Varney M A, Rivera J, Lopez Bernal A, Watson S P
Department of Pharmacology, University of Oxford, U.K.
Biochem J. 1990 Jul 1;269(1):211-6. doi: 10.1042/bj2690211.
We have compared the properties of the [3H]Ins(1,4,5)P3-binding sites from a number of tissues in an attempt to determine if heterogeneity exists within the Ins(1,4,5)P3-receptor family. The binding of Ins(1,4,5)P3 was characterized in detail by using membranes prepared from human uterine smooth muscle and bovine adrenal cortex. Ins(1,4,5)P3 exhibited an approx. 5 times greater affinity for the binding site in adrenal cortex (KD = 9.81 +/- 1.92 nM) compared with uterine smooth muscle (KD = 37.1 +/- 1.8 nM). The binding was dependent on pH in both tissues, with a maximum at pH 8.3; at this pH various inositol phosphates and nucleotides competed for the binding sites with similar potencies on both tissues. However, the binding of Ins(1,4,5)P3 to the uterine smooth-muscle membranes was Ca2(+)-sensitive, whereas that to the bovine adrenal cortex was not; furthermore, heparin displaced the binding of Ins(1,4,5)P3 in the uterus with an IC50 value (concn. of displacer giving 50% inhibition of specific binding) of 3.9 micrograms/ml (2.5, 6.4; lower, upper range), compared with a value of 22 (13, 30) micrograms/ml in adrenal cortex. In view of the ability of Ins(1,4,5)P3 and heparin to distinguish between these binding sites, their effect on other tissues was examined. Ins(1,4,5)P3 showed a similar affinity for receptors located in the bovine cerebellum to those in the bovine adrenal cortex, but heparin displaced Ins(1,4,5)P3 binding with a 5-fold greater affinity from the cerebellum. Ins(1,4,5)P3 had a 2-fold greater affinity for its receptor with human platelets, as compared with human uterus, but heparin was unable to distinguish between these sites. In guinea-pig ileum, Ins(1,4,5)P3 displayed a similar affinity for the receptors in the longitudinal muscle compared with the circular muscle, but heparin could distinguish between these sites. These data show that small differences exist between tissues, but no clear picture is apparent. It is possible that these results reflect tissue-dependent factors such as phosphorylation, the presence of calmedin etc., rather than the presence of receptor subtypes or species difference.
我们比较了多个组织中[3H]肌醇-1,4,5-三磷酸（Ins(1,4,5)P3）结合位点的特性，以确定Ins(1,4,5)P3受体家族中是否存在异质性。通过使用从人子宫平滑肌和牛肾上腺皮质制备的膜，详细表征了Ins(1,4,5)P3的结合情况。与子宫平滑肌（解离常数KD = 37.1±1.8 nM）相比，Ins(1,4,5)P3对肾上腺皮质结合位点的亲和力约高5倍（KD = 9.81±1.92 nM）。两种组织中的结合均依赖于pH，在pH 8.3时达到最大值；在此pH下，各种肌醇磷酸酯和核苷酸在两种组织上以相似的效力竞争结合位点。然而，Ins(1,4,5)P3与子宫平滑肌膜的结合对Ca2+敏感，而与牛肾上腺皮质的结合则不敏感；此外，肝素以3.9微克/毫升（2.5，6.4；下限，上限范围）的半数抑制浓度（IC50值，即引起特异性结合50%抑制的置换剂浓度）置换子宫中Ins(1,4,5)P3的结合，而在肾上腺皮质中的值为22（13，30）微克/毫升。鉴于Ins(1,4,5)P3和肝素区分这些结合位点的能力，研究了它们对其他组织的影响。Ins(1,4,5)P3对牛小脑受体的亲和力与对牛肾上腺皮质受体的亲和力相似，但肝素从牛小脑置换Ins(1,4,5)P3结合的亲和力高5倍。与人类子宫相比，Ins(1,4,5)P3对人血小板受体的亲和力高2倍，但肝素无法区分这些位点。在豚鼠回肠中，Ins(1,4,5)P3对纵肌受体和环肌受体的亲和力相似，但肝素可以区分这些位点。这些数据表明组织之间存在微小差异，但尚无清晰的图景。这些结果可能反映了组织依赖性因素如磷酸化、钙调蛋白的存在等，而非受体亚型的存在或物种差异。
