Ryanodine receptor subtype 2 encodes Ca2+ oscillations activated by acetylcholine via the M2 muscarinic receptor/cADP-ribose signalling pathway in duodenum myocytes
Ryanodine receptor subtype 2 encodes Ca2+ oscillations activated by acetylcholine via the M2 muscarinic receptor/cADP-ribose signalling pathway in duodenum myocytes
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DOI：
10.1242/jcs.02344
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发表时间：
2005-05-15
影响因子：
4
通讯作者：
Morel, JL
中科院分区：
文献类型：
--
作者：
Fritz, N;Macrez, N;Morel, JL
在这项研究中，我们描述了乙酰胆碱激活的信号通路，它编码了大鼠十二指肠肌细胞中的钙振荡。这种振荡在去除外源钙离子后的完整心肌细胞中观察到，在膜电位取消后的通透性细胞中观察到，在肝素(肌醇1，4，5-三磷酸受体的抑制剂)的存在下观察到，但被兰诺定抑制，表明它们依赖于兰诺定受体从细胞内储存的钙释放。M_2受体拮抗剂甲氧曲明可选择性地抑制钙振荡。8-溴环腺苷二磷酸核糖和腺苷二磷酸核糖环化酶抑制剂(ZnCl2和抗CD38抗体)可抑制M受体激活的M2钙振荡。在通透性细胞中，通过测定烟酰胺鸟嘌呤二核苷酸环化产生的环鸟苷二磷核糖(一种荧光化合物)来评价乙酰胆碱对ADP-核糖环化酶活性的刺激作用。由于十二指肠肌细胞表达Ryanodine受体的三种亚型，反义策略显示Ryanodine受体亚型2单独启动乙酰胆碱诱导的钙振荡，环腺苷二磷酸核糖和雷帕霉素(一种诱导12/12.6 kDa FK506结合蛋白和Ryanodine受体解偶联的化合物)也需要启动。雷帕霉素处理后抑制环腺苷二磷核糖诱导的钙振荡，证实两种化合物都与兰尼定受体亚型2相互作用。我们的发现首次表明M2 M受体的激活通过激活环腺苷二磷核糖/FK506结合蛋白/兰诺受体2亚型信号通路而触发十二指肠肌细胞的钙振荡。
In this study, we characterized the signalling pathway activated by acetylcholine that encodes Ca2+ oscillations in rat duodenum myocytes. These oscillations were observed in intact myocytes after removal of external Ca2+, in permeabilized cells after abolition of the membrane potential and in the presence of heparin (an inhibitor of inositol 1,4,5-trisphosphate receptors) but were inhibited by ryanodine, indicating that they are dependent on Ca2+ release from intracellular stores through ryanodine receptors. Ca2+ oscillations were selectively inhibited by methoctramine (a M2 muscarinic receptor antagonist). The M2 muscarinic receptor-activated Ca2+ oscillations were inhibited by 8-bromo cyclic adenosine diphosphoribose and inhibitors of adenosine diphosphoribosyl cyclase (ZnCl2 and anti-CD38 antibody). Stimulation of ADP-ribosyl cyclase activity by acetylcholine was evaluated in permeabilized cells by measuring the production of cyclic guanosine diphosphoribose (a fluorescent compound), which resulted from the cyclization of nicotinamide guanine dinucleotide. As duodenum myocytes expressed the three subtypes of ryanodine receptors, an antisense strategy revealed that the ryanodine receptor subtype 2 alone was required to initiate the Ca2+ oscillations induced by acetylcholine and also by cyclic adenosine diphosphoribose and rapamycin (a compound that induced uncoupling between 12/12.6 kDa FK506-binding proteins and ryanodine receptors). Inhibition of cyclic adenosine diphosphoribose-induced Ca2+ oscillations, after rapamycin treatment, confirmed that both compounds interacted with the ryanodine receptor subtype 2. Our findings show for the first time that the M2 muscarinic receptor activation triggered Ca2+ oscillations in duodenum myocytes by activation of the cyclic adenosine diphosphoribose/FK506-binding protein/ryanodine receptor subtype 2 signalling pathway.