不会刺激依赖Ins P3的基本钙离子信号事件（钙离子瞬变），而当在1 μM卡巴胆碱同时激活M3毒蕈碱受体时应用DAMGO，确实会引发钙离子瞬变。接下来，我们确定μ-阿片受体激活是否可能通过使Ins P3受体对Ins P3敏感来增加[Ca2+]i。DAMGO不会增强由笼锁型Ins P3受体激动剂笼锁型2,3-异亚丙基-Ins P3的闪光光解所引发的[Ca2+]i增加幅度，而Ins P3受体敏化剂硫柳汞（10 μM）确实会增强这种反应。DAMGO也不会延长由笼锁型2,3-异亚丙基-Ins P3的闪光光解所引发的[Ca2+]i增加的衰减速率。此外，在存在细胞膜可渗透的Ins P3受体激动剂Ins P3六（丁酰氧基甲基）酯的情况下，DAMGO不会增加[Ca2+]i。因此，似乎μ-阿片受体不会通过Ins P3受体敏化、增加可释放钙离子池或抑制钙离子从细胞质中移除来增加[Ca2+]i。
