Kawai N, Yamamoto T, Yamamoto H, McCarron R M, Spatz M
Stroke Branch, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892-4128, USA.
Neurochem Int. 1997 Oct;31(4):597-605. doi: 10.1016/s0197-0186(97)00018-1.
This report describes the effects of endothelins (ET-1 and ET-3) on ion transport systems expressed on cultured rat brain capillary endothelial cells (RBEC) and includes investigation of pharmacological properties of ET receptors, their reactivity and induction of signal transduction pathways. ET-1 stimulated IP3 formation and Ca2+ uptake with half-maximal effective concentrations (EC50) of 0.68 and 0.93 nM, respectively; the effects of ET-3 on these responses were much weaker. ET-1-stimulated IP3 formation and Ca2+ uptake were inhibited by an ETA antagonist (BQ123) and a phospholipase C (PLC) inhibitor (U73122), indicating the presence of ETA receptors coupled to PLC. ET-1 stimulated K+ efflux (through a quinine-sensitive mechanism) and K+ uptake (through both ouabain-sensitive and bumetanide-sensitive mechanisms) with EC50 of 0.59 and 0.68 nM, respectively. The potencies of ET-3 on these responses were considerably lower than those of ET-1. By contrast, ET-1 or ET-3 stimulated Na+ uptake with similarly high potencies (EC50 = 0.80 and 1.89 nM, respectively) through EIPA (a Na+/H+ exchange inhibitor)-sensitive mechanisms. ET-stimulated K+ efflux, K+ uptake and Na+ uptake activities were all inhibited by BQ123 (but not by BQ788), suggesting the involvement of ETA (and not ETB) receptors in all these responses. ET-1 stimulated K+ uptake and efflux were inhibited by either U73122 or an intracellular Ca2+ chelator, suggesting that these two responses were mediated via PLC. In contrast, ET stimulation of Na+ uptake was unaffected by PLC inhibition or intracellular Ca2+ chelation. These data suggest the presence of two distinct subtypes of ETA receptors on RBEC; one appears to be a typical ETA receptor which is coupled to PLC and has higher binding affinity for ET-1 than ET-3. The other (ETA-like) receptor is similarly activated by ET-1 and ET-3 with high potencies but is independent of PLC. This possibility was further confirmed by the [125I]ET-1 binding studies demonstrating the presence of high- and low-affinity ET-3 binding sites.
本报告描述了内皮素（ET - 1和ET - 3）对培养的大鼠脑毛细血管内皮细胞（RBEC）上表达的离子转运系统的影响，并包括对ET受体的药理学特性、其反应性及信号转导途径诱导的研究。ET - 1刺激肌醇三磷酸（IP3）形成和钙离子摄取，其半数最大有效浓度（EC50）分别为0.68和0.93纳摩尔；ET - 3对这些反应的作用则弱得多。ET - 1刺激的IP3形成和钙离子摄取被ETA拮抗剂（BQ123）和磷脂酶C（PLC）抑制剂（U73122）抑制，表明存在与PLC偶联的ETA受体。ET - 1刺激钾离子外流（通过一种对奎宁敏感的机制）和钾离子摄取（通过对哇巴因敏感和对布美他尼敏感的机制），EC50分别为0.59和0.68纳摩尔。ET - 3对这些反应的效力明显低于ET - 1。相比之下，ET - 1或ET - 3通过对乙基异丙基氨氯吡啶酸（EIPA，一种钠/氢交换抑制剂）敏感的机制以相似的高效力刺激钠离子摄取（EC50分别为0.80和1.89纳摩尔）。ET刺激的钾离子外流、钾离子摄取和钠离子摄取活性均被BQ123抑制（但不被BQ788抑制），表明所有这些反应中涉及ETA（而非ETB）受体。ET - 1刺激的钾离子摄取和外流被U73122或细胞内钙离子螯合剂抑制，表明这两种反应是通过PLC介导的。相反，ET刺激的钠离子摄取不受PLC抑制或细胞内钙离子螯合的影响。这些数据表明RBEC上存在两种不同亚型的ETA受体；一种似乎是典型的ETA受体，它与PLC偶联，对ET - 1的结合亲和力高于ET - 3。另一种（类ETA）受体被ET - 1和ET - 3以相似的高效力激活，但独立于PLC。[125I]ET - 1结合研究进一步证实了这种可能性，该研究证明存在高亲和力和低亲和力的ET - 3结合位点。
