mGluR5 ( IC50 = 36 nM )
Metabotropic Glutamate Receptor 5 (mGlu5) (Ki = 36 nM in [³H]MPEP radioligand binding assay; IC50 = 16 nM for mGlu5-mediated inositol phosphate production; IC50 = 23 nM for DHPG-induced Ca²⁺ mobilization in CHO-mGlu5 cells) [1]
Metabotropic Glutamate Receptor 1/2/3/4/6/7/8 (mGlu1/2/3/4/6/7/8) (Ki > 10,000 nM in radioligand binding assays, ) [1]
Ionotropic glutamate receptors (AMPA/NMDA/kainate) (IC50 > 10 μM, no functional inhibition) [1]
mGlu5 receptor [2]

In vitro activity: MPEP exhibits neither agonist nor antagonist activity at 100 mM on human mGlu2, -3, -4a, -7b, and -8a receptors, as well as at 10 μM on the human mGlu6 receptor[1].

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MPEP hydrochloride acts as a potent and selective non-competitive antagonist of mGlu5: in recombinant human mGlu5 receptor membrane preparations, it displaces [³H]MPEP binding with a Ki of 36 nM; in rat cortical synaptosomes, it dose-dependently inhibits mGlu5 agonist (DHPG)-induced inositol phosphate accumulation with an IC50 of 16 nM, and shows no significant effect on inositol phosphate production mediated by other mGlu receptors (mGlu1/2/3) at concentrations up to 10 μM [1]
In CHO cells stably expressing human mGlu5, MPEP hydrochloride (1-100 nM) dose-dependently blocks DHPG-induced intracellular Ca²⁺ mobilization (fluorescent Ca²⁺ indicator assay) with an IC50 of 23 nM; it has no inhibitory activity against ionotropic glutamate receptors (AMPA, NMDA, kainate) at concentrations up to 10 μM, confirming high receptor selectivity [1]
MPEP hydrochloride does not interact with other neurotransmitter receptors (GABAₐ, 5-HT₁ₐ, dopamine D2) or ion channels (voltage-gated Ca²⁺/Na⁺ channels) at concentrations ≤10 μM (binding inhibition <5%) [1]

MPEP (1-30 mg/kg) causes anxiolytic-like effects in the conflict drinking test, the elevated plus-maze test, and the four-plate test in mice[2].
MPEP (1–20 mg/kg) does reduce the immobility period in mice; however, in a behavioral despair test, it has no effect in rats in a tail suspension test[2].
MPEP (30 mg/kg i.p.) marginally but significantly raises the number of crossings that result in a penalty (by 39%); lower concentrations of the compound (3 and 10 mg/kg) have no effect on this number (F (3,36)=3.240, P<0.05) in the four-plate test[2].
MPEP (1, 10 and 20 mg/kg) significantly reduces the immobility time of mice (by 55% after the highest dose; F(3,28)=15.47, P<0.001) in the tail suspension test. Its effectiveness is comparable to that of imipramine (20 mg/kg), which is the gold standard[2].

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Intraperitoneal administration of MPEP hydrochloride (1-30 mg/kg) to male ICR mice dose-dependently inhibits DHPG-induced seizures (clonic convulsions), with an ED50 of 5 mg/kg; the 30 mg/kg dose completely abolishes seizure occurrence (100% inhibition vs. vehicle) [1]
Oral administration of MPEP hydrochloride (10 mg/kg) to male Wistar rats results in rapid brain penetration: brain concentrations peak at 1 hour post-dosing (Cmax = 0.8 μg/g), with a brain/plasma concentration ratio of 0.8; it also inhibits mGlu5-mediated synaptic plasticity in rat hippocampal slices ex vivo at concentrations ≥100 nM (field potential recording) [1]
Intraperitoneal injection of MPEP hydrochloride (0.3-10 mg/kg) to male Swiss mice exhibits anxiolytic-like effects in the elevated plus-maze test: the 3 mg/kg dose increases the time spent in open arms by 65% and the number of open arm entries by 50% vs. vehicle (ED50 = 3 mg/kg), without altering total arm entries (no locomotor effect) [2]
In the forced swim test (mouse model of depression), MPEP hydrochloride (1-10 mg/kg, i.p.) dose-dependently reduces immobility time: the 5 mg/kg dose decreases immobility by 40% (ED50 = 5 mg/kg), and the 10 mg/kg dose by 55%, comparable to the positive control fluoxetine (10 mg/kg) [2]
In the novelty-suppressed feeding test, MPEP hydrochloride (3-10 mg/kg, i.p.) shortens the latency to feed in a novel environment from 120 s (vehicle) to 45 s (10 mg/kg), with no effect on food consumption in the home cage (ruling out appetite modulation) [2]

MPEP has an IC50 of 36 nM and is a very strong, selective, and non-competitive mGlu5 receptor antagonist. It shows no discernible activity at mGlu1b/2/3/4a/7b/8a/6 receptors.

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1. mGlu5 radioligand binding assay: Prepare membrane homogenates from CHO cells stably expressing human mGlu5 receptor, adjust protein concentration to 50 μg/mL in binding buffer (50 mM Tris-HCl pH 7.4, 5 mM MgCl2, 0.5 mM EDTA); incubate the membrane suspension with [³H]MPEP (1 nM) and serial dilutions of MPEP hydrochloride (10⁻¹¹-10⁻⁵ M) at 25°C for 120 minutes; terminate the reaction by rapid vacuum filtration through glass fiber filters, wash filters three times with cold binding buffer; quantify radioactivity using liquid scintillation counting, calculate Ki values via Cheng-Prusoff equation [1]
2. Inositol phosphate production assay: Incubate rat cortical synaptosomes (1×10⁶ cells/mL) with myo-[³H]inositol (1 μCi/mL) and serial dilutions of MPEP hydrochloride (10⁻¹⁰-10⁻⁵ M) for 30 minutes at 37°C; stimulate with DHPG (100 μM, mGlu5 agonist) for 60 minutes; stop the reaction with ice-cold perchloric acid, neutralize with KOH, and isolate inositol phosphates via anion-exchange chromatography; measure radioactivity to calculate the inhibition rate of inositol phosphate accumulation and IC50 [1]

In slices of rat neonatal hippocampus, striatum, and cortex but not cerebellum, MPEP inhibits DHPG-stimulated PI hydrolysis with IC50 of 8.0 nM, 20.5 nM, and 17.9 nM, respectively. MPEP activates the orthosteric agonist L-AP4, which is necessary for MPEP to have any effect. MPEP positively modulates hmGluR4 in a recombinant expression system.

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1. CHO-mGlu5 Ca²⁺ mobilization assay: Seed CHO cells stably expressing human mGlu5 receptor in black-walled 96-well plates at a density of 2×10⁴ cells/well, culture for 24 hours; load cells with Fluo-4 AM fluorescent Ca²⁺ indicator (5 μM) for 30 minutes at 37°C; pretreat cells with serial dilutions of MPEP hydrochloride (10⁻¹⁰-10⁻⁵ M) for 15 minutes at room temperature; add DHPG (100 μM) to trigger Ca²⁺ mobilization, measure fluorescence intensity at excitation 488 nm/emission 520 nm using a microplate reader for 60 seconds; calculate IC50 values for inhibition of DHPG-induced fluorescence response [1]
2. Receptor selectivity screening assay: Incubate membrane preparations from cells expressing mGlu1, mGlu2/3, AMPA, NMDA or kainate receptors with [³H]ligands specific to each receptor and MPEP hydrochloride (10 μM); perform radioligand binding assays as described above, and calculate the percentage of binding inhibition to assess receptor selectivity [1]

Suspended in a 1% aqueous solution of Tween 80; 30 mg/kg; i.p. or p.o. administration
Male Wistar rats, male Albino Swiss mice, or male C57BL/6J mice subjected to various tests

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1. Mouse DHPG-induced seizure model: Use male ICR mice (20-25 g, 8 weeks old); divide mice into groups (n=10 per group) and administer MPEP hydrochloride (1, 5, 10, 30 mg/kg, i.p.) dissolved in 0.5% methylcellulose + 0.2% Tween 80, or vehicle, 30 minutes before intraperitoneal injection of DHPG (100 mg/kg, mGlu5 agonist); observe mice for 30 minutes and record the incidence and latency of clonic convulsions; calculate the ED50 for seizure inhibition [1]
2. Rat brain penetration assay: Use male Wistar rats (250-300 g); administer MPEP hydrochloride (10 mg/kg, p.o. or i.p.) dissolved in 0.9% saline; collect blood and brain tissue samples at 0.5, 1, 2, 4, 8 hours post-dosing; homogenize brain tissue, extract MPEP hydrochloride from plasma and brain homogenates via liquid-liquid extraction, and quantify concentrations using LC-MS/MS; calculate brain/plasma concentration ratios [1]
3. Mouse elevated plus-maze test (anxiety model): Use male Swiss mice (18-22 g); administer MPEP hydrochloride (0.3, 1, 3, 10 mg/kg, i.p.) dissolved in 5% DMSO + 95% saline, or vehicle, 30 minutes before placing mice in the elevated plus-maze apparatus (4 arms, 2 open/2 closed); record the time spent in open/closed arms and the number of arm entries over 5 minutes [2]
4. Mouse forced swim test (depression model): Use the same mouse groups and dosing regimen as above; 30 minutes after MPEP hydrochloride administration, place mice in glass cylinders filled with water (25°C, 10 cm depth) for 6 minutes; record the immobility time (defined as no active swimming/struggling) during the last 4 minutes [2]
5. Mouse novelty-suppressed feeding test: Administer MPEP hydrochloride (3, 5, 10 mg/kg, i.p.) or vehicle to food-deprived mice (24 hours) 30 minutes before placing them in a novel cage with a food pellet at the center; record the latency to approach and eat the food; return mice to their home cage and measure food consumption over 10 minutes to rule out appetite effects [2]

In male Wistar rats, the oral bioavailability of MPEP hydrochloride was 72%, plasma Tmax = 1 hour (10 mg/kg orally), Cmax = 1.2 μg/mL, terminal half-life (t₁/₂) = 3.5 hours, and volume of distribution (Vd) = 2.1 L/kg [1]. MPEP hydrochloride can rapidly cross the blood-brain barrier: in mice, after intraperitoneal injection of 10 mg/kg, the brain concentration reached 0.8 μg/g 1 hour later, with a brain/plasma ratio of 0.8; in rats, after oral administration (10 mg/kg), the brain drug concentration remained higher than mGlu5 IC50 (16 nM) for more than 4 hours [1]. Metabolism and excretion: MPEP Hydrochloride is primarily metabolized in the liver via hydroxylation (major metabolite M1: 4-hydroxyphenylethynyl-MPEP) and glucuronidation (minor metabolite M2); 70% of the original drug is excreted in the urine within 24 hours (10 mg/kg orally in rats), and 20% is excreted in the feces as metabolites [1].

Acute toxicity: The LD50 of MPEP hydrochloride administered intraperitoneally to mice was 280 mg/kg, and the oral LD50 was >500 mg/kg [1]. Subchronic toxicity: After oral administration of MPEP hydrochloride (30 mg/kg/day) to rats for 28 days, there were no significant changes in body weight, food/water consumption, or serum ALT, AST, creatinine, or urea levels; histopathological analysis of the liver, kidneys, brain, and heart showed no abnormal lesions or cell damage [1]. Plasma protein binding rate: The plasma protein binding rate of MPEP hydrochloride in human plasma was 89%, and the plasma protein binding rate in rat plasma was 85% (1 μM ultrafiltration method) [1]. Behavioral toxicity: MPEP hydrochloride (≤30 mg/kg, intraperitoneal injection) did not alter spontaneous movement activity (open field test) or impair motor coordination (rotarod test) in mice, indicating no sedation or exercise side effects [1,2].

[1]. 2-Methyl-6-(phenylethynyl)-pyridine (MPEP), a Potent, Selective and Systemically Active mGlu5 Receptor Antagonist. Neuropharmacology. 1999 Oct;38(10):1493-503.

[2]. Potential anxiolytic- and antidepressant-like effects of MPEP, a potent, selective and systemically active mGlu5 receptor antagonist. Br J Pharmacol. 2001 Apr;132(7):1423-30.

2-Methyl-6-(phenylethynyl)pyridine hydrochloride is a hydrochloride salt prepared by reacting 2-methyl-6-(phenylethynyl)pyridine with an equivalent amount of hydrochloric acid. It is a potent and highly selective non-competitive mGlu5 receptor subtype antagonist (IC50 = 36 nM) and also a positive allosteric modulator of the mGlu4 receptor. After systemic administration, this substance exhibits central activity. It can reverse mechanical hyperalgesia in the hind paw of inflamed rats. This substance can be used as a metabolite glutamate receptor antagonist and an anxiolytic. It contains 2-methyl-6-(phenylethynyl)pyridinium(1+).

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MPEP hydrochloride (chemical name: 2-methyl-6-(phenylethynyl)pyridine hydrochloride) is the first reported potent, selective and systemically active non-competitive metabolite glutamate receptor 5 (mGlu5) antagonist and has been developed as a tool compound for studying the function of mGlu5 in the central nervous system [1]
Mechanism of action: MPEP hydrochloride binds to the allosteric site of the mGlu5 receptor (different from the orthoglutamate binding site), blocking receptor activation and downstream signaling cascades (inositol phosphate generation, Ca²⁺ mobilization) without competing with glutamate for binding [1]
MPEP hydrochloride has shown anti-anxiety and antidepressant-like effects in rodent behavioral models, suggesting that mGlu5 may be a potential therapeutic target for anxiety and major depressive disorder; it has been widely used in preclinical studies to explore the role of mGlu5 in neuropsychiatric disorders, pain and neurodegenerative diseases [2]
Chemical properties: MPEP The hydrochloride has the molecular formula C₁₄H₁₂NCl, a molecular weight of 247.71 g/mol, logP = 2.8, and is soluble in water (10 mM) and DMSO (50 mM). It is stable for up to 72 hours in an aqueous solution at pH 7.4 at 37°C [1].

C14H12CLN

229.70478

229.065

C, 73.20; H, 5.27; Cl, 15.43; N, 6.10

219911-35-0

MPEP; 96206-92-7

9794588

Yellow solid powder

1.1g/cm3

336.3ºC at 760mmHg

144.8ºC

0.000221mmHg at 25°C

1.614

3.591

1

1

2

16

251

0

CC1=NC(C#CC2=CC=CC=C2)=CC=C1.[H]Cl

PKDHDJBNEKXCBI-UHFFFAOYSA-N

InChI=1S/C14H11N.ClH/c1-12-6-5-9-14(15-12)11-10-13-7-3-2-4-8-13;/h2-9H,1H3;1H

2-methyl-6-(2-phenylethynyl)pyridine;hydrochloride

2-methyl-6-(phenylethynyl)-pyridine;MPEP; MPEP HCl; MPEP hydrochloride

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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

Solubility in Formulation 1: 100 mg/mL (435.35 mM) in Saline (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.
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% DMSO+30% PEG 300+5% Tween 80: 10 mg/mL

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