Related Experiment Videos
Cyclic AMP-induced desensitization of G-protein-regulated phospholipase C in turkey erythrocyte membranes
1Department of Pharmacology, School of Medicine, University of North Carolina, Chapel Hill 27599, USA.
Abstract:
The interaction of the cyclic AMP and inositol lipid signalling systems was studied in turkey erythrocytes. Elevation of intracellular cyclic AMP concentrations by pretreatment of the cells with forskolin or 8-Br-cAMP resulted in a marked decrease in responsiveness of phospholipase C to G-protein activators in membranes prepared from treated cells. Decreases in responsiveness occurred with a t1/2 of approximately 5 min and were reversible after transfer of desensitized cells to drug-free medium. Pretreatment of the cells with forskolin inhibited inositol phosphate formation in a concentration-dependent manner and addition of the phosphodiesterase inhibitor IBMX 93-isobutyl-1-methylxanthine) during pretreatment increased the capacity of forskolin to desensitize phospholipase C activity. IBMX also produced a similar potentiation of forskolin-stimulated accumulation of cyclic AMP in turkey erythrocytes. Isoproterenol pretreatment of the cells induced, like forskolin, partial inhibition of inositol phosphate generation in response to G-protein activators and to P2y purinoceptor and beta-adrenoceptor agonists. The capacity of isoproterenol to induce desensitization of phospholipase C activity also was increased by the presence of IBMX during pretreatment of the cells. H8 (N-[2-(methylamino)ethyl]-5-isoquinoline-sulfonamide), an inhibitor of cyclic AMP-regulated protein kinase, completely prevented forskolin-induced desensitization but only partially blocked isoproterenol-induced desensitization. These results indicate that the cyclic AMP signalling cascade has a major inhibitory influence on receptor- and G-protein-activated inositol lipid signaling.
Insights
Elevated cyclic AMP levels inhibit inositol lipid signaling in turkey red blood cells. This desensitization of phospholipase C is reversible and mediated by cyclic AMP-dependent protein kinase.
Area of Science:
- Cellular signaling pathways
- Biochemistry
- Pharmacology
Background:
- Cyclic AMP and inositol lipid signaling are crucial cellular communication systems.
- Understanding their interaction is key to deciphering complex cellular responses.
- Dysregulation of these pathways is implicated in various diseases.
Purpose of the Study:
- To investigate the interaction between cyclic AMP and inositol lipid signaling systems.
- To determine the effect of elevated cyclic AMP on phospholipase C activity.
- To elucidate the role of cyclic AMP-dependent protein kinase in this cross-talk.
Main Methods:
- Turkey erythrocytes were pretreated with forskolin or 8-Br-cAMP to elevate cyclic AMP.
- Phospholipase C responsiveness to G-protein activators was measured in cell membranes.
- Inositol phosphate formation was assessed following various agonist stimulations.
- The effect of H8, a protein kinase inhibitor, was evaluated.
Main Results:
- Elevated cyclic AMP significantly decreased phospholipase C responsiveness to G-protein activators.
- This desensitization was concentration-dependent, reversible, and potentiated by phosphodiesterase inhibition (IBMX).
- Both forskolin and isoproterenol induced partial inhibition of inositol phosphate generation.
- H8 completely prevented forskolin-induced desensitization but only partially blocked isoproterenol-induced desensitization.
Conclusions:
- The cyclic AMP signaling cascade exerts a major inhibitory influence on receptor- and G-protein-activated inositol lipid signaling.
- Cyclic AMP-dependent protein kinase plays a significant role in mediating this inhibitory cross-talk.
- These findings provide insights into the complex regulation of cellular signaling networks.