Pharmacological characterization of the human 5-HT(4(d)) receptor splice variant stably expressed Chinese hamster ovary cells

Abstract

1. The recently identified C-terminal splice variant of the human 5-HT4 receptor, the h5-HT(4(d)) receptor, was stably expressed in a CHO cell line at 493±25 fmol mg-1 protein. We analysed its pharmacological properties by measuring binding affinities and 5-HT4 ligand-induced cyclic AMP production. 2. The pharmacological binding profile determined in competition studies with the specific antagonist [3H]-GR113808 revealed a rank order of affinity of 5-HT4 ligands for the h5-HT(4(d)) receptor that was consistent with those previously reported for other 5-HT4 receptor isoforms. 3. In adenylyl cyclase functional assays, the h5-HT(4(d)) receptor displayed equipotent coupling for all 5-HT4 agonists tested (EC50 in the range of 1-6 nM). EC50 values were lower than those previously obtained with the 5-HT(4(e)) receptor stably expressed in CHO cells indicating that the 5-HT(4(d)) receptor was more efficiently coupled to its effector than the 5-HT(4(e)) receptor isoform. Moreover, in terms of agonist efficacy (E(max)), the benzamide derivative, renzapride displayed full agonist properties at the h5-HT(4(d)) receptor (same E(max) as 5-HT) whereas it was previously shown to be a partial agonist at the h5-HT(4(e)) receptor. 4. A constitutive activity of the h5-HT(4(d)) receptor was observed in CHO cells in the absence of any 5-HT4 ligand. Surprisingly, two 5-HT4 ligands, SB204070 and RS39604 which are described as highly potent antagonists in various biological models, revealed partial agonist properties at the h5-HT(4(d)) receptor. 5. We conclude that C-terminal tails of 5-HT4 receptor isoforms may directly influence their functional properties.