Characterization of the effects of four HTR3B polymorphisms on human 5-HT3AB receptor expression and signalling

Background 5-Hydroxytryptamine 3 (5-HT 3 ) receptors mediate the fast excitatory neurotransmission of serotonin. In this study, we have characterized the effects of four naturally occurring, nonsynonymous variants of the human 5-HT 3B subunit on expression and signalling properties of heteromeric 5-HT 3AB receptors. Methods and results 5-HT 3AB receptor signalling was studied in a fluorescence-based cell membrane potential assay and by electrophysiology. Expression levels of cotransfected epitope-tagged 5-HT 3A and 5-HT 3B subunits were determined using enzyme-linked immunosorbent assay and immunocytochemistry. In cells coexpressing 5-HT 3A and 5-HT 3B (I143T) subunits, cell surface expression levels of 5-HT 3B in particular, and also 5-HT 3A were markedly reduced compared with those of wild-type (WT) 5-HT 3AB receptor-expressing cells. Electrophysiological recordings on cells coexpressing 5-HT 3A and 5-HT 3B (I143T) indicated cell surface expression of 5-HT 3AB (I143T) receptors with macroscopic current kinetics similar to those of WT 5-HT 3AB receptors but with 3-fold lower current densities. In the membrane potential assay, 5-HT 3AB (I143T)-transfected cells exhibited signalling properties intermediate to those of WT 5-HT 3AB and 5-HT 3A receptors. Cotransfection of 5-HT 3A , 5-HT 3AB (I143T) and WT 5-HT 3AB subunit cDNAs did not increase cell surface expression of the variant subunit nor did it restore WT 5-HT 3AB receptor signalling completely in the membrane potential assay. In contrast to 5-HT 3B (I143T), the 5-HT 3B variants S156R, V183I and A223T did not give rise to significant changes in 5-HT 3AB receptor expression or signalling properties. Conclusion 5-HT 3B (I143T)-containing 5-HT 3AB receptors display significantly reduced cell surface expression and different signalling properties compared with WT 5-HT 3AB receptors. In contrast, three other 5-HT 3B variants, S156R, V183I and A223T, do not appear to alter 5-HT 3AB receptor expression or signalling.