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Unique allosteric regulation of 5-hydroxytryptamine receptor-mediated signal transduction by oleamide
E A Thomas1, M J Carson, M J Neal
1Department of Molecular Biology, MB-10, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Abstract:
The effects of oleamide, an amidated lipid isolated from the cerebrospinal fluid of sleep-deprived cats, on serotonin receptor-mediated responses were investigated in cultured mammalian cells. In rat P11 cells, which endogenously express the 5-hydroxytryptamine2A (5HT2A) receptor, oleamide significantly potentiated 5HT-induced phosphoinositide hydrolysis. In HeLa cells expressing the 5HT7 receptor subtype, oleamide caused a concentration-dependent increase in cAMP accumulation but with lower efficacy than that observed by 5HT. This effect was not observed in untransfected HeLa cells. Clozapine did not prevent the increase in cAMP elicited by oleamide, and ketanserin caused an approximately 65% decrease. In the presence of 5HT, oleamide had the opposite effect on cAMP, causing insurmountable antagonism of the concentration-effect curve to 5HT, but had no effect on cAMP levels elicited by isoproterenol or forskolin. These results indicate that oleamide can modulate 5HT-mediated signal transduction at different subtypes of mammalian 5HT receptors. Additionally, our data indicate that oleamide acts at an apparent allosteric site on the 5HT7 receptor and elicits functional responses via activation of this site. This represents a unique mechanism of activation for 5HT G protein-coupled receptors and suggests that G protein-coupled neurotransmitter receptors may act like their iontropic counterparts (i.e., gamma-aminobutyric acid type A receptors) in that there may be several binding sites on the receptor that regulate functional activity with varying efficacies.
Insights
Oleamide, a lipid from cerebrospinal fluid, modulates serotonin receptor activity. It affects 5-hydroxytryptamine (5HT) signaling pathways in cultured cells, acting uniquely on 5HT7 receptors.
Area of Science:
- Neuroscience
- Molecular Pharmacology
- Biochemistry
Background:
- Oleamide is an endogenous lipid found in cerebrospinal fluid, implicated in sleep regulation.
- Serotonin receptors (5-hydroxytryptamine receptors, 5HTRs) are crucial G protein-coupled receptors (GPCRs) involved in numerous physiological processes.
- Understanding how lipids modulate neurotransmitter receptor function is key to deciphering complex signaling networks.
Purpose of the Study:
- To investigate the effects of oleamide on serotonin receptor-mediated signal transduction pathways.
- To characterize oleamide's interaction with specific 5-hydroxytryptamine receptor subtypes (5HT2A and 5HT7).
- To elucidate the mechanism of action of oleamide on these receptors, particularly its potential allosteric modulation.
Main Methods:
- Utilized cultured mammalian cell lines (rat P11 and HeLa cells) engineered to express specific serotonin receptor subtypes.
- Measured 5HT-induced phosphoinositide hydrolysis in 5HT2A receptor-expressing cells.
- Assessed cyclic AMP (cAMP) accumulation in 5HT7 receptor-expressing cells in response to oleamide and 5HT, employing receptor antagonists (ketanserin) and agonists (isoproterenol, forskolin).
Main Results:
- Oleamide potentiated 5-hydroxytryptamine (5HT)-induced phosphoinositide hydrolysis in rat P11 cells expressing the 5HT2A receptor.
- In HeLa cells expressing the 5HT7 receptor, oleamide induced a concentration-dependent increase in cAMP accumulation, albeit with lower efficacy than 5HT.
- Oleamide exhibited insurmountable antagonism of 5HT's effect on cAMP in 5HT7 receptor cells, suggesting allosteric modulation, while not affecting isoproterenol or forskolin responses.
Conclusions:
- Oleamide modulates 5HT-mediated signal transduction across different mammalian 5HT receptor subtypes.
- Data suggest oleamide acts at an allosteric site on the 5HT7 receptor, activating it to elicit functional responses.
- This represents a novel activation mechanism for 5HT GPCRs, indicating potential for multiple regulatory binding sites on neurotransmitter receptors, similar to ionotropic receptors.