Section of Biochemical Pharmacology, Department of Neuroscience, University of Cagliari, Cagliari, Italy.
Mol Pharmacol. 2012 Feb;81(2):154-65. doi: 10.1124/mol.111.075267. Epub 2011 Oct 26.
AMP-activated protein kinase (AMPK) and δ-opioid receptors (DORs) are both involved in controlling cell survival, energy metabolism, and food intake, but little is known on the interaction between these two signaling molecules. Here we show that activation of human DORs stably expressed in Chinese hamster ovary (CHO) cells increased AMPK activity and AMPK phosphorylation on Thr172. DOR-induced AMPK phosphorylation was prevented by pertussis toxin, reduced by protein kinase A (PKA) activators, and unaffected by PKA, transforming growth factor-β-activated kinase 1, mitogen-activated protein kinase, and protein kinase C inhibitors. Conversely, the DOR effect was reduced by Ca(2+)/calmodulin-dependent protein kinase kinase (CaMKK) inhibition, apyrase treatment, G(q/11) antagonism, and blockade of P2 purinergic receptors. Apyrase treatment also depressed DOR stimulation of intracellular Ca(2+) concentration, whereas P2 receptor antagonism blocked DOR stimulation of inositol phosphate accumulation. In SH-SY5Y neuroblastoma cells and primary olfactory bulb neurons, DOR activation failed to affect AMPK phosphorylation per se but potentiated the stimulation by either muscarinic agonists or 2-methyl-thio-ADP. Sequestration of G protein βγ subunits (Gβγ) blocked the DOR potentiation of AMPK phosphorylation induced by oxotremorine-M. In CHO cells, the AMPK activator 5-aminoimidazole-4-carboxamide1-β-D-ribonucleoside stimulated AMPK phosphorylation and glucose uptake, whereas pharmacological inhibition of AMPK, expression of a dominant-negative mutant of AMPKα1, and P2Y receptor blockade reduced DOR-stimulated glucose uptake. The data indicate that in different cell systems, DOR activation up-regulates AMPK through a Gβγ-dependent synergistic interaction with G(q/11)-coupled receptors, potentiating Ca(2+) release and CaMKKβ-dependent AMPK phosphorylation. In CHO cells, this coincident signaling mechanism is involved in DOR-induced glucose uptake.
AMP 激活的蛋白激酶 (AMPK) 和 δ 阿片受体 (DOR) 都参与控制细胞存活、能量代谢和食物摄入，但对于这两种信号分子之间的相互作用知之甚少。在这里，我们展示了在中华仓鼠卵巢 (CHO) 细胞中稳定表达的人 DOR 的激活增加了 AMPK 活性和 Thr172 上的 AMPK 磷酸化。百日咳毒素可防止 DOR 诱导的 AMPK 磷酸化，蛋白激酶 A (PKA) 激活剂可减少 DOR 诱导的 AMPK 磷酸化，而 PKA、转化生长因子-β 激活激酶 1、丝裂原激活蛋白激酶和蛋白激酶 C 抑制剂则不受影响。相反，DOR 的作用被 Ca2+/钙调蛋白依赖性蛋白激酶激酶 (CaMKK) 抑制、apyrase 处理、G(q/11) 拮抗剂和 P2 嘌呤能受体阻断所减弱。apyrase 处理还抑制了 DOR 对细胞内 Ca2+浓度的刺激，而 P2 受体拮抗剂则阻断了 DOR 对肌醇磷酸盐积累的刺激。在 SH-SY5Y 神经母细胞瘤细胞和原代嗅球神经元中，DOR 的激活本身不会影响 AMPK 磷酸化，但增强了毒蕈碱激动剂或 2-甲基硫代-ADP 的刺激作用。G 蛋白 βγ 亚基 (Gβγ) 的隔离阻断了 oxotremorine-M 诱导的 DOR 对 AMPK 磷酸化的增强作用。在 CHO 细胞中，AMPK 激活剂 5-氨基咪唑-4-甲酰胺 1-β-D-核糖核苷刺激 AMPK 磷酸化和葡萄糖摄取，而 AMPK 的药理学抑制、AMPKα1 的显性负突变体的表达和 P2Y 受体阻断减少了 DOR 刺激的葡萄糖摄取。数据表明，在不同的细胞系统中，DOR 的激活通过与 G(q/11) 偶联受体协同作用的 Gβγ 依赖性机制上调 AMPK，增强 Ca2+释放和 CaMKKβ 依赖性 AMPK 磷酸化。在 CHO 细胞中，这种并发信号机制参与了 DOR 诱导的葡萄糖摄取。
