Interactions of cholecystokinin and mu opioid receptors

在本研究中，我们共同注射编码μ阿片受体和CCK-B受体的cDNA到非洲爪蟾卵母细胞作为模型系统，以检查受体拮抗作用在单细胞水平。 还注射了内向整流钾通道GIRK的cDNA作为mu受体激活的报告基因。 其他人报道CCK-B受体刺激Ca++激活的Cl-通道。 在我们的手中，这种反应发生在不到20%的卵母细胞中，并且出现过一次，再也没有在卵母细胞中出现过。 CCK-B受体更频繁的反应通过GIRK钾通道偶联，如观察结果所示：（1）电流仅在将编码GIRK的cDNA注入卵母细胞时出现，以及（2）电流被钡阻断。 GIRK对CCK的反应使受体脱敏，需要一个多小时才能完全恢复。 相比之下，由阿片激动剂去甲吗啡引起的脱敏在10分钟内完全恢复。 CCK对去甲吗啡和去甲吗啡对CCK的配对试验表明，两个系统相互交叉脱敏。 初始CCK反应越大，其对mu反应的脱敏作用越大。 此外，CCK试验后立即恢复mu反应的时间接近1小时，这是CCK而不是去甲吗啡脱敏的时间特征。 这种相互抑制作用提示了三种可能的相互作用：（1）两种受体结合在一起形成异源二聚体，其中一个亚基直接调节另一个亚基的敏感性;（2）下游第二信使与受体的相互作用，（G蛋白亚基）介导一种受体的作用，抑制另一种受体;或者（3）一种受体的激活启动了一个整体脱敏过程，作为一种附带反应，该过程部分地使另一种受体脱敏。 我们目前正在努力区分这些假设。

In the present study, we co-injected cDNA encoding both the mu opioid receptor and the CCK-B receptor into Xenopus oocytes as a model system to examine receptor antagonism at the single cell level. The cDNA for the inwardly rectifying potassium channel GIRK was also injected as a reporter for the activation of the mu receptor. Others have reported that the CCK-B receptor stimulates a Ca++ activated Cl- channel. In our hands, this response occurred in less than 20% of the oocytes and, having appeared once, never reappeared in that oocyte. The more frequent response of the CCK-B receptor was coupled through the GIRK potassium channel, as demonstrated by the observations (1) that current only appeared when the cDNA encoding GIRK was injected into an oocyte, and (2) that the current was blocked by barium. The GIRK response to CCK desensitized the receptor and required well over an hour to recover fully. In contrast, recovery from the desensitization caused by an opioid agonist normorphine was complete within ten minutes. Pairing tests of CCK after normorphine and normorphine after CCK showed that the two systems cross-desensitized each other. The greater the initial CCK response, the greater its desensitization of the mu response. Furthermore, the time for recovery of the mu response immediately following a CCK test was close to an hour, the time characteristic of CCK, not normorphine, desensitization. The reciprocal inhibition suggests three possible interactions: (1) that the two receptors bind together as a heterodimer in which one subunit directly regulates the sensitivity of the other; (2) that downstream second messengers (G protein subunits) that mediate actions of one receptor inhibit the other; or (3) that the activation of one receptor initiates a global desensitization process that partially desensitizes the other as a collateral reaction. We are currently working to distinguish among these hypotheses.