Abstract
In view of the therapeutic importance of dopamine D3 and D2 receptors, there remains considerable interest in novel ligands. Herein, we show that the tetrahydroisoquinoline 1H-indole-2-carboxylic acid {4-[2-(cyano-3,4-dihydro-1H-isoquinolin-2-yl)-ethyl]-cyclohexyl}-amide (SB269,652) behaves as an atypical, allosteric antagonist at D3 and D2 receptors. Accordingly, SB269,652 potently (low nanomolar range) abolished specific binding of [3H]nemanopride and [3H]spiperone to Chinese hamster ovary-transfected D3 receptors when radioligands were used at 0.2 and 0.5 nM, respectively. However, even at high concentrations (5 μM), SB269,652 only submaximally inhibited the specific binding of these radioligands when they were employed at 10-fold higher concentrations. By analogy, although SB269,652 potently blocked D3 receptor-mediated activation of Gαi3 and phosphorylation of extracellular-signal-regulated kinase (ERK)1/2, when concentrations of dopamine were increased by 10-fold, from 1 μM to 10 μM, SB269,652 only submaximally inhibited dopamine-induced stimulation of Gαi3. SB269,652 (up to 10 μM) only weakly and partially (by approximately 20–30%) inhibited radioligand binding to D2 receptors. Likewise, SB269,652 only submaximally suppressed D2 receptor-mediated stimulation of Gαi3 and Gαqi5 (detected with the aequorin assay) and phosphorylation of ERK1/2 and Akt. Furthermore, SB269,652 only partially (35%) inhibited the dopamine-induced recruitment of β-arrestin2 to D2 receptors. Finally, Schild analysis using Gαi3 assays, and studies of radioligand association and dissociation kinetics, supported allosteric actions of SB269,652 at D3 and D2 receptors.
Footnotes
↵ The online version of this article (available at http://molpharm.aspetjournals.org) contains supplemental material.
This work was supported in part by Fondo per gli Investimenti della Ricerca di Base (FIRB) [Grant RBIN04CKYN]; Programmi di Ricerca Scientifica di Rilevante Interesse Nazionale (PRIN) [Grant 20085PPEK7]; the National Institutes of Health National Institute on Drug Abuse [Grant DA022413]; the National Institutes of Health National Institute of Mental Health [Grant MH54137]; the Lieber Center for Schizophrenia Research and Treatment; and IDR Servier (to E.S.).
Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
doi:10.1124/mol.110.065755.
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ABBREVIATIONS:
- OSU6162
- (3S)-3-[3-(methylsulfonyl)phenyl]-1-propylpiperidine hydrochloride
- ACR16
- pridopidine
- SB269,652
- 1H-indole-2-carboxylic acid {4-[2-(cyano-3,4-dihydro-1H-isoquinolin-2-yl)-ethyl]-cyclohexyl}-amide
- CHO
- Chinese hamster ovary
- BRET
- bioluminescence resonance energy transfer
- ERK
- extracellular signal-regulated kinase
- TMS
- transmembrane segment
- EL
- extracellular loop
- L741,626
- 3-[4-(4-chlorophenyl)-4-hydroxypiperidin-l-yl]methyl-1H-indole
- GTPγS
- guanosine 5′-O-(3-thio)triphosphate
- S33084
- (+)-trans-3,4,4a,5,6,10b-hexahydro-9-carbamoyl-4-propyl-2H-naphth[1,2-b]-1,4-oxazine]
- SB277
- 011N-{trans-4-[2-(6-cyano-3,4-dihydroisoquinolin-2(1H)-yl)ethyl]cyclohexyl}quinoline-4-carboxamide.
- Received April 21, 2010.
- Accepted July 27, 2010.
- Copyright © 2010 The American Society for Pharmacology and Experimental Therapeutics
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