D3 dopamine receptor

The dopamine D3 receptor gene was identified by Sokoloff and colleagues in 1990. This finding rapidly gained the interest of the scientific community because this unexpected dopamine receptor subtype may play an important role in the antipsychotic activity of neuroleptic drugs. It recognizes most neuroleptics with a high affinity, and its brain distribution is restricted mainly to the ventral part of the striatal complex. However, the characterization and the subsequent identification of functions of the D3 receptor were hampered initially by at least four important factors that are still partially unresolved: (1) the absence of selective drugs that can discriminate between the D2 and D3 receptor subtype functions in vivo, (2) the lack of apparent coupling with GTP-dependent proteins, (3) the absence of effects on second messenger systems, and (4) the low level of expression of this receptor in brain tissue; these factors have contributed to tempering the interest of scientists. However, this situation has begun to change with the identification of [3H]7-hydroxy-N,N-(di-n-propyl)-2-aminotetralin ([3H]7-OH-DPAT), the first selective ligand for the dopamine D3 receptor. Although its binding selectivity for the D3 versus the D2 receptor is somewhat artificial, the potentially important impact of identification of a function for the D3 receptor encouraged scientists to use this aminotetralin compound for in vivo studies with, however, limited success. This commentary is focused on the impact and controversies generated by the use of 7-OH-DPAT and its congeners, on new conceptual views that may arise from this research, and on what partially selective D3 receptor ligands may tell us about dopamine D3 receptor functions.[2]

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We have identified 7-[3H]hydroxy-N,N-di-n-propyl-2-aminotetralin ([3H]7-OH-DPAT) as a selective probe for the recently cloned dopamine D3 receptor and used it to assess the presence of this receptor and establish its distribution and properties in brain. In transfected Chinese hamster ovary (CHO) cells, it binds to D3 receptors with subnanomolar affinity, whereas its affinity is approximately 100-, 1000-, and 10,000-fold lower at D2, D4, and D1 receptors, respectively. Specific [3H]7-OH-DPAT binding sites, with a Kd of 0.8 nM and a pharmacology similar to those at reference D3 receptors of CHO cells, were identified in rat brain. D3 receptors differ from D2 receptors in brain by their lower abundance (2 orders of magnitude) and distribution, restricted to a few mainly phylogenetically ancient areas--e.g., paleostriatum and archicerebellum--as evidenced by membrane binding are autoradiography studies. Native D3 receptors in brain are characterized by an unusually high nanomolar affinity for dopamine and a low modulatory influence of guanyl nucleotides on agonist binding. These various features suggest that D3 receptors are involved in a peculiar mode of neurotransmission in a restricted subpopulation of dopamine neurons [3].

The putative D-3 dopamine receptor agonist 7-OH-DPAT (10 micrograms/kg, s.c.) reduced spontaneous activity in rats, without inducing yawning; higher doses (0.1-10.0 mg/kg, s.c.) stimulated non-stereotyped sniffing, locomotion and chewing, which were attenuated by the selective D-1 antagonist BW 737C (5.0 mg/kg, s.c.) without release of any atypical behaviours. Low doses of 7-OH-DPAT may act on inhibitory D-3 receptors, while higher doses may act at stimulatory D-3 or other "D-2-like" receptors that participate in cooperative but not oppositional interactions with D-1 receptors [1].

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Systemic administration of D2-like dopaminergic-receptor agonists increases yawning behavior. However, only a few studies have been done in animals with pathological conditions. The taiep rat is a myelin mutant with an initial hypomyelination followed by progressive demyelination, being the brainstem one of the most affected areas. In our experiments, we analyzed the effects of systemic administration of the D2-family agonists and antagonists on yawning behavior, and correlated them with the lipid myelin content in the brainstem and other areas in the central nervous system (CNS) in 8 month old male taiep and Sprague-Dawley rats. Subjects were maintained under standard conditions in Plexiglas cages with a 12:12 light-dark cycle, lights on at 0700 and free access to rodent pellets and tap water. Drugs were freshly prepared injected ip at 0800 and subjects were observed for 60 min. When antagonists were used it was administered 15 min before the agonist. Sprague-Dawley and taiep rats significantly increased their yawning frequency after systemic injection of (-)-quinpirole hydrochloride, R(+)-7-Hydroxy-2-(dipropylamino)tetralin hydrobromide (7-OH-DPAT) or trans-(±)-3,4,4a,10b-tetrahydro-4-propyl-2H,5H-[1]benzopyrano [4,3-b]-1,4-oxazin-9-ol hydrochloride ((±)-PD 128,907). Among D2-like agonists used higher effects are obtained with (-)-quinpirole. The effects caused by (-)-quinpirole can be reduced by (-)-sulpiride; and yawning caused by 7-OH-DPAT was decreased by tiapride only in taiep rats. In Sprague-Dawley only (-)-sulpiride is able to decrease (-)-quinpirole-caused yawning. In conclusion, dopaminergic D2-like agonists are still able to cause yawning despite the severe myelin loss in taiep rats. Similarly, patients with various CNS illnesses that affect myelin, such as stroke or multiple sclerosis, are able to yawn suggesting that trigger neurons are still able to command this innate behavior [5].

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Dopamine D3 receptors have been implicated in pathophysiological substrates of schizophrenia, and neuroleptic drugs which are antagonists primarily at D2 receptors possess therapeutic activity in this disorder. In the present study, rats tested for hypomotility induced by 7-OH-DPAT (7OH, a selective D3 agonist) were pretreated with the neuroleptic haloperidol. These animals showed an attenuated agonist-induced suppression of behavior compared with rats receiving 7OH alone. The drug combination also 'normalized' dopamine metabolism in the frontal cortex, as turnover ratios which are typically enhanced by acute neuroleptic administration were no longer significantly increased when 7OH was also given. These observations suggest that the effects of haloperidol in cortical regions regulating limbic locomotor systems may be important for therapeutic efficacy in schizophrenic symptoms generated from a D3 substrate [6].

The compounds used in these studies were the D2-like agonists (−)-quinpirole hydrochloride, R(+)-7-Hydroxy-2-(dipropylamino)tetralin hydrobromide (7-OH-DPAT), or trans-(±)-3,4,4a,10b-tetrahydro-4-propyl-2H,5H-[1]benzopyrano [4,3-b]-1,4-oxazin-9-ol hydrochloride ((±)-PD 128,907), and the D2 antagonists were (−)-sulpiride hydrochloride, 5,6-dimethoxy-2-(di-n-propylamino)indan maleate (U-99194), and tiapride hydrochloride. All drugs were dissolved in sterile water and were freshly prepared at the beginning of each experimental session and administered by intraperitoneal injection (ip). The injection volume for all drugs was adjusted to 1 mL/kg. Sterile water served as the control injection. [5]

[1]. Behavioural effects of the D3 DA receptor agonist 7-OH-DPAT in relation to other D2-like agonists. Neuropharmacology. 1993 May;32(5):509-10.

[2]. Aminotetralin drugs and D3 receptor fuctions. Biochem Pharmacol. 1996 Aug 23;52(4):511-8.

[3]. Identification, characterization, and localization of the DA D3 receptor in rat brain using [3H]-7-hydroxy-N,N-di-n-propyl-2-aminotetralin. Proc Natl Acad Sci U S A. 1992 Sep 1;89(17):8155-9.

[4]. DA receptor pharmacology. Trends Pharmacol Sci. 1994 Jul;15(7):264-70.

[5]. Dopaminergic D2-like agonists produce yawning in the myelin mutant taiep and Sprague-Dawley rats. Pharmacol Biochem Behav. 2012 Jul;102(1):118-23.

[6]. Acute haloperidol attenuates the hypomotility induced with 7-hydroxy-DPAT. Neuroreport. 1997 Feb 10;8(3):611-5.

Dopamine exerts a sustained inhibitory effect on cholinergic neurons that produce yawning (Holmgren and Urbá-Holmgren, 1980; Holmgren et al., 1982; Yamada and Furukawa, 1981). The dose-response curve has been clearly demonstrated to be inverted U-shaped, with the ascending limb mediated by D3 receptors and the descending limb by D2 receptors (Collins et al., 2005; Collins et al., 2007; Baladi et al., 2010; Baladi et al., 2011). Because this study used relatively high doses, only the descending limb of the curve was investigated, as at these doses, grasp-induced tonic-clonic seizures were significantly increased, a characteristic feature of myelin-mutant rats (Eguibar et al., 2010), indicating that taiep rats are insensitive to the effects of D3 and D2 receptor antagonists on yawning. The lack of effect with the D2 receptor antagonist used may be due to the lower frequency of spontaneous yawning. However, thiapride antagonized the increased yawning frequency induced by 7-OH-DPAT in Taiep rats. Because myelin mutants have different sensitivities to D3 receptors, this dose was on the descending limb of the dose-response curve in Sprague-Dawley rats, as previously suggested (Baladi et al., 2010; Baladi et al., 2011; Collins et al., 2007; Collins et al., 2009). Under free-access conditions, Taiep and Sprague-Dawley rats showed different sensitivities to D2-like dopaminergic agonists, which could explain the differences observed when the antagonist was administered before the agonist, as 7-OH-DPAT was on the ascending limb of the dose-response curve in Taiep rats at this dose, while it was likely on the descending limb in Sprague-Dawley rats. The agonist acts on D3 receptors in the former and likely on D2 receptors in the latter (see Table 1 and Figure 2). These differences in response to dopaminergic drugs may be related to age, sex, genetics, and nutritional status, as previously proposed (Baladi et al., 2011; Sevak et al., 2008). Our results clearly indicate that the differences in response to thiabendazole and 7-OH-DPAT are due to the genetic background of taiep rats, which is also the reason for the different yawning frequencies between the two groups of rats. [5] Dopamine receptors are major targets for the treatment of schizophrenia, Parkinson's disease, and Huntington's disease, as discussed in this review by Philip Seeman and Hubert Van Tol. Drugs that selectively target specific subtypes of dopamine receptors can improve treatment efficacy. Most antipsychotic drugs block D2 receptors, and their efficacy is directly related to clinical efficacy, except for clozapine, which is more likely to target D4 receptors. D1 and D2 receptors can enhance each other, possibly through G protein subunits. In schizophrenia, the density of D2 and D3 receptors is increased by 10%, while the density of D4 receptors is increased by 600%. Therefore, D4 receptors may be a target for future antipsychotic drugs. Although antipsychotic drugs initially helped in the discovery of dopamine receptors, the five dopamine receptors currently cloned are facilitating the discovery of selective antipsychotics and anti-Parkinson's drugs. [4]

C16H26BRNO

328.28774

327.12

C, 58.54; H, 7.98; Br, 24.34; N, 4.27; O, 4.87

76135-30-3

11957566

Typically exists as solid at room temperature

4.329

2

2

5

19

237

0

CCCN(CCC)C1CCC2=CC=C(C=C2C1)O.Br

ODNDMTWHRYECKX-UHFFFAOYSA-N

InChI=1S/C16H25NO.BrH/c1-3-9-17(10-4-2)15-7-5-13-6-8-16(18)12-14(13)11-15/h6,8,12,15,18H,3-5,7,9-11H2,1-2H31H

7-Hydroxy-N,N-dipropyl-2-aminotetralin hydrobromide

7-OH DPAT; 7-OH-DPAT HBr; 159795-63-8; (A+-)-7-Hydroxy-2-(di-n-propylamino)tetralin hydrobromide; 633-919-6; 76135-30-3; 7-Hydroxy-DPAT hydrobromide; 7-(Dipropylamino)-5,6,7,8-tetrahydronaphthalen-2-ol hydrobromide; 7-OH-dpat hydrobromide; 2-Naphthalenol, 7-(dipropylamino)-5,6,7,8-tetrahydro-, hydrobromide; 7-OH-DPAT Hydrobromide

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)