Functional Characterization of the Recombinant Human 5-Hydroxytryptamine7(a) Receptor Isoform Coupled to Adenylate Cyclase Stimulation

• Published in: The Journal of pharmacology and experimental therapeutics 1998-11, Vol.287 (2), p.508-514
• Main Authors: Adham, N, Zgombick, J M, Bard, J, Branchek, T A
• Format: Article
• Language: English
• Subjects: Adenylyl Cyclases - metabolism; Animals; Binding, Competitive; Cell Line; Cyclic AMP - metabolism; Enzyme Activation; Humans; Mice; Receptors, Serotonin - metabolism; Serotonin - metabolism; Serotonin Antagonists - pharmacology; Serotonin Receptor Agonists - pharmacology
• ISSN: 0022-3565; 1521-0103
• DOI: 10.1016/S0022-3565(24)37821-8

Functional characterization of the recombinant human 5-hydroxytryptamine 7(a) (h5-HT 7(a) ) receptor isoform was performed using stably transfected LM(tk − ) cells. Expression levels of the h5-HT 7(a) receptor determined from saturation studies using either a labeled agonist ([ 3 H]5-HT) or antagonist ([ 3 H]LSD) were very similar (B max = 160–190 fmol/mg protein), suggesting that all receptors may exist in the high affinity (G protein-coupled) state. In intact cells, 5-HT produced a concentration-dependent elevation of intracellular cAMP levels ([cAMP] i ) with an EC 50 value of 80 nM and a maximal response of 5-fold increase above basal levels. The rank order of agonist potencies in the second messenger assay paralleled their rank order of binding affinities: 5-carboxamidotryptamine > 5-hydroxytryptamine ≥ 5-methoxytryptamine > 8-hydroxy N,N-dipropyl aminotetralin > sumatriptan. Agonist potencies (EC 50 values) to stimulate [cAMP] i were more than 25-fold lower relative to their respective binding affinities ( K i values) obtained in [ 3 H]5-HT competition assays. In contrast, antagonist potencies ( K b values) to block 5-HT-stimulated [cAMP] i were in close agreement with their corresponding K i values. These data may indicate low efficiency of receptor-effector coupling to adenylate cyclase stimulation. Pretreatment of stably transfected cells with cholera toxin abolished the 5-HT-mediated elevation of [cAMP] i , indicating that the 5-HT 7(a) subtype directly interacts with G αs protein(s) to activate adenylate cyclase(s). Clonal cell lines stably expressing h5-HT 7 receptor isoforms will serve as valuable cellular models to study their function and regulation, as well as assist in the development of selective 5-HT 7 receptor agents to uncover the biological roles and potential therapeutic applications of this novel receptor subtype.