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Chronic morphine augments adenylyl cyclase phosphorylation: relevance to altered signaling during
S Chakrabarti1, L Wang, W J Tang
1Department of Biochemistry, State University of New York, Health Science Center at Brooklyn, Brooklyn, New York 11203, USA.
Abstract:
Despite the demonstration that chronic morphine increases phosphorylation of multiple substrate proteins, their identity has, for the most part, remained elusive. Thus far, chronic morphine has not been shown to increase the phosphorylation of any identified effector protein. This is the first demonstration that persistent activation of opioid receptors has profound effects on phosphorylation of adenylyl cyclase (AC). A dramatic increase in phosphorylation of AC (type II family) was observed in ileum longitudinal muscle myenteric plexus preparations obtained from chronic morphine-treated guinea pigs. Analogous results were obtained when AC was immunoprecipitated using two differentially directed AC antibodies. The magnitude of the augmented AC phosphorylation was substantially attenuated by chelerythrine, a protein kinase C-selective inhibitor. These results suggest the potential relevance of increased phosphorylation (protein kinase C-mediated) of AC to opioid tolerant/dependent mechanisms. Because phosphorylation of AC isoforms (type II family) can significantly increase their stimulatory responsiveness to Gsalpha and Gbetagamma, this mechanism could underlie, in part, the predominance of opioid AC stimulatory signaling observed in opioid tolerant/dependent tissue. Moreover, in light of the fact that many G protein-coupled receptors signal through common effector proteins, this effect provides a mechanism for divergent consequences of chronic morphine treatment and could explain the well documented complexity of changes that accompany the opioid tolerant/dependent state.
Insights
Chronic morphine significantly increases adenylyl cyclase (AC) phosphorylation in guinea pig ileum. This protein kinase C-mediated effect may explain opioid tolerance and altered signaling in the nervous system.
Area of Science:
- Neuropharmacology
- Molecular Biology
- G protein-coupled receptor signaling
Background:
- Chronic morphine use leads to opioid tolerance and dependence.
- The specific proteins affected by chronic morphine-induced phosphorylation remain largely unidentified.
- Opioid receptors are G protein-coupled receptors that regulate intracellular signaling pathways.
Purpose of the Study:
- To identify specific effector proteins whose phosphorylation is altered by chronic morphine exposure.
- To investigate the role of adenylyl cyclase (AC) phosphorylation in opioid tolerance.
- To elucidate the signaling mechanisms underlying chronic opioid effects.
Main Methods:
- Chronic morphine administration to guinea pigs.
- Immunoprecipitation of adenylyl cyclase (AC) from ileum longitudinal muscle myenteric plexus.
- Western blot analysis to detect changes in AC phosphorylation.
- Inhibition studies using chelerythrine, a protein kinase C (PKC) inhibitor.
Main Results:
- Chronic morphine treatment significantly increased the phosphorylation of adenylyl cyclase (AC), specifically the type II family.
- Increased AC phosphorylation was observed in the ileum longitudinal muscle myenteric plexus.
- Chelerythrine treatment attenuated the augmented AC phosphorylation, indicating a role for PKC.
- Phosphorylated AC isoforms show increased responsiveness to Gsalpha and Gbetagamma.
Conclusions:
- This study provides the first evidence that chronic opioid receptor activation profoundly affects adenylyl cyclase phosphorylation.
- Increased, protein kinase C-mediated phosphorylation of AC may be a key mechanism in opioid tolerance and dependence.
- This mechanism could explain the enhanced stimulatory signaling through AC observed in opioid-tolerant tissues.
- Altered AC phosphorylation offers a potential explanation for the complex and divergent consequences of chronic morphine treatment.