Intrinsic relative activities of κ opioid agonists in activating Gα proteins and internalizing receptor: Differences between human and mouse receptors.
Intrinsic relative activities of κ opioid agonists in activating Gα proteins and internalizing receptor: Differences between human and mouse receptors.
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DOI：
10.1016/j.ejphar.2015.05.054
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发表时间：
2015-08-15
影响因子：
5
通讯作者：
Liu-Chen LY
中科院分区：
文献类型：
--
作者：
DiMattio KM;Ehlert FJ;Liu-Chen LY
一些研究人员最近发现了偏性阿片受体（KOP受体）激动剂。然而，尚未发表关于可用的KOP受体激动剂对人和小鼠KOP受体（分别为hKOP受体和mKOP受体）的功能选择性的全面研究。在这里，我们研究了超过20种KOP受体激动剂激活G蛋白和内化受体的能力。使用用hKOP受体或mKOP受体稳定转染的克隆neuro-2a小鼠神经母细胞瘤（N2 a）细胞。我们采用激动剂诱导的[35 S]GTPγS结合和KOP受体内化分别作为G蛋白和β-arrestin通路激活的指标。Ehlert及其同事的方法用于量化G蛋白活化（RAi-G）和受体内化（RAi-I）的内在相对活性以及两者之间的功能选择性程度[Log RAi-G-Log RAi-I，RAi-G/RAi-I和偏倚因子]。参数RAi表示激发给定应答的活性受体状态的激动剂亲和力的相对估计。内源性配体强啡肽A（1-17）被指定为平衡配体，偏倚因子为1。有趣的是，我们发现在功能选择性方面存在物种差异。最显著的差异是12-表-鼠尾草素A、U69，593和ICI-199，441。12-表-鼠尾草素A在mKOP受体上高度偏向内化，但在hKOP受体上明显偏向G蛋白。U69，593对mKOP受体的内化偏好性远大于对hKOP受体的内化偏好性。ICI 199，441在mKOP受体上表现出内化倾向，在hKOP受体上表现出G蛋白倾向。所观察到的物种差异的可能机制进行了讨论。
Several investigators recently identified biased opioid receptor (KOP receptor) agonists. However, no comprehensive study of the functional selectivity of available KOP receptor agonists at the human and mouse KOP receptors (hKOP receptor and mKOP receptor, respectively) has been published. Here we examined the ability of over 20 KOP receptor agonists to activate G proteins and to internalize the receptor. Clonal neuro-2a mouse neuroblastoma (N2a) cells stably transfected with the hKOP receptor or mKOP receptor were used. We employed agonist-induced [35S]GTPγS binding and KOP receptor internalization as measures of activation of G protein and β-arrestin pathways, respectively. The method of Ehlert and colleagues was used to quantify intrinsic relative activities at G protein activation (RAi−G) and receptor internalization (RAi−I) and the degree of functional selectivity between the two [Log RAi−G − Log RAi−I, RAi−G/RAi−I and bias factor]. The parameter, RAi, represents a relative estimate of agonist affinity for the active receptor state that elicits a given response. The endogenous ligand dynorphin A (1–17) was designated as the balanced ligand with a bias factor of 1. Interestingly, we found that there were species differences in functional selectivity. The most striking differences were for 12-epi-salvinorin A, U69,593, and ICI-199,441. 12-Epi-salvinorin A was highly internalization-biased at the mKOP receptor, but apparently G protein-biased at hKOP receptor. U69,593 was much more internalization-biased at mKOP receptor than hKOP receptor. ICI199,441 showed internalization-biased at the mKOP receptor and G protein-biased at the hKOP receptor. Possible mechanisms for the observed species differences are discussed.