Expression of the neuropeptide Y Y1 receptor in human embryonic kidney 293 cells: ligand binding characteristics, in situ hybridization and receptor autoradiography.
The human neuropeptide Y Y1 receptor cDNA was transfected into human embryonic kidney 293 (HEK 293) cells and used to determine the selectivity of newly developed Y1 and Y2 radioligands in a model which expresses a single NPY receptor subtype. The Y1 receptor probe, [125I][Leu31,Pro34]PYY, binds with high afinity (KD of 0.4-0.6 nM) to Y1-transfected HEK 293 cells whereas the Y2 radioligand, [125I]PPY3-36 failed to demonstrate any significant labelling. Only non-selective (PYY) or selective Y1 receptor agonists behaved as potent competitors for [125I][Leu31,Pro34]PYY binding in transfected cells. Additionally, the efficacy of the transfection method used was evaluated at both the transcriptional and translational levels. In situ hybridization revealed the heterogeneous distribution of the NPY Y1 receptor mRNA expressed in transfected HEK 293 cells. Similarly, the levels of NPY Y1 binding sites per transfected cell varied as shown using [125I][Leu31,Pro34]PYY receptor autoradiography. Taken together, these results demonstrate that HEK 293 cells transfected with the NPY Y1 receptor cDNA expressed both the related receptor mRNA and protein albeit at different levels depending upon each transfected cell. Additionally, these data further establish the selectivity of the newly developed Y1 and Y2 radioligands.
Tong
Y
Y
Department of Psychiatry, McGill University, Verdun, Qué, Canada.
Dumont
Y
Y
Shen
S H
SH
Herzog
H
H
Shine
J
J
Quirion
R
R
D001345:Autoradiography; D001667:Binding, Competitive; D002460:Cell Line; D006801:Humans; D017403:In Situ Hybridization; D007668:Kidney / Q000737:chemistry* / Q000166:cytology / Q000196:embryology; D011869:Radioligand Assay; D011964:Receptors, Gastrointestinal Hormone / Q000032:analysis*
D011964:Receptors, Gastrointestinal Hormone; C082172:peptide YY receptor