Comparative pharmacology of human dopamine D(2)-like receptor stable cell lines coupled to calcium flux through Galpha(qo5).
The goal of this study was to develop a new approach to study the pharmacology of the dopamine D(4) receptor that could be used in comparative studies with dopamine D(2) and D(3) receptors. Stable HEK-293 cell lines co-expressing recombinant human D(2L), D(3) or D(4) receptors along with Galpha(qo5) cDNA were prepared. Dopamine induced a robust, transient calcium signal in these cell lines with EC(50)s for D(2L), D(3) and D(4) of 18.0, 11.9 and 2.2 nM, respectively. Reported D(4)-selective agonists CP226269 and PD168077 were potent, partial D(4) agonists exhibiting 31-1700-fold selectivity for D(4) over D(3) or D(2). Non-selective D(2)-like agonists apomorphine and quinpirole showed full efficacy but did not discriminate across the three receptors. D(3)-selective agonists 7-hydroxy-DPAT and PD128907 were potent but non-selective D(2)-like agonists. The reported D(3) partial agonist BP-897 exhibited minimal agonist activity at D(3) but was a potent D(3) antagonist and a partial D(4) agonist. Other D(2)-like antagonists, haloperidol, clozapine, and domperidone showed concentration-dependent inhibition of dopamine responses at all three receptors with K(i) ranging from 0.05 to 48.3 nM. The D(3) selective antagonist S33084 and D(4)-selective antagonist L-745870 were highly selective for D(3) and D(4) receptors with K(b) of 0.7 and 0.1 nM, respectively. Stable co-expression of D(2)-like receptors with chimeric Galpha(qo5) proteins in HEK-293 cells is an efficient method to study receptor activation in a common cellular background and an efficient method for direct comparison of ligand affinity and efficacy across human D(2L), D(3) and D(4) receptors.
Moreland
Robert B
RB
Neuroscience Research, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, IL 60064, USA. robert.moreland@abbott.com
Nakane
Masaki
M
Donnelly-Roberts
Diana L
DL
Miller
Loan N
LN
Chang
Renjie
R
Uchic
Marie E
ME
Terranova
Marc A
MA
Gubbins
Earl J
EJ
Helfrich
Rosalind J
RJ
Namovic
Marian T
MT
El-Kouhen
Odile F
OF
Masters
Jeffrey N
JN
Brioni
Jorge D
JD
D000818:Animals; D001692:Biological Transport; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D006224:Cricetinae; D004298:Dopamine; D044385:GTP-Binding Protein alpha Subunits / Q000378:metabolism*; D006801:Humans; D017448:Receptors, Dopamine D2 / Q000378:metabolism*; D050637:Receptors, Dopamine D3; D050638:Receptors, Dopamine D4; D011993:Recombinant Fusion Proteins / Q000378:metabolism
C493593:DRD3 protein, human; C493599:DRD4 protein, human; D044385:GTP-Binding Protein alpha Subunits; D017448:Receptors, Dopamine D2; D050637:Receptors, Dopamine D3; D011993:Recombinant Fusion Proteins; C113776:dopamine D2L receptor; D050638:Receptors, Dopamine D4; D002118:Calcium; D004298:Dopamine