Identification of P2-purinergic receptors

1)PC12嗜铬细胞瘤细胞具有P2受体，P2受体与钙离子内流和儿茶酚胺释放有关。在腺酰环化酶激活剂Forsklin存在下，ATP类似物如ATP和2-甲硫基ATP以浓度依赖的方式抑制环磷酸腺苷的积累。用百日咳毒素完全阻断卡巴胆碱的作用，对ATP的作用没有影响。此外，我们还发现，在胞吐刺激后，ADP核糖化因子从胞浆部分移位到膜上。新型三磷酸腺苷受体的克隆有待研究。2)一氧化氮(NO)调节神经递质的释放。在此之前，我们报道了5-亚硝基-半胱氨酸在体内和体外都能刺激海马区去甲肾上腺素的释放。在PC12细胞中，S亚硝基半胱氨酸对去甲肾上腺素的释放有抑制作用。其他促进环状GMP积累的NO化合物则没有作用。S-亚硝基-半胱氨酸可抑制三磷酸腺苷刺激的钙通道内钙内流，而S-亚硝基-半胱氨酸可刺激细胞内咖啡因敏感的钙池内钙动员。NO从钙池中动员Ca~(2+)不是一个充分的因素，其他刺激释放的因素可能是负调控的。3)已知脂多糖类或细胞因子通过表达诱导型一氧化氮合酶(INOS)来刺激大鼠神经胶质细胞亚硝酸盐的产生。共同加入内皮素可减少刺激剂对iNOS的表达和亚硝酸盐的积累。相反，内皮素或三磷酸腺苷预先处理24 h可促进iNOS的表达。内皮素或三磷酸腺苷的刺激作用分别由ET-B或P2受体介导。蛋白激酶C抑制剂可抑制内皮素和三磷酸腺苷促进的iNOS表达。刺激ATP受体可诱导神经胶质细胞中核因子NFkB的激活。

1) PC12 pheochromocytoma cells have P2 receptors which are coupled to Ca2+influx and catecholamine release. In the presence of forskolin, an activator of adenylyl cyclase, ATP analogs such as ATP and 2-methylthio ATP inhibited cyclic AMP accumulation in a concentration-dependent manner. Treatment with pertussis toxin, which completely abolished the effect of carbachol, had no effect on the action of ATP.In addition, we found that ADP-ribosylation factors translocate to membranes from the cytosol fraction after exocytotic stimulation. The cloning of a new type of ATP receptor remains to be determined.2) Nitric oxide (NO) modulates the release of neurotransmitters. Previously we reported that 5-nitroso-cysteine stimulated noradrenaline release from hippocampus in vivo and in vitro. In PC12 cells, S-nitroso-cysteine did inhibit noradrenaline release. Other NO compounds, which increased cyclic GMP accumulation, had no effect. ATP-stimulated Ca2+ influx via Ca2+ channels was inhibited in PC12 cells treated with S-Nitroso-cysteine, although S-nitroso-cysteine stimulated Ca2+ mobilization from intracellular caffeine-sensitive Ca2+-pools. Ca2+ mobilization by NO from Ca2+ pools was not a sufficient factor, and other factors stimulating release may be regulated negatively.3) Lipopolysaccaride or cytokines are known to stimulate production of nitrite via expression of inducible NO synthase (iNOS) in rat glial cells. Co-addition of endothelin decreased iNOS expression and nitrite accumulation by stimulants. In contrast, pretreatment with endothelin or ATP for 24 h enhanced iNOS expression. The stimulatory effect by endothelin or ATP was mediated by ET-B or P2 receptors, respectively. A protein kinase C inhibitor suppressed endothelin- and ATP-enhanced iNOS expression. Stimulation of ATP receptors induced activation of a nuclear factor, NFkB in glial cells.