G(I)-MEDIATED AND PROTEIN-KINASE-C-MEDIATED HETEROLOGOUS POTENTIATIONOF PHOSPHOLIPASE-C SIGNALING BY G-PROTEIN-COUPLED RECEPTORS

We recently reported that activation of the highly efficient phospholi pase C (PLC) stimulatory m3 muscarinic acetylcholine receptor (mAChR) can induce a long-lasting G(i)-mediated heterologous potentiation of P LC stimulation in human embryonic kidney (HEK) 293 cells, which was ac companied by an increased cellular level of the PLC substrate phosphat idylinositol-4,5-bisphosphate [PtdIns(4,5)P-2]. Here, we examined whet her such a potentiated PLC response is also induced by the rather poor ly PLC stimulatory m2 mAChR and the endogenously expressed purinergic and lysophosphatidic acid receptors. Pretreatment of m2 mAChR-expressi ng HEK 293 cells for 2 min with carbachol, followed by agonist washout and measurement of PLC activity greater than or equal to 40 min later , caused a long-lasting (up to similar to 90 min) heterologous potenti ation of receptor-and G protein-mediated PLC stimulation. A similar he terologous potentiation of receptor-mediated PLC stimulation was induc ed by short term activation of lysophosphatidic acid and purinergic re ceptors. Either of the three receptor agonists increased the cellular level of PtdIns(4,5)P-2 by similar to 50%. The mAChR-induced PLC poten tiation was fully prevented by either pertussis toxin or the protein k inase C (PKC) inhibitors staurosporine and Go 6976, which did not affe ct acute PLC stimulation. On the other hand, the rise in PtdIns(4,5)P- 2 was prevented only by combined treatment of HEK 293 cells with pertu ssis toxin and PKC inhibitors. In conclusion, we demonstrated that act ivation of poorly PLC stimulatory receptors can also induce a long-las ting G(i)-mediated heterologous potentiation of PLC signaling in HEK 2 93 cells and that this novel PLC regulatory process is under the contr ol of PKC.