title: Differences in signal transduction of two 5-HT4 receptor splice variants: compound specificity and dual coupling with Galphas- and Galphai/o-proteins.
abstract: This study documents differences in ligand binding and signal transduction properties between the human (h) 5-hydroxytryptamine (5-HT)4a and h5-HT4b receptor splice variants stably expressed in human embryonic kidney 293 cells. The fraction of the [3H]5-HT high-affinity site relative to the whole receptor population measured with [3H]GR113808 was higher for the h5-HT4a isoform (around 0.4) than for the 5-HT4b isoform (around 0.2) and was independent of the level of expression. The potency and efficacy of reference compounds tested for the cAMP response differed slightly but significantly between both variants. Most remarkably, 5-methoxytryptamine and prucalopride were found more potent on the 5-HT4b variant, whereas SDZ-HTF 919 and SB204070 were more potent on the 5-HT(4a) variant. Guanosine-5'-O-(3-[35S]thio)triphosphate binding on membranes and cAMP assays in whole cells revealed that only the h5-HT4b isoform coupled to Galphai/o-proteins in addition to its well-documented Galphas coupling. In contrast, the h5-HT4a receptor coupled only to Galphas-proteins, however, was able to trigger an increase in the intracellular calcium concentration ([Ca(2+)]i). The observed [Ca(2+)]i increase did not occur through inositol phosphate formation and was not sensitive to Bordetella pertussis toxin, forskolin, or 3-isobutyl-1-methylxanthine (pre)treatment but was due to Ca(2+) influx from the extracellular environment. Interestingly, the Ca(2+) pathway was dependent on high receptor expression levels and was compound-specific, because benzamide-like compounds triggered two to three times higher responses than indoleamines. Taken together, these data provide the first evidence for fine functional differences between C-terminal splice variants of the h5-HT4 receptor, which may contribute to a better understanding of the functional diversity of this receptor class.
authors.affiliation: Department of Receptor Pharmacology, Janssen Research Foundation, B-2340 Beerse, Belgium.
authors.forename: Armelle
authors.initials: A
authors.lastname: Pindon
authors.forename: Geert
authors.initials: G
authors.lastname: van Hecke
authors.forename: Paul
authors.initials: P
authors.lastname: van Gompel
authors.forename: Anne S
authors.initials: AS
authors.lastname: Lesage
authors.forename: Josée E
authors.initials: JE
authors.lastname: Leysen
authors.forename: Mirek
authors.initials: M
authors.lastname: Jurzak
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D017398:Alternative Splicing; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D007295:Inositol Phosphates / Q000378:metabolism; D037342:Pertussis Toxin; D011985:Receptors, Serotonin / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D044422:Receptors, Serotonin, 5-HT4; D012702:Serotonin Antagonists / Q000494:pharmacology; D015398:Signal Transduction; D013449:Sulfonamides / Q000494:pharmacology; D013462:Sulfur Radioisotopes; D014162:Transfection; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
chemical_list: D007211:Indoles; D007295:Inositol Phosphates; D011985:Receptors, Serotonin; D012702:Serotonin Antagonists; D013449:Sulfonamides; D013462:Sulfur Radioisotopes; D010566:Virulence Factors, Bordetella; D044422:Receptors, Serotonin, 5-HT4; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D037342:Pertussis Toxin; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D019205:GTP-Binding Protein alpha Subunits, Gs; D000262:Adenylyl Cyclases; D002118:Calcium; C082871:GR 113808