Related Experiment Videos
Deciphering the role of Gi2 in opioid-induced adenylyl cyclase supersensitization
1Department of Biochemistry and the Biotechnology Research Institute, Hong Kong University of Science and Technology, Kowloon, China.
Abstract:
Prolonged opioid treatment of HEK 293 cells expressing opioid receptors are known to induce adenylyl cyclase supersensitization, a process that requires pertussis toxin (PTX)-sensitive G(i/o) proteins. Here, the role of Gi2 in adenylyl cyclase supersensitization was investigated. A PTX-insensitive G alpha(i2)/z chimera was stably co-expressed with mu-, kappa- or delta-opioid receptors in HEK 293 cells. Functional coupling of G alpha(i2)/z to the opioid receptors was demonstrated by opioid-induced inhibition of adenylyl cyclase and stimulation of ERK1/2 phosphorylation in PTX-treated cells. Chronic opioid treatment of each cell line led to adenylyl cyclase supersensitization but this response was blocked by PTX. Our results demonstrated that although PTX-sensitive G proteins are obligatory for opioid-induced adenylyl cyclase supersensitization, Gi2 alone was insufficient to mediate this response.
Insights
This study investigated G-protein coupled receptor signaling in HEK 293 cells. While G-protein alpha i2 (Gi2) is involved, it alone is insufficient for opioid-induced adenylyl cyclase supersensitization.
Area of Science:
- Cellular and Molecular Pharmacology
- G-Protein Coupled Receptor Signaling
- Neuropharmacology
Background:
- Prolonged opioid treatment of HEK 293 cells induces adenylyl cyclase supersensitization.
- This process relies on pertussis toxin (PTX)-sensitive G(i/o) proteins.
Purpose of the Study:
- To investigate the specific role of the Gi2 protein in mediating adenylyl cyclase supersensitization.
- To determine if Gi2 alone is sufficient for this opioid-induced cellular response.
Main Methods:
- Stable co-expression of a PTX-insensitive G alpha(i2)/z chimera with mu-, kappa-, or delta-opioid receptors in HEK 293 cells.
- Assessment of opioid-induced adenylyl cyclase inhibition and ERK1/2 phosphorylation.
- Evaluation of adenylyl cyclase supersensitization following chronic opioid treatment in PTX-treated and untreated cells.
Main Results:
- Functional coupling of the G alpha(i2)/z chimera to opioid receptors was confirmed via opioid-induced signaling in PTX-treated cells.
- Chronic opioid treatment induced adenylyl cyclase supersensitization in all cell lines.
- This supersensitization response was completely blocked by PTX treatment, indicating reliance on PTX-sensitive G proteins.
Conclusions:
- PTX-sensitive G proteins are essential for opioid-induced adenylyl cyclase supersensitization.
- Gi2, despite its involvement in opioid receptor signaling, is not sufficient on its own to mediate this supersensitization response.