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5-Hydroxytryptamine2A (5-HT2A) receptor desensitization can occur without down-regulation
B L Roth1, E P Palvimaki, S Berry
1Department of Biochemistry, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA.
Abstract:
The connection between agonist-induced desensitization and down-regulation of 5-hydroxytryptamine2A (5-HT2A) receptors was examined in a clonal cell line that stably expresses the 5-HT2A receptor. Brief (2-hr) and prolonged (24-hr) exposure to the agonist quipazine or the agonist 4-iodo-(2,5-dimethoxy)- phenylisopropylamine (DOI) diminished 5-HT2A receptor-mediated phosphoinositide hydrolysis; no change in 5-HT2A receptor number or affinity was measured after 24 hr of exposure to DOI or quipazine. Immunohistochemical studies demonstrated that a 24-hr exposure to DOI did not alter surface 5-HT2A receptor immunoreactivity. Western blot analysis with G alpha q- and G alpha 11-selective antibodies indicate that a 24-hr agonist exposure did not alter the levels of phospholipase C-dependent G proteins. These results suggest that desensitization after prolonged DOI exposure can occur via a process independent of the levels of phospholipase C-coupled G proteins. Studies with a mutant 5-HT2A receptor (F340L) indicated that binding per se is not sufficient for desensitization. Down-regulation of the protein kinase C isozymes alpha and epsilon by overnight exposure to phorbol-12,13-dibutyrate attenuated the intermediate phase (i.e., after 2-6 hr of agonist exposure) of DOI- and quipazine-induced desensitization. These results indicate that the intermediate phase of DOI-induced desensitization is mediated by the alpha- and/or epsilon-protein kinase C isozymes but that neither is involved in the later phase (i.e., after 24 hr of agonist exposure) of desensitization.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Agonist exposure desensitizes serotonin 5-HT2A receptors without changing receptor number. Protein kinase C isozymes mediate early desensitization, but later stages occur independently of these enzymes.
Area of Science:
- Pharmacology
- Cell Biology
- Neuroscience
Background:
- Serotonin 5-HT2A receptors are crucial for neurotransmission.
- Agonist-induced desensitization affects receptor function.
- Understanding desensitization mechanisms is key for therapeutic development.
Purpose of the Study:
- To investigate the mechanisms of agonist-induced desensitization and down-regulation of 5-HT2A receptors.
- To determine the role of G proteins and protein kinase C (PKC) in 5-HT2A receptor desensitization.
Main Methods:
- Utilized a clonal cell line stably expressing 5-HT2A receptors.
- Employed agonist exposure (quipazine, DOI) for varying durations.
- Conducted phosphoinositide hydrolysis assays, immunohistochemistry, Western blot analysis, and studies with mutant receptors and PKC isozymes.
Main Results:
- Agonist exposure (2-24 hr) reduced 5-HT2A receptor-mediated phosphoinositide hydrolysis without altering receptor number or affinity.
- Prolonged agonist exposure did not affect surface receptor immunoreactivity or phospholipase C-coupled G protein levels.
- Down-regulation of PKC isozymes alpha and epsilon attenuated the intermediate phase of desensitization, but not the later phase.
Conclusions:
- 5-HT2A receptor desensitization can occur independently of G protein levels.
- The intermediate phase of desensitization involves PKC alpha and/or epsilon isozymes.
- Later-phase desensitization mechanisms are independent of PKC and G protein levels.