IMMUNOLOGICAL CHARACTERIZATION AND TRANSMEMBRANE TOPOLOGY OF 5-HYDROXYTRYPTAMINE(3) RECEPTORS BY FUNCTIONAL EPITOPE TAGGING

5-Hydroxytryptamine(3) (5-HT3) receptors are the only known monoamine receptors mediating fast excitatory responses in mammalian neurons. Th eir primary structure as well as their electrophysiological and pharma cological properties show a phylogenetic relation to nicotinic acetylc holine, GABA(A), and glycine receptors. As a prototypical member of th is gene superfamily, we investigated the membrane topology of function al homomeric 5-HT3 receptors by using epitope tagging of the channel s ubunits expressed in heterologous systems. Visualization of 5-HT3 rece ptors in transfected COS-7 cells, either in western blot (molecular ma ss 61.2 +/- 0.8 kDa) or in situ, was performed with previously charact erized antibodies recognizing artificial epitopes as well as with anti -fusion protein antibodies directed against a wild-type receptor intra cellular domain, The extracellular location of the distal C-terminal t agged domain demonstrates the presence of a fourth transmembrane domai n in 5-HT3 serotonin-gated channels. In this region, the significant h omology between members of this class of neurotransmitter-gated channe ls suggests strongly that they have a common transmembrane organizatio n basically different from glutamate-gated and ATP-gated channels.