5-HT1B Receptor (pKi = 9.4); 5-HT1D Receptor (pKi = 9.3)
Donitriptan hydrochloride targets the serotonin 5-HT1B and 5-HT1D receptors. It is a potent and high-affinity agonist with pKi values of 9.4 and 9.3 for these receptors, respectively. By activating these receptors, it modulates serotonergic neurotransmission, leading to vasoconstriction. This mechanism is similar to that of other triptans used for migraine, making it a potential antimigraine agent.

In vitro, Donitriptan hydrochloride acts as a potent agonist at 5-HT1B and 5-HT1D receptors. It inhibits forskolin-induced cAMP formation in C6 cells expressing the human receptors, with pD2 values of 8.91 and 9.57, respectively. Its high affinity and intrinsic activity at these receptors make it a valuable tool for studying 5-HT1B/1D receptor function. Standard in vitro assays include receptor binding studies and functional assays measuring cAMP accumulation.

Intracarotid infusions of capsaicin, alpha-CGRP and acetylcholine dose-dependently increased blood flow through the carotid artery. These responses remained unaffected after intravenous (i.v.) infusions of sumatriptan, PNU-142633, PNU-109291 or physiological saline; in contrast, donitriptan significantly attenuated the vasodilator responses to capsaicin, but not those to alpha-CGRP or acetylcholine. Only sumatriptan and donitriptan dose-dependently decreased the carotid blood flow. Interestingly, i.v. administration of the antagonist, SB224289 (5-HT(1B)), but not of BRL15572 (5-HT(1D)), abolished the inhibition by donitriptan.Conclusions and implications: The results suggest that the inhibition produced by donitriptan of capsaicin-induced external carotid vasodilatation is mainly mediated by 5-HT(1B), rather than 5-HT(1D), receptors, probably by a central mechanism [1].

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The aim of the present study was to determine whether donitriptan and sumatriptan decreased jugular venous oxygen saturation and increased carbon dioxide partial pressure in venous blood. However, previous studies conducted with these compounds cannot discriminate whether the decrease of venous oxygen saturation is dependent of cranial vasoconstrictor. In the present study, vehicle (n = 10), donitriptan (2.5, 10, and 40 microg/kg; n = 8) or sumatriptan (630 microg/kg; n = 8) were infused into the carotid artery in the anesthetized rat. Regional blood flows were evaluated in the presence of donitriptan (10 microg/kg; n = 6) or vehicle (n = 6). Jugular venous oxygen saturation was significantly decreased by donitriptan (from 10 microg/kg) with maximal changes of -32.9 +/- 8.0%. Jugular carbon dioxide partial pressure was increased by donitriptan, reaching maximal changes of 17.7 +/- 4.6% (P < 0.05 versus vehicle). Similarly, sumatriptan significantly decreased venous oxygen saturation and increased jugular carbon dioxide partial pressure. These changes induced by donitriptan are abolished by the 5-hydroxytryptamine (5-HT)(1B/1D) receptor antagonist GR 127935 (N-[4-methoxy-3-(4-methyl-1-piperazinyl)phenyl]-2-[-methyl-4(5-methyl-1,2,4)-oxadiazol-3-yl]-(1,1 biphenyl)-4-carboxamide dihydrochloride). In addition, donitriptan was devoid of significant effects on systemic arterial pressure, heart rate, or regional blood flows, including systemic arterial-jugular venous anastomotic, systemic, or cranial. The results demonstrate that donitriptan increases cerebral oxygen consumption by 5-HT(1B/1D) receptor activation in the absence of cranial vasoconstriction.[2]

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The effects of donitriptan on systemic arterial-jugular venous oxygen saturation difference were evaluated in pentobarbitone-anesthetized pigs. Oxygen and carbon dioxide partial pressures in systemic arterial and jugular venous blood as well as hemoglobin oxygen saturation were determined by conventional blood gas analysis. Vehicle (40% polyethyleneglycol in saline, n = 9) or donitriptan (0.01, 0.04, 0.16, 0.63, 2.5, 10, and 40 microg/kg, n = 7) were cumulatively infused over 15 min/dose. The involvement of 5-hydroxytryptamine(1B) (5-HT(1B)) receptors was assessed in the presence of the 5-HT(1B/1D) receptor antagonist, GR 127935. Donitriptan decreased markedly and dose dependently jugular venous oxygen saturation [ED(50) 0.5 (0.3-1.1) microg/kg], in parallel with increases in carotid vascular resistance [ED(50) 0.9 (0.7-1.1) microg/kg]. Since arterial oxygen saturation and partial pressure remained unchanged, donitriptan significantly increased arteriovenous oxygen saturation difference from 0.63 microg/kg (maximal variation: 57 +/- 18%, P < 0.05 compared with vehicle). Unexpectedly, donitriptan from 2.5 microg/kg induced marked and significant increases in carbon dioxide partial pressure (pVCO(2)) in venous blood (maximal increase 18.8 +/- 5.7%; P < 0.05 compared with vehicle). Pretreatment with GR 127935 (0.63 mg/kg, n = 5) abolished the fall in venous oxygen saturation and the increase in carotid vascular resistance and reduced the increases in pVCO(2) induced by donitriptan. The results demonstrate that donitriptan, via 5-HT(1B) receptor activation, decreases the oxygen saturation of venous blood draining the head, concomitantly with cranial vasoconstriction. Since donitriptan also increased pVCO(2), an effect upon cerebral oxygen consumption and metabolism is suggested in addition to cranial vasoconstriction, which may be relevant to its headache-relieving effects[3].

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In vivo, Donitriptan hydrochloride has been studied for its potential as an antimigraine agent. As a brain-penetrant 5-HT1B/1D receptor agonist, it is expected to produce vasoconstriction of cranial blood vessels and inhibit the release of pro-inflammatory neuropeptides, which are key mechanisms in migraine relief. However, comprehensive in vivo efficacy data from published literature are limited. The compound is primarily used as a quantitative benchmark in neurovascular pharmacology.

For non-cell-based receptor binding assays, Donitriptan hydrochloride can be evaluated using membrane preparations from cells expressing human 5-HT1B or 5-HT1D receptors. Radioligand binding displacement experiments are performed using a suitable radiolabeled ligand such as [3H]-5-HT. Membrane homogenates are incubated with increasing concentrations of the test compound and a fixed concentration of the radioligand. Bound radioligand is separated from free by rapid filtration. Non-specific binding is determined in the presence of excess unlabeled 5-HT. pKi values are calculated from displacement curves using nonlinear regression analysis.

For in vitro cellular assays, cells expressing human 5-HT1B or 5-HT1D receptors (e.g., C6 glioma cells) are cultured in appropriate media. For functional assays, the inhibition of forskolin-stimulated cAMP accumulation is measured. Cells are treated with various concentrations of Donitriptan hydrochloride in the presence of forskolin. cAMP levels are measured using ELISA or HTRF-based detection. The compound's ability to inhibit cAMP accumulation is assessed, and pD2 values are calculated from dose-response curves.

Experimental protocol [1]
After the animals (n=59) had been in a stable haemodynamic condition for at least 60 min, baseline values of mean blood pressure, heart rate and external carotid blood flow were determined. Subsequently, the animals were divided into four groups (n=20, 8, 20 and 11). [1]
The first group (n=20) was subdivided into five subgroups (n=4 each) that received consecutive 10-min i.v. infusions of, respectively: (i) sumatriptan (1, 3, 10, 30, 100 and 300 μg kg−1); (ii) donitriptan (0.1, 0.3, 1, 3, 10 and 30 μg kg−1); (iii) PNU-142633 (1, 3, 10, 30, 100 and 300 μg kg−1); (iv) PNU-109291 (0.3, 1, 3, 10, 30 and 100 μg kg−1); and (v) equivalent volumes of physiological saline (0.5 ml min−1 during 10 min; given six times). The above compounds were given consecutively following a cumulative dose-schedule as i.v. infusions (at a rate of 0.5 ml min−1 during 10 min for each dose). [1]
The second group (n=8) received consecutive intracarotid infusions (1 ml min−1, for 1 min) of capsaicin (10, 18, 30 and 56 μg min−1), α-CGRP (0.1, 0.3, 1 and 3 μg min−1) and acetylcholine (0.01, 0.03 and 0.1 μg min−1). Then, this group was subdivided into two subgroups (n=4 each) that received an intracarotid continuous infusion throughout the experiment of, respectively: (i) vehicle (0.3 ml min−1 of physiological saline); and (ii) phenylephrine (1.5 μg min−1, given at a rate of 0.3 ml min−1), which produced a carotid vasoconstriction similar to that elicited by the highest dose of sumatriptan (300 μg kg−1, i.v.) or donitriptan (30 μg kg−1, i.v.) (see first group for details). Twenty minutes after the start of the infusion of physiological saline or phenylephrine, the responses to the above doses of capsaicin, α-CGRP and acetylcholine (in this order) were elicited again as described above during the intracarotid continuous infusion of each compound. [1]
The third group (n=20) received a continuous intracarotid infusion of phenylephrine (1.5 μg min−1) as described previously and, 20 min later, the responses to the above doses of capsaicin, α-CGRP and acetylcholine (in this order) were elicited during the infusion of phenylephrine. Then, this group was subdivided into five subgroups (n=4 each) so that the infusion of phenylephrine was stopped in the first two subgroups (waiting about 60 min for the recovery of baseline external carotid blood flow), whereas it remained continuously infusing throughout the experiment in the remaining three subgroups. Subsequently, by the use of another motor-driven syringe inserted into the femoral vein and infusing at a rate of 0.5 ml min−1 during 10 min (following the procedures described for the first group): (i) the first two subgroups (60 min after stopping the phenylephrine infusion) received, consecutively, cumulative 10 min i.v. infusions of, respectively, sumatriptan (1–300 μg kg−1) and donitriptan (0.1–30 μg kg−1); and (ii) the remaining three subgroups (during phenylephrine infusion) received, consecutively, cumulative 10-min i.v. infusions of, respectively, PNU-142633 (1–300 μg kg−1), PNU-109291 (0.3–100 μg kg−1) and equivalent volumes of physiological saline (0.5 ml min−1 during 10 min; given 6 times). Then, the responses to the above doses of capsaicin, α-CGRP and acetylcholine were reanalysed. It is important to note that, with these procedures, the carotid vasoconstriction was similar in all subgroups before the 1 min intracarotid infusions of capsaicin, α-CGRP and acetylcholine. [1]
Finally, the fourth group (n=11) received an intracarotid continuous infusion of phenylephrine (1.5 μg min−1) and, 20 min later, the responses to the above doses of capsaicin were elicited as described above during the infusion of phenylephrine. At this point, this group was subdivided into three subgroups that received i.v. bolus injections of, respectively, SB224289 (300 μg kg−1; n=4), BRL15572 (300 μg kg−1; n=4) and an equivalent volume of physiological saline (0.15 ml kg−1; n=3). After 10 min, each subgroup received, consecutively, cumulative 10-min i.v. infusions of donitriptan (0.1–30 μg kg−1) as described previously. It is important to note that after the administration of SB224289 the donitriptan-induced vasoconstriction was completely blocked; therefore, in order to maintain the carotid circulation under a vasoconstriction state similar to that observed previously with the administration of this antagonist, the infusion of phenylephrine was maintained at a constant rate (1.5 μg min−1) throughout the experiments in this subgroup. In contrast, as BRL15572 or physiological saline did not modify the donitriptan-induced carotid vasoconstriction, the infusion of phenylephrine was interrupted just before the administration of these compounds (results obtained from preliminary experiments; not shown). Ten minutes after the last i.v. dose of donitriptan (30 μg kg−1) had been given, the responses to the above 1-min intracarotid infusions of capsaicin were elicited again.

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For in vivo animal studies, Donitriptan hydrochloride can be administered to rodents via oral gavage or intraperitoneal injection. In models of migraine, its effects on cranial blood flow and pain responses are assessed. In models of neurovascular function, the compound's effects on vasoconstriction are evaluated. Dosing regimens vary depending on the specific model and desired exposure levels. Blood and tissue samples may be collected for pharmacokinetic analysis.

Donitriptan hydrochloride has a molecular weight of 383.87 and a molecular formula of C20H22ClN5O. It is a brain-penetrant compound. The compound should be stored at -20°C for long-term stability. It is soluble in DMSO and can be formulated for both in vitro and in vivo administration. It is for research use only and is not intended for human consumption.

The toxicity profile of Donitriptan hydrochloride has not been extensively reported. As a 5-HT1B/1D receptor agonist, potential adverse effects may include those associated with vasoconstriction, such as cardiovascular effects. The compound is for research use only and is not intended for human consumption. Standard toxicological evaluation would include acute and repeated-dose toxicity studies.

[1]. Donitriptan, but not sumatriptan, inhibits capsaicin-induced canine external carotid vasodilatation via 5-HT1B rather than 5-HT1D receptors. Br J Pharmacol. 2006 Sep;149(1):82-91.

[2]. Donitriptan decreases jugular venous oxygen saturation in rats in the absence of cranial vasoconstriction: an overlooked mechanism of antimigraine action? J Pharmacol Exp Ther. 2005 Nov;315(2):849-57.

[3]. Donitriptan selectively decreases jugular venous oxygen saturation in the anesthetized pig: further insights into its mechanism of action relevant to headache relief. J Pharmacol Exp Ther . 2003 May;305(2):749-54.

Background and Objectives: Studies have shown that during migraine attacks, the capsaicin-sensitive trigeminal sensory nerve releases calcitonin gene-related peptide (CGRP), leading to intracranial vasodilation and central nociceptive responses; therefore, inhibition of the trigeminal nerve may prevent this vasodilation and terminate migraines. This study aimed to investigate the effects of sumatriptan (a 5-HT(1B/1D) water-soluble agonist), donitriptan (a 5-HT(1B/1D) lipid-soluble agonist), PNU-142633 (a 5-HT(1D) water-soluble agonist), and PNU-109291 (a 5-HT(1D) lipid-soluble agonist) on the external carotid artery's vasodilatory response to capsaicin, α-CGRP, and acetylcholine in dogs. Methods: Fifty-nine vagotomized dogs were anesthetized with sodium pentobarbital. Blood pressure and heart rate were recorded using pressure sensors connected to a cannula inserted into the femoral artery. A pre-calibrated flow probe was placed around the common carotid artery, ligated branches of the internal carotid and occipital arteries, and connected to an ultrasonic flow meter. Thyroid artery cannulation was used for infusion of agonists. [1]

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In summary, the above results suggest that the inhibitory effect of donitriptan on capsaicin-induced external carotid artery vasodilation is mainly mediated by 5-HT1B receptors, possibly through a central mechanism. [1]

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Donitriptan hydrochloride (F-11356, CAS 170911-68-9) is a brain-penetrant 5-HT1B/1D receptor agonist. It exhibits pKi values of 9.4 for 5-HT1B and 9.3 for 5-HT1D receptors. It inhibits forskolin-induced cAMP formation in C6 cells expressing the human receptors. It is a novel 5-HT derivative with potent, selective, and high intrinsic activity at 5-HT1B/1D receptors and could be used as an antimigraine agent. It is available for research purposes only.

C23H26CLN5O2

439.94

439.178

C, 62.79; H, 5.96; Cl, 8.06; N, 15.92; O, 7.27

170911-68-9

Donitriptan;170912-52-4

197705

Typically exists as solid at room temperature

727.1ºC at 760 mmHg

393.6ºC

5.34E-21mmHg at 25°C

3.773

3

5

6

31

618

0

C1=C(C=CC(=C1)N2CCN(CC2)C(=O)COC3=CC4=C(C=C3)NC=C4CCN)C#N.Cl

ZXENQGQAPOYDOJ-UHFFFAOYSA-N

InChI=1S/C23H25N5O2.ClH/c24-8-7-18-15-26-22-6-5-20(13-21(18)22)30-16-23(29)28-11-9-27(10-12-28)19-3-1-17(14-25)2-4-19;/h1-6,13,15,26H,7-12,16,24H2;1H

4-[4-[2-[[3-(2-aminoethyl)-1H-indol-5-yl]oxy]acetyl]piperazin-1-yl]benzonitrile;hydrochloride

Donitriptan hydrochloride; 170911-68-9; Donitriptan monohydrochloride; 4-[4-[2-[[3-(2-aminoethyl)-1H-indol-5-yl]oxy]acetyl]piperazin-1-yl]benzonitrile hydrochloride; F 11356;

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.)