Schmidt M, Lohmann B, Hammer K, Haupenthal S, Nehls M V, Jakobs K H
Institut für Pharmakologie, Universitätsklinikum Essen, D-45122 Essen, Germany.
Mol Pharmacol. 1998 Jun;53(6):1139-48.
We recently reported that activation of the highly efficient phospholipase C (PLC) stimulatory m3 muscarinic acetylcholine receptor (mAChR) can induce a long-lasting Gi-mediated heterologous potentiation of PLC stimulation in human embryonic kidney (HEK) 293 cells, which was accompanied by an increased cellular level of the PLC substrate phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P2]. Here, we examined whether such a potentiated PLC response is also induced by the rather poorly PLC stimulatory m2 mAChR and the endogenously expressed purinergic and lysophosphatidic acid receptors. Pretreatment of m2 mAChR-expressing HEK 293 cells for 2 min with carbachol, followed by agonist washout and measurement of PLC activity >/=40 min later, caused a long-lasting (up to approximately 90 min) heterologous potentiation of receptor- and G protein-mediated PLC stimulation. A similar heterologous potentiation of receptor-mediated PLC stimulation was induced by short term activation of lysophosphatidic acid and purinergic receptors. Either of the three receptor agonists increased the cellular level of PtdIns(4,5)P2 by approximately 50%. The mAChR-induced PLC potentiation was fully prevented by either pertussis toxin or the protein kinase C (PKC) inhibitors staurosporine and Gö 6976, which did not affect acute PLC stimulation. On the other hand, the rise in PtdIns(4,5)P2 was prevented only by combined treatment of HEK 293 cells with pertussis toxin and PKC inhibitors. In conclusion, we demonstrated that activation of poorly PLC stimulatory receptors can also induce a long-lasting Gi-mediated heterologous potentiation of PLC signaling in HEK 293 cells and that this novel PLC regulatory process is under the control of PKC.
我们最近报道，高效磷脂酶C（PLC）刺激性毒蕈碱型乙酰胆碱受体（mAChR）m3的激活可在人胚肾（HEK）293细胞中诱导由Gi介导的PLC刺激的持久异源增强，这伴随着PLC底物磷脂酰肌醇-4,5-二磷酸[PtdIns(4,5)P2]细胞水平的升高。在此，我们研究了这种增强的PLC反应是否也由PLC刺激性较差的m2 mAChR以及内源性表达的嘌呤能和溶血磷脂酸受体诱导。用卡巴胆碱预处理表达m2 mAChR的HEK 293细胞2分钟，然后冲洗激动剂并在≥40分钟后测量PLC活性，可导致受体和G蛋白介导的PLC刺激的持久（长达约90分钟）异源增强。溶血磷脂酸和嘌呤能受体的短期激活也可诱导受体介导的PLC刺激的类似异源增强。三种受体激动剂中的任何一种均可使PtdIns(4,5)P2的细胞水平升高约50%。百日咳毒素或蛋白激酶C（PKC）抑制剂星形孢菌素和Gö 6976可完全阻止mAChR诱导的PLC增强，而这两种抑制剂不影响急性PLC刺激。另一方面，只有通过将HEK 293细胞与百日咳毒素和PKC抑制剂联合处理才能阻止PtdIns(4,5)P2的升高。总之，我们证明了PLC刺激性较差的受体的激活也可在HEK 293细胞中诱导由Gi介导的PLC信号的持久异源增强，并且这种新的PLC调节过程受PKC的控制。
