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.
Neuroscience Research, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park, IL 60064, USA. robert.moreland@abbott.com
Robert B
RB
Moreland
Masaki
M
Nakane
Diana L
DL
Donnelly-Roberts
Loan N
LN
Miller
Renjie
R
Chang
Marie E
ME
Uchic
Marc A
MA
Terranova
Earl J
EJ
Gubbins
Rosalind J
RJ
Helfrich
Marian T
MT
Namovic
Odile F
OF
El-Kouhen
Jeffrey N
JN
Masters
Jorge D
JD
Brioni
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