Nagaoka Miyuki, Nara Masayuki, Tamada Tsutomu, Kume Hiroaki, Oguma Tetsuya, Kikuchi Toshiaki, Zaini Jamal, Moriya Takuya, Ichinose Masakazu, Tamura Gen, Hattori Toshio
Division of Infectious and Respiratory Diseases, Department of Internal Medicine, Tohoku University School of Medicine, Aoba-ku, Sendai, Japan.
Respir Physiol Neurobiol. 2009 Mar 31;166(1):61-7. doi: 10.1016/j.resp.2009.02.002. Epub 2009 Feb 14.
We examined the effects of extracellular adenosine 5'-triphosphate (ATP) on single airway smooth muscle (ASM) cells from porcine trachea using a patch-clamp technique. ATP induced a sustained inward current. Phospholipase C inhibitor U-73122 failed to inhibit the current, suggesting the involvement of P2X receptor. A specific effecter of P2X(4), ivermectin, augmented the current indicating the existence of P2X(4) receptors. Immunohistochemistry and reverse transcription/polymerase chain reaction analysis and Western blot analysis also showed the distribution of the P2X(4) receptors. The inward current was reduced by SKF-96365, an inhibitor of both voltage-dependent Ca(2+) channels (VDCCs) and voltage-independent Ca(2+) channels, although a VDCC antagonist, verapamil, did not affect the current. SKF-96365 caused complete suppression of both the increase in the intracellular Ca(2+) concentration and the contraction of ASM cells induced by ATP. Our results demonstrate that P2X(4) receptors exist on ASM and that the receptors are responsible for Ca(2+) influx. These findings suggest that the Ca(2+) influx regulated by P2X(4) receptors plays an important role in ASM contraction by a pathway distinct from VDCC.
我们使用膜片钳技术研究了细胞外5'-三磷酸腺苷（ATP）对猪气管单个气道平滑肌（ASM）细胞的影响。ATP诱导了持续的内向电流。磷脂酶C抑制剂U-73122未能抑制该电流，提示P2X受体参与其中。P2X(4)的特异性效应物伊维菌素增强了该电流，表明存在P2X(4)受体。免疫组织化学、逆转录/聚合酶链反应分析和蛋白质印迹分析也显示了P2X(4)受体的分布。电压依赖性钙通道（VDCCs）和非电压依赖性钙通道的抑制剂SKF-96365降低了内向电流，尽管VDCC拮抗剂维拉帕米对该电流没有影响。SKF-96365完全抑制了ATP诱导的细胞内钙浓度升高和ASM细胞收缩。我们的结果表明，ASM上存在P2X(4)受体，且这些受体负责钙内流。这些发现提示，由P2X(4)受体调节的钙内流通过一条不同于VDCC的途径在ASM收缩中起重要作用。
