Roy C
Eur J Biochem. 1984 Sep 3;143(2):243-50. doi: 10.1111/j.1432-1033.1984.tb08365.x.
LLC-PK1L cells, a kidney-derived cell line, had sustained growth in a defined medium. When compared to the parent cell line growing with 10% fetal bovine serum, LLC-PK1L cells had about 100-times fewer vasopressin receptors. Upon modifications of the cell culture medium, the vasopressin response of the adenylate cyclase could be increased by more than 10-fold with a parallel increase in vasopressin receptor number. Using cells with high or low receptor densities, the stimulatory and inhibitory effects of N6-L-2-phenylisopropyl-adenosine on the modulation of the adenylate cyclase responsiveness to vasopressin were investigated. When high concentrations of GTP were added, low concentrations of phenylisopropyladenosine inhibited the enzyme, while higher concentrations were found to be stimulatory. The adenylate cyclase activity stimulated by vasopressin could only be inhibited by phenylisopropyladenosine under these conditions in membranes with high receptor density; only the increase in enzyme activity due to high GTP concentration was inhibitable. The analysis of the dependency of the adenylate cyclase activity as a function of the vasopressin concentration showed that, besides reducing the maximum velocity of the system for vasopressin, the addition of phenylisopropyladenosine generated an heterogeneity in the adenylate cyclase response to vasopressin (as judged by a curvilinear Eadie plot). A high-affinity component in the adenylate cyclase response appeared when phenylisopropyladenosine was added. The growth of the cells in a medium containing adenosine deaminase gave results identical to those obtained for control cells. However, growing the cells with both phenylisopropyladenosine and adenosine deaminase abolished the inhibitory effects of the former on the adenylate cyclase and greatly reduced its stimulatory action. Under these conditions, the vasopressin response of the adenylate cyclase was not further regulated by phenylisopropyladenosine. These results indicate a role of adenosine on vasopressin response, especially at low physiological concentrations of the hormone where a high-affinity component of the hormonal response could be demonstrated.
LLC-PK1L细胞是一种源自肾脏的细胞系，在特定培养基中能持续生长。与在含10%胎牛血清的培养基中生长的亲代细胞系相比，LLC-PK1L细胞的血管加压素受体数量约少100倍。对细胞培养基进行改良后，腺苷酸环化酶对血管加压素的反应可增加10倍以上，同时血管加压素受体数量也会相应增加。利用具有高或低受体密度的细胞，研究了N6-L-2-苯基异丙基腺苷对腺苷酸环化酶对血管加压素反应性调节的刺激和抑制作用。当加入高浓度的GTP时，低浓度的苯基异丙基腺苷会抑制该酶，而较高浓度则具有刺激作用。在这些条件下，只有在高受体密度的膜中，血管加压素刺激的腺苷酸环化酶活性才会被苯基异丙基腺苷抑制；只有由于高GTP浓度导致的酶活性增加才是可抑制的。对腺苷酸环化酶活性与血管加压素浓度关系的分析表明，除了降低血管加压素系统的最大反应速度外，加入苯基异丙基腺苷还会使腺苷酸环化酶对血管加压素的反应产生异质性（通过曲线伊迪图判断）。加入苯基异丙基腺苷后，腺苷酸环化酶反应中出现了一个高亲和力成分。在含有腺苷脱氨酶的培养基中培养细胞，得到的结果与对照细胞相同。然而，同时用苯基异丙基腺苷和腺苷脱氨酶培养细胞，消除了前者对腺苷酸环化酶的抑制作用，并大大降低了其刺激作用。在这些条件下，苯基异丙基腺苷不再进一步调节腺苷酸环化酶对血管加压素的反应。这些结果表明腺苷在血管加压素反应中起作用，特别是在激素生理浓度较低时，此时可证明激素反应存在高亲和力成分。
