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Published on: February 20, 2019
Agonist-promoted kappa opioid receptor (KOR) phosphorylation has behavioral endpoint-dependent and sex-specific
Peng Huang1, Chongguang Chen1, Danni Cao1
1Center for Substance Abuse Research (CSAR) & Department of Neural Sciences, Lewis Katz School of Medicine at Temple University, 3500 Broad Street, Philadelphia, PA, 19140, USA.
Abstract:
We reported previously that the selective agonist U50,488H promoted phosphorylation of the mouse kappa opioid receptor (mKOR) in vitro at four residues in the C-terminal domain. In this study, we generated a mutant mouse line in which all the four residues were mutated to Ala (K4A) to examine the in vivo functional significance of agonist-induced KOR phosphorylation. U50,488H promoted KOR phosphorylation in brains of the wildtype (WT), but not K4A, male and female mice. Autoradiography of [3H] 69,593 binding to KOR in brain sections showed that WT and K4A mice had similar KOR distribution and expression levels in brain regions without sex differences. In K4A mice, U50,488H inhibited compound 48/80-induced scratching and attenuated novelty-induced hyperlocomotion to similar extents as in WT mice without sex differences. Interestingly, repeated pretreatment with U50,488H (80 mg/kg, s.c.) resulted in profound tolerance to the anti-scratch effects of U50,488H (5 mg/kg, s.c.) in WT mice of both sexes and female K4A mice, while in male K4A mice tolerance was attenuated. Moreover, U50,488H (2 mg/kg) induced conditioned place aversion (CPA) in WT mice of both sexes and male K4A mice, but not in female K4A mice. In contrast, U50,488H (5 mg/kg) caused CPA in male, but not female, mice, regardless of genotype. Thus, agonist-promoted KOR phosphorylation plays important roles in U50,488H-induced tolerance and CPA in a sex-dependent manner, without affecting acute U50,488H-induced anti-pruritic and hypo-locomotor effects. These results are the first to demonstrate sex differences in the effects of GPCR phosphorylation on the GPCR-mediated behaviors.
Insights
Agonist-induced kappa opioid receptor (KOR) phosphorylation is crucial for developing tolerance and conditioned place aversion, with significant sex-dependent differences observed in these behaviors. This phosphorylation does not impact acute anti-pruritic or hypo-locomotor effects.
Area of Science:
- Neuroscience
- Pharmacology
- Molecular Biology
Background:
- Previous in vitro studies identified four key phosphorylation sites on the mouse kappa opioid receptor (mKOR) in response to the selective agonist U50,488H.
- Understanding the in vivo functional significance of G protein-coupled receptor (GPCR) phosphorylation is essential for elucidating drug mechanisms and behavioral outcomes.
Purpose of the Study:
- To investigate the in vivo functional role of agonist-induced kappa opioid receptor (KOR) phosphorylation.
- To determine the sex-specific effects of KOR phosphorylation on behavioral responses to U50,488H.
Main Methods:
- Generation of a mutant mouse line (K4A) with alanine substitutions at the four identified KOR phosphorylation sites.
- Assessment of U50,488H-induced KOR phosphorylation in wildtype (WT) and K4A mice using autoradiography for receptor binding.
- Evaluation of behavioral responses, including scratching, locomotion, tolerance, and conditioned place aversion (CPA), in WT and K4A mice of both sexes.
Main Results:
- U50,488H induced KOR phosphorylation in WT mice but not in K4A mice, with similar KOR distribution and expression levels observed between genotypes.
- K4A mice exhibited similar acute anti-pruritic and hypo-locomotor responses to U50,488H as WT mice, indicating phosphorylation is not essential for these acute effects.
- Tolerance to U50,488H's anti-scratch effects and U50,488H-induced CPA showed significant sex-dependent differences in K4A mice compared to WT mice, highlighting the role of phosphorylation in these specific behaviors.
Conclusions:
- Agonist-promoted KOR phosphorylation plays a critical role in the development of tolerance and conditioned place aversion (CPA) in a sex-dependent manner.
- These findings represent the first evidence demonstrating sex differences in the impact of GPCR phosphorylation on GPCR-mediated behaviors.
- Selective agonist U50,488H's effects on tolerance and CPA are modulated by KOR phosphorylation, with distinct outcomes observed between male and female mice.
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