The long-term objective of this research is to gain a better understanding of the physiological roles played by 5-HT2c serotonin receptors in the CNS. 5-HT2c receptors are expressed on brain neurons and are members of the G-protein-coupled receptor (GPCR) superfamily. Although little is known about the actual physiological roles played by 5-HT2c receptors in the CNS, they have been implicated in mental disorders, neuroendocrine regulation, CSF production and appetite regulation. Previous studies have shown that mutant, inactive GPCR can decrease the ability of native receptors to bind agonist and activate second messenger systems. Receptor dimerization has been proposed as a possible mechanism. Since mutant, inactive GPCR have been identified in human diseases, we propose studies to determine the mechanism(s) by which inactive receptors have a dominant negative influence on native receptor function. Our preliminary studies indicate that mutant, inactive 5-HT2c receptors have a dominant negative influence on native 5-HT2c receptor function and that 5-HT2c receptors form homodimers.
The specific aims of this grant are designed to 1) identify molecular mechanisms through which inactive 5-HT2c receptors regulate the function of native receptors; 2) demonstrate the formation of 5-HT2c receptor dimers; 3) identify critical regions of the 5-HT2c receptor involved in dimerization; 4) determine if dimerization plays a role in receptor activation; and 5) to determine if a naturally occurring 5-HT2c receptor splice variant, identified in human brain, plays a role in regulating the function of native 5-HT2c receptors in vivo. If receptor dimerization occurs in vivo, it could have important clinical implications in cases where there is one ? mutant and one normal allele for a given receptor. In systems where proteins dimerize to form functional units, the presence of one mutant allele producing an inactive protein could dramatically reduce the ability to form functional protein complexes. The proposed studies will use bioluminescence and fluorescence resonance energy transfer (BRET/FRET) to monitor the effect of inverse agonists on 5-HT2c receptor dimerization. ? ? These studies may have important clinical implications because atypical antipsychotic drugs are potent 5-HT2c receptor inverse agonists. In addition, these studies may enhance our understanding of the functional interaction between atypical antipsychotic drugs and GPCR. Therefore, understanding the role that dimerization plays in GPCR activation is very important to our overall understanding of GPCR function in normal and disease states, and will have important implications for drug design and the development of novel therapeutic agents. ? ?
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