Received for publication September 7, 2007.
Revised November 22, 2007.
Accepted for publication December 13, 2007.

Inverse Agonist and Neutral Antagonist Actions of Antidepressants at Recombinant and Native 5-HT_2C Receptors: Differential Modulation of Cell Surface Expression and Signal Transduction

Abstract

Despite the importance of serotonin (5-HT)_2C receptors in the control of depressive states, actions of antidepressants at these receptors remain poorly characterized. This issue was addressed both in HEK-293 cells co-expressing unedited human (h)5-HT_2CINI receptors and G[alpha]_q protein and in cultured mouse cortical neurons. Indicative of constitutive activity, the inverse agonist, SB206,553, decreased basal inositol phosphate (IP) production in HEK-293 cells. The tetracyclic antidepressants, mirtazapine and mianserin, likewise suppressed basal IP formation. Conversely, the tricyclics, amitriptyline and clomipramine, the m-chlorophenylpiperazine derivatives, trazodone and nefazodone, and the 5-HT reuptake inhibitors, fluoxetine and citalopram, were inactive alone though they blocked 5-HT-induced IP production. Inverse agonist actions of SB206,553 and mirtazapine were abolished by the neutral antagonist, SB242,084, which was inactive alone. As assessed by confocal microscopy and ELISA, prolonged treatment of HEK-293 cells with SB206,553, mirtazapine or mianserin, but not the other antidepressants, enhanced cell surface expression of 5-HT_2C receptors: 5-HT-induced IP production was also increased, and both these actions were blocked by SB242,084. Cortical neurons were shown by RT-PCR to predominantly express constitutively-active 5-HT_2C receptor isoforms. Prolonged pretreatment with SB206,553 or mirtazapine triggered an otherwise-absent, 5-HT-induced elevation in cytosolic Ca²⁺ concentrations. SB242,084, which was inactive alone, abolished these effects of SB206,553 and mirtazapine. In conclusion, the tetracyclic antidepressants, mirtazapine and mianserin, but not other clinically-established antidepressants, suppress constitutive activity at recombinant and native 5-HT_2C receptors. The clinical significance of inverse agonist vs. neutral antagonist properties both during and following drug administration will be of interest to elucidate.

Key words: Serotonin, Phospholipase C's, Alternative splicing/RNA editing, Receptor synthesis/trafficking, Sequestration/Internalization, Recycling, Ca imaging, Immunocytochemistry, Anti-depressants