Structural and functional basis of 5-HT3R modulation by antidepressants

About This Grant

ABSTRACT Serotonin Type 3 Receptors (5-HT3Rs) are a subfamily of cationic pentameric ligand-gated ion channels (pLGICs) that play a critical role in gut-brain signaling. 5-HT3R pathophysiology is associated with neuropsychiatric conditions, including major depressive disorder (MDD). Previous studies have shown that antagonism of the 5-HT3R family has been implicated in the role of current antidepressant therapies such as mirtazapine (MTZ). The 5-HT3R family has five types of subunits (A-E), which influence the molecular binding and activation properties of the receptor. 5-HT3ABR is the only receptor subtype demonstrating tissue-specific expression within the brain. Animal studies have shown that the inhibition of the 5-HT3ABR has beneficial effects on mood and anxiety disorders. Despite the role that 5-HT3Rs play in MDD, there is little structural information on 5-HT3ABR and how antidepressants modulate 5-HT3Rs. As a result, 5-HT3Rs present an appealing target for novel antidepressant drug development. The overall goal of this study is to characterize the mechanistic details through which antidepressants modulate 5-HT3R activation across different subtypes. The proposal has two aims designed to address this deficit. Aim 1 will focus on determining the molecular mechanism by which MTZ inhibits 5-HT3AR. Utilizing a full-length m5-HT3AR construct, high-resolution MTZ-bound cryogenic electron microscopy (cryo-EM) structures will be determined with and without 5-HT. Based on this structural insight, site-directed mutagenesis and two-electrode voltage clamp (TEVC) electrophysiology will be used to assess the interactions necessary for MTZ binding. Residues involved in 5-HT binding are not present within the 5-HT3B subunit; therefore, aim 2 seeks to determine the key interactions required for MTZ and 5-HT binding. Aim 2 will address this by determining high-resolution cryo-EM structures of apo-5-HT3ABR and MTZ-bound 5-HT3ABR in the presence of 5-HT. Electrophysiology will be used to probe the role of each key amino acid and will be assessed for its effect on 5-HT3ABR activation. The results of this work will elucidate key interactions required for MTZ binding and highlight how antidepressants modulate different 5-HT3R subtypes. Overall, these studies will provide a deeper understanding of the role of 5-HT3R in the treatment of MDD. Finally, this work will lay the foundation for future drug development targeting 5-HT3Rs.