Dopamine D1 receptor (K0.5 = 4 nM)

Compute-aided conformational analysis was used to characterize the agonist pharmacophore for D1 dopamine receptor recognition and activation. Dihydrexidine (DHX), a high-affinity full agonist with limited conformational flexibility, served as a structural template that aided in determining a molecular geometry that would be common for other more flexible, biologically active agonists. The intrinsic activity of the drugs at D1 receptors was assessed by their ability to stimulate adenylate cyclase activity in rat striatal homogenates (the accepted measure of D1 receptor activation). In addition, affinity data on 12 agonists including six purported full agonists (dopamine, dihydrexidine, SKF89626, SKF82958, A70108, and A77636), as well as six less efficacious structural analogs, were obtained from D1 dopamine radioreceptor-binding assays. The active analog approach to pharmacophore building was applied as implemented in the SYBYL software package. Conformational analysis and molecular mechanics calculations were used to determine the lowest energy conformation of the active analogs (i.e., full agonists), as well as the conformations of each compound that displayed a common pharmacophoric geometry. It is hypothesized that DHX and other full agonists may share a D1 pharmacophore made up of two hydroxy groups, the nitrogen atom (ca. 7 A from the oxygen of m-hydroxyl) and the accessory ring system characterized by the angle between its plane and that of the catechol ring (except for dopamine and A77636). For all full agonists (DHX, SKF89626, SKF82958, A70108, A77636, and dopamine), the energy difference between the lowest energy conformer and those that displayed a common pharmacophore geometry was relatively small (< 5 kcal/mol). The pharmacophoric conformations of the full agonists were also used to infer the shape of the receptor binding site. Based on the union of the van der Waals density maps of the active analogs, the excluded receptor volume was calculated. Various inactive analogs (partial agonists with D1 K0.5 > 300 nM) subsequently were used to define the receptor essential volume (i.e., sterically intolerable receptor regions). These volumes, together with the pharmacophore results, were integrated into a three-dimensional model estimating the D1 receptor active site topography. [1]

After a training dose of 0.03 mg/kg, SKF-82958 ((±)-SKF 82958) (0.003-0.1 mg/kg; ip) completely replaced SKF-82958, causing a dose-dependent increase in responses on the relevant lever of SKF-82958. Significantly reduced response rates were also obtained by increasing the dose of SKF-82958 [2]. Pilocarpine-induced mandibular movements are considerably inhibited by SKF-82958 (0.5-2.0 mg/kg; intraperitoneal injection) [3].

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We examined the discriminative stimulus effects of the high-efficacy dopamine D(1) receptor agonist (+/-)6-chloro-7, 8-dihydroxy-3-ally1-phenyl-2,3,4,5-tetrahydro-1H-3benzazepine++ + hydrobromide (SKF-82958) in rats trained to discriminate SKF-82958 (0.03 mg/kg) from vehicle in a two-lever food-reinforced drug discrimination task. SKF-82958 produced dose-related increases in responding to the SKF-82958 appropriate lever with full substitution occurring at the training dose. Pretreatment with the dopamine D(1)/D(5) receptor antagonist (-)-trans-6,7,7a,8,9, 13b-hexahydro-3-chloro-2hydroxy-N-methyl-5H-benzo-[d]naphtho -¿2, 1-b¿azepine (SCH-39166) (0.01 mg/kg) attenuated the discriminative stimulus effects of SKF-82958. Pretreatment with the dopamine D(2) receptor antagonist raclopride (0.03 mg/kg) had no effect. The high-efficacy dopamine D(1) receptor agonist R(+)6chloro-7, 8-dihydroxy-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrobromide (SKF-81297) fully substituted for SKF-82958, whereas the low-efficacy dopamine D(1) receptor agonist (+/-)1-phenyl-2,3,4, 5-tetrahydro-(1H)-3-benzazepine-7,8-diol hydrochloride (SKF-38393) produced only partial substitution. The dopamine D(2) receptor agonist trans-(+/-)-4,4a,5,6,7,8,8a, 9-octahydro-5-propyl-1H-propyl-1H-pyrazolo[3,4-g]quinoline dihydrochloride (quinpirole) and the indirect dopamine agonist cocaine did not substitute fully for the SKF-82958 discriminative stimulus cue. These results demonstrate that the high-efficacy dopamine D(1) receptor agonist SKF-82958 can serve as an effective discriminative stimulus in the rat, and that these effects are mediated by a dopamine D(1)-like receptor mechanism. [2]

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SKF 82958 (0.5-2.0 mg/kg IP) reduced the tremulous jaw movements induced by pilocarpine. The suppressive effects of SKF 82958 on jaw movements were dose-dependently reversed by systemic pretreatment with the selective D1 dopamine receptor antagonist SCH 23390 (0.025-0.2 mg/kg IP); SCH 23390 was about 16 times more potent than the D2 antagonist raclopride at reversing the effects of SKF 82958. Intracranial injection of SCH 23390 (0.5-2.0 micrograms/side) into the ventrolateral striatum, the rodent homologue of the human ventral putamen, dose-dependently reversed the reduction of pilocarpine-induced jaw movements produced by SKF 82958. Intracranial injection of SCH 23390 (0.5-2.0 micrograms/side) into the substantia nigra pars reticulata also dose-dependently reversed the reduction by SKF 82958 of pilocarpine-induced jaw movements. Injections of SCH 23390 (2.0 micrograms/side) into control sites dorsal to the striatum or substantia nigra had no effects on the action of SKF 82958. Intranigral (SNr) injections of the GABA-A antagonist bicuculline blocked the suppressive effect of systemically administered SKF 82958 on jaw movement activity. Conclusions: These data suggest that the antiparkinsonian actions of SKF 82958 may be due to stimulation of D1 receptors in the ventrolateral striatum and substantia nigra pars reticulata. In addition, these results indicate that GABA mechanisms in the substantia nigra pars reticulata may be important for the antiparkinsonian effects of D1 agonists. [3]

Animal/Disease Models: Male SD (SD (Sprague-Dawley)) rats [3]
Doses: 0.5-2.0 mg/kg
Route of Administration: intraperitoneal (ip) injection
Experimental Results: Dramatically diminished the number of mandibular tremors caused by 4.0 mg/kg pilocarpine.\n\nDrug discrimination training [2]
\nProcedures that were used to train rats to discriminate injections of SKF-82958 (0.03 mg/kg) from saline were similar to those described by Kosten et al. (1999). Rats were first food restricted and maintained at approximately 85% of their free feeding body weight. During initial training, rats were placed into the operant chambers, where the stimulus light above one of the levers was illuminated to signal the beginning of the session. Lever pressing was shaped progressively until 30 responses [fixed ratio 30 (FR30)] were emitted to obtain one food pellet reinforcer. Once stable responding under the FR30 schedule was achieved on either lever, SKF-82958 discrimination training began. For half of the rats, the left lever was designated as the SKF-82958 appropriate lever and for the other half, the right lever as the SKF-82958 appropriate. On SKF-82958 training days, the drug was administered intraperitoneally (i.p.) and the rats were placed in the operant chamber. The session began 15 min later when the stimulus lights above the levers were illuminated. Every thirtieth response on the SKF-82958-appropriate lever produced a food pellet. On vehicle training days, vehicle was administered i.p. and 15 min later, every 30th response on the vehicle-appropriate lever produced a food pellet. Training sessions ended after 30 min of responding. During training, a double alternation sequence of drug or vehicle injection (SKF, SKF, saline, saline, SKF, etc.) was utilized, with training sessions occurring 5 days per week. Initially, 1.0 mg/kg SKF-82958 was used for the training dose, however, this dose profoundly disrupted lever pressing as shown by rate and response records. The training dose was then lowered to 0.3 mg/kg, where response disruption was also observed. Eventually, the training dose was adjusted to 0.03 mg/kg. At this dose, no overt rate disruption was observed. Discrimination training with this dose of SKF-82958 continued until rats met the following criteria: (1) the first completed FR30 was on the injection-appropriate lever and (2) ≥85% of total responses made were on the injection-appropriate lever for six consecutive sessions.\n

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\n\nDiscrimination testing [2]
\nOnce the above criteria were met, substitution tests began. Tests were usually performed twice a week with the discrimination training occurring on the intervening days. If discrimination performance fell below 85% on these training days, testing was not performed, and rats were given further discrimination training. If rats maintained ≥85% condition-appropriate responding on training days, tests were performed on the following day, otherwise at least two training sessions separated each testing session. Test sessions were identical to training sessions with the exception that rats could obtain food reinforcement by completing 30 responses on either lever.\nSubstitution tests were performed with different doses of SKF-82958 (0.003–0.1 mg/kg), SKF-81297 (0.1–1.0 mg/kg), SKF-38393 (0.3–5.6 mg/kg), trans-(±)-4,4a,5,6,7,8,8a,9-octahydro-5-propyl-1H-propyl-1H-pyrazolo[3,4-g]quinoline dihydrochloride (quinpirole) (0.01–0.1 mg/kg) and cocaine (0.3–5.6 mg/kg). In antagonism studies, combinations of different doses of SKF-82958 and the dopamine D1/D5 receptor antagonist SCH-39166 (0.01 mg/kg) or the dopamine D2 receptor antagonist raclopride (0.03 mg/kg) were studied. The order of substitution and combination agonist/antagonist tests was presented non-systematically across rats.\n

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\n\nDrugs and injections [2]
\nSKF-82958, SKF-81297, SKF-38393, quinpirole, cocaine hydrochloride, and S(−) raclopride l-tartrate, were all purchased from RBI. \nAll compounds were freshly prepared on the day of the experiment in 0.9% NaCl and administered in a volume of 1 ml/kg. To ensure solubility, SKF-82958, SKF-81297, and SKF-38393 were sonicated in a heated water bath for 30 min (1 mg/ml) before dilution. Pretreatment time for each drug was 15 min and drugs were administered via i.p. injection with the exception of SCH-39166 and raclopride. Both SCH-39166 and raclopride were administered subcutaneously with a pretreatment time of 30 min. All doses are expressed as free base.

[1]. Conformational analysis of D1 dopamine receptor agonists: pharmacophore assessment and receptor mapping. J Med Chem. 1996;39(1):285-296.

[2]. The dopamine D(1) receptor agonist SKF-82958 serves as a discriminative stimulus in the rat. Eur J Pharmacol. 2000;388(2):125-131.

[3]. Striatal and nigral D1 mechanisms involved in the antiparkinsonian effects of SKF-82958 (APB): studies of tremulous jaw movements in rats. Psychopharmacology (Berl). 1999;143(1):72-81.

9-Chloro-5-phenyl-3-prop-2-enyl-1,2,4,5-tetrahydro-3-benzozazepine-7,8-diol is a benzozazepine compound. This study demonstrates that the highly potent dopamine D1 receptor agonist SKF-82958 can serve as an effective discrimination stimulus in rats. Pretreatment with the selective dopamine D1/D5 receptor antagonist SCH-39166 attenuated the discrimination stimulus effect of SKF-82958. Conversely, pretreatment with the dopamine D2 receptor antagonist ralapride did not produce a significant antagonistic effect. The antagonistic effect of dopamine D1 receptor antagonists on the SKF-82958 cue is consistent with previous findings that the discriminative stimulatory effects of the dopamine D1 receptor agonists SKF-38393 and SKF-81297 can be selectively attenuated by the dopamine D1 receptor antagonist SCH-23390 (Cunningham et al., 1985; Kamien et al., 1987; Arnt, 1988; Revill et al., 1993). Although this was not tested in this study, it would be meaningful to determine whether the antagonistic effect of SCH-39166 on SKF-82958 can be overcome. Nevertheless, these results clearly indicate that the discriminative stimulatory effect of SKF-82958 is primarily mediated by its interaction with dopamine D1-like receptors. [1]
The specificity of the discriminative stimulus effect of SKF-82958 was further confirmed by substitution tests using the potent and ineffective dopamine D1 receptor agonists SKF-81297 and SKF-38393 and the dopamine D2 receptor agonist quinpirol. SKF-81297 completely substituted for the SKF-82958 cue at moderate doses, while the ineffective dopamine D1 receptor agonist SKF-38393 only partially substituted for it even at doses that significantly reduced the response. In addition, quinpirol failed to reproduce the discriminative stimulus effect of SKF-82958 at the tested doses. In summary, this substitution pattern further confirms the D1-like properties of the discriminative stimulus of SKF-82958. [1]
It is known that potent dopamine D1 receptor agonists can stimulate adenylate cyclase in in vitro rat brain tissue preparations (Stoof and Kebabian, 1981; Izenwasser and Katz, 1993). SKF-82958 exhibits similar potency in stimulating adenylate cyclase as dopamine (O'Boyle et al., 1989; Anderson and Jansen, 1990; Izenwasser and Katz, 1993). In the same study, SKF-81297 produced adenylate cyclase activation levels (81-100%) similar to dopamine and SKF-82958, while SKF-38393 stimulated adenylate cyclase to a lower degree (45-60%). Combined with these current findings, these data suggest that only agonists with similar intrinsic potency can reproduce these effects when using highly potent dopamine receptor agonists as discriminative stimuli, as evidenced by the complete and partial substitution patterns observed with SKF-81297 and SKF-38393, respectively. Conversely, these results are inconsistent with those of Revill et al. (1993), who showed that SKF-38393 could completely replace the highly potent dopamine D1 receptor agonist SKF-81297. A more likely reason is that this stems from the different experimental testing procedures used in the two studies and the inherent differences in the stimulatory properties of the two dopamine D1 receptor agonists (Rosenzweig-Lipson and Bergman, 1993). In this regard, further research (e.g., using different training doses) may help to determine the properties of these compounds as discriminative stimuli. [1]
This study found that even at doses that reduce response rates, the preferred dopamine D2 receptor agonist quetiapine could not replace the SKF-82958 cue, consistent with other findings that dopamine D2-like receptor agonists cannot replace dopamine D1-like receptor agonists and vice versa (Cunningham et al., 1985; Kamien et al., 1987; Arnt, 1988; Revill et al., 1993). In addition, the study found that the discriminative stimulus effect of SKF-82958 could only be reproduced by similar dopamine D1 receptor agonists, but not by dopamine D2 receptor agonists; and the effect was attenuated by the selective dopamine D1/D5 receptor antagonist SCH-39166, but not by the dopamine D2 receptor antagonist ralapride. The study showed that the interoceptive stimulation of SKF-82958 is mediated by interaction with dopamine D1 receptors. [1] In this study, substitution tests using different doses of the indirect dopamine receptor agonist cocaine also failed to completely replace the discriminative stimulus cue of SKF-82958. Higher doses of cocaine (the highest tested dose was 5.6 mg/kg) could completely reproduce the discriminative stimulus effect of SKF-82958, but this test was not performed in this study. However, when the similarly potent dopamine D1 receptor agonist SKF-81297 was used as a discriminative stimulus in squirrel monkeys, neither cocaine nor dextromethorphan could completely replace the discriminative stimulus of SKF-82958 at doses that interfered with the response (Rosenzweig-Lipson and Bergman, 1993). Furthermore, these data are consistent with other findings indicating the opposite: neither highly potent nor ineffective dopamine D1 receptor agonists can reliably replace psychostimulants. For example, in rats, SKF-38393 could not replace cocaine (Barrett and Appel, 1989; Filip and Przegalinski, 1997) or dextromethorphan (Furmidge et al., 1991). Similarly, SKF-81297 cannot fully reproduce the effects of dextroamphetamine (Furmidge et al., 1991; Revill et al., 1993), and dihydroxadine cannot replace cocaine (Witkin et al., 1991). Furthermore, in primates, even at doses that significantly reduce response rates, the dopamine D1 receptor agonists SKF-82958 and SKF-81297 only partially reproduce the effects of cocaine (Spealman et al., 1991). Therefore, based on the results of this and other experiments, the discriminative stimulus-response mechanisms of dopamine D1 receptor agonists (such as SKF-82958) and psychostimulants (such as cocaine) do not appear to completely overlap, and vice versa. [1]
Recent studies have suggested that dopamine D1-like receptor agonists may be an “alternative” drug therapy for cocaine addiction, partly because the potent dopamine D1 receptor agonist SKF-82958 used in this drug identification experiment has been shown to block the recovery of cocaine self-administration behavior in rats after a short regression period (Self et al., 1996a). However, in our experiment, the dose of SKF-82958 used in the aforementioned study resulted in significant response rate disturbances, suggesting that the observed cocaine recovery blockade may be due to behavioral disturbances rather than a selective effect of cocaine reinforcement itself. Furthermore, there is evidence that potent dopamine D1 receptor agonists can be self-administered in rats and primates (Self and Stein, 1992; Self et al., 1996b; Weed and Woolverton, 1995; Weed et al., 1997; Grech et al., 1996), suggesting that they may have an inherent risk of abuse. In contrast, dopamine D1 receptor agonists with lower intrinsic potency are less likely to be self-administered by patients (Woolverton et al., 1984; Katz and Witkin, 1992; Grech et al., 1996; Weed et al., 1997) and can act as functional antagonists in vivo by blocking some of the behavioral effects of cocaine, suggesting that these drugs may play a role in the treatment of cocaine addiction (Katz and Witkin, 1992; Spealman et al., 1997). However, it remains to be determined whether other behavioral effects of dopamine D1 receptor agonists will limit their use. Therefore, it is necessary to conduct specially designed preclinical studies to explore the possibility of using partial dopamine D1 receptor agonists as a drug therapy for cocaine abuse. [1] In summary, this study is the first to demonstrate that the highly potent dopamine D1 receptor agonist SKF-82958 can be used as a reliable discrimination stimulus in rats and that its mechanism of action is related to the dopamine D1-like receptor mechanism. Furthermore, despite some overlap, the stimulatory effect of the direct dopamine D1 receptor agonist SKF-82958 is distinctly different from that of the indirect dopamine receptor agonist cocaine. [2]

C19H20CLNO2

410.73254

329.118

C, 69.19; H, 6.11; Cl, 10.75; N, 4.25; O, 9.70

80751-65-1

SKF-82958 hydrobromide;74115-01-8

1225

Typically exists as solid at room temperature

1.234 g/cm3

473.7ºC at 760 mmHg

240.3ºC

1.616

4.823

2

3

3

23

399

0

OC1=C(O)C=C2C(C3=CC=CC=C3)CN(CC=C)CCC2=C1Cl

HJWHHQIVUHOBQN-UHFFFAOYSA-N

InChI=1S/C19H20ClNO2/c1-2-9-21-10-8-14-15(11-17(22)19(23)18(14)20)16(12-21)13-6-4-3-5-7-13/h2-7,11,16,22-23H,1,8-10,12H2

9-Chloro-5-phenyl-3-prop-2-enyl-1,2,4,5-tetrahydro-3-benzazepine-7,8-diol

(±)-SKF 82958; Chloro-APB; SKF-81297 HBr; SKF-81297; Skf-82958; 80751-65-1; Cl-Apb; SKF 82958; SK&F 82958; SK&F-82958; 6-Chloro-7,8-dihydroxy-3-allyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine; 9-chloro-5-phenyl-3-prop-2-enyl-1,2,4,5-tetrahydro-3-benzazepine-7,8-diol; SKF 81297; SKF81297; (±)-SKF 82958 Chloro-AP

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.)