Chen C C, Chang J, Chen W C
Institute of Pharmacology, College of Medicine, National Taiwan University, Taipei.
Glia. 1996 Mar;16(3):210-7. doi: 10.1002/(SICI)1098-1136(199603)16:3<210::AID-GLIA3>3.0.CO;2-5.
Cultured astrocytes express bradykinin (BK) receptors, which are coupled to phospholipase C (PLC) through G-protein to mediate phosphoinositide (PI) hydrolysis. The regulation of this BK receptor-G protein-PLC pathway by cAMP and endothelin-1 (ET-1) was explored by short-term (20 min) and long-term (24 h) treatment with 100 mu M dibutyryl cyclic AMP (dBcAMP) or 10 nM ET-1. Short-term treatment of cells with dBcAMP had no effect on BK-induced PI hydrolysis; however, long-term treatment resulted in potentiation of the BK response. Similar effects were seen after 10 mu M forskolin pretreatment of the cells. We further explored the site of action of 24 h dBcAMP pretreatment and found that AlF(4)-, ionomycin- or A3187-induced PI hydrolysis was not affected but (3H)BK binding was increased. These results indicate that the site of action of dBcAMP is the BK receptor and Scatchard plot analysis showed that the Bmax was increased but the Kd decreased. Cycloheximide (0.5 mu M) blocked the increase in (3H)BK binding, indicating that new synthesis of receptor protein might occur during 24 h pretreatment with dBcAMP. Twenty minutes pretreatment of cells with ET-1 resulted in desensitization of the ET-1 induced P1 response, while the BK response was unaffected. After 24 h pretreatment with ET-1, desensitization to ET-1 still occurred, while BK-induced PI hydrolysis was markedly potentiated. (3H)BK binding and AlF(4)--induced but not A23187- or ionomycin-induced PI hydrolysis were increased, indicating that the site of action of long-term ET-1 treatment was the BK receptor and G protein; Scatchard analysis showed an increase in Bmax but no effect on Kd. These effects were blocked by cycloheximide, indicating that new synthesis of both receptor protein and G protein might occur during 24 h pretreatment with ET-1. (3H)Thymidine uptake was inhibited or potentiated by dBcAMP and ET-1, respectively. Possible dBcAMP-induced differentiation and ET-1-induced proliferation may contribute to the increased expression of receptor proteins.
培养的星形胶质细胞表达缓激肽（BK）受体，该受体通过G蛋白与磷脂酶C（PLC）偶联，介导磷酸肌醇（PI）水解。通过用100μM二丁酰环磷酸腺苷（dBcAMP）或10 nM内皮素-1（ET-1）进行短期（20分钟）和长期（24小时）处理，探讨了cAMP和内皮素-1（ET-1）对该BK受体-G蛋白-PLC途径的调节作用。用dBcAMP对细胞进行短期处理对BK诱导的PI水解没有影响；然而，长期处理导致BK反应增强。在用10μM福司可林预处理细胞后也观察到类似的效果。我们进一步探讨了24小时dBcAMP预处理的作用位点，发现AlF(4)-离子霉素或A3187诱导的PI水解不受影响，但（3H）BK结合增加。这些结果表明dBcAMP的作用位点是BK受体，Scatchard图分析表明Bmax增加但Kd降低。放线菌酮（0.5μM）阻断了（3H）BK结合的增加，表明在用dBcAMP进行24小时预处理期间可能发生了受体蛋白的新合成。用ET-1对细胞进行20分钟预处理导致ET-1诱导的P1反应脱敏，而BK反应不受影响。用ET-1进行24小时预处理后，对ET-1的脱敏仍然存在，而BK诱导的PI水解明显增强。（3H）BK结合以及AlF(4)-诱导的而非A23187或离子霉素诱导的PI水解增加，表明长期ET-1处理的作用位点是BK受体和G蛋白；Scatchard分析表明Bmax增加但对Kd没有影响。这些作用被放线菌酮阻断，表明在用ET-1进行24小时预处理期间可能发生了受体蛋白和G蛋白的新合成。（3H）胸腺嘧啶摄取分别被dBcAMP抑制或被ET-1增强。可能的dBcAMP诱导的分化和ET-1诱导的增殖可能有助于受体蛋白表达的增加。
