mGlu4 Receptor ( EC50 = 7500 nM )
Lu AF21934 is a selective positive allosteric modulator (PAM) of the metabotropic glutamate receptor 4 (mGlu4). The human mGlu4 receptor PAM EC50 is 500 nM, with 120% modulation and a 5-fold glutamate shift. [3]
It also shows activity as a PAM at the mGlu6 receptor with an EC50 of 7 µM. [3]
In a broad cross-reactivity panel, it showed antagonist activity at the adenosine A2A receptor (Ki = 7 µM) and the serotonin 5-HT2B receptor (Ki = 2 µM). [3]

In electrophysiological studies on mouse frontal cortex slices, application of DOI (10 µM) increased the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) in layer V pyramidal neurons. Lu AF21934 (1 µM), when applied concurrently with DOI, reversibly suppressed this DOI-induced increase in sEPSC frequency, but did not affect the mean amplitude, suggesting a presynaptic site of action. The effect was U-shaped with the highest decrease observed at 1 µM. [3]

Lu AF21934 treatment shows a dose-dependent anxiolytic-like effect in the stress-induced hyperthermia, four-plate, and marble-burying tests. Flumazenil (10 mg/kg), a benzodiazepine receptor antagonist, inhibits Lu AF21934's (5 mg/kg) anti-hyperthermic effect in the SIH test. This effect is not serotonin dependent. In the tail suspension test, Lu AF21934 does not have antidepressant-like effects on mice; however, it does reduce the basal locomotor activity of mice that have not become accustomed to activity cages[1]. While Lu AF21934 (0.5–5 mg/kg sc) does not affect tremor, harmaline-induced hyperactivity is reversed at doses of 0.5 and 2.5 mg/kg. At a dose of 2.5 mg/kg, Lu AF21934 amplifies the first 30 minutes of the measurement's inhibitory effect of harmaline on AP1 and exploratory activity, and it offsets the harmaline-induced rise in basic activity over the next 30 to 90 minutes[2]. Hyperactivity brought on by MK-801 or amphetamine is dose-dependently inhibited by Lu AF21934 (0.1–5 mg/kg). It also makes head twitches more aggressive and raises the frequency of DOI-induced spontaneous excitatory postsynaptic currents in brain slices[3].

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Subcutaneous administration of Lu AF21934 (0.5–2 mg/kg) reversed hyperactivity induced by MK-801 (0.3 mg/kg, i.p.) in Albino Swiss mice. Lower (0.1 mg/kg) and higher doses (5, 10 mg/kg) were not effective. [3]
It reversed hyperactivity induced by amphetamine (3 mg/kg, i.p.) at doses of 0.1, 0.5, and 2 mg/kg (s.c.), but not at 5 and 10 mg/kg. [3]
It decreased the number of head twitches induced by DOI (2.5 mg/kg, i.p.) in mice at doses of 2, 5, and 10 mg/kg (s.c.) by 45%, 38.5%, and 35% respectively, but not at 0.5 mg/kg. This effect was absent in mGlu4 receptor knockout (mGlu4-/-) mice, confirming target specificity. [3]
At 0.5 mg/kg (s.c.), it reversed MK-801 (0.1 mg/kg, s.c.)-induced deficits in the social interaction test in rats, increasing both the number and duration of social episodes. Doses of 0.2 and 1 mg/kg were not effective. [3]
At 1 and 2 mg/kg (s.c.), it reversed MK-801 (0.1 mg/kg, s.c.)-induced deficits in the spatial delayed alternation task in rats, a model of cognitive disturbance. [3]
When administered alone at behaviorally active doses (0.5–5 mg/kg, s.c.), it did not significantly affect the locomotor activity of mice habituated to activity cages. [3]

Rats: The 20% (2-hydropropyl)-b-cyclodextrin-dispersed Lu AF21934 is applied subcutaneously (s.c.) 60 minutes prior to the test. Acute administration of Lu AF21934 (2, 5, 10 and 15 mg/kg, s.c.) and diazepam (5 mg/kg, i.p.) occurs one hour prior to the Vogel's conflict test. In all experiments, groups of eight to ten animals are used to measure the effects of each drug[1].
Mice: Sixty minutes prior to the test, Lu AF21934 is applied subcutaneously (s.c.) after being diluted in 20% (2-hydropropyl)-b-cyclodextrin. The mice are inserted carefully into the box and given fifteen seconds to explore. Once the mouse has moved from one plate to another, the experimenter electrifies the entire floor, causing the mouse to visibly flee. No more shocks are given for the next three seconds if the animal keeps running[1].

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MK-801/Amphetamine-induced Hyperactivity (Mice): Male Albino Swiss mice were placed individually in locomotor activity cages for a 30-minute acclimatization. They were then injected subcutaneously (s.c.) with Lu AF21934 or vehicle (20% (2-hydroxypropyl)-β-cyclodextrin, HPBCD). After 60 minutes, they received an intraperitoneal (i.p.) injection of MK-801 (0.3 mg/kg) or amphetamine (3 mg/kg) and were returned to the cages. Ambulation scores were recorded for 80 minutes. [3]
DOI-induced Head Twitch (Mice): Mice were habituated in a glass cage for 30 minutes. Lu AF21934 or vehicle (HPBCD) was administered s.c. 60 minutes before an i.p. injection of DOI (2.5 mg/kg). Immediately after DOI injection, head twitches were counted over a 20-minute session. [3]
Social Interaction Test (Rats): Male Wistar rats, pair-housed from separate cages, received an s.c. injection of MK-801 (0.1 mg/kg). Lu AF21934 or vehicle (HPBCD) was administered s.c. 60 minutes before the test, which occurred 3.5 hours after MK-801. Social interaction (sniffing, chasing, etc.) between two rats in a novel arena was video-recorded and analyzed for total time and number of episodes. [3]
Spatial Delayed Alternation Task (Rats): Water-deprived male Wistar rats were trained in a T-maze to alternate arms for a sucrose reward. On test days, rats received an s.c. injection of Lu AF21934 or vehicle (HPBCD) 60 minutes before the test session. MK-801 (0.1 mg/kg, s.c.) was administered to induce deficits. The number of correct alternations in 10 trials was recorded. [3]
Electrophysiology (Mice): Albino Swiss mice were decapitated, frontal cortices were dissected and sliced. Whole-cell voltage-clamp recordings were made from layer V pyramidal neurons. After baseline recording, DOI (10 µM) was applied via superfusate for 10 minutes to increase sEPSC frequency, followed by co-application of DOI and Lu AF21934 for 15 minutes. sEPSC frequency and amplitude were analyzed. [3]

One hour after subcutaneous injection of a single dose of 10 mg/kg (dissolved in 20% HPBCD aqueous solution) into rats, the brain tissue concentration was measured to be 2422 ng/g, the plasma concentration to be 2763 ng/mL, and the brain/plasma concentration ratio to be 0.9. The cerebrospinal fluid (CSF) concentration was 95 ng/mL. [3] The rat plasma protein binding rate (rPPB) was 95.6%, and the rat brain tissue free fraction (f_{u, brain}) was 3.0%. [3] The rat intrinsic clearance rate (CL_int) was 13 mL/min. [3]

[1]. Anxiolytic- but not antidepressant-like activity of Lu AF21934, a novel, selective positive allosteric modulator of the mGlu? receptor. Neuropharmacology. 2013 Mar;66:225-35.

[2]. Lu AF21934, a positive allosteric modulator of mGlu4 receptors, reduces the harmaline-induced hyperactivity but not tremor in rats. Neuropharmacology. 2014 Aug;83:28-35.

[3]. The antipsychotic-like effects of positive allosteric modulators of metabotropic glutamate mGlu4 receptors in rodents. Br J Pharmacol. 2013 Aug;169(8):1824-39.

Lu AF21934 is a chemically unique, highly selective, and blood-brain barrier-crossing allosteric regulator of the mGlu4 receptor. [3]
Its enantiomer, Lu AF21935, is inactive against the mGlu4 receptor (PAM EC50 >10,000 nM) and showed no activity in the MK-801-induced ADHD test, suggesting that the in vivo effects of Lu AF21934 are mediated by enhancing mGlu4 receptor activity. [3]
This study proposes that its antipsychotic-like effects may be mediated by presynaptic mGlu4 receptors located at glutamatergic nerve endings, the activation of which can inhibit excessive glutamate release caused by NMDA receptor blockade or 5-HT2A receptor activation. [3]

C14H16CL2N2O2

315.1950

314.058

C, 53.35; H, 5.12; Cl, 22.49; N, 8.89; O, 10.15

1445605-23-1

66553157

White to off-white solid powder

1.4±0.1 g/cm3

577.6±50.0 °C at 760 mmHg

303.1±30.1 °C

0.0±1.6 mmHg at 25°C

1.615

2.62

2

2

3

20

378

2

ClC1=C(C([H])=C([H])C(=C1[H])N([H])C([C@@]1([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[C@@]1([H])C(N([H])[H])=O)=O)Cl

AVKZWCJNDWOBAM-ZJUUUORDSA-N

InChI=1S/C14H16Cl2N2O2/c15-11-6-5-8(7-12(11)16)18-14(20)10-4-2-1-3-9(10)13(17)19/h5-7,9-10H,1-4H2,(H2,17,19)(H,18,20)/t9-,10+/m1/s1

(1R,2S)-2-N-(3,4-dichlorophenyl)cyclohexane-1,2-dicarboxamide

Lu-AF21934; Lu-AF-21934; Lu-AF 21934; Lu AF21934; Lu AF-21934; Lu AF 21934; LuAF21934; LuAF-21934; LuAF 21934

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)

DMSO: ≥ 80 mg/mL (~253.8 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (7.93 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (7.93 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
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.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (7.93 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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