HORMONAL MODULATION OF BONE ADENYLATE CYCLASE

本研究的长期目标是了解 糖皮质激素过多、甲状腺功能亢进 激素过量和雌激素缺乏导致骨质减少， 镁缺乏导致骨骼抵抗吸收 刺激。 在拟议的研究中，我们将测试假设， 这些条件引起的变化，在监管性质的 骨腺苷酸环化酶 其中大约三分之二 调查将涉及详细的特征， 腺苷酸环化酶存在于从骨中获得的膜中， 正常成年豚鼠 这将需要：1）确定 PTH、异丙肾上腺素和前列腺素E2激动剂的作用 激活骨腺苷酸环化酶，对Mg++的Ka和对 高、低亲和力钙抑制的Ki值 先前在酶复合物中鉴定的位点; 2） 确定酶复合物中Mg++ 结合激活发生;和3）识别内的位点 其中Ca++结合抑制腺苷酸的酶复合物 环化酶活性和机制的研究， 出现Ca++抑制。 重构方法， 来自火鸡红细胞和S49细胞周期小鼠的质膜 淋巴瘤细胞和骨膜的胆酸盐提取物将 在这两项研究中使用。 可能发挥的作用 抑制性鸟嘌呤核苷酸调节蛋白，Ni，在 还将评价腺苷酸环化酶的钙抑制。 骨膜将用百日咳毒素预处理， 钙抑制参数 腺苷酸环化酶活性在处理的膜相比， 对照膜。 一旦获得这一基线数据， 腺苷酸环化酶的调节特性 Mg++和激动剂激活和Ca++抑制将 在从暴露于以下物质的动物中获得的骨膜中进行了检查： 体内：1）糖皮质激素过量，2）甲状腺激素过量，3） 雌激素缺乏和4）镁缺乏。 此外，本发明还提供了一种方法， 糖皮质激素或甲状腺激素过量的影响， 雌激素对镁缺乏的相对量 刺激性和抑制性鸟嘌呤核苷酸调节蛋白， 将测定骨膜中的Ns和Ni。

The long-term objectives of this research is to understand the mechanisms by which glucocorticoid hormone excess, thyroid hormone excess and estrogen deficiency lead to osteopenia and magnesium deficiency leads to skeletal resistance to resorptive stimuli. In the proposed study we will test the hypothesis that these conditions induce changes in the regulatory properties of bone adenylate cyclase. Approximately two-thirds of this investigation will involve the detailed characterization of the adenylate cyclase present in membranes obtained from bones of normal adult guinea pigs. This will entail: 1) determination of the effects of PTH, isoproterenol, and prostaglandin E2, agonists which activate bone adenylate cyclase, on the Ka for Mg++ and on the Ki for calcium of the high and low affinity calcium inhibition sites previously identified in the enzyme complex; 2) identification of the sites within the enzyme complex where Mg++ binding activation occurs; and 3) identification of the sites within the enzyme complex where Ca++ binding inhibits adenylate cyclase activity and investigation of the mechanism by which Ca++ inhibition occurs. Reconstitution methodology employing plasma membranes from turkey erythrocytes and S49 cyc-murine lymphoma cells and cholate extracts from bone membranes will be used in these latter two studies. The possible role of the inhibitory guanine nucleotide regulatory protein, Ni, in the calcium inhibition of the adenylate cyclase will also be evaluated. Bone membranes will be pretreated with pertussis toxin to inactivate Ni and the parameters of calcium inhibition of the adenylate cyclase activity in the treated membranes compared to control membranes. Once this baseline data has been obtained, the regulatory characteristics of adenylate cyclase with respect to Mg++ and agonist activation and Ca++ inhibition will be examined in bone membranes obtained from animals exposed in vivo to: 1) glucocorticoid excess, 2) thyroid hormone excess, 3) estrogen deficiency, and 4) magnesium deficiency. In addition, the effects of glucocorticoid or thyroid hormone excess and estrogen on magnesium deficiency on the relative amounts of the stimulatory and inhibitory guanine nucleotide regulatory proteins, Ns and Ni, in the bone membranes will be determined.