Chueh S H, Hsu L S, Song S L
Department of Biochemistry, National Defense Medical Center, Taipei, Taiwan, R.O.C.
Mol Pharmacol. 1994 Mar;45(3):532-9.
The ATP signaling mechanism in neuroblastoma x glioma hybrid NG108-15 cells differentiated by exposure to dibutyryl-cAMP was characterized. In cells loaded with fura-2, ATP rapidly raised the cytosolic Ca2+ concentration ([Ca2+]i); the magnitude of the rise was inversely proportional to the extracellular Na+ concentration. Large increases in cytosolic Na+ concentration, measured with the fluorescent Na+ indicator sodium-binding benzofuran isophthalate, were dose-dependently elicited by ATP. ATP also evoked the entry of ethidium bromide into cells, and this process was inhibited by Mg2+. Inositol-1,4,5-trisphosphate (IP3) generation induced by ATP was totally blocked by removal of extracellular Ca2+, but residual IP3 generation still remained in nondifferentiated cells. In addition, ATP produced a concentration-, time-, and Mg(2+)-dependent biphasic uptake of 45Ca2+. A range of nucleotides and ATP analogues, including CTP, UTP, and GTP, induced only 9-29% of the ATP response. However, adenosine 5'-thiotriphosphate evoked 79% of ATP-induced 45Ca2+ uptake. 45Ca2+ uptake elicited by ATP could be potently blocked by purinoceptor antagonists, but other tested reagents less effectively blocked the action of ATP. When bradykinin was used as an agonist, the [Ca2+]i rise was transient and was insensitive to the extracellular Na+ concentration. Na+ influx, entry of ethidium bromide, and 45Ca2+ uptake were unaffected by bradykinin. Furthermore, bradykinin-evoked IP3 generation was insensitive to extracellular Ca2+. Neither ATP nor bradykinin had any effect on cAMP levels within cells. These data suggest that ATP induces a [Ca2+]i rise in differentiated NG108-15 cells via two distinct Ca2+ influx mechanisms, i.e., a receptor-operated cation channel and pores formed by ATP4-. These mechanisms are distinct from those elicited by bradykinin.
对经二丁酰环磷腺苷（dibutyryl - cAMP）诱导分化的神经母细胞瘤x胶质瘤杂交NG108 - 15细胞中的ATP信号传导机制进行了表征。在装载了fura - 2的细胞中，ATP迅速升高胞质Ca2 +浓度（[Ca2 +]i）；升高幅度与细胞外Na +浓度呈反比。用荧光Na +指示剂邻苯二甲酸苯并呋喃钠（sodium - binding benzofuran isophthalate）测量发现，ATP能剂量依赖性地引起胞质Na +浓度大幅升高。ATP还能促使溴化乙锭进入细胞，且该过程受到Mg2 +的抑制。ATP诱导的肌醇 - 1,4,5 - 三磷酸（IP3）生成在去除细胞外Ca2 +后完全被阻断，但在未分化细胞中仍有残余的IP3生成。此外，ATP产生了浓度、时间和Mg(2 +)依赖性的45Ca2 +双相摄取。一系列核苷酸和ATP类似物，包括CTP、UTP和GTP，仅诱导出ATP反应的9 - 29%。然而，腺苷5'-硫代三磷酸（adenosine 5'-thiotriphosphate）诱发了ATP诱导的45Ca2 +摄取的79%。ATP诱导的45Ca2 +摄取可被嘌呤受体拮抗剂有效阻断，但其他测试试剂对ATP作用的阻断效果较差。当使用缓激肽作为激动剂时，[Ca2 +]i升高是短暂的，且对细胞外Na +浓度不敏感。缓激肽不影响Na +内流、溴化乙锭进入细胞以及45Ca2 +摄取。此外，缓激肽诱发的IP3生成对细胞外Ca2 +不敏感。ATP和缓激肽对细胞内cAMP水平均无任何影响。这些数据表明，ATP通过两种不同的Ca2 +内流机制，即受体操纵的阳离子通道和由ATP4 -形成的孔道，诱导分化的NG108 - 15细胞中[Ca2 +]i升高。这些机制与缓激肽引发的机制不同。
