Characterisation of agonist binding on human 5-HT 2C receptor isoforms

Abstract

The 5-HT 2C receptor is expressed in different isoforms as a result of mRNA editing. Both INI (unedited) and VSV (a fully edited version) isoforms are abundant in rat brain. The VSV isoform lacks the high affinity recognition site for 5-HT, which may be caused by low efficiency coupling to G-proteins. In this study we have investigated the pharmacology of the agonist binding site of these two isoforms of the 5-HT 2C receptor. The VSV isoform was expressed in Chinese hamster ovary cells (CHO) and the INI isoform in both Chinese hamster ovary cells and human embryonic kidney cells (HEK-293). Saturation analysis using [ 3H]5-HT revealed high and low affinity recognition sites on the INI isoform in both cell types whilst the VSV isoform did not have the high affinity binding site for [ 3H]5-HT. Displacement studies were undertaken using [ 3H]5-HT to label the receptors. In these studies the affinity of agonists (5-HT, Ro600175 (( S)-2-(6-Chloro-5-fluoroindol-1-yl)-1-methylethylamine), MK212 (6-Chloro-2-(piperazinyl) pyrazine), mCPP (1-( m-chlorophenyl)-piperazine), TfMPP ( N-( m-trifluoromethylphenyl)piperazine), DOI (1-(2,5-Dimethoxy-4-iodophenyl)-2-aminopropane), DOB (1-(4-bromo-2,5-dimethoxyphenyl)-2-aminopropane) and 8OH-DPAT (8-hydroxy-2-(di- N-propylamino)tetralin) was higher at the INI isoform, whilst antagonist affinity (ketanserin and mesulergine) did not change between the two receptor isoforms. There were no differences between the INI isoform expressed in the CHO and HEK-293. This suggests that the INI isoform of the 5-HT 2C receptor is pharmacologically similar to the VSV form of the 5-HT 2C receptor but that it couples more efficiently to G-proteins.