Isolation of the serotoninergic 5-HT4(e) receptor from human heart and comparative analysis of its pharmacological profile in C6-glial and CHO cell lines.
RT - PCR technique was used to clone the human 5-HT(4(e)) receptor (h5-HT(4(e))) from heart atrium. We showed that this h5-HT(4(e)) receptor splice variant is restricted to brain and heart atrium. Recombinant h5-HT(4(e)) receptor was stably expressed in CHO and C6-glial cell lines at 347 and 88 fmol mg(-1) protein, respectively. Expression of h5-HT(4(e)) receptors at the cell membrane was confirmed by immunoblotting. The receptor binding profile, determined by competition with [(3)H]-GR113808 of a number of 5-HT(4) ligands, was consistent with that previously reported for other 5-HT(4) receptor isoforms. Surprisingly, we found that the rank order of potencies (EC(50)) of 5-HT(4) agonists obtained from adenylyl cyclase functional assays was inversely correlated to their rank order of affinities (K(i)) obtained from binding assays. Furthermore, EC(50) values for 5-HT, renzapride and cisapride were 2 fold lower in C6-glial cells than in CHO cells. ML10302 and renzapride behaved like partial agonists on the h5-HT(4(e)) receptor. These results are in agreement with the reported low efficacy of the these two compounds on L-type Ca(2+) currents and myocyte contractility in human atrium. A constitutive activity of the h5-HT(4(e)) receptor was observed in CHO cells in the absence of any 5-HT(4) ligand and two 5-HT(4) antagonists, GR113808 and ML10375, behaved as inverse agonists. These data show that the h5-HT(4(e)) receptor has a pharmacological profile which is close to the native h5-HT(4) receptor in human atrium with a functional potency which is dependent on the cellular context in which the receptor is expressed.
Mialet
J
J
Laboratoire de Cardiologie Cellulaire et Moleculaire, INSERM U-446, Universite de Paris-Sud, Faculte de Pharmacie, F-92296 Chatenay-Malabry, France.
Berque-Bestel
I
I
Eftekhari
P
P
Gastineau
M
M
Giner
M
M
Dahmoune
Y
Y
Donzeau-Gouge
P
P
Hoebeke
J
J
Langlois
M
M
Sicsic
S
S
Fischmeister
R
R
Lezoualc'h
F
F
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D000918:Antibody Specificity; D001667:Binding, Competitive; D016466:CHO Cells / Q000378:metabolism; D003001:Cloning, Molecular; D006224:Cricetinae; D005910:Glioma / Q000235:genetics / Q000378:metabolism; D006325:Heart Atria / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D009206:Myocardium / Q000737:chemistry*; D009928:Organ Specificity; D051381:Rats; D011985:Receptors, Serotonin / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology*; D044422:Receptors, Serotonin, 5-HT4; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012702:Serotonin Antagonists / Q000494:pharmacology*; D017366:Serotonin Receptor Agonists / Q000494:pharmacology*
D011985:Receptors, Serotonin; D012702:Serotonin Antagonists; D017366:Serotonin Receptor Agonists; D044422:Receptors, Serotonin, 5-HT4